Patent Publication Number: US-2022226396-A1

Title: Antibacterial agents &amp; methods

Description:
TECHNICAL FIELD 
     The invention relates to means for carrying out conjugation between bacteria, and in particular the invention relates to carrier bacteria comprising antimicrobial agents and methods of use. A carrier bacterium is capable of conjugative transfer of DNA encoding the agent to a target cell. 
     The invention further relates to growth or feed conversion ratio (FCR) promotion in animals. The invention further relates to killing  Salmonella  or inhibiting the growth or proliferation of  Salmonella. The invention further relates to killing Pseudomonas  or inhibiting the growth or proliferation of  Pseudomonas , such as useful for promoting growth, dry weight, wet weight or crop production of plants. 
     BACKGROUND 
     DNA sequences controlling extra-chromosomal replication (ori) and transfer (tra) are distinct from one another; i.e., a replication sequence generally does not control plasmid transfer, or vice-versa. Replication and transfer are both complex molecular processes that make use of both plasmid- and host-encoded functions. Bacterial conjugation is the unidirectional and horizontal transmission of genetic information from one bacterium to another. The genetic material transferred may be a plasmid or it may be part of a chromosome. Bacterial cells possessing a conjugative plasmid contain a surface structure (the sex pilus) that is involved in the coupling of donor and recipient cells, and the transfer of the genetic information. Conjugation involves contact between cells, and the transfer of genetic traits can be mediated by many plasmids. Among all natural transfer mechanisms, conjugation is the most efficient. For example, F plasmid of  E. coli , pCFlO plasmid of  Enterococcus faecalis  and pXO16 plasmid of  Bacillus thuringiensis  employ different mechanisms for the establishment of mating pairs, the sizes of mating aggregates are different, and they have different host ranges within gram-negative (F) as well as gram-positive (pCFlO and pXO16) bacteria. Their plasmid sizes are also different; 54, 100 and 200 kb, respectively. Remarkably, however, those conjugation systems have very important characteristics in common: they are able to sustain conjugative transfer in liquid medium and transfer efficiencies close to 100% are often reached in a very short time. Thus, the conjugative process permits the protection of plasmid DNA against environmental nucleases, and the very efficient delivery of plasmid DNA into a recipient cell. Conjugation functions are naturally plasmid encoded. Numerous conjugative plasmids (and transposons) are known, which can transfer associated genes within one species (narrow host range) or between many species (broad host range). Transmissible plasmids have been reported in numerous Gram-positive genera, including but not limited to pathogenic strains of  Streptococcus, Staphylococcus, Bacillus, Clostridium  and  Nocardia . The early stages of conjugation generally differ in Gram-negative and Gram-positive bacteria. The role of some of the transfer genes in conjugative plasmids from Gram-negative bacteria are to provide pilus-mediated cell-to-cell contact, formation of a conjugation pore and related morphological functions. The pili do not appear to be involved in initiating conjugation in Gram-positive bacteria. 
     SUMMARY OF THE INVENTION 
     The invention provides: 
     A method (eg, a non-medical or a medical) for enhancing the growth or weight of a subject, wherein the subject comprises bacterial target cells, the method comprising administering to the subject a first episomal DNA encoding an antibacterial agent that is toxic to the target cells, wherein first DNA is transferred into target cells and the agent is expressed, thereby killing target cells in the subject or reducing the growth or proliferation of target cells and enhancing growth or weight of the subject. 
     In an embodiment, there is provided: 
     A method (eg, a non-medical or a medical) for enhancing the growth or weight of a subject, wherein the method comprises the administration of a plurality of carrier cells to the subject, wherein the subject comprises bacterial target cells and each carrier cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the agent, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells and enhancing growth or weight of the subject. 
     Advantageously, the FCR is improved (ie, enhanced), ie, the FCR number is lowered, in the subject. When the method of the invention is carried out on a group of subjects (eg, a group of animals, such as livestock animals) the FCR is lowered in an individual in the group or the average FCR is lowered in the group. Lowering may be assessed as a comparison with FCR prior to administration of the carrier cell(s) or compared to an average for animals of the same species, age group and when fed on comparable or the same diet. The skilled addressee will be familiar with standard FCRs, such as for livestock animals, eg, piglets, pigs, sheep, cattle (dairy or meat cattle), fish, shellfish, poultry (eg, chickens (broiler or egg-layer hens), geese, ducks or turkeys). 
     The invention also provides: 
     A carrier cell, wherein the cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a bacterial target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the antibacterial agent, hereby killing the target cell, wherein the target cell is a  Salmonella  cell and the carrier cell is an Enterobacteriaceae cell. 
     A composition comprising a plurality of carrier cells for use in a method comprising administration of the cells to a subject to treat an infection by pathogenic bacterial target cells, wherein each carrier cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the agent, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells, wherein the target cells are  Salmonella  cells and the carrier cells are Enterobacteriaceae cells. 
     A non-medical method of killing zoonotic bacterial target cells in an animal (optionally a livestock animal), the method comprising administering to the animal a plurality of carrier cells, wherein each carrier cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the agent, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells, wherein the target cells are  Salmonella  cells and optionally the carrier cells are Enterobacteriaceae cells. 
     A DNA (optionally for use in the method of the invention), wherein the DNA is capable of being introduced into a target cell, wherein the DNA encodes a plurality of guide RNAs or crRNAs of a CRISPR/Cas system wherein the guide RNAs or crRNAs are operable with Cas nuclease in the target cell to recognise a plurality of protospacer sequences comprised by the target cell genome; wherein
         (a) the protospacer sequences comprise one or more pathogenic island nucleotide sequences of the target cell genome;   (b) the protospacer sequences comprise one or more invasion gene sequences of the target cell genome;   (c) the protospacer sequences comprise one or more secretion system gene sequences of the target cell genome; and/or   (d) the protospacer sequences comprise one or more nucleotide sequences of genes selected from A gene selected from avrA, sptP, sicP, sipA, sipD, sipC, sipB, sicA, invB, ssaE, sseA, sseB, sscA, sseC, sseD, sseE, sscB, sseF, sseG, mgtC, cigR, pipA, pipB, pipC, sopB and pipD (optionally selected from invB, sicP, sseE, pipA, pipB, pipC, hilA, marT and sopB of  Salmonella , and orthologues or homologues thereof.       

     The also invention provides the following configurations:— 
     In a First Configuration 
     A carrier bacterial cell comprising a first episomal DNA, the DNA encoding a nucleic acid sequence of interest (NSI) or encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into the target cell for expression therein of the agent, optionally wherein
         (a) the carrier cell comprises a second DNA which is different from the first DNA, wherein the second DNA comprises or encodes a first factor required for replication of the first DNA;   (b) the first DNA does not comprise or encode said first factor, wherein the first DNA is non-self-replicative in the absence of the first factor, but is able to replicate in the carrier cell in the presence of the first factor provided by the second DNA;   (c) wherein the cell comprises genes encoding one or more conjugation factors sufficient to carry out conjugative transfer of the first DNA into a target bacterial cell.       

     In embodiments, the invention usefully recognizes the benefit of using antibacterial agents that act by target sequence recognition in the target cell genome but not in the carrier cell, which frees up the ability for the first DNA to be freely replicated in the carrier cell without toxicity to the carrier cell. 
     In a Second Configuration 
     A DNA encoding an NSI or encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic to a carrier cell that is capable of carrying the DNA for conjugative transfer into the target cell, wherein the first DNA does not comprise or encode a first factor required for replication of the first DNA, and wherein the first DNA is devoid of a component required for conjugative transfer of the first DNA into a target bacterial cell. 
     In a Third Configuration 
     A method for enhancing growth or weight of a human or animal subject, wherein the method comprises the administration of a plurality of carrier cells according to the invention to a microbiota of the subject, wherein the microbiota comprises target cells and first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells (eg,  Salmonella  cells) in the subject or reducing the growth or proliferation of target cells. 
     In a Fourth Configuration 
     A method for enhancing growth or weight of a plant, wherein the method comprises the administration of a plurality of carrier cells according to the invention to a microbiota of the plant, wherein the microbiota comprises target cells and first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells (eg,  Pseudomonas  cells) in the plant or reducing the growth or proliferation of target cells. 
     In a Fifth Configuration 
     A method for reducing a biofilm comprised by a subject or comprised on a surface, wherein the biofilm comprises target cells (eg,  Pseudomonas  cells), wherein the method comprises the administration of a plurality of carrier cells according to the invention to the biofilm, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the biofilm or reducing the growth or proliferation of target cells. 
     In a Sixth Configuration 
     A method of replicating a first DNA (eg, comprised by conjugative plasmids) to produce a plurality of copies of said DNA, the method comprising culturing a plurality of carrier bacterial cells according to the invention, wherein the first DNA is replicated in the cells; and optionally isolating a plurality of copies of the first DNA from carrier cells. 
     In a Seventh Configuration 
     A method of killing a plurality of target bacterial cells or reducing the growth or proliferation thereof, the method comprising
         (a) obtaining a sample of the carrier cells according to the invention or obtainable by the method of the sixth configuration;   (b) contacting the sample of carrier cells with the plurality of target bacterial cells to allow conjugation between carrier cells and target cells; and   (c) allowing copies of first DNA to be transferred by conjugative transfer from carrier cells to target cells, wherein the antibacterial agent is provided in target cells and target cells are killed or the growth or proliferation of target cells is reduced;   (d) wherein the first DNA is not (or not substantially) replicable in the target cells.       

     In an Eighth Configuration 
     A pharmaceutical composition, livestock growth promoting composition, zoonosis control agent, biocidal agent for administration to livestock, soil improver, herbicide, plant fertilizer, food or food ingredient sterilizing composition, dental composition, personal hygiene composition or disinfectant composition (eg, for domestic or industrial use) comprising a plurality of carrier cells according to the invention. 
     In an Ninth Configuration 
     In a first aspect:— 
     A method of promoting the growth of an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal a guided nuclease system or a component thereof, and introducing the system or component into target bacteria comprised by the animal, wherein the guided nuclease is capable of recognising and modifying (eg, cutting) a target nucleotide sequence comprised by the target bacteria, whereby target bacteria are killed or the growth or proliferation of target bacteria are inhibited and the growth of the animal is promoted. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of inhibiting the growth of the animal. Thus, the method reduces the burden of such bacteria in the animal and promotes growth. 
     In a second aspect:— 
     A method of enhancing feed conversion ratio (FCR) in an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal a guided nuclease system or a component thereof, and introducing the system or component into target bacteria comprised by the animal, wherein the guided nuclease is capable of recognising and modifying (eg, cutting) a target nucleotide sequence comprised by the target bacteria, whereby target bacteria are killed or the growth or proliferation of target bacteria are inhibited and the FCR of the animal is increased. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of increasing the FCR of the animal. Thus, the method reduces the burden of such bacteria in the animal and enhances FCR (ie, reduces FCR number). 
     In a third aspect:— 
     A method of promoting the growth of an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal an antibacterial agent that is toxic to  Salmonella  bacteria, wherein  Salmonella  target bacteria comprised by the animal are exposed to the agent and are killed or the growth or proliferation of target bacteria are inhibited and the growth of the animal is promoted. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of inhibiting the growth of the animal. Thus, the method reduces the burden of such bacteria in the animal and promotes growth. 
     In a fourth aspect:— 
     A method of enhancing feed conversion ratio (FCR) in an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal an antibacterial agent that is toxic to  Salmonella  bacteria, wherein  Salmonella  target bacteria comprised by the animal are exposed to the agent and are killed or the growth or proliferation of target bacteria are inhibited and the FCR of the animal is enhanced. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of increasing the FCR of the animal. Thus, the method reduces the burden of such bacteria in the animal and enhances FCR (ie, reduces FCR number). 
     In an Tenth Configuration 
     In a first aspect:— 
     A method of promoting growth, dry weight, wet weight or crop production of a plant, the plant comprising a microbiota that comprises target bacteria, the method comprising contacting the microbiota with an antibacterial agent that is toxic to the target bacteria, wherein target bacteia are killed or the growth or proliferation of target bacteria is inhibited and the growth, dry weight, wet weight or crop production of the plant is increased. 
     In a second aspect:— 
     Use of an antibacterial agent that is toxic to target bacteria in a method wherein target bacteria comprised by a microbiota of a plant are contacted with the agent, whereby target bacteria are killed or the growth or proliferation of target bacteria is inhibited, for promoting growth, dry weight, wet weight or crop production of the plant. 
     In a third aspect:— 
     A method of increasing crop yield of a plant, the plant comprising a microbiota that comprises target bacteria, the method comprising contacting the microbiota with an antibacterial agent that is toxic to the target bacteria, wherein target bacteia are killed or the growth or proliferation of target bacteria is inhibited and the growth, dry weight, wet weight or crop production of the plant is increased. 
     In a fourth aspect:— 
     Use of an antibacterial agent that is toxic to target bacteria in a method wherein target bacteria comprised by a microbiota of a plant are contacted with the agent, whereby target bacteria are killed or the growth or proliferation of target bacteria is inhibited, for increasing crop yield of the plant. 
     In a fifth aspect:— 
     A method of increasing leaf chlorophyll of a plant, the plant comprising a microbiota that comprises target bacteria, the method comprising contacting the microbiota with an antibacterial agent that is toxic to the target bacteria, wherein target bacteia are killed or the growth or proliferation of target bacteria is inhibited and the leaf chlorophyll of the plant is increased. 
     In a sixth aspect:— 
     Use of an antibacterial agent that is toxic to target bacteria in a method wherein target bacteria comprised by a microbiota of a plant are contacted with the agent, whereby target bacteria are killed or the growth or proliferation of target bacteria is inhibited, for increasing leaf chlorophyll of the plant. 
     In a seventh aspect:— 
     A method of increasing greening of a plant, the plant comprising a microbiota that comprises target bacteria, the method comprising contacting the microbiota with an antibacterial agent that is toxic to the target bacteria, wherein target bacteia are killed or the growth or proliferation of target bacteria is inhibited and greening of the plant is increased. 
     In a eighth aspect:— 
     Use of an antibacterial agent that is toxic to target bacteria in a method wherein target bacteria comprised by a microbiota of a plant are contacted with the agent, whereby target bacteria are killed or the growth or proliferation of target bacteria is inhibited, for increasing greening of the plant. 
     In a ninth aspect:— 
     A method of decreasing a biofilm (eg, a leaf biofilm) comprised by a plant, the plant comprising a biofilm that comprises target bacteria, the method comprising contacting the microbiota with an antibacterial agent that is toxic to the target bacteria, wherein target bacteia are killed or the growth or proliferation of target bacteria is inhibited and the biofilm of the plant is decreased. 
     In a tenth aspect:— 
     Use of an antibacterial agent that is toxic to target bacteria in a method wherein target bacteria comprised by a biofilm (eg, a leaf biofilm) of a plant are contacted with the agent, whereby target bacteria are killed or the growth or proliferation of target bacteria is inhibited, for inhibiting the biofilm of the plant. 
     Optionally, the target bacteria are  Pseudomonas  bacteria, such as  P. syringae  or  P. aeruginosa  bacteria or any other  Pseudomonas  bacteria disclosed herein. 
     Optionally, the agent is a guided nuclease system or a component thereof, eg, any such system or component disclosed herein for modifying (eg, cutting) a target nucleic acid sequence comprised by target bacteria. 
     Optionally, the plant is any plant disclosed herein. 
     Optionally, the chlorophyll is a chlorophyll a and/or chlorophyll b. 
     Optionally, the agent is comprised by a carrier cell of the present invention and said contacting comprises contacting the cell with the carrier cell. 
    
    
     
       BRIEF DESCRIPTION OF FIGURES 
         FIG. 1 . PCRs performed on recipient colonies. From the left to the right: 1) Plasmid pFSMobC* (control). 2) Plasmid pFSMobC* in S17 (control). 3) Colony from transconjugants plate (Cm30+Nal25). 4) Colony n° 2 from transconjugants plate (Cm30+ Nal25); 
         FIG. 2 : Effect of viability of pFSMobSal7 delivered by conjugation on  Salmonella enteritidis  FS26; 
         FIG. 3 . Numbers of  Salmonella  in the caecum at 7 days post infection. N=15 birds per group; 
         FIG. 4 . Frequency of detection of  Salmonella  in the caecum at 7 days post infection. N=15 birds per group; and 
         FIG. 5 . Post mortem carcass weight of birds at 6 weeks of age. 
         FIG. 6 . Numbers (CFU/g) of  Salmonella  in crop digesta 7 days post-challenge; N=9 birds; P=0.03. 
     
    
    
     DETAILED DESCRIPTION 
     The invention relates to means for carrying out conjugation between bacteria, and in particular the invention relates to carrier bacteria comprising antimicrobial agents and methods of use. A carrier bacterium is capable of conjugative transfer of DNA encoding the agent to a target cell. 
     The invention further relates to growth, weight or feed conversion ratio (FCR) promotion in animals. 
     The invention further relates to killing  Salmonella  or inhibiting the growth or proliferation of  Salmonella.    
     Thus, there is provided: 
     A method (eg, a non-medical or a medical) for enhancing the growth or weight of a subject, wherein the subject comprises bacterial target cells, the method comprising administering to the subject a first episomal DNA encoding an antibacterial agent that is toxic to the target cells, wherein first DNA is transferred into target cells and the agent is expressed, thereby killing target cells in the subject or reducing the growth or proliferation of target cells and enhancing growth or weight of the subject. 
     Advantageously, the FCR is enhanced in the subject. Optionally, the FCR is enhanced (ie, lowered) by at least 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10%, eg from 2 to 6% or from 2 to 5% (such as 2, 3, 4, 5 or 6%) compared to the FCR of the subject prior to administration of the agent or carrier cells. Optionally, the FCR is enhanced by at least 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10%, eg from 2-5% (such as 2, 3, 4, or 5%) compared to the FCR of a control subject of the same species and sex that has not received administration of the agent or carrier cells. In an embodiment, the subject is a bird, poultry bird, chicken, turkey, goose or duck (preferably a chicken) and the FCR number is lowered by an amount from 0.03 to 0.07, eg, from 0.04 to 0.06, eg, 0.04, 0.05 or 0.06. Thus, for example, compared to a control bird, the FCR of the control bird is 1.7 (g feed/g bird) and after a subject bird that has been treated using the invention has a FCR of 1.64 to 1.66. The control bird (eg, chicken) of the same species, sex and age as the subject of the invention and fed on the same diet (feed), but does not receive the treatment of the invention. The birds may be from the same flock. In an embodiment, therefore, a bird (eg, chicken) has a FCR of 1.64 to 1.66, wherein the subject has been treated by the method of the invention. 
     In an example, the method is carried out on a group of subjects (eg, subjects of the same species, such as a group of livestock animals, eg, a poultry flock or herd of cattle or sheep). In this example, the average FCR of the group is enhanced by the method (ie, the FCR number is lowered) compared to a control group of animals of the same species, subspecies or type, or compared to the average FCR of the group prior to treatment with the method. A control group may be a group of animals of the same species, with the same average age, same proportion of males and females and fed on the same diet as the group of the invention. For example, the group of the invention and control group are both poultry (eg, chicken) flocks, eg, with common ancestry one or two or three generations back. The flock may be a flock of broiler chickens or hen-layer hens. For example, the group of the invention and control group are both beef cattle herds, or dairy herds, eg, with common ancestry one or two or three generations back. 
     FCR, FE &amp; ECI 
     In animal husbandry, feed conversion ratio (FCR) is a ratio measuring the efficiency with which the bodies of livestock convert animal feed into the desired output. For dairy cows, for example, the output is milk, whereas in animals raised for meat (such as beef cows, pigs, chickens, and fish) the output is the flesh, that is, the body mass gained by the animal, represented either in the final mass of the animal or the mass of the dressed output. FCR is the mass of the input divided by the output (thus mass of feed per mass of milk or meat). In some sectors, Feed Efficiency (FE), which is the output divided by the input (i.e. the inverse of FCR), is used. These concepts are also closely related to efficiency of conversion of ingested foods (ECI). FCR is widely used in swine and poultry production, while FE is used more commonly with cattle. Being a ratio the FCR is dimensionless, that is, it is not affected by the units of measurement used to determine the FCR. Animals that have a low FCR are considered efficient users of feed. 
     Preferably, the enhancement produced by the invention is an increase in body mass of the subject. Instead of enhancement of FCR, the invention may be a method for enhancing Feed Efficiency (FE) in the subject, or enhancing the Efficiency of Conversion of Ingested Foods (ECI) in the subject. 
     FCR may be calculated using feed dry mass, or may be calculated on an as-fed wet mass basis. 
     In an embodiment, the FCR, FE or ECI number is changed by 0.2, 0.5, 1, 1.5 or 2. For example, the FCR is reduced by 1, 1.5 or 2. 
     Conversion Ratios for Livestock 
     Beef Cattle 
     As of 2013 in the US, an FCR calculated on live weight gain of 4.5-7.5 was in the normal range with an FCR above 6 being typical. Thus, in an example of the invention the or each animal is a beef cow and the FCR number in the animal (or average FCR of the animals) is reduced by the method to below 6 (calculated using live weight of the animal), eg, below 5.5, 5, 4.5, 4 or 3.5 (optionally, said FCR or average FCR is reduced to an FCR from 3.5 to 5.9, eg, from 4 to 5.5. 
     Dairy Cattle 
     In the US, the price of milk is based on the protein and fat content, so the FCR is often calculated to take that into account. Using an FCR calculated just on the weight of protein and fat in milk obtained from the animal(s), as of 2011 an FCR of 13 was poor, and an FCR of 8 was very good. Thus, in an example of the invention the or each animal is a dairy cow and the FCR number in the animal (or average FCR of the animals) is reduced by the method to below 11, 10, 9 or 8, (and optionally greater than 5 or 6) wherein the FCR is the FCR of milk obtained from the animal(s) that have been treated by the method of the invention, the FCR being calculated using the combined weight of protein and fat in the milk. 
     Another method for dealing with pricing based on protein and fat, is using energy-corrected milk (ECM), which adds a factor to normalize assuming certain amounts of fat and protein in a final milk product; that formula is (0.327×milk mass)+(12.95×fat mass)+(7.2×protein mass). In the dairy industry, Feed Efficiency (ECM/intake) is often used instead of FCR (intake/ECM); an FE less than 1.3 is considered problematic. Thus, in an example of the invention the or each animal is a dairy cow and the FE number in the animal (or average FE of the animals) is increased by the method to above 1.3, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.5 or 3, (and optionally no greater than 4 or 5). Optionally, the FE is increased above 1.3, 1.5, 1.6 and no greater than 1.7. 
     FE based simply on the weight of milk is also used; an FE between 1.30 and 1.70 is normal. Thus, in an example of the invention the or each animal is a dairy cow and the FE number in the animal (or average FE of the animals) is increased by the method to above 1.3, 1.5, 1.6, 1.7, 1.8, 1.9, 2, 2.5 or 3, (and optionally no greater than 4 or 5). Optionally, the FE is increased above 1.3, 1.5, 1.6 and no greater than 1.7. 
     Pigs 
     As of 2011, pigs used commercially in the UK and Europe had an FCR, calculated using weight gain, of about 1 as piglets and ending about 3 at time of slaughter. As of 2012 in Australia and using dressed weight for the output, a FCR calculated using weight of dressed meat of 4.5 was fair, 4.0 was considered “good”, and 3.8, “very good”. In the US as of 2012, commercial pigs had FCR calculated using weight gain, of 3.46 for while they weighed between 240 and 250 pounds, 3.65 between 250 and 260 pounds, 3.87 between 260 and 270 lbs, and 4.09 between 280 and 270 lbs. Thus, in an example of the invention the or each animal is a piglet and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 1 or less, the FCR being calculated using weight gain of the animal(s). Thus, in an example of the invention the or each animal is a pig up to 3 months old and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 1 or less, the FCR being calculated using weight gain of the animal(s). Thus, in an example of the invention the or each animal is a pig greater than 3 months old, but up to 6 or 7 months&#39; old, and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 3 or less (eg, from 1 to 3), the FCR being calculated using weight gain of the animal(s). A piglet may be from 1.5 to 3 months&#39; of age; a pig for slaughter may be greater than 3, but up to 6 months&#39; of age. Thus, in an example of the invention the or each animal is a pig whose weight is between 240 and 250 pounds and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 3.5 or less (eg, 3.4 or less, but optionally no less than 3 or 2.5), the FCR being calculated using weight gain of the animal(s). Thus, in an example of the invention the or each animal is a pig whose weight is between 250 and 260 pounds and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 3.7 or less (eg, 3.65 or less, but optionally no less than 3 or 3.5), the FCR being calculated using weight gain of the animal(s). Thus, in an example of the invention the or each animal is a pig whose weight is between 260 and 270 pounds and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 3.9 or less (eg, 3.8 or less, but optionally no less than 3.7 or 3.5), the FCR being calculated using weight gain of the animal(s). Thus, in an example of the invention the or each animal is a pig whose weight is between 280 and 270 pounds and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 4.1 or less (eg, 4 or less, but optionally no less than 3.8 or 3.9), the FCR being calculated using weight gain of the animal(s). 
     Sheep 
     Some data for sheep illustrate variations in FCR. A FCR (kg feed dry matter intake per kg live mass gain) for lambs is often in the range of about 4 to 6. Thus, in an example of the invention the or each animal is a sheep and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 6 or less (eg, from 4 to 6), the FCR being calculated using kg feed dry matter intake per kg live mass gain of the animal(s). 
     Poultry 
     As of 2011 in the US, broiler chickens typically have an FCR of 1.6 based on body weight gain, and mature in 39 days. At around the same time the FCR based on weight gain for broilers in Brazil was 1.8. Thus, in an example of the invention the or each animal is a poultry bird (eg, a broiler chicken) and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 1.9, 1.8, 1.7, 1.6 or less (eg, from 1.9 to 1.5; or 1.8 to 1.6), the FCR being calculated based on body weight gain. Optionally, the FCR is calculated when each animal is 35-40, eg, 39 days old. 
     For hens used in egg production in the US, as of 2011 the FCR was about 2, with each hen laying about 330 eggs per year. Thus, in an example of the invention the or each animal is a poultry bird (eg, an egg-laying hen, eg, a chicken) and the FCR number in the animal (or average FCR of the animals) is reduced by the method to 2 or less (eg, from 2 to 1.5), the FCR being calculated based on body weight gain. Thus, in an example of the invention the or each animal is a poultry bird (eg, an egg-laying hen, eg, a chicken) and the number of eggs layed by the animal (or average number of eggs layed by the animals) is more than 330 eggs per year (or pro rated for a different period), eg, more than 340, 350 or 400 eggs per year (or pro rated period). 
     Carnivorous Fish 
     The FIFO ratio (or Fish In-Fish Out ratio) is a conversion ratio applied to aquaculture, where the first number is the mass of harvested fish used to feed farmed fish, and the second number is the mass of the resulting farmed fish. FIFO is a way of expressing the contribution from harvested wild fish used in aquafeed compared with the amount of edible farmed fish, as a ratio. Fishmeal and fish oil inclusion rates in aquafeeds have shown a continual decline over time as aquaculture grows and more feed is produced, but with a finite annual supply of fishmeal and fish oil. Calculations have shown that the overall fed aquaculture FIFO declined from 0.63 in 2000 to 0.33 in 2010, and 0.22 in 2015. In 2015, approximately 4.55 kg of farmed fish was produced for every 1 kg of wild fish harvested and used in feed. The fish used in fishmeal and fish oil production are not used for human consumption, but with their use as fishmeal and fish oil in aquafeed they contribute to global food production. Thus, in an example the method of the invention enhances (ie, increases) the FIFO ratio when the animal if a fish (eg, a salmon, tilapia or catfish). For example, the FIFO is increased to 0.3, 0.4, 0.5, 0.6, 0.7 or 0.8, 1, 1.5 or 2 or more. For example, the fish is a salmon or catfish and the FIFO is raised above 1, 1.5 or 2 (and optionally no more than 2 or 2.5). For example, the fish is a talapia and the FIFO is raised above 1.5 or 2 (and optionally no more than 2 or 2.5). 
     Enhancement of growth or weight may be an enhancement (increase) in milk production or yield, eg, an average increase for a group of the invention compared with a control group. A control group may be a group of animals of the same species, with the same average age, same proportion of males and females and fed on the same diet as the group of the invention. For example, here the subjects may be dairy cattle. 
     Enhancement of growth or weight may be an enhancement (increase) in egg production or yield, eg, an average increase for a group of the invention compared with a control group. A control group may be a group of animals of the same species, with the same average age, same proportion of males and females and fed on the same diet as the group of the invention. For example, here the subjects may be hen-layer chickens. 
     Enhancement of growth or weight may be an enhancement (increase) in meat production or yield, eg, an average increase for a group of the invention compared with a control group. A control group may be a group of animals of the same species, with the same average age, same proportion of males and females and fed on the same diet as the group of the invention. For example, here the subjects may be dairy cattle, poultry (eg, chickens), fish, shellfish, sheep or pigs. 
     Enhancement of growth or weight may be an enhancement (increase) in fat production or yield, eg, an average increase for a group of the invention compared with a control group. A control group may be a group of animals of the same species, with the same average age, same proportion of males and females and fed on the same diet as the group of the invention. For example, here the subjects may be dairy cattle, poultry (eg, chickens), fish (eg, salmon, talapia or catfish), shellfish, sheep or pigs. 
     Enhancement of growth or weight may be an enhancement (increase) in fur or hide production or yield, eg, an average increase for a group of the invention compared with a control group. A control group may be a group of animals of the same species, with the same average age, same proportion of males and females and fed on the same diet as the group of the invention. For example, here the subjects may be cows. 
     In an example, the subject is a shellfish. The shellfish may be selected from Shrimp, crayfish, crab, lobster, clam, scallop, oyster, prawn and mussel. 
     The subject may be any subject disclosed herein. The subject may be an animal, such as a livestock animal, eg, a bird (such as a poultry bird; or a chicken or a turkey) or swine. Alternatively, the subject may be a human, eg, a human suffering from an eating disorder (such as anorexia) or who is underweight, eg, wherein the human has a body mass index (BMI) less than 18.5, 18, 17, 16 or 15. 
     For example, the human has mild anorexia (ie, the human has a BMI&lt;17.5), moderate anorexia (ie, the human has a BMI of from 16 to 16.99), severe anorexia (ie, the human has a BMI of from 15 to 15.99) or extreme anorexia (ie, the human has a BMI&lt;15). 
     In an alternative, the subject is a plant, eg, and the target bacteria are plant pathogen bacteria. In an example, the target baceteria are  Pseudomonas , eg,  P. syringae  or  P. aeruginosa.    
     In an alternative, the target cells are archaeal cells. For example the target cells are  methanobacterium  cells. 
     For example the target cells are methanogen cells. For example, the target cells comprise one or more species of cell selected from:
           Methanobacterium bryantii        Methanobacterium formicum        Methanobrevibacter arboriphilicus        Methanobrevibacter gottschalkii        Methanobrevibacter ruminantium        Methanobrevibacter smithii        Methanococcus chunghsingensis        Methanococcus burtonii        Methanococcus aeolicus        Methanococcus deltae        Methanococcus jannaschii        Methanococcus maripaludis        Methanococcus vannielii        Methanocorpusculum labreanum        Methanoculleus bourgensis  ( Methanogenium olentangyi  &amp;  Methanogenium bourgense )     Methanoculleus marisnigri        Methanoflorens stordalenmirensis [34]     Methanofollis liminatans        Methanogenium cariaci        Methanogenium frigidum        Methanogenium  organophilum     Methanogenium wolfei        Methanomicrobium mobile        Methanopyrus kandleri        Methanoregula boonei        Methanosaeta concilii        Methanosaeta thermophila        Methanosarcina acetivorans        Methanosarcina barkeri        Methanosarcina mazei        Methanosphaera stadtmanae        Methanospirillium hungatei        Methanothermobacter defluvii  ( Methanobacterium defluvii )     Methanothermobacter thermautotrophicus  ( Methanobacterium thermoautotrophicum )     Methanothermobacter thermoflexus  ( Methanobacterium thermoflexum )     Methanothermobacter wolfei  ( Methanobacterium wolfei )     Methanothrix sochngenii          

     Optionally, the target cells are not pathogenic to the subject, for example when the method is a non-medical method. In an example, the method is a cosmetic method. 
     For example the target cells are methane-producing cells, and optionally the subject is a livestock animal, preferably a ruminant, or a cow (eg, a beef or dairy cattle). By reducing methane-producing cells in such animal, the invention may in one embodiment enhance the weight of the animal (eg, enhance the yield of meat from the animal) and/or enhance the yield of milk or another product of the animal, such as fur or fat. 
     In an example, the target cells are selected from  E. coli, Salmonella  and  Campylobacter  cells. In an example, the target cells are  E. coli, Salmonella  or  Campylobacter  cells. In an example, each animal is a chicken (eg, a broiler or hen-layer) and the target cells are  Salmonella  or  Campylobacter  cells. In an example, each animal is a cow (eg, a beef or dairy cow) and the target cells are mehanogen cells. 
     In an example, the target cells are selected from  Mycoplasma  (eg,  Mycoplasma mycoides  (eg,  Mycoplasma mycoides  subsp.  Mycoides ),  Mycoplasma leachii  or  Mycoplasma bovis ),  Brucella abortus, Listeria monocytogenes, Clostridium  (eg,  Clostridium chauvoei  or  Clostridium septicum ),  Leptospira  (eg,  L. canicola, L. icterohaemorrhagiae, L. grippotyphosa, L. hardjo or L. Pomona ),  Mannheimia haemolytica, Trueperella pyogenes, Mycobacterium bovis, Campylobacter  spp. (eg,  Campylobacter jejuni  or  Campylobacter coli ),  Bacillus anthracis, E. coli  (eg,  E. coli  O157:H7) or  Pasteurella multocida  (eg,  Pasteurella multocida  B:2, E:2, A:1 or A:3). In the example, optionally the subject or animal is a livestock animal, such as a cow, sheep, goat or chicken (preferably a cow). 
     Optionally, eg, wherein the subject is an animal (eg, a livestock animal or a wild animal), the target cells are zoonotic bacterial cells, such as cells of a species selected from  Bacillus anthracis, Mycobacterium bovis  (eg, wherein the animal is a cow),  Campylobacter  spp (eg, wherein the animal is a poultry animal),  Mycobacterium marinum  (eg, wherein the animal is a fish), Shiga toxin-producing  E. Coli  (eg, wherein the animal is a ruminant),  Listeria  spp (eg, wherein the animal is a cow or sheep),  Chlamydia abortus  (eg, wherein the animal is a sheep),  Coxiella burnetii  (eg, wherein the animal is a cow, sheep or goat),  Salmonella  spp (eg, wherein the animal is a poultry animal),  Streptococcus suis  (eg, wherein the animal is a pig) and  Corynebacterium  (eg,  C. ulcerans ) (eg, wherein the animal is a cow). 
     In an example, a plurality of carrier cells as described herein (eg, carrier cells of any configuration, aspect, example or embodiment described herein) is administered to the subject, wherein the carrier cells comprise the DNA encoding the agent. 
     In an example, each animal is a chicken (eg, a broiler or hen-layer) and the target cells are  Salmonella  or  Campylobacter  cells. In an example, each animal is a cow (eg, a beef or dairy cow) and the target cells are mehanogen cells. 
     In an embodiment, therefore, there is provided: 
     A method (eg, a non-medical or a medical) for enhancing the growth or weight of a subject, wherein the method comprises the administration of a plurality of carrier cells to the subject, wherein the subject comprises bacterial target cells and each carrier cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the agent, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells and enhancing growth or weight of the subject. 
     Optionally, the target cells are  Salmonella  cells. In an example, the target cells comprise  S. enterica  and/or  S. typhimurium  cells; optionally wherein the  S. enterica  is  S. enterica  subspecies  enterica. Optionally, the method kills a plurality of different S. enterica  subspecies  enterica  serovars; optionally wherein each serovar is selected from the group consisting of  Typhimurium, Enteritidis , Virchow, Montevideo, Heidelberg, Hadar, Binza, Bredeney,  Infantis , Kentucky, Seftenberg, Mbandaka, Anatum, Agona and Dublin. Optionally, the method kills  S. enterica  subspecies  enterica  serovars  Typhimurium, Infantis  and  Enteritidis . Optionally, the method kills  S. enterica  subspecies  enterica  serovars  Typhimurium  and  Enteritidis . Optionally, the method kills  S. enterica  subspecies  enterica  serovars  Typhimurium  and  Infantis . Optionally, the method kills  S. enterica  subspecies  enterica  serovars  Enteritidis  and  Infantis . The most prevalent serovars in chicken are  Salmonella Enteritidis, Salmonella Infantis  and  Salmonella Typhimurium . In general, similar serovars of  Salmonella  are found in infected humans and chicken ( S. Enteritidis  and  S. Typhimurium ). By killing  Salmonella  in livestock animals, the invention is useful for reducing the pool of zoonotic bacteria that are available for transmission to humans (such as by eating the livestock or products made thereofrom, such as meat or dairy products for human consumption). 
     Advantageously, the carrier cells are Enterobacteriaceae cells, optionally  E. coli  cells. As exemplified by the Examples below, the inventors have surprisingly found that (i) antibacterial agents can be efficiently transferred from such cells to  Salmonella  target cells both in vivo and in vitro; and (ii) 100% killing of  Salmonella  was possible when the agent is transferred by conjugation from Enterobacteriacae cells to  Salmonella  cells. Furthermore, surprisingly DNA encoding a guided nuclease antibacterial was capable of killing 18  Salmonella  spp. Serotypes, including the clinically- and zoonotically-important  Typhimurium, Infantis  and  Enteritidis . Optionally, the method kills  S enterica  subspecies  enterica  serovars  Typhimurium  and  Enteritidis  serovars. Optionally, the Enterobacteriaciae cells are cells of an Enterobacteriaciae species shown in Table 5. 
     Importantly, reduction of target cells was observed in the GI tract of the subject. Thus, optionally the method reduces target cells in the gastrointestinal tract of the animal; optionally the method reduces target cells in the jejunum, ileum, colon, liver, spleen or caecum of the animal; optionally wherein the animal is a bird and the method reduces target cells in the caecum of the bird. This may be important to reduce spread of zoonotic or other deterimental target strains in the faeces of the subjects, such as livestock animals. Thus, in an example the method is carried out on a group of subjects (eg, a herd or flock, such as a herd of swine or a flock of birds), wherein spread of cells of the target species is reduced in the group. The inventors have also demonstrated this in a flock of poultry where  Salmonella  were killed using the invention, wherein a guided nuclease was used to cut a plurality of protospacer sequences in target cells, thereby killing the cells and reducing spread thereof in the flock. 
     Thus, in an example the method is carried out on a group (optionally a flock or herd) of animals, wherein some or all of the animals comprise target cells, wherein spread of cells of the target species is reduced in the group; or wherein spread is reduced from the group to a second group of animals. 
     Optionally, the first DNA is comprised by a plasmid, for example wherein the plasmid comprises a RP4 origin of transfer (oriT). The plasmid may be any type of plasmid disclosed herein. 
     The agent may be any antibacterial agent disclosed herein, preferably a guided nuclease that is programmed to cut one or more target sequences in target cells. A suitable nuclease may be a TALEN, meganuclease, zinc finger nuclease or Cas nuclease. For example, the agent comprises one or more components (eg, a Cas nuclease and/or a guide RNA or a crRNA) of a CRISPR/Cas system that is operable in a target cell to cut a protospacer sequence comprised by the target cell, optionally wherein the target cells comprise first and second strains of a bacterial species and each strain comprises the protospacer sequence, wherein cells of the strains are killed. For example, the system is operable to cut at least 3 different protospacer sequences comprised by the cell genome. Optionally, each or some of said protospacer sequences is comprised by a pathogenicity island that is comprised by the cell. As shown in the Examples, this is highly effective for target cell killing. Optionally, the agent is operable to cut a plurality of different protospacer sequences comprised by the target cell genome. Optionally, the agent comprises one or more components of a CRISPR/Cas system that is operable in a target cell to cut at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 different protospacer sequences comprised by the target cell genome (eg, comprised by the target cell chromosome). 
     In an embodiment, the agent
         (a) comprises a guided nuclease that is capable of recognising and modifying a target nucleic acid sequence, wherein the target sequence is comprised by an endogenous chromosome or episome of the target cells but is not comprised by the carrier cells, wherein the nuclease modifies the chromosome or episome to kill the target cells or inhibit the growth or proliferation of the target cells; and/or   (b) encodes a guide RNA or crRNA of a CRISPR/Cas system that operates with a Cas nuclease in the target cells to cut a protospacer sequence comprised by the target cells.       

     There is also provided: 
     A carrier cell (optionally for use in a method of the invention), wherein the cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a bacterial target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the antibacterial agent, hereby killing the target cell, wherein the target cell is a  Salmonella  cell and the carrier cell is an Enterobacteriaceae cell. 
     There is also provided: 
     A composition comprising a plurality of carrier cells for use in a method comprising administration of the cells to a subject to treat an infection by pathogenic bacterial target cells, wherein each carrier cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the agent, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells, wherein the target cells are  Salmonella  cells and the carrier cells are Enterobacteriaceae cells. In an alternative, the method treats or reduces a symptom of an infection by pathogenic target cells. 
     Any administration of cells to a subject herein may be by oral administration. Any administration of cells to a subject herein may preferably be by administration to the GI tract. Any administration of cells to a subject herein may be by systemic, intranasal or inhaled administration. 
     There is also provided: 
     A non-medical method of killing zoonotic bacterial target cells in an animal, the method comprising administering to the animal a plurality of carrier cells, wherein each carrier cell is a bacterial cell comprising a first episomal DNA encoding an antibacterial agent that is toxic to a target cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into a target cell for expression therein of the agent, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells, wherein the target cells are  Salmonella  cells and optionally the carrier cells are Enterobacteriaceae cells. 
     The animal may be any animal disclosed herein, eg, a livestock animal, domesticated animal or wild animal (eg, a bat or bird)). 
     Optionally, any method herein reduces  Salmonella  in the gastrointestinal tract of the subject. 
     Optionally, the target cells comprise different  Salmonella  spp. types that are killed. 
     In any aspect, configuration, example, concept or embodiment, the carrier cell, target cell(s) or DNA is respectively a carrier cell, target cell(s) or DNA as defined in any other aspect, configuration, example, concept or embodiment. 
     The invention also provides: 
     A DNA, wherein the DNA is capable of being introduced into a target cell, wherein the DNA encodes a plurality of guide RNAs or crRNAs of a CRISPR/Cas system wherein the guide RNAs or crRNAs are operable with Cas nuclease in the target cell to recognise a plurality of protospacer sequences comprised by the target cell genome, optionally wherein the target cell is a  Salmonella  cell or a cell of a species disclosed in Table 5; and
         (a) the protospacer sequences comprise one or more pathogenic island nucleotide sequences of the target cell genome;   (b) the protospacer sequences comprise one or more invasion gene sequences of the target cell genome;   (c) the protospacer sequences comprise one or more secretion system gene sequences of the target cell genome; and/or   (d) the protospacer sequences comprise one or more nucleotide sequences of genes selected from avrA, sptP, sicP, sipA, sipD, sipC, sipB, sicA, invB, ssaE, sseA, sseB, sscA, sseC, sseD, sseE, sscB, sseF, sseG, mgtC, cigR, pipA, pipB, pipC, sopB and pipD (optionally selected from invB, sicP, sseE, pipA, pipB, pipC, hilA, marT and sopB) of  Salmonella  (eg,  S. enterica ) or orthologues or homologues of said genes.       

     Homologue: A gene, nucleotide or protein sequence related to a second gene, nucleotide or protein sequence by descent from a common ancestral DNA or protein sequence. The term, homologue, may apply to the relationship between genes separated by the event of or to the relationship between genes separated by the event of genetic duplication. 
     Orthologue: Orthologues are genes, nucleotide or protein sequences in different species that evolved from a common ancestral gene, nucleotide or protein sequence by speciation. Normally, orthologues retain the same function in the course of evolution. 
     By “orthologues or homologues of said genes” it is meant that a protospacer may be a sequence comprised by a gene of the target cell genome, wherein the gene is an orthologue or homologue of a  Salmonella  gene selected from avrA, sptP, sicP, sipA, sipD, sipC, sipB, sicA, invB, ssaE, sseA, sseB, sscA, sseC, sseD, sseE, sscB, sseF, sseG, mgtC, cigR, pipA, pipB, pipC, sopB and pipD (optionally selected from invB, sicP, sseE, pipA, pipB, pipC, hilA, marT and sopB). 
     Optionally the DNA is the first DNA for use in the method of the invention. Preferably, the target cell is a  Salmonella  cell, eg, a  S. enterica  cell, such as a  Salmonella enterica  subsp.  enterica  serovar  Enteritidis  cell. 
     Optionally any  Salmonella  herein is  Salmonella enterica  subsp.  enterica  serovar  Typhimurium  str. LT2. 
     Optionally, the target cell is a cell of a species disclosed in Table 5 other than a  Salmonella  species and the sequences of (d) are sequences of genes selected from orthologues or homologues of avrA, sptP, sicP, sipA, sipD, sipC, sipB, sicA, invB, ssaE, sseA, sseB, sscA, sseC, sseD, sseE, sscB, sseF, sseG, mgtC, cigR, pipA, pipB, pipC, sopB and pipD (optionally selected from invB, sicP, sseE, pipA, pipB, pipC, hilA, marT and sopB). 
     of  Salmonella  (eg,  S. enterica ). 
     The DNA may comprise one or more CRISPR spacers, wherein each spacer consists of a nucleotide sequence of a secretion system gene (optionally a type III protein secretion system or a secretion system of SPI-1 or SPI-2 of  Salmonella ); or a T3SS locus gene; or with up to 10, 9, 8, 7, 6, 5, 4, 3 or 2 nucleotide differences therefrom. 
     Optionally, the DNA (eg, DNA of the carrier cell) encodes a plurality of guide RNAs or crRNAs of a CRISPR/Cas system wherein the guide RNAs or crRNAs are operable with Cas nuclease in the target cell to recognise a plurality of protospacer sequences comprised by the target cell genome, wherein the target cell is a  Salmonella  cell and the protospacer sequences comprise one or more nucleotide sequences of genes selected from invB, sicP and sseE. For example, the protospacer sequences comprise nucleotide sequences of genes invB, sicP and sseE. In an example, the DNA also encodes a Cas, eg, a Cas9, Cas3, Cpf1, Cas12, Cas13, CasX or CasY. In an embodiment, the Cas is a Type I, II, III, IV, V or VI Cas, preferably a Type I or II Cas. In an example, the DNA also encodes a Cas3 and cognate Cascade proteins (eg, CasA, B, C, D and E). Optionally, the Cas (and Cascade of present) are  E. coli  Cas (and Cascade). 
     Optionally, the gene of the target cell encodes a chaperone or secreted effector protein, eg, such a protein encoded by a pathogenicity island or type III protein secretion system (optionally a T3SS locus, such as a secretion system of SPI-1 and SPI-2 of  Salmonella ). 
     The DNA may comprise one or more CRISPR spacers, wherein each spacer consists of 20-40, 25-35, or 30-35 consecutive nucleotides of a gene comprised by the genome of the target cell; eg,
         (a) a gene selected from avrA, sptP, sicP, sipA, sipD, sipC, sipB, sicA, invB, ssaE, sseA, sseB, sscA, sseC, sseD, sseE, sscB, sseF, sseG, mgtC, cigR, pipA, pipB, pipC, sopB and pipD of  Salmonella  or a homologue or orthologue thereof;   (b) a gene comprised by a pathogenicity island that is comprised by the target cell genome;   (c) a secretion system (eg, a type III protein secretion system) gene comprised by the target cell genome       

     Optionally, the DNA is comprised by a plasmid which comprises an origin of transfer (oriT) and an origin of replication (oriV) that is operable for replication of the DNA in a bacterial host cell. 
     Optionally, the first DNA is comprised by a plasmid, wherein the plasmid comprises a RP4 origin of transfer (oriT) and/or a p15A origin of replication. 
     Optionally, the DNA comprises SEQ ID NO: 15, optionally wherein the DNA is comprised by a plasmid in a carrier bacterial cell for conjugation to a  Salmonella  target cell. In an example, the DNA of the invention is comprised by a conjugative plasmid or phagemid. 
     In an example, the DNA comprises CRISPR repeat and spacer sequences, wherein
         (e) the repeat sequences each comprise SEQ ID NO: 16; and/or   (f) the spacer sequences comprise one, two or three sequences selected from SEQ ID NOs: 17-19 and complement sequences thereof;
 
optionally wherein the DNA is comprised by a plasmid in a carrier bacterial cell for conjugation to a  Salmonella  target cell.
       

     Optionally, the DNA comprises (optionally in 5′ to 3′ order) SEQ ID NO: 17, SEQ ID NO: 18 and SEQ ID NO: 19. Optionally, the DNA comprises one or more (eg, at least 3) spacer sequences shown in Table 6. 
     In an example, the DNA encodes a Cas3 and optionally one or more Cascade proteins (eg, one or more of CasA, B, C, D and E). In an embodiment, the first DNA encodes a Cas3 and CasA, B, C, D and E. In an embodiment, the first DNA encodes an  E. coli  Cas3 and CasA, B, C, D and E. Optionally, the guided nuclease (eg, Cas3) is a Type I-A, -B, -C, -D, -E, -F or -U Cas. 
     In an example, the agent in any configuration, example, option or embodiment herein, the agent comprises one or more components of a CRISPR/Cas system that is operable in the target cell to cut a protospacer sequence comprised by the target cell. 
     In an example, the system is operable to cut at least 3 different protospacer sequences comprised by the target cell genome. In an embodiment, each or some of said protospacer sequences is comprised by a pathogenicity island that is comprised by the target cell. 
     In an example, the agent in any configuration, example, option or embodiment herein
         (a) comprises a guided nuclease that is capable of recognising and modifying a target nucleic acid sequence, wherein the target sequence is comprised by an endogenous chromosome or episome of the target cells but is not comprised by the carrier cells, wherein the nuclease modifies the chromosome or episome to kill the target cells or inhibit the growth or proliferation of the target cells; and/or   (b) encodes a guide RNA or crRNA of a CRISPR/Cas system that operates with a Cas nuclease in the target cell to cut a protospacer sequence comprised by the target cell.       

     Optionally, the DNA comprises a constitutive promoter for expression of the guide RNAs or crRNAs. Optionally, the DNA comprises a constitutive promoter for expression of a Cas nuclease that is operable in a target cell with the guide RNAs or crRNAs to modify (eg, cut) protospacer sequences of the target cell genome. 
     Optionally, the Cas, Cascade proteins, gRNAs and crRNAs are  E. coli  K12 (MG1655) Cas, Cascade proteins, gRNAs and crRNAs respectively. Optionally, the DNA is devoid of nucleotide sequences encoding Cas1 and Cas2 proteins. 
     The invention also relates to carrier bacteria encoding desired protein or RNA (eg, encoding an antimicrobial agent) and methods of use. In embodiments, the agent can be transferred into target cells by conjugation between carrier cells (to which the agent is not toxic) and the target cells, whereby the agent is toxic to the target cells and kills the target cells. In other embodiments, the growth or proliferation of target cells is reduced (eg, by at least 40, 50, 60, 70, 80, or 90% compared to growth in the absence of the agent). Each carrier cell comprises episomal DNA encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic (or is less toxic) to the carrier cell. The invention finds application, for example, in controlling or killing target bacteria that are pathogenic to humans, animals or plants. The invention finds application, for example, in controlling or killing zoonotic target bacteria comprised by an animal (eg, a livestock animal). For example, the carrier cells may be comprised by a medicament for treating or preventing a disease or condition in a human or animal; a growth promoting agent for administration to animals for promoting growth thereof; killing zoonositic bacteria in the animals; for administration to livestock as a pesticide; a pesticide to be applied to plants; or a plant fertilizer. 
     An advantage of the invention is that the carrier cells may be used as producer cells in which DNA encoding the antibacterial agent can be replicated. Another advantage of an example of the invention is that further replication of the DNA can be avoided in target cells that do not comprise one or more factors that are required for such replication. Thus, a scheme can be envisaged wherein the factor(s) are present in the carrier cells (and DNA encoding the agent is replicated to provide many copies for subsequent conjugative transfer into target cells) and in the target cells the DNA is not further replicated, which is useful for containing the action of the antibacterial agent, such as in environments or in a human or animal body or in (or on) a plant a Containment may be required to avoid undesired killing of non-target cells or to provide control generally over dosing and/or the administration of the killing activity regime. 
     The first DNA is replicable in the carrier cell (but not in the target cell), in which case it is important that the agent is not toxic to the carrier cell, whilst it is toxic to the target cell. In certain embodiments, the invention uses sequence-specific killing of the target cell to achieve this selectivity. 
     To this end, in an example the first DNA encodes a guided nuclease that is operable in the target cell to recognize and cut a target sequence of a target cell chromosome, thereby killing the cell or wherein the growth or proliferation of the cell is reduced. This is advantageous over the use of other types of toxic agent, which are less discriminate in their action, being able to kill several species or strain (eg, potentially also being toxic to the carrier cell to some degree). By using a guided nuclease (eg, a TALEN or Cas nuclease), these can be programmed to recognize a target sequence that is present in the target cell genome (eg, comprised by a chromosome or episome of the target cell), but is absent in the genome of the carrier cell. 
     Thus, in this case replication of the first DNA can freely happen in the carrier cell without risk of killing the cell or reducing its growth or proliferation due to the encoded agent and replication of sequences encoding the agent. Thus, in an example, where the first DNA encodes a guided nuclease, the guided nuclease is capable of recognizing and cutting a target nucleic acid sequence comprised by the genome (eg, chromosome) of the target cell, wherein the target cell is absent in the carrier cell. 
     A particularly useful example is where the first DNA encodes a Cas nuclease (eg, a Cas9 or Cas3) that is operable with a guide RNA or crRNA in the target cell, wherein the RNA is operable to guide the Cas to the target sequence, wherein the Cas modifies (eg, cuts) the target sequence and the target cell is killed or target cell growth or proliferation is inhibited. In one embodiment, the first DNA encodes the Cas and the guide RNA or crRNA. In another embodiment, the first DNA encodes the guide RNA or crRNA, but does not encode a cognate Cas. In this embodiment, the RNA is operable in the target cell with an endogenous Cas encoded by the target cell genome, wherein the RNA is operable to guide the Cas to the target sequence, wherein the Cas modifies (eg, cuts) the target sequence and the target cell is killed or target cell growth or proliferation is inhibited. In this se sense, the agent may comprise a component of a CRISPR/Cas system (eg, a Cas nuclease, Cascade Cas, crRNA, guide RNA or tracrRNA). Thus, the invention usefully recognizes the benefit of using antibacterial agents that act by target recognition in the target cell but not in the carrier cell, which opens up the ability for the first DNA to freely replicated in the carrier cell without significant toxicity to the carrier cell. 
     Example Plasmids 
     A method of delivery of any agent, such as a CRISPR-Cas system (or a component thereof) can be by bacterial conjugation, a natural process whereby a donor bacterium transfers DNA from itself to a recipient bacterium. Donor bacteria elaborate a surface structure, the pilus which can be considered to be like a syringe or drinking straw through which the DNA is delivered. The donor pilus binds to the surface of a receptive recipient and this event triggers the process of DNA transfer. Plasmids are suitable for this conjugative process, where the plasmid comprises DNA encoding the agent of the invention. 
     DNA transfer by conjugation may only take place with a ‘susceptible recipient’ but does not generally occur with a recipient carrying a similar type of plasmid. Because conjugation is via pilus bridge, it is possible for that bridge to attach itself not to a recipient but to the donor bacterium. This could result in a futile cycle of transfer of the plasmid DNA to itself. Plasmids thus naturally encode incompatibility factors. One is a surface arrayed protein that prevents the pilus binding to bacterium displaying that surface protein such as itself or any other bacterium carrying the same plasmid. Additionally, plasmids naturally encode another incompatibility system that closely regulates the copy number of the plasmid inside a bacterium. Thus, should a conjugation event manage to evade surface exclusion and start to transfer DNA by conjugation, the recipient will prevent that plasmid establishing as it already maintains the current copy number and will not accept and maintain a further unwanted additional copy. 
     In an example of the invention, the DNA encoding the agent is comprised by a plasmid. In an embodiment, the plasmid is a member of a plasmid incompatibility group, wherein the target cell does not comprise a plasmid of said group. Optionally, the plasmid of the invention is a member of the incompatibility group P (ie, the plasmid is an incP plasmid).  Salmonella  very rarely carry incP plasmids, so this incP plasmid is useful where the target cell is a  Salmonella  cell. For example within the Enterobacteriaceae the following is a non-exclusive list of potential plasmids that could use for delivery: IncFI, IncFII, IncFIII, IncFIV, IncFV, IncM, Inc9, InclO, Incl, IncA, IncB, IncC, IncH, Incla, InclIc, IncI2, Incly, IncJ, IncL, IncN, Inc2e, IncO, IncP, IncS, IncT and/or IncW. Thus, optionally, the target cell is an Enterobacteriaceae cell and the DNA of the invention is comprised by a plasmid, wherein the plasmid is selected from an IncFI, IncFII, IncFIII, IncFIV, IncFV, IncM, Inc9, InclO, Incl, IncA, IncB, IncC, IncH, Incla, InclIc, IncI2, Incly, IncJ, IncL, IncN, Inc2e, IncO, IncP, IncS, IncT and IncW plasmid. 
     In an example, the carrier cell of the invention comprises two or more plasmids, each plasmid comprising a DNA of the invention that encodes an antibacterial agent, wherein a first of said plasmids is a member of a first incompatibility group, wherein the target cell does not comprise a plasmid of said first group, and wherein a second of said plasmids is a member of a second incompatibility group, wherein the target cell does not comprise a plasmid of said second group. For example, a carrier cell may comprise an incP plasmid encoding an anti-target cell CRISPR-Cas system or a component thereof (eg, encoding a first crRNA or guide RNA that targets a first protospacer sequence of the target cell genome) and wherein the carrier cell further comprises an incF1 plasmid encoding an anti-target cell CRISPR-Cas system or a component thereof (eg, encoding a second crRNA or guide RNA that targets a second protospacer sequence of the target cell genome), the protospacers comprising different nucleotide sequences. For example, the protospacers are comprised by different genes of the target cell genome. For example the protospacers are comprised by one or more pathogenicity islands of the target cell genome. Optionally, the target cell is an Enterobacteriaceae cell. Optionally, the carrier cell comprises a group of plasmids comprising 2, 3, 4, 5, 6 or more different types of plasmid, wherein each plasmid is capable of being conjugatively transferred into a target cell, wherein the plasmids encode different agents or different components of an antibacterial agent. For example, the plasmids encode different cRNAs or gRNAs that target different protospacers comprised by the target cell genome. For example, the group of plasmids comprises up to n different types of plasmid, wherein the plasmids are members of up to n different incompatibility groups, eg, groups selected from IncFI, IncFII, IncFII, IncFIV, IncFV, IncM, Inc9, InclO, Incl, IncA, IncB, IncC, IncH, IncIa, InclIc, IncI2, Incly, IncJ, IncL, IncN, Inc2e, IncO, IncP, IncS, IncT and IncW. For example, n=2, 3, 4, 5, 6, 7, 8, 9 or 10. 
     For example, the carrier cell comprises (i) a first plasmid that encodes a first type of CRISPR/Cas system that targets a first protospacer comprised by the target cell genome, or encodes a component of said system; and (ii) a second plasmid that encodes a second type of CRISPR/Cas system that targets a second protospacer comprised by the target cell genome, or encodes a component of said system, wherein the first and second types are different. For example, the first type is a Type I system, and the second type is a Type II system (eg, the first plasmid encodes a Cas3, Cascade and a crRNA or guide RNA that is operable with the Cas3 and Cascade in the target cell to modify the first protospacer; and the second plasmid encodes a Cas9 and a crRNA or guide RNA that is operable with the Cas9 in the target cell to modify the second protospacer). In an alternative, the Cas3 and Cascade are encoded by an endogenous target cell gene, wherein the first plasmid encodes the crRNA or guide RNA that is operable with the endogenous Cas3 and Cascade in the target cell to modify the first protospacer. In an alternative, the Cas9 is encoded by an endogenous target cell gene, wherein the second plasmid encodes the crRNA or guide RNA that is operable with the endogenous Cas9 in the target cell to modify the second protospacer. Optionally, the Cas3 and Cascade are encoded by endogenous genes of the target cell and the Cas9 is encoded by the second plasmid. 
     Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type I CRISPR/Cas system (or component thereof, eg, a Cas3 or a crRNA or a gRNA) and a second plasmid encoding a Type III CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type I CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type IV CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type I CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type V CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type I CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type VI CRISPR/Cas system (or a component thereof). 
     Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type II CRISPR/Cas system (or component thereof, eg, a Cas9 or a crRNA or a gRNA) and a second plasmid encoding a Type III CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type II CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type IV CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type II CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type V CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type II CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type VI CRISPR/Cas system (or a component thereof). 
     Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type V CRISPR/Cas system (or component thereof, eg, a Cas12a or a crRNA) and a second plasmid encoding a Type III CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type V CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type IV CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type V CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type V CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment a first plasmid encoding a Type V CRISPR/Cas system (or component thereof) and a second plasmid encoding a Type VI CRISPR/Cas system (or a component thereof). 
     Instead of a Type I and Type II system, the invention alternatively provides in an embodiment first and second plasmids, each encoding a Type I CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment first and second plasmids, each encoding a Type II CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment first and second plasmids, each encoding a Type III CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment first and second plasmids, each encoding a Type IV CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment first and second plasmids, each encoding a Type V CRISPR/Cas system (or a component thereof). Instead of a Type I and Type II system, the invention alternatively provides in an embodiment first and second plasmids, each encoding a Type VI CRISPR/Cas system (or a component thereof). 
     Optionally, the plasmids are members of different incompatibility groups, eg, groups selected from IncFI, IncFII, IncFIII, IncFIV, IncFV, IncM, Inc9, InclO, Incl, IncA, IncB, IncC, IncH, IncIa, InclIc, IncI2, Incly, IncJ, IncL, IncN, Inc2e, IncO, IncP, IncS, IncT and IncW. In an example here, the target cell is an Enterobacteriaceae cell. 
     Embodiments 
     Thus, by way of example the invention provides the following Embodiments.
     1. A carrier bacterial cell comprising a first episomal DNA, the DNA comprising a nucleic acid of interest (NSI) that encodes a protein of interest (POI) or RNA of interest (ROI) for expressing the POI or ROI in a target bacterial cell, the carrier cell being capable of conjugative transfer of the DNA into the target cell for expression therein of the POI, ROI or agent, wherein
       (a) the carrier cell comprises a second DNA which is different from the first DNA, wherein the second DNA comprises or encodes a first factor required for replication of the first DNA;   (b) the first DNA does not comprise or encode said first factor, wherein the first DNA is non-self-replicative in the absence of the first factor, but is able to replicate in the carrier cell in the presence of the first factor provided by the second DNA;   (c) wherein the carrier cell comprises genes encoding one or more conjugation factors sufficient to carry out conjugative transfer of the first DNA into a target bacterial cell.   
       

     In a first alternative, Embodiment 1 provides:— 
     A carrier bacterial cell comprising a first episomal DNA, the DNA comprising a nucleic acid of interest (NSI) that encodes a protein of interest (POI) or RNA of interest (ROI) for expressing the POI or ROI in a target bacterial cell, the carrier cell being capable of conjugative transfer of the DNA into the target cell for expression therein of the POI or ROI. 
     In a second alternative, Embodiment 1 provides:— 
     A carrier bacterial cell comprising a first episomal DNA, the DNA encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into the target cell for expression therein of the agent. 
     In an example, the agent comprises a guided nuclease that is capable of recognizing and cutting a target nucleic acid sequence comprised by the target cell genome, wherein the target sequence is not comprised by the carrier cell. In this sense, therefore, the antibacterial agent is toxic to a target bacterial cell but is not toxic to the carrier cell. 
     For example, there is provided:— 
     A carrier bacterial cell comprising a first episomal DNA, the DNA encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into the target cell for expression therein of the agent, wherein the agent is a component of a CRISPR/Cas system that is operable in the target cell to modify a target nucleic acid sequence comprised by the target cell genome (eg, comprised by the target cell chromosome). 
     Another example provides:— 
     A carrier bacterial cell comprising a first episomal DNA, the DNA encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic to the carrier cell, the carrier cell being capable of conjugative transfer of the DNA into the target cell for expression therein of the agent, wherein
         (a) the carrier cell comprises a second DNA which is different from the first DNA, wherein the second DNA comprises or encodes a first factor required for replication of the first DNA;   (b) the first DNA does not comprise or encode said first factor, wherein the first DNA is non-self-replicative in the absence of the first factor, but is able to replicate in the carrier cell in the presence of the first factor provided by the second DNA;   (c) wherein the carrier cell comprises genes encoding one or more conjugation factors sufficient to carry out conjugative transfer of the first DNA into a target bacterial cell.       

     An example also provides:— 
     A carrier bacterial cell comprising a plasmid comprising a first DNA, the DNA encoding a guided nuclease that is capable of recognising and modifying a target nucleic acid sequence, the carrier cell being capable of conjugative transfer of the plasmid into the target cell for expression therein of the nuclease, wherein the target sequence is comprised by an endogenous chromosome or episome of the target cell but is not comprised by the carrier cell, wherein the nuclease is capable of modifying (eg, cutting) the chromosome or episome to kill the target cell or inhibit the growth or proliferation of the target cell. 
     Optionally, the nuclease is a Cas nuclease, meganuclease, zinc finger nuclease or TALEN. 
     Optionally, the nuclease is a Cas nuclease of a Type I, II, III, IV or V CRISPR system. 
     An example also provides:— 
     A carrier bacterial cell comprising a plasmid comprising a first DNA, the DNA encoding a component of a CRISPR/Cas system that is capable of recognising and modifying a target nucleic acid sequence, the carrier cell being capable of conjugative transfer of the plasmid into the target cell for expression therein of the component, whereby the expressed component forms part of a said system in the target cell, wherein the target sequence is comprised by an endogenous chromosome or episome of the target cell but is not comprised by the carrier cell, wherein the system is capable of modifying (eg, cutting) the chromosome or episome to kill the target cell or inhibit the growth or proliferation of the target cell. 
     In an aspect, the component is a guide RNA or crRNA that is capable of hybridising to the target sequence of the target cell. They system may be a Type I, II, III, IV or V CRISPR system. 
     Optionally,
         (a) the carrier cell comprises a second DNA which is different from the first DNA, wherein the second DNA comprises or encodes a first factor required for replication of the first DNA;   (b) the first DNA does not comprise or encode said first factor, wherein the first DNA is non-self-replicative in the absence of the first factor, but is able to replicate in the carrier cell in the presence of the first factor provided by the second DNA;   (c) wherein the carrier cell comprises genes encoding one or more conjugation factors sufficient to carry out conjugative transfer of the first DNA into a target bacterial cell.       

     The second DNA may be comprised by a chromosome or episome (eg, plasmid) of the carrier cell. Any method of the invention may use carrier cell(s) of any of these examples. Advantageously, the cell is for treating or preventing a target cell infection in a human or an animal subject (eg, a chicken, cow, pig, fish or shellfish). Advantageously, the carrier cell is a cell of a species that is probiotic to said subject or is probioitic to humans or animals (eg, chickens). For example, the carrier cell is a probiotic  E. coli  cell. For example, the carrier cell is a probiotic  Bacillus  cell. In an example, the target cell is a cell of a species that is pathogenic to said subject, or is pathogenic to humans or aniumals (eg, chickens). Advantageously, the first DNA encodes one or more guide RNAs or one or more crRNAs that are capable of hybridizing in the target cell to respective target nucleic acid sequence(s), wherein the target sequence(s) are comprised by an endogenous chromosome and/or endogenous episome of the target cell. For example, the first DNA encodes 2, 3, 4, 5, 6, 7, 7, 9, or 10 (or more than 10) different gRNAs or different crRNAs that hybridise to a respective target sequence, wherein the target sequences are different from each other. For example, 3 different gRNAs or crRNAs are encoded by the first DNA. For example, 2 different gRNAs or crRNAs are encoded by the first DNA. For example, 3 different gRNAs or crRNAs are encoded by the first DNA. For example, 4 different gRNAs or crRNAs are encoded by the first DNA. For example, 3 different gRNAs or crRNAs are encoded by the first DNA. For example, 5 different gRNAs or crRNAs are encoded by the first DNA. For example, 6 different gRNAs or crRNAs are encoded by the first DNA. For example, 7 different gRNAs or crRNAs are encoded by the first DNA. For example, 8 different gRNAs or crRNAs are encoded by the first DNA. For example, 9 different gRNAs or crRNAs are encoded by the first DNA. For example, 10 different gRNAs or crRNAs are encoded by the first DNA. For example, 11 different gRNAs or crRNAs are encoded by the first DNA. For example, 12 different gRNAs or crRNAs are encoded by the first DNA. For example, 13 different gRNAs or crRNAs are encoded by the first DNA. In an example, the target cells are  Salmonella  cells (eg, wherein the subject is a chicken). In an example, the target cells are  E. coli  cells. In an example, the target cells are  Campylobacter  cells (eg, wherein the subject is a chicken). In an example, the target cells are Edwardsiella cells (eg, wherein the subject is a fish or shellfish, eg, a catfish or a shrimp or prawn). In an example, the target cells are  E. coli  cells. 
     In an alternative herein, the carrier and target cells are archaeal cells. In an alternative herein, the carrier and target cells are yeast cells and “conjugation” is to be read instead as referring to yeast “mating” and, eg, the second DNA is comprised by a chromosome of the carrier yeast cell. 
     In a preferred example, the NOI encodes an antibacterial agent that is toxic to a target bacterial cell but is not toxic to the carrier cell. In an example, the POI is an antibiotic agent, an antibody, an antibody chain or an antibody variable domain. In an example, the ROI is a guide RNA or a crRNA that is operable in the target cell with a cognate Cas (eg, a Cas nuclease to target and cut a protospacer sequence comprised by a chromosome or episome of the target cell). In an example the RNA is a siRNA that is capable of hybridizing to an endogenous target nucleic acid sequence of the target cell to silence transcription and/or translation thereof. 
     The the carrier cell comprises a chromosome or second episomal DNA which is different from the first DNA. Thus, for example the second DNA is comprised by a chromosome of the carrier cell. In an other example, the second DNA is comprised by a plasmid of the carrier cell.
     2. The carrier cell of Embodiment 1, wherein
       (d) the first DNA is devoid of a component required for conjugative transfer of the first DNA into a target bacterial cell; and   (e) the carrier cell comprises said component, wherein the component is comprised by or encoded by the second DNA or a third DNA comprised by the carrier cell.   
       

     The third DNA may be comprised by a chromosome or episome (eg, plasmid). Where explicitly recited in Embodiments 2 onwards, the component is referring to the component required for conjugative transfer, as the context makes clear.
     3. The carrier cell of Embodiment 2, wherein the component is comprised by a Mpf or Dtr module. For example, the module is a RK2, RP4 or R6K tra module. Optionally, the component is comprised by a Mpf (mating pair formation) module, eg, a RK2 or RP4 tra1 module or a homologue thereof. Optionally, the component is comprised by a Dtr (DNA-transfer replication) module, eg, a RK2 or RP4 tra2 module or a homologue thereof.   4. The carrier cell of Embodiment 2, wherein the component is encoded by an operon of a Mpf (eg, tra1) or Dtr (eg, tra2) module.   

     For example, the module is a RK2, RP4 or R6K tra module. Optionally, the module is a Mpf (mating pair formation) module, eg, a RK2 or RP4 tra1 module or a homologue thereof. Optionally, the module is a Dtr (DNA-transfer replication) module, eg, a RK2 or RP4 tra2 module or a homologue thereof. 
     The Mpf operon of RP4 or RK2 is the tra2 operon (comprising the trb genes trbBCDEFGHIJKL) together with gene traF. The gene traF is comprised in RK2 or RP4 in a tra1 operon (along with traJXIHG). Thus, in an embodiment, the component is a gene selected from trbB, trbC, trbD, trbE, trbF, trbG, trbH, trbI, trbJ, trbK, trbL and traF. In an embodiment the first DNA is devoid of two or more genes selected from RP4 trbB, trbC, trbD, trbE, trbF, trbG, trbH, trbI, trbJ, trbK, trbL and traF or homologues thereof. In an embodiment the first DNA is devoid of two or more genes selected from RK2 trbB, trbC, trbD, trbE, trbF, trbG, trbH, trbI, trbJ, trbK, trbL and traF or homologues thereof. In an example, the component is a traK, traL or traM gene. In an embodiment the first DNA is devoid of two or more genes selected from RP4 traK, traL and traM or homologues thereof. In an embodiment the first DNA is devoid of two or more genes selected from RK2 traK, traL and traM or homologues thereof. Optionally, in these embodiments the first DNA is comprised by a RP4 or RK2-type plasmid. 
     R6K Mpf is encoded by the sltX1tivB1B2B3-4B5B6B7B8B9B10B11 operon that also includes clpX1 of the Dtr module. Thus, in an embodiment, the component is a gene comprises by the RK6 sltX1tivB1B2B3-4B5B6B7B8B9B10B11 operon. In an embodiment, the the component is clpX1 of the RK6 Dtr module. In an embodiment, the component is a gene selected from clpX1 and dtrX1rlxX1. In an embodiment, the component is a gene selected from a RK6 clpX1 and dtrX1rlxX1 or homologues thereof. Optionally, in these embodiments the first DNA is comprised by a RK6-type plasmid. 
     The carrier cell of any preceding Embodiment, wherein the carrier cell chromosome and/or an episome of the carrier cell (other than an episome comprising the first DNA) comprises an expressible tra1 and/or tra2 module or a homologue thereof. 
     Any episome herein may be a plasmid.
     5. The carrier cell of any preceding Embodiment, wherein the carrier cell chromosome and/or an episome of the carrier cell (other than an episome comprising the first DNA) comprises an expressible operon of a tra1 and/or tra2 module or a homologue thereof.   6. The carrier cell of any one of Embodiments 3 to 6, wherein the component is tra1 component.   7. The carrier cell of any one of Embodiments 3 to 6, wherein the component is tra2 component.   8. The carrier cell of any one of Embodiments 2 to 7, wherein the carrier cell chromosome encodes the first factor and comprises said component.   9. The carrier cell of any preceding Embodiment, wherein the first factor is a rep protein, optionally wherein the protein is encoded by pir or trfA or a homologue thereof.   10. The carrier cell of any preceding Embodiment, wherein the first DNA is comprised by a RK2 or R6K plasmid.   11. The carrier cell of any preceding Embodiment, wherein the first DNA comprises an oriV of a RK2 or R6K plasmid, or a homologue thereof.   12. The carrier cell of any preceding Embodiment, wherein the first DNA comprises an oriT of a RK2 or R6K plasmid, or a homologue thereof.   13. The carrier cell of any preceding Embodiment, wherein the first DNA is comprised by a plasmid.   14. The carrier cell of any preceding Embodiment, wherein the second DNA is comprised by a plasmid or is comprised by a chromosome of the carrier cell.   15. The carrier cell of any preceding Embodiment, wherein the agent comprises one or more components of a CRISPR/Cas system that is operable in the target cell to cut a protospacer sequence comprised by the target cell, eg, wherein the protospacer sequence is comprised by the cell chromosome.   

     In an embodiment, the cutting herein kills the target cell. In another embodiment, the cutting inhibits the growth or proliferation of the target cell.
     16. The carrier cell of any preceding Embodiment, wherein the agent encodes a guide RNA or crRNA of a CRISPR/Cas system that is operable with a Cas nuclease in the target cell to cut a protospacer sequence comprised by the target cell, eg, wherein the protospacer sequence is comprised by the cell chromosome.   

     In an example, the target cell is a  Salmonella  cell and the protospacer is comprised by a pipA, pipB, pipC, hilA, sicP, mart or sopB gene. In an example, the protospacer is comprised by a gene that is a homologue or orthologue of a  Salmonella  sicP, sseF, pipA, pipB, pipC, hilA, sicP, mart or sopB gene.
     17. The carrier cell of any preceding Embodiment, wherein the first DNA comprises a gene that encodes a product, wherein the product is essential for survival or proliferation of the carrier cell when in an environment that is devoid of the product, wherein the carrier cell chromosome does not comprise an expressible gene encoding the product and optionally the first DNA is the only episomal DNA comprised by the carrier cell that encodes the product.   18. The carrier cell of Embodiment 17, wherein the gene is selected from an aroA, argH, hisD, leuB, lysA, metB, proC, thrC, pheA, tyrA, trpC and pflA gene; or wherein the gene is an anti-toxin gene and optionally the first DNA encodes a cognate toxin.   19. The carrier cell of any preceding Embodiment, wherein the carrier cell is an  E. coli  (eg, Nissle, F18 or S17  E. coli  strain),  Bacillus  (eg,  B subtilis ),  Enterococcus  or  Lactobacillus  cell. Optionally, the carrier cell is a cell of a human, chicken pig, sheep, cow, fish (eg, catfish or salmon) or shellfish (eg, shrimp or lobster) commensal bacterial strain (eg, a commensal  E. coli  strain).   20. The carrier cell of any preceding Embodiment, wherein the carrier cell is for administration to a microbiota of a human or animal subject for medical use.   

     For example, the medical use is for treating or preventing a disease disclosed herein. For example, the medical use is for treating or preventing a condition disclosed herein.
     21. The carrier of Embodiment 20, wherein the medical use is for the treatment or prevention of a disease or condition mediated by said target cells.   22. The carrier of any one of Embodiments 1 to 20 for administration to an animal for enhancing growth or weight of the animal.   

     In alternative, the administration is to a human for enhancing the growth or weight of the human. Optionally, the enhancing is not a medical therapy. Optionally, the enhancing is a medical therapy.
     23. The carrier of any one of Embodiments 20 to 22, wherein the use comprises the administration of a plurality of carrier cells to a microbiota (eg, a gut microbiota) of the subject, wherein the microbiota comprises target cells and first DNA is transferred into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the subject or reducing the growth or proliferation of target cells.   24. A DNA encoding an antibacterial agent that is toxic to a target bacterial cell but is not toxic to a carrier cell, wherein the first DNA comprises an origin of replication but does not comprise or encode a first factor required for replication of the first DNA, and wherein the first DNA comprises an origin of transfer but is devoid of a component required for conjugative transfer of the first DNA into a target bacterial cell, wherein when in the presence of the component the DNA is capable of conjugative transfer into the target cell.   

     Thus, when the DNA is in a carrier cell, the DNA is capable of conjugative transfer into a target cell.
     25. The DNA of Embodiment 24, wherein the DNA is according to the DNA recited in any one of Embodiments 1 to 23.   26. The DNA of Embodiment 24 or 25, wherein the cells are according to the cells recited in any one of Embodiments 1 to 23.   27. A method for enhancing growth or weight of an animal subject (eg, a chicken), wherein the method comprises the administration of a plurality of carrier cells according to any one of Embodiments 1 to 23 to a microbiota of the subject, wherein the microbiota comprises target cells and first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells (eg,  Salmonella  cells) in the subject or reducing the growth or proliferation of target cells.   

     In alternative, the administration is to a human for enhancing the growth or weight of the human. Optionally, the enhancing is not a medical therapy. Optionally, the enhancing is a medical therapy.
     28. A method for enhancing growth or weight of a plant (eg, a tomato plant), wherein the method comprises the administration of a plurality of carrier cells according to any one of Embodiments 1 to 23 to a microbiota of the plant, wherein the microbiota comprises target cells and first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells (eg,  Pseudomonas  cells) in the plant or reducing the growth or proliferation of target cells.   

     The plant may be any plant disclosed herein. For example a plant herein in any configuration or embodiment of the invention is selected from a tomato plant, a potato plant, a wheat plant, a corn plant, a maize plant, an apple tree, a bean-producing plant, a pea plant, a beetroot plant, a stone fruit plant, a barley plant, a hop plant and a grass. For example, the plant is a tree, eg, palm, a horse chestnut tree, a pine tree, an oak tree or a hardwood tree. For example the plant is a plant that produces fruit selected from strawberries, raspberries, blackberries, reducrrants, kiwi fruit, bananas, apples, apricots, avoocados, cherries, oranges, clementines, satsumas, grapefruits, plus, dates, figs, limes, lemons, melons, mangos, pears, olives or grapes. Optionally, the plant is a dicotyledon. Optionally, the plant is a flowering plant. Optionally, the plant is a monocotyledon. 
     In any configuration, embodiment or example herein, the target bacteria are  P. syringae  bacteria (eg, comprised by a plant).  Pseudomonas syringae  pv.  syringae  is a common plant-associated bacterium that causes diseases of both monocot and dicot plants worldwide. In an example the targt bacteria are  P. syringae  bacteria of a pathovar selected from P. s. pv.  aceris , P. s. pv.  aptata , P. s. pv.  atrofaciens,  
         P. s. pv.  dysoxylis , P. s. pv.  japonica , P. s. pv.  lapsa , P. s. pv.  panici , P. s. pv.  papulans , P. s. pv.  pisi,      P. s. pv.  syringae  and P. s. pv.  morsprunorum.      P. s. pv.  aceris  attacks maple Acer species.   P. s. pv.  actinidiae  attacks kiwifruit  Actinidia deliciosa.      P. s. pv.  aesculi  attacks horse chestnut  Aesculus hippocastanum , causing bleeding canker.   P. s. pv.  aptata  attacks beets  Beta vulgaris.      P. s. pv.  atrofaciens  attacks wheat  Triticum aestivum.      P. s. pv.  dysoxylis  attacks the kohekohe tree  Dysoxylum spectabile.      P. s. pv.  japonica  attacks barley  Hordeum vulgare.      P. s. pv.  lapsa  attacks wheat  Triticum aestivum.      P. s. pv.  panici  attacks  Panicum  grass species.   P. s. pv.  papulans  attacks crabapple  Malus sylvestris  species.   P. s. pv.  phaseolicola  causes halo blight of beans.   P. s. pv.  pisi  attacks peas  Pisum sativum.      P. s. pv.  syringae  attacks  Syringa, Prunus , and  Phaseolus  species.   P. s. pv.  glycinea  attacks soybean, causing bacterial blight of soybean.       

     In an example, the target bacteria are  P. syringae  selected from a serovar recited in a bullet point in the immediately preceding paragraph and the bacteria are comprised by a plant also mentioned in that bullet point. 
     In an example, the weight is dry weight. For example, the method is for increasing dry weight (eg, within 1 or 2 weeks of said administration). Optionally, the increase is an increase of at least 5, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20% compared to a control plant of the same species or strain to which the administration if carrier cells has not taken place, wherein all plants are kept under the same environmental conditions. For example, such an increase is within 1, 2, 3, 4, 5, 6, or 8 weeks following the first administration of the carrier cells. In an example, the method is for increasing the dry weight of a leaf and/or fruit of the plant, such as a tomato plant. 
     In an example, the weight is wet weight. For example, the method is for increasing wet weight (eg, within 1 or 2 weeks of said administration). Optionally, the increase is an increase of at least 5, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20% compared to a control plant of the same species or strain to which the administration if carrier cells has not taken place, wherein all plants are kept under the same environmental conditions. For example, such an increase is within 1, 2, 3, 4, 5, 6, or 8 weeks following the first administration of the carrier cells. In an example, the method is for increasing the dry weight of a leaf and/or fruit of the plant, such as a tomato plant.
     29. The method of Embodiment 28, wherein the microbiota is comprised by a leaf, trunk, root or stem of the plant.   

     In an example, in any configuration or embodiment herein the target bacteria (or taraget cell) is comprised by a microbiota of a plant. In an example, the microbiota is comprised by a leaf. In an example, the microbiota is comprised by a xylem. In an example, the microbiota is comprised by a phloem. In an example, the microbiota is comprised by a root. In an example, the microbiota is comprised by a tuber. In an example, the microbiota is comprised by a bulb. In an example, the microbiota is comprised by a seed. In an example, the microbiota is comprised by an exocarp, epicarp, mesocarp or endocarp. In an example, the microbiota is comprised by a fruit, eg, a simple fruits; aggregate fruits; or multiple fruits. In an example, the microbiota is comprised by a seed or embryo, eg, by a seed coat; a seed leaf; cotyledons; or a radicle. In an example, the microbiota is comprised by a flower, eg, comprised by a peduncle; sepal: petals; stamen; filament; anther or pistil. In an example, the microbiota is comprised by a root; eg, a tap root system, or a fibrous root system. In an example, the microbiota is comprised by a leaf or leaves, eg, comprised by a leaf blade, petiole or stipule. In an example, the microbiota is comprised by a stem, eg, comprised by bark, epidermis, phloem, cambium, xylem or pith.
     30. A method for reducing a biofilm comprised by a subject or comprised on a surface, wherein the biofilm comprises target cells (eg,  Pseudomonas  cells), wherein the method comprises the administration of a plurality of carrier cells according to any one of Embodiments 1 to 23 to the biofilm, wherein first DNA is transferred from carrier cells into target cells for expression therein to produce the antibacterial agent, thereby killing target cells in the biofilm or reducing the growth or proliferation of target cells.   

     In an example “reducing a biofilm” comprises reducing the coverage area of the biofilm. In an example “reducing a biofilm” comprises reducing the proliferation of the biofilm. In an example “reducing a biofilm” comprises reducing the durability of the biofilm. In an example “reducing a biofilm” comprises reducing the spread of the biofilm (eg, in or on the subject, eg, spread to the environment containing the subject).
     31. The method of Embodiment 30, wherein the subject is a human or animal.   

     For example, the biofilm is comprised by a lung of the subject, eg, wherein the target cells are  Pseudomonas  (eg,  P. aeruginosa ) cells. This may be useful wherein the subject is a human suffering from a lung disease or condition, such as pneumonia or cystic fibrosis. For example, the biofilm is comprised by an animal or human organ disclosed herein. For example, the biofilm is comprised by a microbiota of a human or animal disclosed herein.
     32. The method of Embodiment 30, wherein the subject is a plant (eg, any plant disclosed herein).   

     Optionally, in this Embodiment the target cells are  Pseudomonas syringae  cells.
     33. The method of Embodiment 31 or 32, wherein the biofilm is comprised by a leaf, trunk, root or stem of the plant.   

     In an example, in any configuration or embodiment herein the target bacteria (or taraget cell) is comprised by a biofilm of a plant. In an example, the biofilm is comprised by a leaf. In an example, the biofilm is comprised by a xylem. In an example, the biofilm is comprised by a phloem. In an example, the biofilm is comprised by a root. In an example, the biofilm is comprised by a tuber. In an example, the biofilm is comprised by a bulb. In an example, the biofilm is comprised by a seed. In an example, the biofilm is comprised by an exocarp, epicarp, mesocarp or endocarp. In an example, the biofilm is comprised by a fruit, eg, a simple fruits; aggregate fruits; or multiple fruits. In an example, the biofilm is comprised by a seed or embryo, eg, by a seed coat; a seed leaf; cotyledons; or a radicle. In an example, the biofilm is comprised by a flower, eg, comprised by a peduncle; sepal: petals; stamen; filament; anther or pistil. In an example, the biofilm is comprised by a root; eg, a tap root system, or a fibrous root system. In an example, the biofilm is comprised by a leaf or leaves, eg, comprised by a leaf blade, petiole or stipule. In an example, the biofilm is comprised by a stem, eg, comprised by bark, epidermis, phloem, cambium, xylem or pith.
     34. The method of any one of Embodiments 30 to 33 (eg, 30), wherein the surface is a surface ex vivo, such as a surface comprised by a domestic or industrial apparatus or container.   35. A method of replicating a first DNA to produce a plurality of copies of said DNA, the method comprising culturing a plurality of carrier bacterial cells according to any one of Embodiments 1 to 23, wherein the first DNA is replicated in the cells.   

     Optionally, the method further comprises isolating first DNA after said culturing. The skilled addresse is familiar with techniques and conditions for culturing cells that can be used.
     36. The method of Embodiment 35, comprising
       (a) obtaining a sample of the carrier cells after culturing;   (b) contacting the sample of carrier cells with a plurality of target bacterial cells to allow conjugation between carrier cells and target cells; and   (c) allowing copies of first DNA to be transferred by conjugative transfer from carrier cells to target cells, wherein the antibacterial agent is provided in target cells and target cells are killed or the growth or proliferation of target cells is reduced;   (d) wherein the first DNA is not replicable in the target cells.   
       

     For example, the first DNA is not substantially replicable in the target cells, eg, less than 10, 5, 4, 3, 2 or 1% replication). 
     The first DNA is not (or not substantially) replicable in the target cells due to the absence of the first factor or sequence encoding it in the target cells.
     37. A method of killing a plurality of target bacterial cells or reducing the growth or proliferation thereof, the method comprising
       (a) obtaining a sample of the carrier cells according to any one of Embodiments 1 to 23 or obtainable by the method of Embodiment 35;   (b) contacting the sample of carrier cells with the plurality of target bacterial cells to allow conjugation between carrier cells and target cells; and   (c) allowing copies of first DNA to be transferred by conjugative transfer from carrier cells to target cells, wherein the antibacterial agent is provided in target cells and target cells are killed or the growth or proliferation of target cells is reduced;   (d) wherein the first DNA is not (or not substantially) replicable in the target cells.   
       38. The method of Embodiment 36 or 37, wherein the target cells are comprised by a biofilm, eg, a biofilm as disclosed herein.   39. A method for containing the action of an antibacterial agent in an environment, wherein the agent is toxic to target bacterial cells, the method comprising carrying out the method of Embodiment 37 or 38 and the agent is said agent recited of the Embodiment.   40. The method of Embodiment 39, wherein the environment is comprised by a human or animal subject and the target cells are comprised by a biofilm of the subject, wherein the method comprises administering the sample of carrier cells to the biofilm of the subject, wherein the carrier cells are contacted with the target cells in step (b), wherein the method is a contained method for treating or preventing a disease or condition mediated by target cells in the subject.   41. The carrier cell, DNA or method of any preceding Embodiment, wherein the target bacteria are  Salmonella, Pseudomonas, Escherichia, Klebsiella, Campylobacter, Helicobacter, Acinetobacter, Enterobacteriacea, Clostridium, Staphylococcus  or  Streptococcus  bacteria.   42. The carrier cell, DNA or method of any preceding Embodiment, wherein the target bacteria are  Salmonella enterica  bacteria.   

     For example, the target bacteria are selected from the group consisting of  Salmonella enterica  subsp.  enterica , serovars  Typhimurium, Enteritidis , Virchow, Montevideo, Hadar and Binza.
     43. The carrier cell, DNA or method of any one of Embodiments 1 to 41, wherein the target bacteria are  Pseudomonas  (eg,  P. syringae  or  P. aeruginosa ) bacteria.   44. The carrier cell or method of any one of Embodiments 1 to 41, wherein the target bacteria are  E coli  bacteria.   

     Optionally, the target bacteria are enterohemorrhagic  E. coli  (EHEC),  E. coli  Serotype O157:H7 or Shiga-toxin producing  E. coli  (STEC)). In an example, the taraget bacteria are selected from
         Shiga toxin-producing  E. coli  (STEC) (STEC may also be referred to as Verocytotoxin-producing  E. coli  (VTEC);   Enterohemorrhagic  E. coli  (EHEC) (this pathotype is the one most commonly heard about in the news in association with foodborne outbreaks);   Enterotoxigenic  E. coli  (ETEC);   Enteropathogenic  E. coli  (EPEC);   Enteroaggregative  E. coli  (EAEC);   Enteroinvasive  E. coli  (EIEC); and   Diffusely adherent  E. coli  (DAEC).       

     Enterohemorrhagic  Escherichia coli  (EHEC) serotype O157:H7 is a human pathogen responsible for outbreaks of bloody diarrhoea and haemolytic uremic syndrome (HUS) worldwide. Conventional antimicrobials trigger an SOS response in EHEC that promotes the release of the potent Shiga toxin that is responsible for much of the morbidity and mortality associated with EHEC infection. Cattle are a natural reservoir of EHEC, and approximately 75% of EHEC outbreaks are linked to the consumption of contaminated bovine-derived products. EHEC causes disease in humans but is asymptomatic in adult ruminants. Characteristics of  E. coli  serotype O157:H7 (EHEC) infection includes abdominal cramps and bloody diarrhoea, as well as the life-threatening complication haemolytic uremic syndrome (HUS). Currently there is a need for a treatment for EHEC infections (Goldwater and Bettelheim, 2012). The use of conventional antibiotics exacerbates Shiga toxin-mediated cytotoxicity. In an epidemiology study conducted by the Centers for Disease Control and Prevention, patients treated with antibiotics for EHEC enteritis had a higher risk of developing HUS (Slutsker et al., 1998). Additional studies support the contraindication of antibiotics in EHEC infection; children on antibiotic therapy for hemorrhagic colitis associated with EHEC had an increased chance of developing HUS (Wong et al., 2000; Zimmerhackl, 2000; Safdar et al., 2002; Tarr et al., 2005). Conventional antibiotics promote Shiga toxin production by enhancing the replication and expression of stx genes that are encoded within a chromosomally integrated lambdoid prophage genome. The approach of some configurations of present invention rely on nuclease cutting. Stx induction also promotes phage-mediated lysis of the EHEC cell envelope, allowing for the release and dissemination of Shiga toxin into the environment (Karch et al., 1999; Matsushiro et al., 1999; Wagner et al., 2002). Thus, advantageously, these configurations of the invention provide alternative means for treating EHEC in human and animal subjects. This is exemplified below with surprising results on the speed and duration of anti-EHEC action produced by nuclease action (as opposed to conventional antibiotic action). 
     In an example, the subject (eg, a human or animal) is suffering from or at risk of haemolytic uremic syndrome (HUS), eg, the subject is suffering from an  E. coli  infection, such as an EHEC  E. coli  infection.
     45. A pharmaceutical composition, livestock growth promoting composition, soil improver, herbicide, plant fertilizer, food or food ingredient sterilizing composition, dental composition, personal hygiene composition or disinfectant composition (eg, for domestic or industrial use) comprising a plurality of carrier cells according to any one of Embodiments 1 to 23.   

     Herein, a carrier cell is, eg, a probiotic cell for administration to a human or animal subject. For example, the carrier cell is commensal in a microbiome (eg, gut or blood microbiome) of a human or animal subject, wherein the carrier is for administration to the subject. In an example, a carrier cell is a bacterial cell (and optionally the target cell is a bacterial cell). In an example, a carrier cell is an archaeal cell (and optionally the target cell is an archaeal cell) Optionally, the carrier cell is a gram-positive bacterial cell and the target cell is a gram-positive bacterial cell. 
     Optionally, the carrier cell is a gram-positive bacterial cell and the target cell is a gram-negative bacterial cell. 
     Optionally, the carrier cell is a gram-negative bacterial cell and the target cell is a gram-positive bacterial cell. 
     Optionally, the carrier cell is a gram-negative bacterial cell and the target cell is a gram-negative bacterial cell. 
     Optionally, the carrier cell is a  Bacillus  bacterial cell and the target cell is a gram-positive bacterial cell. 
     Optionally, the carrier cell is a  Bacillus  bacterial cell and the target cell is a gram-netative bacterial cell. 
     Optionally, the carrier cell is a  Bacillus  bacterial cell and the target cell is a  Salmonella  bacterial cell. 
     Optionally, the carrier cell is a  Bacillus  bacterial cell and the target cell is an  E. coli  bacterial cell. 
     Optionally, the carrier cell is an  E. coli  bacterial cell and the target cell is a  Pseudomonas  bacterial cell. 
     Optionally, the carrier cell is an  E. coli  bacterial cell and the target cell is a gram-positive bacterial cell. 
     Optionally, the carrier cell is an  E. coli  bacterial cell and the target cell is a gram-netative bacterial cell. 
     Optionally, the carrier cell is an  E. coli  bacterial cell and the target cell is a  Salmonella  bacterial cell. 
     Optionally, the carrier cell is an  E. coli  bacterial cell and the target cell is an  E. coli  bacterial cell. 
     Optionally, the carrier cell is an  E. coli  bacterial cell and the target cell is a  Pseudomonas  bacterial cell. 
     A  Bacillus  cell herein is optionally a  B subtilis  cell. 
     Optionally, the carrier cell is a probiotic or commensal  E. coli  bacterial cell for administration to a human or animal subject. Optionally, the carrier cell is a probiotic or commensal  Bacillus  bacterial cell for administration to a human or animal subject. 
     Herein, optionally the first DNA is comprised by a plasmid, eg, a closed circular DNA. 
     In an embodiment, in an example the first DNA is dsDNA. In an embodiment, in an example the first DNA is ssDNA. 
     In an alternative configuration, the first DNA is instead a first RNA. 
     Optionally, the target cell is a  Salmonella  cell (eg, wherein the carrier cell is an  E. coli  cell), eg, a  Salmonella enterica  subsp.  enterica , eg, a  Salmonella enterica  subsp.  enterica  serovar  Typhimurium, Enteritidis , Virchow, Montevideo, Hadar or Binza. 
     For example, the target bacteria are selected from the group consisting of  S. enterica; S. typhimurium; P. aeruginosa; E. coli; K. pneumoniae; C. jujeni; H. pylori; A. baumanii; C. difficile; S. aureus; S. pyogenes  or  S. thermophilus.    
     In an example, the target cell is a cell of a species that causes nosocomial infection in humans. 
     Optionally, the target cell is comprised by an animal (eg, poultry animal (such as chicken), swine, cow, fish (eg, catfish or salmon) or shellfish (eg, prawn or lobster)) microbiome. Optionally, the microbiome is a gut microbiome. For example, the target cell is a  Salmonella  cell comprised by a chicken gut biofilm. For example, the target cell is a  Salmonella  cell comprised by a chicken gut biofilm sample ex vivo. 
     In an embodiment, the first DNA comprises a bacterial oriV and/or an oriT. In an embodiment, the first DNA is comprised by a plasmid, wherein the plasmid comprises and oriV and/or an oriT. 
     The first factor may be a protein or RNA. For example, the first factor is pir or trfA. In an example the first factor is operable with an oriV comprised by the first DNA for replication thereof. 
     In an embodiment, the first DNA is comprised by a plasmid, wherein the plasmid comprises an oriV and does not encode any replication protein (eg, pir or trfA) that is operable with the oriV to initiate replication of the plasmid. 
     In an embodiment, the first DNA is devoid of a component required for conjugative transfer of the first DNA into a target bacterial cell. Optionally, the component is a protein. Optionally, the component is a RNA. Optionally, the component is a tra1 component. Optionally, the component is a tra2 component. In an example, the carrier cell comprises said component, wherein the component is comprised by or encoded by the carrier cell chromosome, the second DNA or a third DNA in the carrier cell. Preferably, the component is comprised by or encoded by the carrier cell chromosome. 
     Optionally, the component required for conjugative transfer is a RP4 plasmid component, eg, a component of a RP4 tra module (eg, tra1 or tra2 module). 
     Optionally, the component required for conjugative transfer is a RK2 plasmid component, eg, a component of a RK2 tra module (eg, tra1 or tra2 module). 
     Optionally, the component required for conjugative transfer is a R6K plasmid component, eg, a component of a R6K tra module (eg, tra1 or tra2 module). 
     Optionally, the first DNA is comprised by a plasmid that does not comprise an antibiotic resistance marker gene and/or a plasmid addiction system gene. As explained in the Examples section, this may be advantageous in the rare case of an identical IncP plasmid being present in the target cell that will provide trfA (or other component required for conjugation where that component is not encoded by the plasmid comprising the first DNA), both plasmids will compete for the available TrfA (or other component), resulting in loss of one plasmid, and thus will be quickly lost from any offspring target cells. 
     Optionally, the plasmid comprising the first DNA further comprises an anti-restriction gene encoding a product for inhibiting a restriction enzyme of the target cell (eg, an Type I restriction enzyme). as anti-restriction genes that inhibit Type I restriction enzymes (eg, the anti-restriction gene product is an ocr of T7, klcA of RK2, ard or ardB). 
     Additionally or alternatively, the plasmid comprises a gene encoding an essential component of a type IV secretion system, wherein the chromosome of the carrier cell comprises genes encoding the remainder of the secretion system, wherein the essential component is required for conjugative transfer of the plasmid from the carrier cell to the target cell. For example, with the exception of a tra1 or tra2 gene encoding the essential component, the chromosome comprises all of the remainder of traKLM, traJXIHGF genes of RK2 tra1 and all of the remainder of trbBCEFGHJL genes of RK2 tra2 (preferably when the plasmid is a RK2-type plasmid). In an alternative example, with the exception of a gene encoding the essential component, the chromosome comprises all of the reaminder of RK6 homologues of traKLM, traJXIHGF genes of RK2 tra1 and all of the remainder of RK6 homologues of trbBCEFGHJL genes of RK2 tra2 (preferably when the plasmid is a RK6-type plasmid). In an alternative example, with the exception of a gene encoding the essential component, the chromosome comprises all of the reaminder of RP4 homologues of traKLM, traJXIHGF genes of RK2 tra1 and all of the remainder of RP4 homologues of trbBCEFGHJL genes of RK2 tra2 (preferably when the plasmid is a RP4-type plasmid). 
     For example, with the exception of a tra1 or tra2 gene encoding the essential component, the chromosome comprises all of the remainder of a DNA fragment of the RK2 plasmid from traF to traM and all of the remainder of a DNA fragment of the RK2 plasmid from trbB to trK (preferably when the plasmid is a RK2-type plasmid). For example, with the exception of a tra1 or tra2 gene encoding the essential component, the chromosome comprises all of the remainder of the RK2 plasmid genes from traF to traM and all of the remainder of the RK2 plasmid genes from trbB to trK (preferably when the plasmid is a RK2-type plasmid). For example, the chromosome comprises (in the following order 5′ to 3′, or 3′ to 5′) the following RK2 tra1 genes: traFGHIXJKLM, or traKLM, or traJXIHGF. For example, the chromosome comprises (in the following order 5′ to 3′, or 3′ to 5′) the following RK2 tra2 genes: trbBCDEFGHIJKL or trbBCEFGHJL. 
     For example, with the exception of a gene encoding the essential component, the chromosome comprises all of the remainder of a DNA fragment of the RK6 plasmid from a homologue of RK2 traF to a homologue of RK2 traM and all of the remainder of a DNA fragment of the RK6 plasmid from a homologue of RK2 trbB to a homologue of a RK2 trK (preferably when the plasmid is a RK6-type plasmid). For example, with the exception of a gene encoding the essential component, the chromosome comprises all of the remainder of the RK6 plasmid genes from a homologue of RK2 traF to a homologue of RK2 traM and all of the remainder of the RK6 plasmid genes from a homologue of RK2 trbB to a homologue of RK2 trK (preferably when the plasmid is a RK6-type plasmid). For example, the chromosome comprises (in the following order 5′ to 3′, or 3′ to 5′) RK6 homologues of the following RK2 tra1 genes: traFGHIXJKLM, or traKLM, or traJXIHGF. For example, the chromosome comprises (in the following order 5′ to 3′, or 3′ to 5′) RK6 homologues of the following RK2 tra2 genes: trbBCDEFGHIJKL or trbBCEFGHJL. 
     For example, with the exception of a gene encoding the essential component, the chromosome comprises all of the remainder of a DNA fragment of the RP4 plasmid from a homologue of RK2 traF to a homologue of RK2 traM and all of the remainder of a DNA fragment of the RP4 plasmid from a homologue of RK2 trbB to a homologue of a RK2 trK (preferably when the plasmid is a RP4-type plasmid). For example, with the exception of a gene encoding the essential component, the chromosome comprises all of the remainder of the RP4 plasmid genes from a homologue of RK2 traF to a homologue of RK2 traM and all of the remainder of the RP4 plasmid genes from a homologue of RK2 trbB to a homologue of RK2 trK (preferably when the plasmid is a RP4-type plasmid). For example, the chromosome comprises (in the following order 5′ to 3′, or 3′ to 5′) RP4 homologues of the following RK2 tra1 genes: traFGHIXJKLM, or traKLM, or traJXIHGF. For example, the chromosome comprises (in the following order 5′ to 3′, or 3′ to 5′) RP4 homologues of the following RK2 tra2 genes: trbBCDEFGHIJKL or trbBCEFGHJL. 
     Preferably, the second DNA or a chromosome comprising the second DNA is devoid of a oriT. Preferably, the third DNA or a chromosome comprising the third DNA is devoid of a oriT. For example, the oriT is only comprised by the first DNA (or plasmid comprising this) and is not comprised by any other DNA in the carrier cell. Thus, transfer is confined just to the first DNA or its plasmid. 
     The first DNA in all embodiments and configurations may preferably not be comprised by a runaway replication plasmid. Naturally occurring plasmids are present within host cells at a characteristic concentration (referred to herein as a particular plasmid “copy number”). Mutations that destroy the elements of the control cause an over-replication phenotype that manifests itself by an increase in the plasmid copy number (“copy-up” phenotype). In extreme cases of copy-up mutations, plasmid replication is completely unchecked due to the loss of copy control mechanisms. This is referred to as “runaway plasmid replication” or simply “runaway replication” and the plasmid engaging in such runaway replication is a “runaway replication plasmid”. 
     In an example, the invention relates to a composition comprising a plurality of carrier cells of the invention (eg, wherein copies of the first DNA are comprised by respective plasmids). Optionally, all of the carrier cells comprise identical said first and second DNAs. Optionally, the plurality comprises a first sub-population of carrier cells (first cells) and a second sub-population of carrier cells (second cells) wherein the first cells comprise identical first DNAs and the second cells comprise identical first DNAs (which are different from the first DNAs of the first cells). For example, the former DNAs comprise a NSI that is different from the NSI comprised by the other DNAs. For example, the first DNAs encode a first guide RNA or crRNA and the second DNAs encode a second guide RNA or crRNA, wherein the first guide RNA/crRNA is capable of hybridizing to a first protospacer sequence in first target cells; and the second guide RNA/crRNA is capable of hybridizing to a second protospacer sequence in second target cells, wherein the protospacers are different. Optionally, the first target cells are different from the second target cells. Optionally, the first target cells are of the same species or strain as the second target cells. Alternatively, the first target cells are of species or strain that is different from the species or strain of the second target cells (in this way a cocktail of carrier cells is provided, eg, for administration to a human or animal or plant, to target and kill a plurality of target cells of different species or strains). 
     In an example, the or each first DNA comprises a plurality (eg, a first and a second) NSIs wherein a first NSI is different from a second NSI (eg, they encode different proteins or RNAs, such as different guide RNAs or crRNAs). In an example, the or each first DNA comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 different types of NSIs. In an example, the or each first DNA comprises NSIs encoding 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 different guide RNAs. In an example, the or each first DNA comprises NSIs encoding 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 different crRNAs. In an example, the or each first DNA comprises NSIs encoding at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 different guide RNAs. In an example, the or each first DNA comprises NSIs encoding at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 different crRNAs. 
     Optionally, the composition is comprised by a liquid (eg, an aqueous liquid or in water), the composition comprising the carrier cells at an amount of from 1×10 3  to 1×10 10  (eg, from 1×10 4  to 1×10 10 ; from 1×10 4  to 1×10 9 ; from 1×10 4  to 1×10 8 ; from 1×10 4  to 1×10 7 ; from 1×10 3  to 1×10 10 ; from 1×10 3  to 1×10 9 ; from 1×10 3  to 1×10 8 ; from 1×10 3  to 1×10 7 ; from 1×10 6  to 1×10 10 ; from 1×10 5  to 1×10 9 ; from 1×10 5  to 1×10 8 ; from 1×10 5  to 1×10 7 ; from 1×10 6  to 1×10 10 ; from 1×10 6  to 1×10 9 ; from 1×10 6  to 1×10 8 ; or from 1×10 6  to 1×10 7 ) cfu/ml. For example, the liquid is a beverage, such for human or animal consumption. For example, the beverage is a livestock beverage, eg, a poultry beverage (ie, a beverage for consumption by poultry, such as chicken). 
     In an example, the composition is a dietary (eg, dietary supplement) composition for consumption by humans or animals. In an example, the composition is a slimming composition for consumption by humans or animals. In an example, the composition is a growth promotion composition for consumption by humans or animals. In an example, the composition is a body building composition for consumption by humans. In an example, the composition is a probiotic composition for consumption by humans or animals. In an example, the composition is a biocidal composition for consumption by humans or animals. In an example, the composition is a pesticidal composition for consumption by humans or animals. In an example, the composition is a zoonosis control composition for consumption by animals. 
     In an example, the composition comprises vitamins in addition to the carrier cells. In an example, the composition comprises vitamin A, B (eg, B12), C, D, E and/or K in addition to the carrier cells. In an example, the composition comprises lipids in addition to the carrier cells. In an example, the composition comprises carbohydrates in addition to the carrier cells. In an example, the composition comprises proteins and/or amino acids in addition to the carrier cells. In an example, the composition comprises minerals in addition to the carrier cells. In an example, the composition comprises metal ions (eg, Mg 2+ , Cu 2+  and/or Zn 2+ ) in addition to the carrier cells. In an example, the composition comprises sodium ions, potassium ions, magnesium ions, calcium ions, manganese ions, iron ions, cobalt ions, copper ions, zinc ions and/or molybdenum ions. 
     In an example, the composition is a plant fertilizer composition. In an example, the composition is a herbicide. In an example, the composition is a pesticide composition for application to plants. 
     In any embodiment or example, where appropriate: The plants are, for example, crop plants. The plants are, for example, wheat. The plants are, for example, corn. The plants are, for example, maize. The plants are, for example, fruiting plants. The plants are, for example, vegetable plants. The plants are, for example, tomato plants. The plants are, for example, potato plants. The plants are, for example, grass plants. The plants are, for example, flowering plants. The plants are, for example, trees. The plants are, for example, shrubs. 
     In an example, the composition is for environmental application, wherein the environment is an outdoors environment (eg, application to a field or waterway or reservoir). 
     In an example, the composition is comprised by a food or food ingredient (eg, for human or animal consumption). In an example, the composition is comprised by a beverage or beverage ingredient (eg, for human or animal consumption). 
     In an example the target cell(s) are human biofilm cells, eg, wherein the biofilm is a gut, skin, lung, eye, nose, ear, gastrointestinal tract (GI tract), stomach, hair, kidney, urethra, bronchiole, oral cavity, mouth, liver, heart, anus, rectum, bladder, bowel, intestine, penis, vagina or scrotum biofilm. In an example the target cell(s) are animal biofilm cells, eg, wherein the biofilm is a gut, skin, lung, eye, nose, ear, gastrointestinal tract (GI tract), caecum, jejunum, ileum, colon, stomach, hair, feather, scales, kidney, urethra, bronchiole, oral cavity, mouth, liver, spleen, heart, anus, rectum, bladder, bowel, intestine, penis, vagina or scrotum biofilm. For example, the biofilm is a bird (eg, chicken) caecum biofilm. For example, the biofilm is a bird (eg, chicken) gastrointestinal tract (GI tract), caecum, jejunum, ileum, colon or stomach biofilm. 
     In an example, any method herein is ex vivo. In an example, a method herein is in vivo. In an example, a method herein is in vitro. In an example, a method herein is carried out in an environment, eg, in a domestic (such as in a house), industrial (such as in a factory) or agricultural environment (such as in a field). In an example, a method herein is carried out in or on a container; or on a surface. 
     In an example, the NSI (or a RNA product thereof) is capable of recombination with the target cell chromosome or an episome comprised by the target cell to modify the chromosome or episome. Optionally, this is carried out in a method wherein the chromosome or episome is cut (eg, at a predetermined site using a guided nuclease, such as a Cas, TALEN, zinc finger nuclease or meganuclease) and simultaneously or sequentially the first DNA is introduced into the target cell by conjugation with the carrier cell and the NSI or a sequence thereof is inserted into the chromosome or episome at or adjacent the cut site. 
     In an example the first DNA comprises one or more components of a CRISPR/Cas system operable to perform protospacer cutting in the target cell (eg, wherein the protospacer comprises 10-20, 10-30, 10-40, 10-100, 12-15 or 12-20 consecutive nucleotides that are capable of hybridizing in the target cell with a crRNA or gRNA encoded by the NSI). 
     For example, the system is a Type I, II, III, IV or V CRISPR/Cas system. 
     In an example, the NSI encodes a Cas9 (and optionally a second, different, Cas, such as a Cas3, Cas9, Cpf1, Cas13a, Cas13b or Cas10). In an example, the NSI encodes a Cas3 (and optionally a second, different, Cas, such as a Cas3, Cas9, Cpf1, Cas13a, Cas13b or Cas10). In an example, the NSI encodes a Cas selected from a Cas3, Cas9, Cpf1, Cas13a, Cas13b and Cas10. Additionally or alternatively, the first DNA (eg, the NSI) encodes a guide RNA or crRNA or tracrRNA. For example, the guide RNA or crRNA or tracrRNA is cognate to (ie, operable with in the target cell) the first Cas. 
     In an example, a Cas herein is a Cas9. In an example, a Cas herein is a Cas3. The Cas may be identical to a Cas encoded by the target bacteria. 
     In an embodiment, the presence in the target bacterium of the NSI or its encoded protein or RNA mediates target cell killing, or downregulation of growth or propagation of target cells. In an embodiment, the presence in the target bacterium of the NSI or its encoded protein or RNA mediates switching off of expression of one or more RNA or proteins encoded by the target cell genome, or downregulation thereof. 
     In an embodiment, the presence in the target bacterium of the NSI or its encoded protein or RNA mediates upregulation of growth or propagation of the target cell. In an embodiment, the presence in the target bacterium of the NSI or its encoded protein or RNA mediates switching on of expression of one or more RNA or proteins encoded by the target cell genome, or upregulation thereof. 
     In an embodiment, the NSI encodes a component of a CRISPR/Cas system that is toxic to the target bacterium. 
     In an embodiment, the first DNA is comprised by a plasmid or shuttle vector. In an embodiment, the second DNA is comprised by a vector (eg, a plasmid or shuttle vector), helper phage (eg, a helper phagemid) or is integrated in the genome of a host bacterial cell. 
     Optionally, the target cell is devoid of a functional endogenous CRISPR/Cas system before transfer therein of the first DNA, eg, a first DNA comprising component of an exogenous CRISPR/Cas system that is functional in the target cell and toxic to the target cell. An embodiment provides an antibacterial composition comprising a plurality of carrier cells of the invention, wherein each target cell is optionally according to this paragraph, for administration to a human or animal subject for medical use. 
     In an example, the composition of the invention is a herbicide, pesticide, insecticide, plant fertilizer or cleaning agent. 
     Optionally, target bacteria herein are comprised by a microbiome of the subject, eg, a gut microbiome. Alternatively, the microbiome is a skin, scalp, hair, eye, ear, oral, throat, lung, blood, rectal, anal, vaginal, scrotal, penile, nasal or tongue microbiome. 
     In an example the subject (eg, human or animal) is further administered a medicament simultaneously or sequentially with the carrier cell administration. In an example, the medicament is an antibiotic, antibody, immune checkpoint inhibitor (eg, an anti-PD-1, anti-PD-L1 or anti-CTLA4 antibody), adoptive cell therapy (eg, CAR-T therapy) or a vaccine. 
     In an embodiment, the NSI encodes a guided nuclease, such as a Cas nuclease, TALEN, zinc finger nuclease or meganuclease. Thus, the toxic agent may comprise a guided nuclease, such as a Cas nuclease, TALEN, zinc finger nuclease or meganuclease. Optionally, the NSI encodes a restriction nuclease that is capable of cutting the chromosome of the target cell. 
     Optionally, the composition is a pharmaceutical composition for use in medicine practised on a human or animal subject. 
     In an example, the animal is a livestock or companion pet animal (eg, a cow, pig, goat, sheep, horse, dog, cat or rabbit). In an example, the animal is an insect (an insect at any stage of its lifecycle, eg, egg, larva or pupa). In an example, the animal is a protozoan. In an example, the animal is a cephalopod. 
     Optionally, the composition is a herbicide, pesticide, food or beverage processing agent, food or beverage additive, petrochemical or fuel processing agent, water purifying agent, cosmetic additive, detergent additive or environmental (eg, soil) additive or cleaning agent. 
     The invention also provides: A target bacterial cell or a plurality of target bacterial cells each comprising a said first DNA. 
     For example the carrier bacteria are  Lactobacillus  (eg,  L. reuteri  or  L. lactis ),  E. coli, Bacillus  or  Streptococcus  (eg,  S. thermophilus ) bacteria. Usefully, the carrier can provide protection for the first DNA from the surrounding environment. The use of a carrier may be useful for oral administration or other routes where the carrier can provide protection for the first DNA from the acid stomach or other harsh environments in the subject. Furthermore, the carrier can be formulated into a beverage, for example, a probiotic drink, eg, an adapted Yakult (trademark), Actimel (trademark), Kevita (trademark), Activia (trademark), Jarrow (trademark) or similar drink for human consumption. 
     Optionally, the carrier cell(s) or composition are for administration to a human or animal subject for medical use, comprising killing target bacteria using the agent or expression product of the NSI, wherein the target bacteria mediate as disease or condition in the subject. In an example, when the subject is a human, the subject is not an embryo. In an example, the carrier cells are probiotic in the subject. 
     The invention also provides: A method of killing target bacterial cells in an environment, optionally wherein the method is not practised on a human or animal body, wherein the method comprises exposing the environment to the carrier cell(s) or composition of the invention and allowing the product of the NSI to be expressed in the target cells, wherein the target bacteria are killed in the presence of said product. For example, the product encodes a CRISPR/Cas system or component thereof, such as a system or component disclosed herein. Thus, the system may be capable of recognisisng and cutting a chromosomal protopspacer sequence of the target cell, whereby the target cell is killed. Optionally, in a further step killed target cells are isolated. 
     The invention also provides: Use of the composition or cell(s) of the invention, in the manufacture of an antibacterial agent that kills target bacteria, for the treatment of a disease or condition in a human or animal subject comprising the target bacteria. 
     Optionally, the environment is a microbiome of soil; a plant, part of a part (e.g., a leaf, fruit, vegetable or flower) or plant product (e.g., pulp); water; a waterway; a fluid; a foodstuff or ingredient thereof; a beverage or ingredient thereof; a medical device; a cosmetic; a detergent; blood; a bodily fluid; a medical apparatus; an industrial apparatus; an oil rig; a petrochemical processing, storage or transport apparatus; a vehicle or a container. 
     Optionally, the environment is an ex vivo bodily fluid (e.g., urine, blood, blood product, sweat, tears, sputum or spit), bodily solid (e.g., faeces) or tissue of a human or animal subject that has been administered the composition. 
     Optionally, the environment is an in vivo bodily fluid (e.g., urine, blood, blood product, sweat, tears, sputum or spit), bodily solid (e.g., faeces) or tissue of a human or animal subject that has been administered the composition. 
     In an embodiment, the first DNA is comprised by a phagemid or cloning vector (eg, a shuttle vector, eg, a pUC vector). 
     In an embodiment, the second DNA is comprised by the bacterial carrier cell chromosome. 
     Optionally, the toxic agent comprises one or more components of a CRISPR/Cas system, eg, a DNA sequence encoding one or more components of Type I Cascade (eg, CasA). 
     Optionally, the toxic agent comprises a DNA sequence encoding guided nuclease, such as a Cas nuclease, TALEN, zinc finger nuclease or meganuclease. 
     In an example, the carrier cell(s) or composition are comprised by a medical container, eg, a syringe, vial, IV bag, inhaler, eye dropper or nebulizer. In an example, the carrier cell(s) or composition are comprised by a sterile container. In an example, the carrier cell(s) or composition are comprised by a medically-compatible container. In an example, the carrier cell(s) or composition are comprised by a fermentation vessel, eg, a metal, glass or plastic vessel. In an example, the carrier cell(s) or composition are comprised by an agricultural apparatus. In an example, the carrier cell(s) or composition are comprised by food production or processing apparatus. In an example, the carrier cell(s) or composition are comprised by a horticultural apparatus. In an example, the carrier cell(s) or composition are comprised by a farming apparatus. In an example, the carrier cell(s) or composition are comprised by petrochemicals recovery or processing apparatus. In an example, the carrier cell(s) or composition are comprised by a distillation apparatus. In an example, the carrier cell(s) or composition are comprised by cell culture vessel (eg, having a capacity of at least 50, 100, 1000, 10000 or 100000 litres). Additionally or alternatively, the target cell(s) are comprised by any of these apparatus etc. 
     In an example, the carrier cell(s) or composition are comprised by a medicament, e.g in combination with instructions or a packaging label with directions to administer the medicament by oral, IV, subcutaneous, intranasal, intraocular, vaginal, topical, rectal or inhaled administration to a human or animal subject. In an example, the carrier cell(s) or composition are comprised by an oral medicament formulation. In an example, the carrier cell(s) or composition are comprised by an intranasal or ocular medicament formulation. In an example, the carrier cell(s) or composition are comprised by a personal hygiene composition (eg, shampoo, soap or deodorant) or cosmetic formulation. In an example, th the carrier cell(s) or composition are comprised by a detergent formulation. In an example, the carrier cell(s) or composition are comprised by a cleaning formulation, eg, for cleaning a medical or industrial device or apparatus. In an example, the carrier cell(s) or composition are comprised by foodstuff, foodstuff ingredient or foodstuff processing agent. 
     In an example, the carrier cell(s) or composition are comprised by beverage, beverage ingredient or beverage processing agent. In an example, the carrier cell(s) or composition are comprised by a medical bandage, fabric, plaster or swab. In an example, the carrier cell(s) or composition are comprised by a herbicide or pesticide. In an example, the carrier cell(s) or composition are comprised by an insecticide. 
     In an example, the CRISPR/Cas component(s) are component(s) of a Type I CRISPR/Cas system. In an example, the CRISPR/Cas component(s) are component(s) of a Type II CRISPR/Cas system. In an example, the CRISPR/Cas component(s) are component(s) of a Type III CRISPR/Cas system. In an example, the CRISPR/Cas component(s) are component(s) of a Type IV CRISPR/Cas system. In an example, the CRISPR/Cas component(s) are component(s) of a Type V CRISPR/Cas system. In an example, the CRISPR/Cas component(s) comprise a Cas9-encoding nucleotide sequence (eg,  S pyogenes  Cas9,  S. aureus  Cas9 or  S. thermophilus  Cas9). In an example, the CRISPR/Cas component(s) comprise a Cas3-encoding nucleotide sequence (eg,  E. coli  Cas3 , C. dificile  Cas3 or  Salmonella  Cas3). In an example, the CRISPR/Cas component(s) comprise a Cpf-encoding nucleotide sequence. In an example, the CRISPR/Cas component(s) comprise a CasX-encoding nucleotide sequence. In an example, the CRISPR/Cas component(s) comprise a CasY-encoding nucleotide sequence. 
     In an example, each carrier cell encodes a CRISPR/Cas component or protein of interest from a nucleotide sequence (NSI) comprising a promoter that is operable in the target bacteria. 
     Optionally, target bacteria are gram negative bacteria (eg, a  spirilla  or  vibrio ). Optionally, target bacteria are gram positive bacteria. Optionally, target bacteria are  mycoplasma , chlamydiae, spirochete or  mycobacterium  bacteria. Optionally, target bacteria are  Streptococcus  (eg,  pyogenes  or  thermophilus ). Optionally, target bacteria are  Staphylococcus  (eg,  aureus , eg, MRSA). Optionally, target bacteria are  E. coli  (eg, O157: H7), eg, wherein the Cas is encoded by the vecor or an endogenous target cell Cas nuclease (eg, Cas3) activity is de-repressed. Optionally, target bacteria are  Pseudomonas  (eg,  syringae  or  aeruginosa ). Optionally, target bacteria are Vibro (eg,  cholerae  (eg, O139) or  vulnificus ). Optionally, target bacteria are  Neisseria  (eg,  gonnorrhoeae  or  meningitidis ). Optionally, target bacteria are  Bordetella  (eg, pertussis). Optionally, target bacteria are  Haemophilus  (eg,  influenzae ). Optionally, target bacteria are  Shigella  (eg,  dysenteriae ). Optionally, target bacteria are  Brucella  (eg,  abortus ). Optionally, target bacteria are  Francisella  host. Optionally, target bacteria are  Xanthomonas . Optionally, target bacteria are  Agrobacterium . Optionally, target bacteria are  Erwinia . Optionally, target bacteria are  Legionella  (eg,  pneumophila ). Optionally, target bacteria are  Listeria  (eg,  monocytogenes ). Optionally, target bacteria are  Campylobacter  (eg,  jejuni ). Optionally, target bacteria are  Yersinia  (eg,  pestis ). Optionally, target bacteria are  Borelia  (eg,  burgdorferi ). Optionally, target bacteria are  Helicobacter  (eg,  pylori ). Optionally, target bacteria are  Clostridium  (eg,  dificile  or  botulinum ). Optionally, target bacteria are  Erlichia  (eg,  chaffeensis ). Optionally, target bacteria are  Salmonella  (eg,  typhi  or  enterica , eg, serotype  typhimurium , eg, DT 104). Optionally, target bacteria are  Chlamydia  (eg,  pneumoniae ). Optionally, target bacteria are Parachlamydia host. Optionally, target bacteria are  Corynebacterium  (eg,  amycolatum ). Optionally, target bacteria are  Klebsiella  (eg,  pneumoniae ). Optionally, target bacteria are  Enterococcus  (eg,  faecalis  or  faecim , eg, linezolid-resistant). Optionally, target bacteria are  Acinetobacter  (eg,  baumannii , eg, multiple drug resistant). 
     Further examples of target cells are as follows:— 
     1. Optionally the target bacteria are  Staphylococcus aureus  cells, eg, resistant to an antibiotic selected from methicillin, vancomycin, linezolid, daptomycin, quinupristin, dalfopristin and teicoplanin.
 
2. Optionally the target bacteria are  Pseudomonas aeruginosa  cells, eg, resistant to an antibiotic selected from cephalosporins (eg, ceftazidime), carbapenems (eg, imipenem or meropenem), fluoroquinolones, aminoglycosides (eg, gentamicin or tobramycin) and colistin.
 
3. Optionally the target bacteria are  Klebsiella  (eg,  pneumoniae ) cells, eg, resistant to carbapenem.
 
4. Optionally the target bacteria are  Streptococcus  (eg,  thermophilus, pneumoniae  or  pyogenes ) cells, eg, resistant to an antibiotic selected from erythromycin, clindamycin, beta-lactam, macrolide, amoxicillin, azithromycin and penicillin.
 
5. Optionally the target bacteria are  Salmonella  (eg, serotype  Typhi ) cells, eg, resistant to an antibiotic selected from ceftriaxone, azithromycin and ciprofloxacin.
 
6. Optionally the target bacteria are  Shigella  cells, eg, resistant to an antibiotic selected from ciprofloxacin and azithromycin.
 
7. Optionally the target bacteria are  Mycobacterium tuberculosis  cells, eg, resistant to an antibiotic selected from Resistance to isoniazid (INH), rifampicin (RMP), fluoroquinolone, amikacin, kanamycin and capreomycin and azithromycin.
 
8. Optionally the target bacteria are  Enterococcus  cells, eg, resistant to vancomycin.
 
9. Optionally the target bacteria are Enterobacteriaceae cells, eg, resistant to an antibiotic selected from a cephalosporin and carbapenem.
 
10. Optionally the target bacteria are  E. coli  cells, eg, resistant to an antibiotic selected from trimethoprim, itrofurantoin, cefalexin and amoxicillin.
 
11. Optionally the target bacteria are  Clostridium  (eg,  dificile ) cells, eg, resistant to an antibiotic selected from fluoroquinolone antibiotic and carbapenem.
 
12. Optionally the target bacteria are  Neisseria gonnorrhoea  cells, eg, resistant to an antibiotic selected from cefixime (eg, an oral cephalosporin), ceftriaxone (an injectable cephalosporin), azithromycin and tetracycline.
 
13. Optionally the target bacteria are  Acinetoebacter baumannii  cells, eg, resistant to an antibiotic selected from beta-lactam, meropenem and a carbapenem.
 
14. Optionally the target bacteria are  Campylobacter  (eg,  jejuni ) cells, eg, resistant to an antibiotic selected from ciprofloxacin and azithromycin.
 
15. Optionally, the target cell(s) produce Beta (β)-lactamase (eg, ESBL-producing  E. coli  or ESBL-producing  Klebsiella ).
 
16. Optionally, the target cell(s) are bacterial cells that are resistant to an antibiotic recited in any one of examples 1 to 14.
 
     In an example, the target cell(s) is a cell of a species selected from  Shigella, E. coli, Salmonella, Serratia, Klebsiella, Yersinia, Pseudomonas  and  Enterobacter.    
     Optionally, the composition comprises carrier cells that are each or in combination capable of conjugative transfer of first DNAs into target cells of species selected from two or more of  Shigella, E coli, Salmonella, Serratia, Klebsiella, Yersinia, Pseudomonas  and  Enterobacter.    
     In an example, the reduction in growth or proliferation of carrier cells is at least 50, 60, 70, 80, 90 or 95%. Optionally, the composition or carrier cell(s) are administered simultaneously or sequentially with an an antibiotic that is toxic to the target cells. For example, the antibiotic can be any antibiotic disclosed herein. 
     Optionally, the expression of the NSI is under the control of an inducible promoter that is operable in the target cell. Optionally, the expression of the NSI is under the control of a constitutive promoter that is operable in the target cell. 
     In embodiments, the first DNA (eg, comprised by a plasmid) contains a screenable or selectable marker gene. For example, the selectable marker gene is an antibiotic resistance gene. 
     The carrier bacteria can be bacteria of a species or genus selected from those appearing in Table 5. For example, the species is found in warm-blooded animals (eg, livestock vertebrates). For example, the species is found in humans. For example, the species is found in plants. Preferably, non-pathogenic bacteria that colonize the non-sterile parts of the human or animal body (e.g., skin, digestive tract, urogenital region, mouth, nasal passages, throat and upper airway, ears and eyes) are utilized as carrier cells, and in an example the methodology of the invention is used to combat a target cell bacterial infection of such a part of the body of a human or animal. In another embodiment, the infection is systemic infection. Examples of particularly preferred carrier bacterial species include, but are not limited to: non-pathogenic strains of  Escherichia coli  ( E. coli  F18, S17 and  E. coli  strain Nissle), various species of  Lactobacillus  (such as  L. casei, L. plantarum, L. paracasei, L. acidophilus, L. fermentum, L. zeae  and  L. gasseri ), or other nonpathogenic or probiotic skin- or GI colonizing bacteria such as  Lactococcus , Bifidobacteria, Eubacteria, and bacterial mini-cells, which are anucleoid cells destined to die but still capable of transferring plasmids (see; e.g., Adler et al., Proc. Natl. Acad. Sci. USA 57; 321-326, 1970; Frazer and Curtiss III, Current Topics in Microbiology and Immunology 69: 1-84, 1975; U.S. Pat. No. 4,968,619 to Curtiss III). In some embodiments, the target recipient cells are pathogenic bacteria comprised by a human, animal or plant, eg, on the skin or in the digestive tract, urogenital region, mouth, nasal passage, throat and upper airway, eye(s) and ear(s). Of particular interest for targeting and eradication are pathogenic strains of  Pseudomonas aeruginosa, Escherichia coli, Staphylococcus pneumoniae  and other species,  Enterobacter  spp.,  Enterococcus  spp. and  Mycobacterium tuberculosis . In an example, the target cell genus or species is any genus or species listed in Table 5. 
     The present invention finds use with a wide array of settings or environments, eg, in therapeutic, agricultural, or other settings, including, but not limited to, those described in U.S. Pat. Nos. 6,271,359, 6,261,842, 6,221,582, 6,153,381, 6,106,854, and 5,627,275. Others are also discussed herein, and still others will be readily apparent to those of skill in the art. 
     Numerous types of plasmids comprising the first DNA are suitable for use in the present invention. In view of this, one of skill in the art will appreciate that a single carrier bacterial strain might harbor more than one type of such plasmid (eg, differing in the antibacterial agent that they encode). Further, in another example two or more different carrier bacterial strains, each containing one or more such plasmids, may be combined for a multi-target effect, ie, for killing two or more different target species or strains, or for killing the cells of the same species or strain of target cell. 
     The present invention finds utility for treatment of humans and in a variety of veterinary, agronomic, horticultural and food processing applications. For human and veterinary use, and depending on the cell population or tissue targeted for protection, the following modes of administration of the carrier bacteria of the invention are contemplated: topical, oral, nasal, ocular, aural, pulmonary (e.g., via an inhaler), ophthalmic, rectal, urogenital, subcutaneous, intraperitoneal and intravenous. The bacteria may be supplied as a pharmaceutical composition, in a delivery vehicle suitable for the mode of administration selected for the patient being treated. The term “patient” or “subject” as used here refers to humans or animals (animals being particularly useful as models for clinical efficacy of a particular donor strain, for example, or being farmed or livestock animals). Commercially-relevant animals are chicken, turkey, duck, catfish, salmon, cod, herring, lobster, shrimp, prawns, cows, sheep, goats, pigs, goats, geese or rabbits. 
     For example, to deliver the carrier bacteria to the gastrointestinal tract or to the nasal passages, the preferred mode of administration may be by oral ingestion or nasal aerosol, or by feeding (alone or incorporated into the subject&#39;s feed or food and/or beverage, such as drinking water). In this regard, the carrier cells may be comprised by a food of livestock (or farmed or companion animal), eg, the carrier bacteria are comprised by a feed additive for livestock. Alternatively, the additive is a beverage (eg, water) additive for livestock. It should be noted that probiotic bacteria, such as  Lactobacillus acidophilus , are sold as gel capsules containing a lyophilized mixture of bacterial cells and a solid support such as mannitol. When the gel capsule is ingested with liquid, the lyophilized cells are re-hydrated and become viable, colonogenic bacteria. Thus, in a similar fashion, carrier bacterial cells of the present invention can be supplied as a powdered, lyophilized preparation in a gel capsule, or in bulk, eg, for sprinkling onto food or beverages. The re-hydrated, viable bacterial cells will then populate and/or colonze sites throughout the upper and/or lower gastrointestinal system, and thereafter come into contact with the target bacteria. 
     For topical applications, the carrier bacteria may be formulated as an ointment or cream to be spread on the affected skin surface. Ointment or cream formulations are also suitable for rectal or vaginal delivery, along with other standard formulations, such as suppositories. The appropriate formulations for topical, vaginal or rectal administration are well known to medicinal chemists. The present invention will be of particular utility for topical or mucosal administrations to treat a variety of bacterial infections or bacterially related undesirable conditions. Some representative examples of these uses include treatment of (1) conjunctivitis, caused by  Haemophilus  sp., and corneal ulcers, caused by  Pseudomonas aeruginosa ; (2) otititis externa, caused by  Pseudomonas aeruginosa ; (3) chronic sinusitis, caused by many Gram-positive cocci and Gram-negative rods, or for general decontamination of bronchii; (4) cystic fibrosis, associated with  Pseudomonas aeruginosa ; (5) enteritis, caused by  Helicobacter pylori  (eg, to treat or prevent gastric ulcers),  Escherichia coli, Salmonella typhimurium, Campylobacter  or  Shigella  sp.; (6) open wounds, such as surgical or non-surgical, eg, as a prophylactic measure; (7) burns to eliminate  Pseudomonas aeruginosa  or other Gram-negative pathogens; (8) acne, eg, caused by  Propionobacter acnes ; (9) nose or skin infection, eg, caused by metlncillin resistant  Staphylococcus aureus  (MSRA); (10) body odor, eg, caused by Gram-positive anaerobic bacteria (i.e., use of carrier cells in deodorants); (11) bacterial vaginosis, eg, associated with  Gardnerella vaginalis  or other anaerobes; and (12) gingivitis and/or tooth decay caused by various organisms. 
     In one example, the target cells are  E. coli  cells and the disease or condition to be treated in a human is a uterine tract infection or a ventilator associated infection, eg, pneumonia. 
     In other embodiments, the carrier cells of the present invention find application in the treatment of surfaces for the removal or attenuation of unwanted target bacteria, for example use in a method of treating such a surface or an environment comprising target bacteria, wherein the method comprises contacting the surface or environment with carrier bacteria of the invention, allowing conjugative transfer of the first DNA of the invention from the carrier to the target bacteria, and allowing the antibacterial agent to kill target cells. For example, surfaces that may be used in invasive treatments such as surgery, catheterization and the like may be treated to prevent infection of a subject by bacterial contaminants on the surface. It is contemplated that the methods and compositions of the present invention may be used to treat numerous surfaces, objects, materials and the like (e.g., medical or first aid equipment, nursery and kitchen equipment and surfaces) to control bacterial contamination thereon. 
     Pharmaceutical preparations or other compositions comprising the carrier bacteria may be formulated in dosage unit form for ease of administration and uniformity of dosage. Dosage unit form, as used herein, refers to a physically discrete unit of the pharmaceutical preparation appropriate for the patient or plant or environment or surface undergoing treatment. Each dosage should contain a quantity of the carrier bacteria calculated to produce the desired antibacterial effect in association with the selected carrier. Procedures for determining the appropriate dosage unit are well known to those skilled in the art. Dosage units may be proportionately increased or decreased based on the weight of a patient, plant, surface or environment. Appropriate concentrations for achieving eradication of pathogenic target cells (eg, comprised by a tissue of the patient) may be determined by dosage concentration curve calculations, as known in the art. 
     Other uses for the carrier bacteria of the invention are also contemplated. These include a variety agricultural, horticultural, environmental and food processing applications. For example, in agriculture and horticulture, various plant pathogenic bacteria may be targeted in order to minimize plant disease. One example of a plant pathogen suitable for targeting is  Erwinia  (eg,  E. amylovora , the causal agent of fire blight). Similar strategies may be utilized to reduce or prevent wilting of cut flowers. For veterinary or animal farming, the carrier cells of the invention may be incorporated into animal feed (chicken, swine, poultry, goat, sheep, fish, shellfish or cattle feed) to reduce bio-burden or to eliminate certain pathogenic organisms (e.g.,  Salmonella , such as in chicken, turkey or other poultry). In other embodiments, the invention may be applied on meat or other foods to eliminate unwanted or pathogenic bacteria (e.g.,  E. coli O 157:1H7 on meat, or  Proteus  spp., one cause of “fishy odour” on seafood). 
     Environmental utilities comprise, for example, engineering carrier bacteria, eg,  Bacillus thurengiensis  and one of its conjugative plasmids, to deliver and conditionally express an insecticidal agent in addition to or instead of an antibacterial agent (e.g., for the control of mosquitos that disseminate malaria or West Nile virus). In such applications, as well as in the agricultural and horticultural or other applications described above, formulation of the carrier bacteria as solutions, aerosols, or gel capsules are contemplated. 
     In preferred embodiments of the present invention, certain features are employed in the DNA, plasmids and carrier cells of the invention to minimize potential risks associated with the use of engineered DNA or genetically modified organisms in the environment. For instance, eg, in environmentally sensitive circumstances, it may be advantageous to utilise non-self-transmissible DNA or plasmids. Instead, the DNA or plasmids will be mobilisable by conjugative machinery but will not be self-transmissible. This may be accomplished in some embodiments by integrating into the carrier cell chromosome all or some of the tra genes whose products are necessary for the assembly of conjugative machinery. In such embodiments, DNA or plasmids of the invention are configured to possess an origin of transfer (oriT) but not the tra genes that are provided on the cell&#39;s chromosome. This feature prevents the recipient killer cell, before or even after it dies, from transferring the killer DNA or plasmid further. Another biosafety feature comprises utilizing conjugation systems with predetermined host-ranges. Certain elements are known to function only in few related bacteria (narrow-host-range) and others are known to function in many unrelated bacteria (broad-host-range or promiscuous) (del Solar et al., Mol. Microbiol. 32: 661-666, 1996; Zatyka and Thomas, FEMS Microbiol. Rev. 21: 291-319, 1998). Also, many of those conjugation systems can function in either gram-positive or gram-negative bacteria but generally not in both (del Solar, 1996, supra; Zatyka and Thomas, 1998). 
     Inadvertant proliferation of antibiotic resistance may be minimized in embodiments by avoiding the use of antibiotic resistance markers on the DNA or plasmids of the invention which are conjugatively transferred into target cells. In an alternative approach, the gene responsible for the synthesis of an amino acid (i.e. serine) can be mutated, generating the requirement for this amino acid in the donor. Such mutant bacteria will prosper on media lacking serine provided that they contain a plasmid with the ser gene whose product is needed for growth. Thus, the invention contemplates the advantageous use of plasmids containing the ser gene or another nutritional genetic marker. These markers will permit selection and maintenance of the DNA or plasmids in carrier cells. Another biosafety approach comprises the use of restriction-modification systems to modulate the host range of the DNA or plasmids. Conjugation and plasmid establishment are expected to occur more frequently between taxonomically related species in which plasmid can evade restriction systems and replicate. Type II restriction endonucleases make a double-strand break within or near a specific recognition sequence of duplex DNA. Cognate modification enzymes can methylate the same sequence and protect it from cleavage. Restriction-modification systems (RM) are ubiquitous in bacteria and archaebacteria but are absent in eukaryotes. Some of RM systems are plasmid-encoded, while others are on the bacterial chromosome (Roberts and Macelis, Nucl. Acids Res. 24: 223-235, 1998). Restriction enzymes cleave foreign DNA such as viral or plasmid DNA when this DNA has not been modified by the appropriate modification enzyme. In this way, cells are protected from invasion of foreign DNA. Thus, by using a carrier strain producing one or more methylases, cleavage by one or more restriction enzymes could be evaded. Site-directed mutagenesis is used to produce plasmid DNA that is either devoid of specific restriction sites or that comprises new sites, protecting or making plasmid DNA vulnerable, respectively against endonucleases. Broad-host range plasmids (eg. RP4) may evade restriction systems simply by not having many of the restriction cleavage sites that are typically present on narrow-host plasmids (Willkins et al., 1996, J. Mol. Biol 258, 447-456). Preferred embodiments of the present invention also utilize environmentally safe bacteria as carriers. For example, delivery of DNA vaccines by attenuated intracellular gram-positive and gram-negative bacteria has been reported (Dietrich et al., 2001 Vaccine 19, 2506-2512; Grillot-Courvalin et al, 1999 Current Opinion in Biotech. 10, 477-481). In addition, the donor strain can be one of thousands of harmless bacteria that colonize the non-sterile parts of the body (e.g., skin, gastrointestinal, urogenital, mouth, nasal passages, throat and upper airway systems). Examples of preferred donor (ie, carrier) bacterial species are set forth hereinabove. 
     In another strategy, non-dividing, non-growing carrier cells are utilized instead of living cells. Minicells and maxicells are well studied model systems of metabolically active but nonviable bacterial cells. Minicells lack chromosomal DNA and are generated by special mutant cells that undergo cell division without DNA replication. If the cell contains a multicopy plasmid, many of the minicells will contain plasmids. Minicells neither divide nor grow. However, minicells that possess conjugative plasmids are capable of conjugal replication and transfer of plasmid DNA to living recipient cells. (Adler et al., 1970, supra; Frazer and Curtiss, 1975, supra; U.S. Pat. No. 4,968,619, supra). Maxicells can be obtained from a strain of  E. coli  that carries mutations in the key DNA repair pathways (recA, uvrA and phr). Because maxicells lack so many DNA repair functions, they die upon exposure to low doses of UV. Importantly, plasmid molecules (e.g., pBR322) that do not receive an UV hit continue to replicate. Transcription and translation (plasmid-directed) can occur efficiently under such conditions (Sancar et al., J. Bacteriol. 137: 692-693, 1979), and the proteins made prior to irradiation should be sufficient to sustain conjugation. This is supported by the following two observations: i) that streptomycin-killed cells remain active donors, and ii) that transfer of conjugative plasmids can occur in the presence of antibiotics that prevent de novo gene expression (Heineman and Ankenbauer, 1993, J. Bacteriol. 175, 583-588; Cooper and Heineman, 2000. Plasmid 43, 171-175). Accordingly, UV-treated maxicells will be able to transfer plasmid DNA to live recipients. It should also be noted that the conservation of recA and uvrA genes among bacteria should allow maxicells of donor strains other than  E. coli  to be obtained. 
     Also contemplated for use in the invention are any of the modified bacteria that cannot function because they contain temperature-sensitive mutation(s) in genes that encode for essential cellular functions (e.g., cell wall, protein synthesis, RNA synthesis, as described, for example, in U.S. Pat. No. 4,968,619, supra). 
     In an alternative, archaea are used instead of bacteria for the carrier cells. Additionally or alternatively, the target cells are archaeal cells. 
     As used herein, the term “carrier cell” includes dividing and/or non-dividing bacterial cells (minicells and maxicells), or conditionally non-functional cells. 
     In an example the first DNA is comprised by an engineered RK2 plasmid (ie, a RK2 plasmid that has been modified by recombinant DNA technology or a progeny of such a modified plasmid). Plasmid RK2 is a promiscuous plasmid that can replicate in 29 (and probably many more) gram-negative species (Guiney and Lanka, 1989, p 27-54. In C. M. Thomas (ed) Promiscous plasmids in gram-negative bacteria. London, Ltd London United Kingdom). Plasmid RK2 is a 60-kb self-transmissible plasmid with a complete nucleotide sequence known (Pansegrau et al., 1994, J. Mol. Biol. 239, 623-663). A minimal replicon derived from this large plasmid has been obtained that is devoid of all its genes except for a trfA gene, that encodes plasmid&#39;s Rep protein called TrfA, and an origin of vegetative replication oriV For a review of RK2 replication and its control by TrfA protein, see Helinski et al., 1996 (In  Escherichia coli  and  Salmonella  Cellular and Molecular Biology, Vol. 2 (ed. F. Neidhardt, et al., 2295-2324, ASM Press, Washington D.C.). 
     In an example the first DNA is comprised by an engineered R6K plasmid (ie, a R6K plasmid that has been modified by recombinant DNA technology or a progeny of such a modified plasmid). 
     The present invention is optionally for an industrial or domestic use, or is used in a method for such use. For example, it is for or used in agriculture, oil or petroleum industry, food or drink industry, clothing industry, packaging industry, electronics industry, computer industry, environmental industry, chemical industry, aerospace industry, automotive industry, biotechnology industry, medical industry, healthcare industry, dentistry industry, energy industry, consumer products industry, pharmaceutical industry, mining industry, cleaning industry, forestry industry, fishing industry, leisure industry, recycling industry, cosmetics industry, plastics industry, pulp or paper industry, textile industry, clothing industry, leather or suede or animal hide industry, tobacco industry or steel industry. 
     The present invention is optionally for use in an industry or the environment is an industrial environment, wherein the industry is an industry of a field selected from the group consisting of the medical and healthcare; pharmaceutical; human food; animal food; plant fertilizers; beverage; dairy; meat processing; agriculture; livestock farming; poultry farming; fish and shellfish farming; veterinary; oil; gas; petrochemical; water treatment; sewage treatment; packaging; electronics and computer; personal healthcare and toiletries; cosmetics; dental; non-medical dental; ophthalmic; non-medical ophthalmic; mineral mining and processing; metals mining and processing; quarrying; aviation; automotive; rail; shipping; space; environmental; soil treatment; pulp and paper; clothing manufacture; dyes; printing; adhesives; air treatment; solvents; biodefence; vitamin supplements; cold storage; fibre retting and production; biotechnology; chemical; industrial cleaning products; domestic cleaning products; soaps and detergents; consumer products; forestry; fishing; leisure; recycling; plastics; hide, leather and suede; waste management; funeral and undertaking; fuel; building; energy; steel; and tobacco industry fields. 
     In an example, the first DNA comprises a CRISPR array that targets target bacteria, wherein the array comprises one, or two or more different spacers (eg, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50 or more spacers) for targeting the genome of target bacteria. 
     In an example, the target bacteria are comprised by an environment as follows. In an example, the environment is a microbiome of a human, eg, the oral cavity microbiome or gut microbiome or the bloodstream. In an example, the environment is not an environment in or on a human. In an example, the environment is not an environment in or on a non-human animal. In an embodiment, the environment is an air environment. In an embodiment, the environment is an agricultural environment. In an embodiment, the environment is an oil or petroleum recovery environment, eg, an oil or petroleum field or well. In an example, the environment is an environment in or on a foodstuff or beverage for human or non-human animal consumption. In an example, the environment is a maritime environment, eg, in seawater or on a boat (eg, in ship or boat ballast water). 
     In an example, the environment is a a human or animal microbiome (eg, gut, vaginal, scalp, armpit, skin or oral cavity microbiome). In an example, the target bacteria are comprised by a human or animal microbiome (eg, gut, vaginal, scalp, armpit, skin or oral cavity microbiome). 
     In an example, the carrier bacteria or composition of the invention are administered intranasally, topically or orally to a human or non-human animal, or is for such administration. The skilled person aiming to treat a microbiome of the human or animal will be able to determine the best route of administration, depending upon the microbiome of interest. For example, when the microbiome is a gut microbiome, administration can be intranasally or orally. When the microbiome is a scalp or armpit microbiome, administration can be topically. When the microbiome is in the mouth or throat, the administration can be orally. 
     In an example, the environment is harboured by a beverage or water (eg, a waterway or drinking water for human consumption) or soil. The water is optionally in a heating, cooling or industrial system, or in a drinking water storage container. 
     In an example, the carrier and/or target bacteraia are Firmicutes selected from  Anaerotruncus, Acetanaerobacterium, Acetitomaculum, Acetivibrio, Anaerococcus, Anaerofilum, Anaerosinus, Anaerostipes, Anaerovorax, Butyrivibrio, Clostridium, Capracoccus, Dehalobacter, Dialister, Dorea, Enterococcus, Ethanoligenens, Faecalibacterium, Fusobacterium, Gracilibacter, Guggenheimella, Hespellia, Lachnobacterium, Lachnospira, Lactobacillus, Leuconostoc, Megamonas, Moryella, Mitsuokella, Oribacterium, Oxobacter, Papillibacter, Proprionispira, Pseudobutyrivibrio, Pseudoramibacter, Roseburia, Ruminococcus, Sarcina, Seinonella, Shuttleworthia, Sporobacter, Sporobacterium, Streptococcus, Subdoligranulum, Syntrophococcus, Thermobacillus, Turibacter  and Weisella. 
     In an example, the carrier bacteria, composition, use or method is for reducing pathogenic infections or for re-balancing gut or oral biofilm eg, for treating or preventing obesity or disease in a human or animal; or for treating or preventing a GI condition (such as Crohn&#39;s disease, IBD or colitis). For example, the DNA, carrier bacteria, composition, use or method is for knocking-down  Salmomnella, Campylobacter, Erwinia, Xanthomonous, Edwardsiella, Pseudomonas, Klebsiella, Pectobacterium, Clostridium dificile  or  E. coli  bacteria in a gut biofilm of a human or animal or a plant, preferably in a human or animal. 
     In an example, the animal is a chicken, eg, and the target bacteria are  Salmonella  or  Campylobacter . In an example, the animal is a fish (eg, catfish or salmon) or shellfish (eg, prawn or lobster), eg, and the target bacteria are Edwardsiella. In an example, the plant is a potato plant and, eg, the target bacteria are  Pectobacterium . In an example, the plant is a cabbage plant and, eg, the target bacteria are  Xanthomonous  (eg,  X. campestris ). In an example, the plant is a marijuana plant and, eg, the targt bacteria are  Pseudomonas  (eg,  P. cannabina  or  P. amygdali ),  Agrobacterium  (eg,  A. tumefaciens ) or  Xanthomonas  (eg,  X. campestris ). In an example, the plant is a hemp plant and, eg, the targt bacteria are are  Pseudomonas  (eg,  P. cannabina  or  P. amygdali ),  Agrobacterium  (eg,  A. tumefaciens ) or  Xanthomonas  (eg,  X. campestris ). 
     In an example, the disease or condition is a cancer, inflammatory or autoimmune disease or condition, eg, obesity, diabetes IBD, a GI tract condition or an oral cavity condition. 
     Optionally, the environment is comprised by, or the target bacteria are comprised by, a gut biofilm, skin biofilm, oral cavity biofilm, throat biofilm, hair biofilm, armpit biofilm, vaginal biofilm, rectal biofilm, anal biofilm, ocular biofilm, nasal biofilm, tongue biofilm, lung biofilm, liver biofilm, kidney biofilm, genital biofilm, penile biofilm, scrotal biofilm, mammary gland biofilm, ear biofilm, urethra biofilm, labial biofilm, organ biofilm or dental biofilm. Optionally, the environment is comprised by, or the target bacteria are comprised by, a plant (eg, a tobacco, crop plant, fruit plant, vegetable plant or tobacco, eg on the surface of a plant or contained in a plant) or by an environment (eg, soil or water or a waterway or aqueous liquid). 
     In an example, the carrier cell(s) or composition is for treating a disease or condition in an animal or human, wherein the disease or condition. In an example, the disease or condition is caused by or mediated by an infection of target cells comprised by the subject or patient. In an example, the disease or condition is associated with an infection of target cells comprised by the subject or patient. 
     In an example, a symptom of the disease or condition is an infection of target cells comprised by the subject or patient. 
     Optionally, the disease or condition of a human or animal subject is selected from
         (a) A neurodegenerative disease or condition;   (b) A brain disease or condition;   (c) A CNS disease or condition;   (d) Memory loss or impairment;   (e) A heart or cardiovascular disease or condition, eg, heart attack, stroke or atrial fibrillation;   (f) A liver disease or condition;   (g) A kidney disease or condition, eg, chronic kidney disease (CKD);   (h) A pancreas disease or condition;   (i) A lung disease or condition, eg, cystic fibrosis or COPD;   (j) A gastrointestinal disease or condition;   (k) A throat or oral cavity disease or condition;   (l) An ocular disease or condition;   (m) A genital disease or condition, eg, a vaginal, labial, penile or scrotal disease or condition;   (n) A sexually-transmissible disease or condition, eg, gonorrhea, HIV infection, syphilis or  Chlamydia  infection;   (o) An ear disease or condition;   (p) A skin disease or condition;   (q) A heart disease or condition;   (r) A nasal disease or condition   (s) A haematological disease or condition, eg, anaemia, eg, anaemia of chronic disease or cancer;   (t) A viral infection;   (u) A pathogenic bacterial infection;   (v) A cancer;   (w) An autoimmune disease or condition, eg, SLE;   (x) An inflammatory disease or condition, eg, rheumatoid arthritis, psoriasis, eczema, asthma, ulcerative colitis, colitis, Crohn&#39;s disease or IBD;   (y) Autism;   (z) ADHD;   (aa) Bipolar disorder;   (bb) ALS [Amyotrophic Lateral Sclerosis];   (cc) Osteoarthritis;   (dd) A congenital or development defect or condition;   (ee) Miscarriage;   (ff) A blood clotting condition;   (gg) Bronchitis;   (hh) Dry or wet AMD;   (ii) Neovascularisation (eg, of a tumour or in the eye);   (jj) Common cold;   (kk) Epilepsy;   (ll) Fibrosis, eg, liver or lung fibrosis;   (mm) A fungal disease or condition, eg, thrush;   (nn) A metabolic disease or condition, eg, obesity, anorexia, diabetes, Type I or Type II diabetes.   (oo) Ulcer(s), eg, gastric ulceration or skin ulceration;   (pp) Dry skin;   (qq) Sjogren&#39;s syndrome;   (rr) Cytokine storm;   (ss) Deafness, hearing loss or impairment;   (tt) Slow or fast metabolism (ie, slower or faster than average for the weight, sex and age of the subject);   (uu) Conception disorder, eg, infertility or low fertility;   (vv) Jaundice;   (ww) Skin rash;   (xx) Kawasaki Disease;   (yy) Lyme Disease;   (zz) An allergy, eg, a nut, grass, pollen, dust mite, cat or dog fur or dander allergy;   (aaa) Malaria, typhoid fever, tuberculosis or cholera;   (bbb) Depression;   (ccc) Mental retardation;   (ddd) Microcephaly;   (eee) Malnutrition;   (fff) Conjunctivitis;   (ggg) Pneumonia;   (hhh) Pulmonary embolism;   (iii) Pulmonary hypertension;   (jjj) A bone disorder;   (kkk) Sepsis or septic shock;   (lll) Sinusitus;   (mmm) Stress (eg, occupational stress);   (nnn) Thalassaemia, anaemia, von Willebrand Disease, or haemophilia;   (ooo) Shingles or cold sore;   (ppp) Menstruation;   (qqq) Low sperm count.       

     Neurodegenerative or CNS Diseases or Conditions for Treatment or Prevention by the Invention 
     In an example, the neurodegenerative or CNS disease or condition is selected from the group consisting of Alzheimer disease, geriopsychosis, Down syndrome, Parkinson&#39;s disease, Creutzfeldt-jakob disease, diabetic neuropathy, Parkinson syndrome, Huntington&#39;s disease, Machado-Joseph disease, amyotrophic lateral sclerosis, diabetic neuropathy, and Creutzfeldt Creutzfeldt-Jakob disease. For example, the disease is Alzheimer disease. For example, the disease is Parkinson syndrome. 
     In an example, wherein the method of the invention is practised on a human or animal subject for treating a CNS or neurodegenerative disease or condition, the method causes downregulation of Treg cells in the subject, thereby promoting entry of systemic monocyte-derived macrophages and/or Treg cells across the choroid plexus into the brain of the subject, whereby the disease or condition (eg, Alzheimer&#39;s disease) is treated, prevented or progression thereof is reduced. In an embodiment the method causes an increase of IFN-gamma in the CNS system (eg, in the brain and/or CSF) of the subject. In an example, the method restores nerve fibre and/or reduces the progression of nerve fibre damage. In an example, the method restores nerve myelin and/or reduces the progression of nerve myelin damage. In an example, the method of the invention treats or prevents a disease or condition disclosed in WO2015136541 and/or the method can be used with any method disclosed in WO2015136541 (the disclosure of this document is incorporated by reference herein in its entirety, eg, for providing disclosure of such methods, diseases, conditions and potential therapeutic agents that can be administered to the subject for effecting treatment and/or prevention of CNS and neurodegenerative diseases and conditions, eg, agents such as immune checkpoint inhibitors, eg, anti-PD-1, anti-PD-L1, anti-TIM3 or other antibodies disclosed therein). 
     Cancers for Treatment or Prevention by the Method 
     Cancers that may be treated include tumours that are not vascularized, or not substantially vascularized, as well as vascularized tumours. The cancers may comprise non-solid tumours (such as haematological tumours, for example, leukaemias and lymphomas) or may comprise solid tumours. Types of cancers to be treated with the invention include, but are not limited to, carcinoma, blastoma, and sarcoma, and certain leukaemia or lymphoid malignancies, benign and malignant tumours, and malignancies e.g., sarcomas, carcinomas, and melanomas. Adult tumours/cancers and paediatric tumours/cancers are also included. 
     Haematologic cancers are cancers of the blood or bone marrow. Examples of haematological (or haematogenous) cancers include leukaemias, including acute leukaemias (such as acute lymphocytic leukaemia, acute myelocytic leukaemia, acute myelogenous leukaemia and myeloblasts, promyeiocytic, myelomonocytic, monocytic and erythroleukaemia), chronic leukaemias (such as chronic myelocytic (granulocytic) leukaemia, chronic myelogenous leukaemia, and chronic lymphocytic leukaemia), polycythemia vera, lymphoma, Hodgkin&#39;s disease, non-Hodgkin&#39;s lymphoma (indolent and high grade forms), multiple myeloma, Waldenstrom&#39;s macroglobulinemia, heavy chain disease, myeiodysplastic syndrome, hairy cell leukaemia and myelodysplasia. 
     Solid tumours are abnormal masses of tissue that usually do not contain cysts or liquid areas. Solid tumours can be benign or malignant. Different types of solid tumours are named for the type of cells that form them (such as sarcomas, carcinomas, and lymphomas). Examples of solid tumours, such as sarcomas and carcinomas, include fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteosarcoma, and other sarcomas, synovioma, mesothelioma, Ewing&#39;s tumour, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, lymphoid malignancy, pancreatic cancer, breast cancer, lung cancers, ovarian cancer, prostate cancer, hepatocellular carcinoma, squamous eel! carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, medullary thyroid carcinoma, papillary thyroid carcinoma, pheochromocytomas sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, Wilms&#39; tumour, cervical cancer, testicular tumour, seminoma, bladder carcinoma, melanoma, and CNS tumours (such as a glioma (such as brainstem glioma and mixed gliomas), glioblastoma (also known as glioblastoma multiforme) astrocytoma, CNS lymphoma, germinoma, medu!loblastoma, Schwannoma craniopharyogioma, ependymoma, pineaioma, hemangioblastoma, acoustic neuroma, oligodendroglioma, menangioma, neuroblastoma, retinoblastoma and brain metastases). 
     Autoimmune Diseases for Treatment or Prevention by the Method 
     
         
         
           
             1. Acute Disseminated Encephalomyelitis (ADEM) 
             2. Acute necrotizing hemorrhagic leukoencephalitis 
             3. Addison&#39;s disease 
             4. Agammaglobulinemia 
             5. Alopecia areata 
             6. Amyloidosis 
             7. Ankylosing spondylitis 
             8. Anti-GBM/Anti-TBM nephritis 
             9. Antiphospholipid syndrome (APS) 
             10. Autoimmune angioedema 
             11. Autoimmune aplastic anemia 
             12. Autoimmune dysautonomia 
             13. Autoimmune hepatitis 
             14. Autoimmune hyperlipidemia 
             15. Autoimmune immunodeficiency 
             16. Autoimmune inner ear disease (AIED) 
             17. Autoimmune myocarditis 
             18. Autoimmune oophoritis 
             19. Autoimmune pancreatitis 
             20. Autoimmune retinopathy 
             21. Autoimmune thrombocytopenic purpura (ATP) 
             22. Autoimmune thyroid disease 
             23. Autoimmune urticaria 
             24. Axonal &amp; neuronal neuropathies 
             25. Balo disease 
             26. Behcet&#39;s disease 
             27. Bullous pemphigoid 
             28. Cardiomyopathy 
             29. Castleman disease 
             30. Celiac disease 
             31. Chagas disease 
             32. Chronic fatigue syndrome 
             33. Chronic inflammatory demyelinating polyneuropathy (CIDP) 
             34. Chronic recurrent multifocal osteomyelitis (CRMO) 
             35. Churg-Strauss syndrome 
             36. Cicatricial pemphigoid/benign mucosal pemphigoid 
             37. Crohn&#39;s disease 
             38. Cogans syndrome 
             39. Cold agglutinin disease 
             40. Congenital heart block 
             41. Coxsackie myocarditis 
             42. CREST disease 
             43. Essential mixed cryoglobulinemia 
             44. Demyelinating neuropathies 
             45. Dermatitis herpetiformis 
             46. Dermatomyositis 
             47. Devic&#39;s disease (neuromyelitis optica) 
             48. Discoid lupus 
             49. Dressler&#39;s syndrome 
             50. Endometriosis 
             51. Eosinophilic esophagitis 
             52. Eosinophilic fasciitis 
             53. Erythema nodosum 
             54. Experimental allergic encephalomyelitis 
             55. Evans syndrome 
             56. Fibromyalgia 
             57. Fibrosing alveolitis 
             58. Giant cell arteritis (temporal arteritis) 
             59. Giant cell myocarditis 
             60. Glomerulonephritis 
             61. Goodpasture&#39;s syndrome 
             62. Granulomatosis with Polyangiitis (GPA) (formerly called Wegener&#39;s Granulomatosis) 
             63. Graves&#39; disease 
             64. Guillain-Barre syndrome 
             65. Hashimoto&#39;s encephalitis 
             66. Hashimoto&#39;s thyroiditis 
             67. Hemolytic anemia 
             68. Henoch-Schonlein purpura 
             69. Herpes gestationis 
             70. Hypogammaglobulinemia 
             71. Idiopathic thrombocytopenic purpura (ITP) 
             72. IgA nephropathy 
             73. IgG4-related sclerosing disease 
             74. Immunoregulatory lipoproteins 
             75. Inclusion body myositis 
             76. Interstitial cystitis 
             77. Juvenile arthritis 
             78. Juvenile diabetes (Type 1 diabetes) 
             79. Juvenile myositis 
             80. Kawasaki syndrome 
             81. Lambert-Eaton syndrome 
             82. Leukocytoclastic vasculitis 
             83. Lichen planus 
             84. Lichen sclerosus 
             85. Ligneous conjunctivitis 
             86. Linear IgA disease (LAD) 
             87. Lupus (SLE) 
             88. Lyme disease, chronic 
             89. Meniere&#39;s disease 
             90. Microscopic polyangiitis 
             91. Mixed connective tissue disease (MCTD) 
             92. Mooren&#39;s ulcer 
             93. Mucha-Habermann disease 
             94. Multiple sclerosis 
             95. Myasthenia gravis 
             96. Myositis 
             97. Narcolepsy 
             98. Neuromyelitis optica (Devic&#39;s) 
             99. Neutropenia 
             100. Ocular cicatricial pemphigoid 
             101. Optic neuritis 
             102. Palindromic rheumatism 
             103. PANDAS (Pediatric Autoimmune Neuropsychiatric Disorders Associated with  Streptococcus ) 
             104. Paraneoplastic cerebellar degeneration 
             105. Paroxysmal nocturnal hemoglobinuria (PNH) 
             106. Parry Romberg syndrome 
             107. Parsonnage-Turner syndrome 
             108. Pars planitis (peripheral uveitis) 
             109. Pemphigus 
             110. Peripheral neuropathy 
             111. Perivenous encephalomyelitis 
             112. Pernicious anemia 
             113. POEMS syndrome 
             114. Polyarteritis nodosa 
             115. Type I, II, &amp; III autoimmune polyglandular syndromes 
             116. Polymyalgia rheumatica 
             117. Polymyositis 
             118. Postmyocardial infarction syndrome 
             119. Postpericardiotomy syndrome 
             120. Progesterone dermatitis 
             121. Primary biliary cirrhosis 
             122. Primary sclerosing cholangitis 
             123. Psoriasis 
             124. Psoriatic arthritis 
             125. Idiopathic pulmonary fibrosis 
             126. Pyoderma gangrenosum 
             127. Pure red cell aplasia 
             128. Raynauds phenomenon 
             129. Reactive Arthritis 
             130. Reflex sympathetic dystrophy 
             131. Reiter&#39;s syndrome 
             132. Relapsing polychondritis 
             133. Restless legs syndrome 
             134. Retroperitoneal fibrosis 
             135. Rheumatic fever 
             136. Rheumatoid arthritis 
             137. Sarcoidosis 
             138. Schmidt syndrome 
             139. Scleritis 
             140. Scleroderma 
             141. Sjogren&#39;s syndrome 
             142. Sperm &amp; testicular autoimmunity 
             143. Stiff person syndrome 
             144. Subacute bacterial endocarditis (SBE) 
             145. Susac&#39;s syndrome 
             146. Sympathetic ophthalmia 
             147. Takayasu&#39;s arteritis 
             148. Temporal arteritis/Giant cell arteritis 
             149. Thrombocytopenic purpura (TTP) 
             150. Tolosa-Hunt syndrome 
             151. Transverse myelitis 
             152. Type 1 diabetes 
             153. Ulcerative colitis 
             154. Undifferentiated connective tissue disease (UCTD) 
             155. Uveitis 
             156. Vasculitis 
             157. Vesiculobullous dermatosis 
             158. Vitiligo 
             159. Wegener&#39;s granulomatosis (now termed Granulomatosis with Polyangiitis (GPA). 
           
         
       
    
     Inflammatory Diseases for Treatment or Prevention by the Method 
     
         
         
           
             1. Alzheimer 
             2. ankylosing spondylitis 
             3. arthritis (osteoarthritis, rheumatoid arthritis (RA), psoriatic arthritis) 
             4. asthma 
             5. atherosclerosis 
             6. Crohn&#39;s disease 
             7. colitis 
             8. dermatitis 
             9. diverticulitis 
             10. fibromyalgia 
             11. hepatitis 
             12. irritable bowel syndrome (IBS) 
             13. systemic lupus erythematous (SLE) 
             14. nephritis 
             15. Parkinson&#39;s disease 
             16. ulcerative colitis. 
           
         
       
    
     Growth Promoters &amp; Lowering Food Conversion Ratios 
     The Examples demonstrate that target bacteria can be targeted using an antibacterial agent to promote growth and enhance FCR in poultry. In the Example, a guided nuclease system was used to specifically target  Salmonella  in the poultry. 
     In a first aspect, there is provided:— 
     A method of promoting the growth of an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal a guided nuclease system or a component thereof, and introducing the system or component into target bacteria comprised by the animal, wherein the guided nuclease is capable of recognising and modifying (eg, cutting) a target nucleotide sequence comprised by the target bacteria, whereby target bacteria are killed or the growth or proliferation of target bacteria are inhibited and the growth of the animal is promoted. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of inhibiting the growth of the animal. Thus, the method reduces the burden of such bacteria in the animal and promotes growth. 
     In a second aspect there is provided:— 
     A method of enhancing feed conversion ratio (FCR) in an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal a guided nuclease system or a component thereof, and introducing the system or component into target bacteria comprised by the animal, wherein the guided nuclease is capable of recognising and modifying (eg, cutting) a target nucleotide sequence comprised by the target bacteria, whereby target bacteria are killed or the growth or proliferation of target bacteria are inhibited and the FCR of the animal is enhanced. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of increasing the FCR of the animal. Thus, the method reduces the burden of such bacteria in the animal and enhances FCR (ie, reduces FCR number). 
     In a third aspect there is provided:— 
     A method of promoting the growth of an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal an antibacterial agent that is toxic to  Salmonella  bacteria, wherein  Salmonella  target bacteria comprised by the animal are exposed to the agent and are killed or the growth or proliferation of target bacteria are inhibited and the growth of the animal is promoted. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of inhibiting the growth of the animal. Thus, the method reduces the burden of such bacteria in the animal and promotes growth. 
     In a fourth aspect there is provided:— 
     A method of enhancing feed conversion ratio (FCR) in an animal (eg, a livestock animal, eg, a poultry animal), the method comprising administering to the animal an antibacterial agent that is toxic to  Salmonella  bacteria, wherein  Salmonella  target bacteria comprised by the animal are exposed to the agent and are killed or the growth or proliferation of target bacteria are inhibited and the FCR of the animal is enhanced. 
     The method is a non-medical method and the presence of target bacteria in the animal is capable of increasing the FCR of the animal. Thus, the method reduces the burden of such bacteria in the animal and enhances FCR (ie, reduces FCR number). 
     In any of these aspects, optionally the animal is a livestock animal. Optionally, the animal is a bird, eg, a poultry bird, eg, a chicken, turkey, goose or duck. Preferably, the animal is a chicken. 
     In any of these aspects, optionally the bacteria are Enterobacteriaciae bacteria, eg,  Salmonella . For example, the bacteria are  Salmonella enterica, typhimurium  or  enteritidis . For example, the  Salmonella  is any  Salmonella  species or strain disclosed herein. 
     For example, the system, component or agent is supplied to the animal in an animal feed and/or beverage (eg, mixed in drinking water). When supplied in a beverage, the system, component or agent may be comprised by carrier bacteria, wherein the carrier bacteria are comprised in the beverage at an amount of from 1×10 3  to 1×10 10  (eg, from 1×10 4  to 1×10 10 ; from 1×10 4  to 1×10 9 ; from 1×10 4  to 1×10 8 ; from 1×10 4  to 1×10 7 ; from 1×10 3  to 1×10 10 ; from 1×10 3  to 1×10 9 ; from 1×10 3  to 1×10 8 ; from 1×10 3  to 1×10 7 ; from 1×10 5  to 1×10 10 ; from 1×10 5  to 1×10 9 ; from 1×10 5  to 1×10 8 ; from 1×10 5  to 1×10 7 ; from 1×10 6  to 1×10 10 ; from 1×10 6  to 1×10 9 ; from 1×10 6  to 1×10 8 ; or from 1×10 6  to 1×10 7 ) cfu/ml. When supplied in a beverage, the system, component or agent may be comprised by carrier bacteria, wherein the carrier bacteria are comprised in the beverage at an amount of at least 1×10 8  cfu/ml, eg, wherein the animal is a poultry bird, such as a chicken. 
     Optionally, the guided nuclease is any guided nuclease disclosed herein, eg, a Cas, TALEN, meganuclease or a zinc finger nuclease. In an example, the component is a crRNA or guide RNA that is operable in target cells with a cognate Cas nuclease. The Cas nuclease can be any Cas nuclease disclosed herein. The Cas nuclease may be an endogenous Cas of the target cells or may be encoded by an exogenous nucleic acid that is administered to the animal. 
     Systems, components and agents of the invention may be introduced into target bacteria by bacterial conjugation (eg, conjugative transfer from a carrier cell to the target cell) or by phage wherein the phage transduce into the target cells nucleic acid encoding the system, component or agent. 
     Optionally, the target bacteria are comprised by any microbiota disclosed herein that is found in animals. Preferably, the microbiota is a gut microbiota (eg, a gut microbiota of a chicken). 
     The livestock animal can be any livestock animal disclosed herein, eg, a chicken, pig, cow, sheep, farmed fish (such as salmon or catfish) or farmed shellfish (eg, lobster, prawn or shrimp). 
     It will be understood that particular embodiments described herein are shown by way of illustration and not as limitations of the invention. The principal features of this invention can be employed in various embodiments without departing from the scope of the invention. Those skilled in the art will recognize, or be able to ascertain using no more than routine study, numerous equivalents to the specific procedures described herein. Such equivalents are considered to be within the scope of this invention and are covered by the claims. All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which this invention pertains. All publications and patent applications and all US equivalent patent applications and patents are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference. The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and/or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.” The use of the term “or” in the claims is used to mean “and/or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and/or.” Throughout this application, the term “about” is used to indicate that a value includes the inherent variation of error for the device, the method being employed to determine the value, or the variation that exists among the study subjects. 
     As used in this specification and claim(s), the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps 
     The term “or combinations thereof” or similar as used herein refers to all permutations and combinations of the listed items preceding the term. For example, “A, B, C, or combinations thereof is intended to include at least one of: A, B, C, AB, AC, BC, or ABC, and if order is important in a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continuing with this example, expressly included are combinations that contain repeats of one or more item or term, such as BB, AAA, MB, BBC, AAABCCCC, CBBAAA, CABABB, and so forth. The skilled artisan will understand that typically there is no limit on the number of items or terms in any combination, unless otherwise apparent from the context. 
     Any part of this disclosure may be read in combination with any other part of the disclosure, unless otherwise apparent from the context. 
     All of the compositions and/or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the compositions and/or methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims. 
     EXAMPLES 
     Example 1: Target Gene Sequence Assessment &amp; crRNA Array Design 
     List of Putative Target Genes in  Salmonella    
     Through careful analysis and decision making, we compiled the following target genes for assessment by cutting using CRISPR/Cas components delivered by conjugative plasmids from carrier cells as per the invention. 
     1) pipA Pathogenicity island encoded protein: SPI5
 
2) mViM putative virulence factor
 
3) mViN putative virulence factor
 
4) phoP Transcribes genes expressed under low Mg+ concentration (OmpR family); virulence transcriptional regulatory protein PHOP
 
5) hilA Invasion genes transcription activator
 
6) BigA putative surface-exposed virulence protein
 
7) sugR ATP binding protein; Pathogenicity island encoded protein: SPI3
 
8) rhuM Pathogenicity island encoded protein: SPI3
 
9) pipC invasion gene E protein/Pathogenicity island encoded protein: SPI5
 
10) pipB Pathogenicity island encoded protein: SPI5
 
11) sicP chaperone related to virulence
 
12) sopB invasion gene D protein/Pathogenicity island encoded protein: SPI5
 
13) marT putative transcriptional regulatory protein/Pathogenicity island encoded protein: SPI3
 
     Target Selection 
     Initial target selection of  Salmonella -specific genes identified 13 putative candidates located almost exclusively in  Salmonella  pathogenicity islands (listed at the start of this example). After close inspection, the list of targets was reduced to seven genes, namely pipA, pipB, pipC, hilA, marT, sicP and sopB, that we found importantly showed no significant sequence homology to other genes from Enterobacteria, thus enabling high specificity of targeting  Salmonella  cells in vivo and in microbiomes containing non-target Enterobacteria of different (non- Salmonella ) species. Target sites for  S. pyogenes  Cas9 defined by presence of the PAM motif (NGG) were extracted using the respective function available in molecular biology software. A number of 6 to 14 putative target sites were identified in each of the seven selected genes. 
     crRNA Design and Synthesis 
     To select target sites most likely to result in efficient restriction of dsDNA, all potential sequences were screened for the presence of nucleotides at key position that have been described to have positive or negative effects on the efficiency of double strand cuts introduced by Cas9 in the available literature. Three publications reported such data, although on sgRNA:
         (1) Doench et al. 2014, Nature Biotechnology   (2) Gagnon et al. 2014, PLoS One   (3) Liu et al. 2016, Scientific Reports       

     One target sequence for each gene that adhered to most of the parameters described influencing restriction efficiency in each of the three publications was selected to be incorporated into a crRNA array, resulting in three arrays to be tested. crRNA arrays were designed based on the sequence data from  S. pyogenes  (published in Deltcheva et al. 2011, Nature). The following sequences were constructed: SEQ ID NOs: 1-3. 
     Two of these sequences encoding crRNAs were assembled with sequences encoding cognate tracrRNA and Cas9. 
     Cas9 Constructs (tracrRNA-Cas9-crRNA) 
     The selected sequences encoding crRNAs were combined with different versions of the tracrRNA-Cas9 modules, resulting in a varied set of plasmids. 
     For constitutive expression of all components (tracrRNA, Cas9 and crRNA), a tracr-Cas fragment based on plasmid pCas9 (Jiang et al. 2013, Nature Biotechnology; sequence presented as SEQ ID NO: 4) was amplified from  S. pyogenes  gDNA. We will join this fragment with the available crRNA-encoding fragments and clone into high copy number general cloning plasmid pJet1.2. This series of construct is named pFS1. 
     To determine whether expression of this constitutive CRISPR/Cas construct in a low copy number background will work as efficiently, we will move the system described above into a low copy number plasmid. pFS2 constructs are therefore based on the low copy number pSC101 origin from pZS21MCS (Expressys). 
     To control expression of at least one of the CRISPR/Cas components by means of an inducible promoter, we will replace the constitutive Cas9 promoter with the TetR regulated promoter PLtetO-1. The resulting plasmids of pFS3 are based on the low copy number pSC101 origin of pZS21MCS as well. The introduced changes will be as follows:
         Spacing between RBS and Start ATG has been shortened to 7 bp (TAAATAC)   Stop codon has been changed from a less effective  TGA C to a tandem  TAATAA T sequence       

     Lastly, we will produce a mobilisable version of the constitutively expressed plasmids like pFS1 and pFS2 mentioned above. To this end, we will clone the origin of transfer (oriT) of plasmid RP4 into an intermediate copy number plasmid (p15A origin) carrying a chloramphenicol resistance. This construct should be able to be mobilised and transferred by a chromosomally integrated conjugation machinery as present in RP4 and other mobilisable plasmids. 
     Further Constructs 
     To determine the efficiency of dsDNA restriction by the selected crRNAs prior to a final selection and transformation into  Salmonella , we will aim to clone the target genes (or substantial parts thereof) into a compatible plasmid. The plasmid selected is pZA31MCS (Expressys) with a p15A origin compatible to the pFS1/pFS2/pFS3 constructs described above. Efficiency can then be tested in  E. coli  by loss of chloramphenicol resistance (or loss of fluorescence if a yfp derivative is present as a reporter). The best performing target sequences can then be used in future, more targeted plasmids. 
     We made the following plasmids:—
     pFSmob-C tracrRNA/Cas9—mob—pZA31MCS   pFSmobSal7 tracrRNA/Cas9—Salcr7-3—mob—pZA31MCS   pFSmobSal3 tracrRNA/Cas9—Salcr3.2—mob—pZA31MCS   pFSmob-C* control based on Salcr7-3—mob—pZA31MCS   

     Plasmid Details 
     
         
         Plasmid Name: pFSmob-C 
         Description: p15A origin plasmid based on pZA31MCS (Expressys) carrying 4768 bp insert (tracrRNA-Cas9-oriT/mob) 
         Plasmid Length: 6685 bp 
         Sequence verified: Yes; Insert sequence verified by primer walking sequencing 
         Additional Notes: No crRNA array present; Control plasmid 
       
    
     Plasmid Details 
     
         
         Plasmid Name: pFSmobSal7 
         Description: p15A origin plasmid based on pZA31MCS (Expressys) carrying 5598 bp insert (tracrRNA-Cas9-crRNA-oriT/mob) 
         Plasmid Length: 7515 bp 
         Sequence verified: Yes; Insert sequence verified by primer walking sequencing 
       
    
     Plasmid Details 
     
         
         Plasmid Name: pFSmobSal3 
         Description: p15A origin plasmid based on pZA31MCS (Expressys) carrying 5341 bp insert (tracrRNA-Cas9-crRNA-oriT/mob) 
         Plasmid Length: 7258 bp 
         Sequence verified: Yes; Insert sequence verified by primer walking sequencing 
       
    
     Plasmid Details 
     
         
         Plasmid Name: pFSmob-C* 
         Description: p15A origin plasmid based on pZA31MCS (Expressys) 
         Plasmid Length: 7348 bp 
         Sequence verified: Yes; Insert sequence verified by primer walking sequencing 
       
    
     Example 2: Selectively Removal of Unwanted  Salmonella  Strains by Conjugation Using a Guided Nuclease 
     Purpose of the Study 
     The objective of this study was to selectively remove unwanted  Salmonella  strains by conjugation experiment. We mobilised the pFSMobSal7 plasmid from  E. coli  S17 to  Salmonella  spp. This plasmid contains tracr-Cas9 the crRNA-encoding array (Sal7-3) and the origin of transfert (oriT). 
     SUMMARY 
     As a proof of concept the conjugation experiment was first done from  E. coli  to  E. coli  before mobilising to  Salmonella  spp. The plasmid pFSMobSal7 (chloramphenicol resistant, CmR) was first transformed into a carrier strain that allows the mobilisation. In this experiment, the strain that produced the pilus was  E. coli  S17 (ATCC® 47055TM). The conjugative functions are provided by an RP4 plasmid integrated into the chromosome. The recipient cell should be Nalidixic acid resistant (25 μg/ml), here we used  E. coli  JM109 (ATCC® 53323TM). The  Salmonella  targeted was FS26 from our  Salmonella  spp collection. The results showed a good efficiency of conjugation from  E. coli  S17 to  E. coli  JM109 and from  E. coli  S17 to  Salmonella enteritidis  (FS26). After conjugation to  Salmonella  as a recipient the killing with pFSMob-3 was efficient. 
     Introduction 
     The pFSmobSal7 plasmid was originated from the intermediated copy number pZA31MCS (Expressys). An origin of transfer (OriT) sequence was synthesised (called mob2) and cloned into pZA31MCS by assembly method in addition to the tracr-Cas9 and the crRNA array fragments. 
     Before starting the conjugation experiment,  E. coli  S17 strains were made electro-competent by a standard protocol (O&#39;Challahan and Charbit, 1990). Plasmids pFSMobsal7 and pFSMobC* were then transformed into  E. coli  S17 by electroporation and selected in chloramphenicol (Cm30, 30 μg/mL). 
     pFSMobSal7 and pFSMobC* were mobilised from the donor  E. coli  S17 to the recipient  E. coli  JM109 and to  Salmonella  spp that are Nalidixic resistant (NalR) and chloramphenicol sensitive (CmS) by mating on a filter followed by an incubation at 37° C. The mixtures were then plated in Cm30, Nal25 and Cm30+Nal25. The transconjugants were selected in double antibiotics Cm30+Nal25 (Phornphisutthimas et al, 2007). 
     Note that before starting the conjugation experiment, the efficiency of killing of pFSMob plasmids were first tested by transformation in  Salmonella.    
     Methodology 
     Material: 
     
         
         
           
             Donor strain:  E. coli  S17 transformed with pFSMobSal7/pFSMobC* 
             Recipient strains: JM109/ Salmonella enteritidis  FS26 
             Luria Bertani Broth (LB) (Product number L3522, Sigma-Aldrich), LB Cm30 g/ml 
             Plates of LB agar 1% 
             Plates of LB agar 1% Cm30 g/ml 
             Plates of LB agar 1% NaL25 g/ml 
             Plates of LB agar 1% Cm30+NaL25 
             Whatman® membrane filters nylon, pore size 0.45 m to 1 m, diameter 25 mm. Reference number 28420770 supplied by Sigma-Aldrich 
           
         
       
    
     Experiment 1: Mobilisation of pFSMobC* from  E. coli  S17 to  E. coli  JM109 
     As a proof of concept, the conjugation experiment was done first from  E. coli  S17 containing a control plasmid (pFSMobC*) to  E. coli  JM109. This experiment followed the protocol from of Phornphisutthimas (Phornphisutthimas et al, 2007):
         1. Grow overnight cultures of donors pFSMobC*(S17) from our collection in LB+Cm30 and recipient strain JM109 in LB broth.   2. In the morning pre-warm the plates of LB agar (1%) for at least 30 minutes before starting the experiment   3. Mix the donor strains with the recipient at the ratio of 1:1 as following:   4. 50 W of pFSMobC*(S17)+50 μl of JM109   5. 50 W of pFSMobC*(S17)+50 μl of LB   6. 50 W of LB+50 μl of JM109   7. For each reaction, put a filter membrane (0.45 μM to 1 μM) on the top of the pre-warmed LB agar plate (1%)   8. Load each sample of mating on a filter   9. Leave the plates 3 hours at 37° C. without shaking, the LB will be absorbed and the bacteria should stay on the filter   10. After incubation, put each filter into a universal tube with lml LB broth and vortex well   11. Plate 100 μl (And serial dilution) on Cm30, Nal25 and Cm30+Nal25, leave O.N at 37° C.       

     The plates of Cm30 give the number of donors. Plates of Nal25 give the number of recipient and plates of Nal25+Cm30 give the number of transconjugants (i.e plasmid mobilised from donor to recipient). Plates with donors and recipient alone (no mating) are negative controls 
     Results 
     1) After 18 hours at 37° C. colonies were individually counted at different dilutions, when countable, and the number of colonies was expressed as cfu/mL. No colonies were recovered on negative control plates, as expected (Data not shown). The results (Table 1) show a good efficiency of conjugation from  E. coli  S17 to  E. coli  JM109. 
     
       
         
           
               
             
               
                 TABLE 1 
               
             
            
               
                   
               
               
                 Conjugation of pFSMobC* from  E. coli  S17 to  E. coli  JM109. Results 
               
               
                 obtained from mating on filter after 3 hours of incubation at 37° C. 
               
            
           
           
               
               
               
               
               
            
               
                   
                 Number of 
                 Number of 
                 Number of 
                   
               
               
                 Conjugative 
                 donors per ml 
                 recipient per ml 
                 transconjugants per 
               
               
                 plasmid 
                 S17 (Cm30)  a   
                 JM109 (Nal25)  a   
                 ml (Cm30 + Nal25)  a   
                 Efficiency  b   
               
               
                   
               
               
                 pFSMobC* 
                 9.46*10 4   
                 1.004*10 5   
                 2.35*10 4   
                 2.4*10 −1   
               
               
                   
               
               
                   a  All the controls have been checked. 
               
               
                   b  The efficiency of conjugation is the number of transconjugants per donor cell 
               
            
           
         
       
     
     2) The mobilisation of pFSmobC* plasmid in the recipient strain was confirmed by PCR. Primers spCas9-6 forward (5′-ATTGTTTGTGGAGCAGCATAAGC)(SEQ ID NO: 8) and mob2 reverse (5′-GCCTCTAGCACGCGTACCATGGGAT) (SEQ ID NO: 9) were used with an annealing temperature of 50° C. Two colonies from plates with transconjugants have been tested by PCR. Positive controls were also included ( FIG. 1 ). 
     Conclusion 1 
     The conjugation experiment from  E. coli  to  E. coli  has shown a good efficiency of mobilisation. Further optimisation tests on  E. coli  conjugation showed a higher efficiency of conjugation when the mating was incubated for 6 hours at 37° C. with a ratio of 1:4. 
     Experiment 2: Conjugation of pFSMobSal7 from  E. coli  S17 to  Salmonella  Spp 
     This experiment is following the protocol from of Phornphisutthimas (Phornphisutthimas et al, 2007) and Carraro et al, 2017 with some modifications:
         1. Grow overnight cultures of pFSMobSal7 (S17) and the control pFSMobC*(S17) from our collection in LB+Cm30 and recipient strain  Salmonella enteritidis  FS26 in LB broth.   2. In the morning, pre-warm the plates of LB agar (1%) for at least 30 minutes before starting the experiment   3. Mix the donor strains with the recipient at the ratio of 1:4 as following:   4. 100 μl of pFSMobC*(S17)+400 μl of  Salmonella  FS26   5. 100 μl of pFSMobSal7(S17)+400 μl of  Salmonella  FS26   6. 100 μl of pFSMobC*(S17)+400 μl of LB   7. 100 μl of pFSMobSal7 (S17)+400 μl of LB   8. 100 μl of LB+400 μl of  Salmonella      9. Spin at 2000×g for 3 minutes   10. Resuspend in 200 μl of LB   11. Spin at 2000×g for 3 minutes   12. Resuspend in 25 μl of LB   13. For each reaction, put a filter membrane (0.45 μM to 1 μM) on the top of the pre-warmed LB agar plate (1%)   14. Load each sample of mating on a filter   15. Leave the plates 6 hours at 37° C. without shaking, the LB will be absorbed and the bacteria should stay on the filter   16. After incubation, put each filter into a universal tube with 1 ml LB broth and vortex well   17. Plate 100 μl (And serial dilution) on Cm30, Nal25 and Cm30+Nal25, leave O.N at 37° C.       

     The plates of Cm30 plates give the number of donors. Plates of Nal25 give the number of recipient and plates of Nal25+Cm30 give the number of transconjugants (i.e plasmid mobilised from donor to recipient). 
     Results 
     
         
         
           
             After 18 hours at 37° C. colonies were individually counted at dilution −3, and the number of colonies was expressed as cfu/mL (Table 2). No colonies were recovered on negative control plates, as expected (Data not shown). 
           
         
       
    
     
       
         
           
               
             
               
                 TABLE 2 
               
             
            
               
                   
               
               
                 Conjugation experiment results of pFSMob plasmids from  E. coli  S17 to  Salmonella   
               
               
                   enteritidis  FS26. Results obtained from mating of a filter for 6 hours at 37° C. 
               
            
           
           
               
               
               
               
               
               
            
               
                   
                 Donor 
                 Recipient 
                   
                 Number of 
                   
               
               
                 Conjugative 
                 S17/ml 
                 FS26/ml 
                 Number of 
                 transconjugants/ 
                 Conjugation 
               
               
                 plasmid 
                 (Cm30) 
                 (Nal25) 
                 transconjugants 
                 ml (Cm30 + Nal25) 
                 efficiency a   
               
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 pFSMobC* 
                 6.6*10 6   
                 Full 
                 72 
                 7.2*10 5   
                 1.09*10 −1   
               
               
                 pFSMobSal7 
                 4.7*10 6   
                 Full 
                 0 
                 0 
                 0 
               
               
                   
               
            
           
         
       
         
         
           
             In comparison with the plasmid control pFSMobC*, these data show that the plasmid pFSMobSal7 delivered to the target strain by conjugation is able to remove by 100% the strain  Salmonella enteritidis  (FS26) ( FIG. 2 ). 
           
         
       
    
     Conclusion 
     The study showed that the conjugation is a good method of delivery from  E. coli  S17 to  Salmonella enteritidis  (FS26) and that pFSMobSal7 is still efficiently killing when delivered to the recipient by conjugation. Conjugation was able to remove (kill) 100% of the  Salmonella.    
     REFERENCES 
     
         
         Carraro N, Durand R, Rivard N, Anquetil C, Barrette C, Humbert M, Burrus V. (2017).  Salmonella  genomic island 1 (SGI1) reshapes the mating apparatus of IncC conjugative plasmids to promote self-propagation. PLOS Gen. 
         O&#39;Challahan D, Charbit A. (1990). High efficiency transformation of  Salmonella typhimurium  and  Salmonella typhi  by electroporation. Mol. Gen. Genet., 223; 156-8. 
         Phornphisutthimas S, Thamchaipenet A, and Panijpan B. (2007). Conjugation in  Escherichia coli. Biochem and Mol Bio Education Vol.  35, No. 6, 440-445 
       
    
     Example 3: Pan-Serotype Conjugative Transfer and Killing of  Salmonella    
     Purpose of the Study 
     The purpose of this study was to test the in vitro ability of pFSmobSal7 and pFSmobSal3 plasmids to selectively remove  Salmonella enterica  subsp.  enterica  serotypes by conjugation. 
     Summary 
     Eighteen strains of  Salmonella enterica  subsp.  enterica , serovars  Typhimurium  (4),  Enteritidis  (1), Virchow (1), Montevideo (1), Heidelberg (1), Hadar (1), Binza (1), Bredeney (1),  Infantis  (1), Kentucky (1), Seftenberg (1), Mbandaka (1), Anatum (1), Agona (1) and Dublin (1), were selected from our  Salmonella  spp. collection (Table 3). Bacteria were made competent and transformed with two versions of pFSmob plasmid, i.e. pFSmobSal7 and pFSmobSal3, two versions of pFSmob control plasmid, i.e. pFSmob-C* and pFSmob-C, and pZA31MCS (Expressys) as further control. After electroporation and 2 hrs of incubation in SOC Outgrowth Medium (NEB, B90205, LOT-10019857) at 37° C., different dilutions of the original solution were plated out on 1% LB agar supplemented by chloramphenicol (Cm 30 , 30 g/mL). Plates were incubated at 37° C. for 18 hrs. Results were interpreted as colony forming unit/mL (CFU/mL). 
     Introduction 
     The pFSmobSal7 and pFSmobSal3 plasmids carry on their crRNA arrays 7 target genes (pipA, pipB, pipC, hilA, sicP, mart, sopB), and 3 target genes (pipC, hilA, mart), respectively. Positive control pFSmob-C originated from pFSmobSal7 plasmid was used, in particular pFSmob-C does not contain the crRNA array. Control pFSmob-C originated from the intermediated copy number pZA31MCS (Expressys) used as a further control. Selection of positive colonies was performed on chloramphenicol (Cm 30 , 30 g/mL). It was previously showed the in vitro ability of pFSmobSal7 to selectively remove by transformation  Salmonella enterica  serovar  Typhimurium, Enteritidis , Virchow, Montevideo, Hadar and Binza. The purpose of this study was to assess all the  Salmonella  spp. serotypes present in our collection, in order to evaluate the in vitro ability of both pFSmobSal7 and pFSmobSal3 plasmids to selectively unwanted bacteria; in order to collect significant results, 18 different serotypes were selected from our collection and tested by electroporation (Table 3). Plasmids pFSmob-C, pFSmob-C* and pZA31MCS (Expressys) were used as controls. 
     Methodology 
     Bacterial strains were recovered from frozen stock kept at −80° C., and cultured at 37° C. for 24 hrs (Edwards and Ewing, 1986). Susceptibility to Cm 30  (30 g/mL) was tested using an agar dilution method in accordance with EUCAST clinical breakpoint tables. Bacteria were made competent using O&#39;Challahan and Charbit protocol (O&#39;Challahan and Charbit, 1990). All the selected strains were transformed with 100 ng of pFSmobSal7, pFSmobSal3, pFSmob-C, pFSmob-C*, and pZA31MCS (Expressys); a negative control was also included (competent cells without the addition of DNA) (Table 4). After 18 hrs of incubation, countable colonies (number of colonies between 30 and 300) were expressed as number of colonies per mL (CFU/mL) (Table 4). 
     
       
         
           
               
             
               
                 TABLE 3 
               
             
            
               
                   
               
               
                   Salmonella  spp. collection. Eighteen strains of  Salmonella enterica   
               
               
                 subsp.  enterica , serovars Typhimurium (4), Enteritidis (1), 
               
               
                 Virchow (1), Montevideo (1), Heidelberg (1), Hadar (1), Binza 
               
               
                 (1), Bredeney (1), Infantis (1), Kentucky (1), Seftenberg (1), 
               
               
                 Mbandaka (1), Anatum (1), Agona (1) and Dublin (1), were selected 
               
               
                 from our  Salmonella  spp. collection. 
               
            
           
           
               
               
               
            
               
                   
                 Collection no 
                   Salmonella  sertypes 
               
               
                   
                   
               
               
                   
                 FS2 
                 Typhimurium 
               
               
                   
                 FS3 
                 Typhimurium 
               
               
                   
                 FS10 
                 Typhimurium 
               
               
                   
                 FS11 
                 Typhimurium 
               
               
                   
                 FS11 
                 Typhimurium 
               
               
                   
                 FS26 
                 Enteritidis 
               
               
                   
                 FS35 
                 Virchow 
               
               
                   
                 FS38 
                 Montevideo 
               
               
                   
                 FS41 
                 Heidelberg 
               
               
                   
                 FS42 
                 Hadar 
               
               
                   
                 FS43 
                 Binza 
               
               
                   
                 FS44 
                 Bredeney 
               
               
                   
                 FS50 
                 Infantis 
               
               
                   
                 FS58 
                 Kentucky 
               
               
                   
                 FS60 
                 Seftenberg 
               
               
                   
                 FS61 
                 Mbandaka 
               
               
                   
                 FS62 
                 Anatum 
               
               
                   
                 FS64 
                 Agona 
               
               
                   
                 FS67 
                 Dublin 
               
               
                   
                   
               
            
           
         
       
     
     Results 
     All the bacteria selected were found to be susceptible to Cm 30  according to EUCAST breakpoints (EUCAST, 2013). After electroporation and plating of different dilutions (undiluted, 10 −1 , 10 −2 ) on LB agar plates (Fisher Bioreagents, BP1425-500, LOT-171784) supplemented with Cm 30  (30 g/mL) (Sigma), plates were incubated for 18 hrs at 37° C. Bacteria were considered transformed when they grew on Cm 30  (30 g/mL) agar plates. Countable colonies (between 30 and 300) were expressed as CFU/ml (Table 4). No colonies were recovered on negative control plates, as expected. All the serotypes but one (FS67, S. Dublin) were efficiently transformed with pZA31MCS (Expressys). Almost all the serotypes were efficiently transformed with the two control plasmids (pFSmob-C* and pFSmob-C) with a number of colonies between 10 2  and 10 6  CFU/mL. 
     With regard to pFSmobSal7 and pFSmobSal3, the killing effect occurred in all the serotypes and it was found to be similar. In the majority of the serotypes, a decreasing in the number of colonies was relatively evident in comparison to the ones transformed with the control plasmids. No significant differences were found between pFSmobSal7 and pFSmobSal3 in their killing effect, except in one strain (FS38, S. Montevideo). 
     
       
         
           
               
             
               
                 TABLE 4 
               
             
            
               
                   
               
               
                 Transformation of 18  Salmonella  spp. serotypes. Eighteen  Salmonella   
               
               
                 spp. Serotypes were transformed with pZA31MCS (Expressys), pFSmob-C*, pFSmob- 
               
               
                 C, pFSmobSal7, and pFSmobSal3. Almost all the serotypes were efficiently 
               
               
                 transformed with pZA31MCS (Expressys), pFSmob-C*, and pFSmob-C, with some 
               
               
                 exceptions. No significant differences were observed in the transformation 
               
               
                 with pFSmobSal7 and pFSmobSal3. Strain FS67, S. Dublin, was found to be 
               
               
                 not transformable (NT*) with all the plasmids used in this study. 
               
            
           
           
               
               
               
               
               
               
               
            
               
                 Collection 
                 Serotype 
                 pZA31MCS 
                 pFSmob-C* 
                 pFSmob-C 
                 pFSmobSal7 
                 pFSmobSal3 
               
               
                   
               
               
                 FS2 
                 Typhimurium 
                 4.8*10{circumflex over ( )}4  
                 1*10{circumflex over ( )}5 
                 4*10{circumflex over ( )}4 
                 0 
                 6*10{circumflex over ( )}1 
               
               
                 FS3 
                 Typhimurium 
                  5*10{circumflex over ( )}5 
                 1*10{circumflex over ( )}5 
                 3*10{circumflex over ( )}3 
                     1*10{circumflex over ( )}1 
                 6*10{circumflex over ( )}1 
               
               
                 FS10 
                 Typhimurium 
                  5*10{circumflex over ( )}5 
                 0 
                 1.1*10{circumflex over ( )}2     
                 0 
                 0 
               
               
                 FS11 
                 Typhimurium 
                 4.7*10{circumflex over ( )}4  
                 7*10{circumflex over ( )}2 
                 4.1*10{circumflex over ( )}2     
                 0 
                 0 
               
               
                 FS26 
                 Enteritidis 
                 &gt;1*10{circumflex over ( )}6 
                 9*10{circumflex over ( )}5 
                 &gt;1*10{circumflex over ( )}6  
                     3*10{circumflex over ( )}1 
                 4*10{circumflex over ( )}1 
               
               
                 FS35 
                 Virchow 
                 &gt;1*10{circumflex over ( )}6 
                 2.8*10{circumflex over ( )}3     
                 1.3{circumflex over ( )}10{circumflex over ( )}4 
                 0 
                 0 
               
               
                 FS38 
                 Montevideo 
                 &gt;1*10{circumflex over ( )}6 
                 2.1*10{circumflex over ( )}4     
                 4.2*10{circumflex over ( )}4     
                 1.4*10{circumflex over ( )}2 
                 0 
               
               
                 FS41 
                 Heidelberg 
                 1.1*10{circumflex over ( )}4  
                 4.1*10{circumflex over ( )}2     
                 6*10{circumflex over ( )}2 
                 0 
                 0 
               
               
                 FS42 
                 Hadar 
                 &gt;1*10{circumflex over ( )}6 
                 7*10{circumflex over ( )}2 
                 0 
                 0 
                 0 
               
               
                 FS43 
                 Binza 
                 1.7*10{circumflex over ( )}4  
                 7*10{circumflex over ( )}1 
                 1*10{circumflex over ( )}1 
                 0 
                 0 
               
               
                 FS44 
                 Bredeney 
                 &gt;1*10{circumflex over ( )}6 
                 1*10{circumflex over ( )}5 
                 9*10{circumflex over ( )}4 
                 1.7*10{circumflex over ( )}3 
                 3*10{circumflex over ( )}3 
               
               
                 FS50 
                 Infantis 
                  1*10{circumflex over ( )}4 
                 1.8*10{circumflex over ( )}2     
                 2*10{circumflex over ( )}3 
                 0 
                 4*10{circumflex over ( )}1 
               
               
                 FS58 
                 Kentucky 
                 1.7*10{circumflex over ( )}4  
                 7.5*10{circumflex over ( )}3     
                 3.6*10{circumflex over ( )}3     
                 0 
                 0 
               
               
                 FS60 
                 Seftenberg 
                 &gt;1*10{circumflex over ( )}6 
                 1.2*10{circumflex over ( )}5     
                 &gt;1*10{circumflex over ( )}5  
                     2*10{circumflex over ( )}1 
                 2.4*10{circumflex over ( )}2     
               
               
                 FS61 
                 Mbandaka 
                  2*10{circumflex over ( )}2 
                 2*10{circumflex over ( )}1 
                 0 
                 0 
                 0 
               
               
                 FS62 
                 Anatum 
                 1.4*10{circumflex over ( )}3  
                 4*10{circumflex over ( )}1 
                 0 
                 0 
                 0 
               
               
                 FS64 
                 Agona 
                 &gt;1*10{circumflex over ( )}6 
                 &gt;1*10{circumflex over ( )}6  
                 3*10{circumflex over ( )}4 
                 7.8*10{circumflex over ( )}2 
                 3*10{circumflex over ( )}3 
               
               
                 FS67 
                 Dublin 
                 NT* 
                 NT 
                 NT 
                 NT 
                 NT 
               
               
                   
               
            
           
         
       
     
     Conclusion 
     All the  Salmonella  spp. serotypes used in this experiment were efficiently made competent and transformed with pZA31MCS (Expressys), pFSmob-C and pFSmob-C* used as controls.  Salmonella Typhimurium  serotype seems not to be always transformable by pFSmob-C*, as observed previously. More in general, no particular differences occurred between the two pFSmob-derived control plasmids used in this study (between 10 2  and 10 6  CFU/mL). On the other hand, the killing effect of pFSmobSal7 and pFSmobSal3 was found to be similar (between 101 and 103). In conclusion, in this study it was demonstrated that pFSmobSal7 and pFSmobSal3 can selectively remove different  Salmonella  spp. serotypes in comparison with the control plasmids. 
     REFERENCE 
     
         
         Edwards P R, Ewing W H. (1986). Identification of Enterobateriaceae (4 th  ed.). Elsevier, New York. EUCAST, European Committee on Antimicrobial Susceptibility Testing. (2013). Breakpoint tables for interpretation of MICs and zone diameters. Available at: http://www.eucast.org (last accessed Aug. 16, 2013, Version 3.1. 
         O&#39;ChallahanD, Charbit A. (1990). High efficiency transformation of  Salmonella typhimurium  and  Salmonella typhi  by electroporation. Mol. Gen. Genet., 223; 156-8. 
       
    
     Example 4: Feed Conversion Ratio Improvement in Chickens. Combined Second 2 Week Efficacy, 2 Week Safety and 6 Week Productivity Trial 
     Purpose of study
         To determine the effect antibacterial conjugative plasmids (pFSmobSal7) on the health, welfare and productivity of chickens over 6 weeks.       

     Methodology 
     Ross 308 birds were housed under controlled biosecure conditions and given water and standard commercial rations ad libitum. 
     Birds were dosed continually with either:
         1. No addition to water (60 birds)   2. Strain S17 @ 10 8  cfu/ml drinking water (30 birds)   3. Strain S17 containing pFSmob-C* @ 10 8  cfu/ml drinking water (30 birds)   4. Strain S17 containing pFSmobSal7 @ 10 8  cfu/ml drinking water (60 birds)   5. Strain S17 containing pFSmobSal7 @ 10 9  cfu/ml drinking water (30 birds)   6. Strain S17 containing pFSmobSal7 @ 10 10  cfu/ml drinking water (30 birds)       

     Bacterial strains were recovered from frozen stocks kept at −78 C and cultured on LB agar for 24 h at 37 C. Cultures were prepared daily in LB broth containing antibiotic to prevent loss of plasmid with shaking at 180 rpm for 16 h at 37 C then centrifuged for 10 min at 4000×g. Medium was removed and the pellet resuspended in PBS then diluted in PBS to give solution for dosage to chickens. 
     In parallel, a group of 30 birds was dosed orally with 0.5 ml 10 5  CFU/mL  Salmonella Enteritidis  strain FS26 (Folium) on day 1. Birds were checked for  Salmonella  colonisation at day 3 of the experiment by cloacal swab using ISO 6759 methods (1). On day 5 of the experiment 3 of these verified  Salmonella -colonised birds (seeder birds) were marked and added to each of groups 1-6. Birds were weighed weekly and feed consumption and mortality recorded. 
     Fifteen birds from each group were euthanised on days 12 and 19 (7 and 14 days post mixing with seeder birds). Caeca were removed for examination for  Salmonella  by ISO 6759. Hock and pad marks were recorded. Samples of 1 g liver and caecal contents were snap frozen. 
     Thirty birds from each of group 1 and 4 were then monitored for behaviour and weight weekly until day 42, when they were euthanised and examined as for birds above. 
     Results were recorded on paper or dictated via telephone or radio in biosecure accommodation and transcribed to Microsoft Excel™. For the purposes of data transformation birds with no bacteria detected were allocated a count of 1 bacterium per g. Counts and weight were log transformed and statistical analysis conducted using GraphPad Prism™. Data was assessed for normality of distribution using a D&#39;Agostino and Pearson omnibus normality test and as non-normal was analysed using a Kruskall-Wallis test with Dunn&#39;s multiple comparison test post hoc. Differences in proportions of birds was analysed using Fisher&#39;s exact test. 
     Results 
     All seeder birds were colonised with  Salmonella  by day 3. Those used to seed infections in the test groups had counts of &gt;10 5  cfu/g faeces. 
     At day seven 15 birds per group were euthanised and  Salmonella  in the caecum enumerated ( FIG. 3 ).  FIG. 4  shows data presented a birds positive/negative for  Salmonella.    
     Birds dosed with S17-pFSmobSal7 (lowest dose) and pFSmob-C* had significantly lower numbers of  Salmonella  in the caecum than controls ( FIG. 3 ) and were significantly less likely to have detectable  Salmonella  in the caecum at day 7 post mixing with seeder birds. 
     No mortality was seen in any group; no morbidity was observed and high dosage of S17-mobSal7 was tolerated well. Birds were scored for positive and negative behaviours, but no difference was seen between groups. 
     Thirty birds from each of groups 1 and 4 were monitored further until 42 days of age. During this period again, no significant differences in behaviour were observed. Feed consumption between groups was not significantly different and weights did not differ significantly between groups measured in life. At post mortem  Salmonella  was not detectable in the caecum by direct count; no hock or pad marks were observed. Median carcass weight of birds in the S17-pFSmobSal7 group was 205 g greater than in the control group, a significant increase ( FIG. 5 ). As this was associated with equivalent feed consumption between groups, the S17-pFSmobSal7 group had a significantly improved feed conversion ratio (FCR). 
     REFERENCES 
     
         
         ISO. 2007. ISO 6579:2007: Microbiology of food and animal feeding stuffs—Horizontal method for the detection of  Salmonella  spp. (ISO 6579:2002+Amd 1:2007). Geneva, Switzerland 
       
    
     Example 5: Non-Replicative Conjugative Plasmid with Conditionally Essential Gene Marker 
     To meet regulatory requirements, any plasmid used will preferably be devoid of an antibiotic selection marker and will be non-replicative in the vast majority, if not all, cells except for the host cell (ie, carrier cell). To render the plasmid non-replicative, the replication system of broad host range plasmid RK2, a fairly low copy number plasmid, will be utilised. RK2 replication is dependent on the presence of a replication protein, TrfA encoded by trfA on RK2, and binding of TrfA to the vegetative origin (oriV) of the plasmid and does not utilize the host machinery for replication initiation. Physical separation of trfA from the plasmid and incorporation of this gene into the chromosome will make the resulting plasmid carrying RK2 oriV dependent on a host encoded function. This will prevent the plasmid from actively replicating in a target strain after delivery via conjugation. In the rare case of an identical IncP plasmid being present in the target cell that will provide trfA, both plasmids will compete for the available TrfA, resulting in loss of one plasmid. As our plasmid in this example further does not carry an advantageous antibiotic resistance marker or similar and lacks plasmid addiction systems, it will be quickly lost from the offspring. 
     As a selection marker in the absence of an antibiotic resistance, the aroA gene encoding for an enzyme in the biosynthesis pathway of aromatic amino acids will be used. This is a conditionally essential gene that is essential when the host cell is grown in the absence of available aromatic amino acids or an intermediate from the reaction catalysed by aroA. Moving the aroA gene from the host genome to the plasmid backbone will provide this selection marker. The aroA gene of the chromosome will be replaced in the host (carrier cell) strain by a copy of trfA. 
     An aroA knock-out and replacement of the chromosomal copy by a trfA expression cassette in a  E. coli  lab strain DH10B will be made to generate a test strain of bacteria for testing the plasmid. The trfA expression cassette to be used has the sequence of SEQ ID NO: 10. 
     The final host (carrier) strain, a commensal  E. coli  isolate from chicken, will undergo the same strain construction procedure. 
     The aroA coding sequence (SEQ ID NO: 11) will be amplified with a similar promoter and terminator region and assembled into the plasmid along with modules for the Cas9, tracrRNA, crRNA and the oriV of RK2. 
     Additionally, alternative modules can be tested in this plasmid configuration such as anti-restriction genes that inhibit Type I restriction enzymes (ocr of T7, klcA of RK2, ardA from conjugative plasmids/transposons, ardB from conjugative plasmids) to improve DNA stability after transfer into target cells. 
     Further or alternatively, a module encoding an essential part of the type IV secretion system present for conjugation will located on the plasmid as this will lead to a non-functional transfer system present in the cell if the plasmid is not present. To facilitate conjugation by the host cell, the type IV secretion system present on plasmid RK2 will have to be integrated into the host genome. The essential components of this system are encoded within three operons on two locations of the plasmid, tra1 and tra2. Tral encodes the the traKLM and traJXIHGF operons, tra2 the trbBCEFGHJL genes. During the integration process, the regulatory region present in tra1 located between the traKLM and traJXIHGF operons will be modified. This sequence contains the origin of transfer (oriT), harbouring the binding site for the TraJ protein responsible for initiating transfer of the DNA. If this binding site is not altered by mutagenesis to prevent TraJ binding, transfer of chromosomal DNA fragments during the conjugation process will be initiated, leading to transfer of unwanted genetic information. 
     Sequences of the RK2 tra1 and tra2 modules to be used are in SEQ ID NOs 13 and 14. 
     Example 6: Anti- Salmonella  Plasmid Construction &amp; Testing 
     Summary 
     The type I-E Cas system from  E. coli  K12 (MG1655) is a RNA guide-directed DNase machinery, known as Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) and CRISPR-associated proteins system (CRISPR-Cas), which represents an adaptive immune system for prokaryotes that targets invading foreign genetic material for degradation. The  Escherichia coli  type I-E Cascade system is made up of different Cas proteins (casABCDE12 and cas3) and recognizes a wide variety of PAM sequences with varying degrees of efficacy. The main PAM sequences are: 5′-AAG, AGG, ATG, GAG-3′. Type I-E Cas system from  E. coli  K12 was used to selectively remove  Salmonella  species. The system was modified as in  E. coli  MG1655 the main components of the cas protein complex are expressed from two transcription units, cas3 and casABCDE12, respectively. The modifications introduced during the plasmid construction process included the exchange of the native, regulated promoter element of the casABCDE operon with the constitutive J23114 promoter and replacing the cas1 and cas2 genes with the cas3 gene including its native ribosomal binding site. These changes, in effect, created a constitutively expressed CRISPR-Cas module that was subsequently tested for functionality by addition of a crRNA array for specific targeting of  Salmonella  species. 
     Two versions of Guided Biotic® plasmids were constructed differing in origin of replication (oriV) and antibiotic selection marker. The resulting plasmids were named pFS-Sal-08-rm and pFS-Sal-09-rm and were tested in vitro to selectively inhibit growth of  Salmonella enterica  subsp.  enterica  serovar  Enteritidis  by conjugative DNA transfer. In the conjugation experiments, pFS-EcoCas3-01-rm and pFS-EcoCas3-03-rm were used as control plasmids for  Salmonella -specific growth inhibition Plasmids were constructed, each comprising nucleotide sequences encoding Type I-E  E. coli  Cas3 and cognate casA, B, C, D and E; a CRISPR array that is operable with the Cas3 and comprises a first spacer that is complementary to a sequence of an invB gene of  S. enterica , a second spacer that is complementary to a sequence of a sicP gene of  S. enterica  and a third spacer that is complementary to a sequence of a sseE gene of  S. enterica ; an RP4 oriT; a p15A ori; an  E. coli  proA gene; and an  E. coli  proB gene. See Table 7 for more details. 
     Methodology 
     crRNA Array Design 
     Target gene selection, to design the CRISPR spacers, was focused on the  Salmonella -specific pathogenicity islands (SPIs), which are major virulence factors for  Salmonella . Criteria for selection were conservation within  Salmonella enterica  and a low to very low occurrence and conservation of the respective gene in other bacteria. These analyses were performed using the Basic Local Alignment Search Tool (BLAST). Cut-off for number of hits in non- Salmonella  species was set at 1000. Level of conservation was considered as high if a search produced &gt;1000 hits. Between 100 and 1000 was medium, between 20 and 100 low, very low if &lt;20 hits. Specifically, genes associated with the type III protein secretion systems, T3SS, of SPI-1 and SPI-2 were explored as targets, taking care to include gene candidates that are conserved across  Salmonella  species but exclude those that are conserved across other bacteria species. Initially, the search focused on secreted effectors and chaperones, avoiding regulators and functional components of the T3SS. 
     The three selected highly conserved gene targets sicP, invB and sseE are all part of the T3SS loci encoded by  Salmonella  pathogenicity islands (SPIs), SPI-1 in the case of sicP and invB (both chaperones), and SPI-2 in the case of sseE (secreted effector). They all occur in only one copy in  Salmonella enterica  subsp.  enterica  serovar  Typhimurium  str. LT2 and this is the case for all  Salmonella enterica  although their duplication in some strains cannot be ruled out. 
     Candidate target genes (a) and, subsequently, spacers (b) were searched on Blast to make sure they are specific for  Salmonella  and to minimize off-targets. Search was performed using RefSeq Database, which contains around 10600  Salmonella enterica  genomes:
         a) Blast search parameters for gene selection:
             Salmonella  conservation: RefSeq DB restricted to  Salmonella enterica  (taxid:28901), megablast, max hits: 20000, standard parameters   Occurrence outside  Salmonella : RefSeq DB restricted to bacteria (taxid:2) excluding  Salmonella  (taxid:590), Blastn, max hits 1000, standard parameters   
           b) Blast search parameters for spacer sequences (including PAMs):
             Salmonella  conservation: RefSeq DB restricted to  Salmonella enterica  (taxid:28901), blastn, max hits:20000, standard parameters   Occurrence outside  Salmonella : RefSeq DB restricted to bacteria (taxid:2) excluding  Salmonella enterica  (taxid:28901), Blastn, max hits 1000, standard parameters but expect value set to 100 to catch sequences with low but still significant homology   
               

     A number of spacers targeting different  Salmonella enterica  genes were initially screened in silico, selected, cloned and individually tested in vitro for their efficacy of targeting  Salmonella enterica  subsp.  enterica  serovar  Enteritidis  FS26 (see Summary on target gene and spacer selection and Rep-21: Construction and in vitro testing of spacer reporter systems). The three best performing spacer sequences active against  Salmonella  species were selected and combined into a short crRNA array. The spacer sequences were derived from three highly conserved genes present in  Salmonella  pathogenicity islands (SPIs), invB, sicP and sseE. In brief, a limited potential spacer sequences characterized by the presence of a strong consensus 5′-PAM sequence capable of initiating DNA restriction were selected and ranked according to sequence features associated with efficient target DNA restriction, conservation within  Salmonella  isolates and lack of DNA homology outside of  Salmonella enterica . Three spacer sequences meeting these criteria were finally selected and incorporated into a functional crRNA array. The finalised crRNA array was ordered as a synthetic gene sequence inserted into a standard cloning plasmid (GeneArt, Thermo Fisher Scientific). 
     Sequences of selected genes are shown in SEQ ID NOs: 20-22. In each gene, spacers are highlighted in bold and respective PAMs in italics (in the case of invB, the spacer was selected on the negative DNA strand (anti-sense) so the PAM appears 3′ to the spacer. PAM sequences used for spacer selection were AAG, ATG, AGG and GAG. Blast searches for the spacers with their respective PAMs was conducted (Table 10). 
     The non- Salmonella  hits were further analysed for their conservation level (spacers accepted when level of conservation in off-targets was &lt;75%). Sequence selection also depended on where the mismatches were located in the off-target hit, ruling out the ones showing conserved PAM and seed (first 8pb of the spacer). One additional criterion for our selection also included the type of bacteria where the hit was present, if present in a pathogen, the spacer was still considered as a possible candidate. 
     Each spacer was individually tested in vitro for their efficacy of targeting  Salmonella enterica  subsp.  enterica  serovar  Enteritidis  FS26, before combining them together in the array present in the plasmid pFS-Sal-09-rm. 
     Plasmid Assembly 
     For the homology-based DNA assembly, individual DNA modules were carrying 5′ and/or 3′ extensions with exact homology to the adjacent module. The extensions were either introduced during a PCR amplification step of the module or were incorporated during the design of synthetic DNA modules. The constitutive J23114 promoter was introduced. 
     CasA-E: The 4464 bp casA-casB-casC-casD-casE module was amplified from  E. coli  K12 (MG1655) genomic DNA. 
     Cas3: The 2709 bp Cas3 module was amplified. 
     crRNA: The crRNA array module Sal-crRNA 1 (546 bp), carries a 25 bp homology extension to the 3′-end of the Cas3 module. The sequence of the Sal-crRNA Array1 is shown in SEQ ID NO: 15, with the −10 region of the promoter in italic, direct repeats underlined and the spacer regions corresponding to the selected  Salmonella  target sequences of invB, sicP and seeE, in bold. 
     pFS-Sal-08-Rm and pFS-Sal-09-Rm Validation and In Vitro Testing by Conjugation 
     Having confirmed the sequences of both constructs, the plasmids were transformed into a carrier strain that allowed plasmid mobilization via conjugation to assess the efficacy of the Cas3 system to selectively kill  Salmonella  in presence of the Sal-crRNA array 1. In this study,  E. coli  strain S17-1 ΔTn7 was used. 100 ng of pFS-Sal-08-rm and pFS-Sal-09-rm were used for transformation of 50 l electrocompetent cells of  E. coli  S17-1 ΔTn7 by electroporation and selected on Kanamycin and Chloramphenicol, respectively. Transformation with both plasmids resulted in a good transformation efficiency of  E. coli  S17-1 ΔTN7 (10-10 6  transformed CFU/mL). Control plasmids, pFS-EcoCas3-01-rm and pFS-EcoCas3-03, lacking the crRNA array, were also transformed in the S17-1 ΔTn7 carrier strain to be used as controls in FS26 transformation for pFS-Sal-09-rm and pFS-Sal-08-rm. In order to confirm the mobilization capability of the new plasmids, S17-1 carrier cells, were first used to conjugate  E. coli  JM109 cells (nalidixic acid, Nal, resistant). Results, reported in Table 8, show a similar conjugation efficiency between pFS-EcoCas3-01 and pFS-Sal-09-rm and between pFS-EcoCas3-03 and pFS-Sal-08-rm, in  E. coli  JM109. Conjugation efficiency is calculated dividing the number (CFU/mL) of transconjugants (selected on plates with double antibiotics) by the number of recipients (selected on plate with Nal). 
     Subsequently,  E. coli  carrier strain S17-1 ΔTn7, previously transformed with the plasmids, were used to assess whether there is a significant growth inhibition of  Salmonella Enteritidis  FS26 by conjugation of pFS-Sal-08-rm compared to the control plasmid pFS-EcoCas3-03-rm and of pFS-Sal-09-rm compared to pFS-EcoCas3-01-rm. Conjugation was performed from  E. coli  S17-1 ΔTn7 into nalidixic acid (Nal) resistant  Salmonella Enteritidis  FS26. Results showed a significant reduction (CFU/mL) in S.  Enteritidis  strain FS26 conjugated with both pFS-Sal-08-rm and pFS-Sal-09-rm compared to the respective control plasmids (Table 9). 
     In conclusion, inhibition of  Salmonella  growth by both plasmids carrying the Sal-crRNA array 1 generated in this study indicated that the modified Cas module is functional and was able to express all the individual components of the Type I-E Cascade complex, the Cas3 nuclease and the crRNA array without negatively affecting growth of non-target  E. coli  host strains 517-1 and JM109. We demonstrated the ability of the  E. coli  type I-E Cas system-based plasmids to be transferred by conjugation from the  E. coli  S17-1 carrier strain to S.  Enteritidis  strain FS26. We observed a significant inhibition of growth in the  Salmonella  transconjugants (&gt;99.9%) of the selected strain through conjugation of both pFS-Sal-08-rm and pFS-Sal-09-rm in comparison to the control used. These data therefore highlighted the potential of these  E. coli  type I-E Cascade based constructs for the removal of unwanted bacteria, such as for zoonotic control. 
     Example 7: Test of Stability and Efficacy of a Folium  E. coli -Based Product in Chickens Fed a Diet Containing  Salmonella    
     In Example 4, we tested a Guided Biotic® plasmid contained in  E. coli  carrier cells of S17 strain. In this present Example 7, we instead tested a Guided Biotic® plasmid (GB plasmid pFS-Sal-09-proAB-rm, Example 6 &amp; Table 7) contained in a different strain (Strain X) of  E. coli  carrier. A helper plasmid pCon_aroA carried a functional copy of the conjugation machinery of plasmid RP4 (traJXIHGF-traKLM-trbBCDEFGHIJKL) that was found to facilitate mobilization of oriT-containing plasmids from the host cell to a recipient cell in vitro. 
     Purpose of Study 
     
         
         
           
             To determine the stability of  E. coli  Strain X with the Guided Biotic® in drinking water over 24 hours. 
             To enumerate intestinal and organ contamination with  Salmonella  given in-feed. 
             To determine the efficacy of  E. coli  Strain X containing an active GB plasmid in chickens challenged with  Salmonella  in-feed. 
           
         
       
    
     Summary 
     
         
         
           
               E. coli  Strain X was found at the expected level of 5×log-8 CFU/mL immediately after dosing into deionised water containing stabilizers, and 24 hrs had dropped to 8×log-7 CFU/mL. 
             Following challenge in-feed (10 4  CFU/g feed) for 24 hrs, low levels (˜log-2-5 CFU/g) of  Salmonella  were found in the crop and caeca on days 1, 3 and 7 after challenge. Lower levels (˜log-1 CFU/g) were also found in ileal contents, liver and spleen 7 days post-challenge. 
             The active Guided Biotic®,  E. coli  Strain X containing an active GB plasmid, reduced (P&lt;0.03)  Salmonella  counts in the crop 7 days post-challenge by ˜log-1 CFU/g. 
           
         
       
    
     Methodology 
     Ross 308 birds (30) were housed under controlled biosecurity conditions and given water and a standard commercial ration ad libitum. Each experimental group was held in a separate pen with wood shaving bedding. Birds had a 18 h light/6 h dark lighting regime. Temperature and humidity were kept between the standard levels shown in Table 11. 
     Treatment Groups: 
     
         
         
           
             Low Control—no addition to water, 10 4  CFU  Salmonella  per gram feed for 24 hrs on day 7 (15 birds) 
             GB-Sal— E. coli  Strain X Guided Biotic® containing active GB plasmid (pFS-Sal-09-proAB-rm) at 10 8  CFU/mL in drinking water from days 1-14, 10 4  CFU  Salmonella  per gram feed for 24 hrs on day 7 (15 birds) 
           
         
       
    
     Guided Biotic® strains were recovered daily from frozen stocks kept at −78° C. and cultured on LB agar plus Chloramphenicol (30 g/mL) for 24 h at 37° C. Strain X with active GB plasmid strains were prepared in Terrific broth, grown with shaking at 180 rpm for 16 h at 37° C. After measuring OD 600  to estimate cell numbers, cells were centrifuged and resuspended in deionised water containing stabilizers to give solutions for dosage to chickens (10 8  CFU/mL in drinking water). Vac Pac containing blue dye was included 3 hrs before collection on sampling days, to confirm water and GB intake. Supplemented water samples were collected on GB addition and 24 hrs later on days 4-14 of the trial, for enumeration for GB. Water samples were decimally diluted in PBS and then plated on to MacConkey agar number 3 (Oxoid CM0115), then incubated at 24 h at 37° C. 
       Salmonella  strain (FS26, with Nalidixic acid marker) was recovered from frozen stocks kept at −78° C. and cultured on LB agar for 24 h at 37° C. Test cultures were prepared daily in LB broth grown with shaking at 180 rpm for 16 h at 37° C. After measuring OD 600  to estimate cell numbers, cells were centrifuged and resuspended and diluted in phosphate-buffered saline to give solutions of 1×10 7  CFU/ml which was dripped on to the feed at a rate of 1 ml bacterial suspension to 100 g feed while the feed was mixed thoroughly. Four feed samples were collected for enumeration of  Salmonella  content. 
     Animal Sample Collection and Plating 
     No bird mortality was recorded. Lighting level, additions of bedding, temperature, humidity and stocking density were logged and did not differ between treatments. Three birds from each group on days 1 &amp; 3 and nine birds on day 7 after challenge were euthanised and samples taken from the crop, ileal, caecal, liver and spleen. Euthanasia, post-mortem and dissection times were recorded. Feathers were spray wetted with water. Birds were opened with a disposable scalpel that was discarded after opening and gloves were changed. Samples of 1 g liver and crop, ileal and caecal contents were taken and snap frozen in liquid nitrogen. Scalpels were disposed of as each organ was taken. Samples were homogenised in 9 volumes of phosphate-buffered saline, decimally diluted in phosphate-buffered saline and examined, with agars incubated for 16-18 h at 37° C. Samples that were negative in direct plating were examined for  Salmonella  by enrichment in 9 volumes of selenite cystine broth (Oxoid CM0699) for 16-18 h at 37° C. The enriched broth was subsequently streaked on to XLD agar (CM0469) containing antibiotics as Table 12 and incubated at 37° C. for 16-18 h. 
     Birds with no bacteria detected in direct counts were allocated a count of 0 bacterium per g, while those negative in direct counts but positive in the enhanced method were allocated 50 CFU/g. Results were recorded on paper or dictated via telephone or radio in biosecure accommodation and transcribed to Microsoft Excel. Data were analysed in GraphPad Prism. Data were assessed for normality of distribution using a D&#39;Agostino and Pearson omnibus normality test and as non-normal was analysed using a Mann-Whitney U test. Paired data were analysed using a paired T-test. 
     Results 
     1.  Salmonella  In-Feed
         The presence of  Salmonella  in-feed (average 9×log-4 CFU/g) was as expected.       

     2. Guided Biotic® Levels in Drinking Water
         The analysed GB level in drinking water immediately on mixing (average 5×log-8 CFU/mL) was as planned. Samples taken 24 hrs later confirmed an average drop to 8×log-7 CFU/mL.       

     3. Effects of  Salmonella  Dose
         Overall, low levels of  Salmonella  were found in the crop (˜log 1-3 CFU/g) and caeca (˜log 4-5 CFU/g) on days 1 and 3 after challenge. No  Salmonella  was found in ileal, liver or spleen samples at these times.   At day 7 post challenge similar levels of  Salmonella  were again found in the crop and caeca, as well as lower levels (˜log 1) in the ileum, liver and spleen.       

     4. Effects of Active Guided Biotic (Control vs GB)
         GB reduced (P&lt;0.03)  Salmonella  counts in the crop 7 days post-challenge ( FIG. 6 ).   There was a weak trend for GB to reduce  Salmonella  in the crop (P=0.40) and caeca (P=0.50) on day 3, and the ileum (P=0.58) and caeca (P=0.60) on day 7.       

     
       
         
           
               
             
               
                 TABLE 5 
               
               
                   
               
               
                 Example Bacteria 
               
               
                 Optionally, the carrier cells are selected from this Table and/or the target cells are selected from this Table (eg, wherein the carrier and 
               
               
                 target cells are of a different species; or of the same species but are a different strain or the carrier cells are engineered but the target cells are 
               
               
                 wild-type or vice versa). For example the carrier cells are  E coli  cells and the target cells are  C dificile ,  E coli ,  Akkermansia ,  Enterobacteriacea , 
               
               
                   Ruminococcus , Faecalibacterium ,  Firmicutes ,  Bacteroidetes ,  Salmonella ,  Klebsiella ,  Pseudomonas ,  Acintenobacter  or  Streptococcus  cells. 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 
                   Abiotrophia 
                 
                 
                   Acidocella 
                 
                 
                   Actinomyces 
                 
                 
                   Alkalilimnicola 
                 
                 
                   Aquaspirillum 
                 
               
               
                 
                   Abiotrophia defectiva 
                 
                 
                   Acidocella aminolytica 
                 
                 
                   Actinomyces bovis 
                 
                 
                   Alkalilimnicola ehrlichii 
                 
                 
                   Aquaspirillum polymorphum 
                 
               
               
                 
                   Acaricomes 
                 
                 
                   Acidocella facilis 
                 
                 
                   Actinomyces denticolens 
                 
                 
                   Alkaliphilus 
                 
                 
                   Aquaspirillum 
                 
               
               
                 
                   Acaricomes phytoseiuli 
                 
                 
                   Acidomonas 
                 
                 
                   Actinomyces europaeus 
                 
                 
                   Alkaliphilus oremlandii 
                 
                 
                   putridiconchylium 
                 
               
               
                 
                   Acetitomaculum 
                 
                 
                   Acidomonas methanolica 
                 
                 
                   Actinomyces georgiae 
                 
                 
                   Alkaliphilus transvaalensis 
                 
                 
                   Aquaspirillum serpens 
                 
               
               
                 
                   Acetitomaculum ruminis 
                 
                 
                   Acidothermus 
                 
                 
                   Actinomyces gerencseriae 
                 
                 
                   Allochromatium 
                 
                 
                   Aquimarina 
                 
               
               
                 
                   Acetivibrio 
                 
                 
                   Acidothermus cellulolyticus 
                 
                 
                   Actinomyces 
                 
                 
                   Allochromatium vinosum 
                 
                 
                   Aquimarina latercula 
                 
               
               
                 
                   Acetivibrio cellulolyticus 
                 
                 
                   Acidovorax 
                 
                 
                   hordeovulneris 
                 
                 
                   Alloiococcus 
                 
                 
                   Arcanobacterium 
                 
               
               
                 
                   Acetivibrio ethanolgignens 
                 
                 
                   Acidovorax anthurii 
                 
                 
                   Actinomyces howellii 
                 
                 
                   Alloiococcus otitis 
                 
                 
                   Arcanobacterium 
                 
               
               
                 
                   Acetivibrio multivorans 
                 
                 
                   Acidovorax caeni 
                 
                 
                   Actinomyces hyovaginalis 
                 
                 
                   Allokutzneria 
                 
                 
                   haemolyticum 
                 
               
               
                 
                   Acetoanaerobium 
                 
                 
                   Acidovorax cattleyae 
                 
                 
                   Actinomyces israelii 
                 
                 
                   Allokutzneria albata 
                 
                 
                   Arcanobacterium pyogenes 
                 
               
               
                 
                   Acetoanaerobium noterae 
                 
                 
                   Acidovorax citrulli 
                 
                 
                   Actinomyces johnsonii 
                 
                 
                   Altererythrobacter 
                 
                 
                   Archangium 
                 
               
               
                 
                   Acetobacter 
                 
                 
                   Acidovorax defluvii 
                 
                 
                   Actinomyces meyeri 
                 
                 
                   Altererythrobacter ishigakiensis 
                 
                 
                   Archangium gephyra 
                 
               
               
                 
                   Acetobacter aceti 
                 
                 
                   Acidovorax delafieldii 
                 
                 
                   Actinomyces naeslundii 
                 
                 
                   Altermonas 
                 
                 
                   Arcobacter 
                 
               
               
                 
                   Acetobacter cerevisiae 
                 
                 
                   Acidovorax facilis 
                 
                 
                   Actinomyces neuii 
                 
                 
                   Altermonas haloplanktis 
                 
                 
                   Arcobacter butzleri 
                 
               
               
                 
                   Acetobacter cibinongensis 
                 
                 
                   Acidovorax konjaci 
                 
                 
                   Actinomyces odontolyticus 
                 
                 
                   Altermonas macleodii 
                 
                 
                   Arcobacter cryaerophilus 
                 
               
               
                 
                   Acetobacter estunensis 
                 
                 
                   Acidovorax temperans 
                 
                 
                   Actinomyces oris 
                 
                 
                   Alysiella 
                 
                 
                   Arcobacter halophilus 
                 
               
               
                 
                   Acetobacter fabarum 
                 
                 
                   Acidovorax valerianellae 
                 
                 
                   Actinomyces radingae 
                 
                 
                   Alysiella crassa 
                 
                 
                   Arcobacter nitrofigilis 
                 
               
               
                 
                   Acetobacter ghanensis 
                 
                 
                   Acinetobacter 
                 
                 
                   Actinomyces slackii 
                 
                 
                   Alysiella filiformis 
                 
                 
                   Arcobacter skirrowii 
                 
               
               
                 
                   Acetobacter indonesiensis 
                 
                 
                   Acinetobacter baumannii 
                 
                 
                   Actinomyces turicensis 
                 
                 
                   Aminobacter 
                 
                 
                   Arhodomonas 
                 
               
               
                 
                   Acetobacter lovaniensis 
                 
                 
                   Acinetobacter baylyi 
                 
                 
                   Actinomyces viscosus 
                 
                 
                   Aminobacter aganoensis 
                 
                 
                   Arhodomonas aquaeolei 
                 
               
               
                 
                   Acetobacter malorum 
                 
                 
                   Acinetobacter bouvetii 
                 
                 
                   Actinoplanes 
                 
                 
                   Aminobacter aminovorans 
                 
                 
                   Arsenophonus 
                 
               
               
                 
                   Acetobacter nitrogenifigens 
                 
                 
                   Acinetobacter calcoaceticus 
                 
                 
                   Actinoplanes auranticolor 
                 
                 
                   Aminobacter niigataensis 
                 
                 
                   Arsenophonus 
                 
               
               
                 
                   Acetobacter oeni 
                 
                 
                   Acinetobacter gerneri 
                 
                 
                   Actinoplanes brasiliensis 
                 
                 
                   Aminobacterium 
                 
                 
                   nasoniae 
                 
               
               
                 
                   Acetobacter orientalis 
                 
                 
                   Acinetobacter haemolyticus 
                 
                 
                   Actinoplanes consettensis 
                 
                 
                   Aminobacterium mobile 
                 
                 
                   Arthrobacter 
                 
               
               
                 
                   Acetobacter orleanensis 
                 
                 
                   Acinetobacter johnsonii 
                 
                 
                   Actinoplanes deccanensis 
                 
                 
                   Aminomonas 
                 
                 
                   Arthrobacter agilis 
                 
               
               
                 
                   Acetobacter pasteurianus 
                 
                 
                   Acinetobacter junii 
                 
                 
                   Actinoplanes derwentensis 
                 
                 
                   Aminomonas paucivorans 
                 
                 
                   Arthrobacter albus 
                 
               
               
                 
                   Acetobacter pornorurn 
                 
                 
                   Acinetobacter lwoffi 
                 
                 
                   Actinoplanes digitatis 
                 
                 
                   Ammoniphilus 
                 
                 
                   Arthrobacter aurescens 
                 
               
               
                 
                   Acetobacter senegalensis 
                 
                 
                   Acinetobacter parvus 
                 
                 
                   Actinoplanes durhamensis 
                 
                 
                   Ammoniphilus oxalaticus 
                 
                 
                   Arthrobacter chlorophenolicus 
                 
               
               
                 
                   Acetobacter xylinus 
                 
                 
                   Acinetobacter radioresistens 
                 
                 
                   Actinoplanes ferrugineus 
                 
                 
                   Ammoniphilus oxalivorans 
                 
                 
                   Arthrobacter citreus 
                 
               
               
                 
                   Acetobacterium 
                 
                 
                   Acinetobacter schindleri 
                 
                 
                   Actinoplanes globisporus 
                 
                 
                   Amphibacillus 
                 
                 
                   Arthrobacter crystallopoietes 
                 
               
               
                 
                   Acetobacterium bakii 
                 
                 
                   Acinetobacter soli 
                 
                 
                   Actinoplanes humidus 
                 
                 
                   Amphibacillus xylanus 
                 
                 
                   Arthrobacter cumminsii 
                 
               
               
                 
                   Acetobacterium carbinolicum 
                 
                 
                   Acinetobacter tandoii 
                 
                 
                   Actinoplanes italicus 
                 
                 
                   Amphritea 
                 
                 
                   Arthrobacter globiformis 
                 
               
               
                 
                   Acetobacterium dehalogenans 
                 
                 
                   Acinetobacter tjernbergiae 
                 
                 
                   Actinoplanes liguriensis 
                 
                 
                   Amphritea balenae 
                 
                 
                   Arthrobacter 
                 
               
               
                 
                   Acetobacterium fimetarium 
                 
                 
                   Acinetobacter towneri 
                 
                 
                   Actinoplanes lobatus 
                 
                 
                   Amphritea japonica 
                 
                 
                   histidinolovorans 
                 
               
               
                 
                   Acetobacterium malicum 
                 
                 
                   Acinetobacter ursingii 
                 
                 
                   Actinoplanes missouriensis 
                 
                 
                   Amycolatopsis 
                 
                 
                   Arthrobacter ilicis 
                 
               
               
                 
                   Acetobacterium paludosum 
                 
                 
                   Acinetobacter venetianus 
                 
                 
                   Actinoplanes palleronii 
                 
                 
                   Amycolatopsis alba 
                 
                 
                   Arthrobacter luteus 
                 
               
               
                 
                   Acetobacterium tundrae 
                 
                 
                   Acrocarpospora 
                 
                 
                   Actinoplanes philippinensis 
                 
                 
                   Amycolatopsis albidoflavus 
                 
                 
                   Arthrobacter methylotrophus 
                 
               
               
                 
                   Acetobacterium wieringae 
                 
                 
                   Acrocarpospora corrugata 
                 
                 
                   Actinoplanes rectilineatus 
                 
                 
                   Amycolatopsis azurea 
                 
                 
                   Arthrobacter mysorens 
                 
               
               
                 
                   Acetobacterium woodii 
                 
                 
                   Acrocarpospora 
                 
                 
                   Actinoplanes regularis 
                 
                 
                   Amycolatopsis coloradensis 
                 
                 
                   Arthrobacter nicotianae 
                 
               
               
                 
                   Acetofilamentum 
                 
                 
                   macrocephala 
                 
                 
                   Actinoplanes 
                 
                 
                   Amycolatopsis lurida 
                 
                 
                   Arthrobacter nicotinovorans 
                 
               
               
                 
                   Acetofilamentum rigidum 
                 
                 
                   Acrocarpospora pleiomorpha 
                 
                 
                   teichomyceticus 
                 
                 
                   Amycolatopsis mediterranei 
                 
                 
                   Arthrobacter oxydans 
                 
               
               
                 
                   Acetohalobium 
                 
                 
                   Actibacter 
                 
                 
                   Actinoplanes utahensis 
                 
                 
                   Amycolatopsis rifamycinica 
                 
                 
                   Arthrobacter pascens 
                 
               
               
                 
                   Acetohalobium arabaticum 
                 
                 
                   Actibacter sediminis 
                 
                 
                   Actinopolyspora 
                 
                 
                   Amycolatopsis rubida 
                 
                 
                   Arthrobacter 
                 
               
               
                 
                   Acetomicrobium 
                 
                 
                   Actinoalloteichus 
                 
                 
                   Actinopolyspora halophila 
                 
                 
                   Amycolatopsis sulphurea 
                 
                 
                   phenanthrenivorans 
                 
               
               
                 
                   Acetomicrobium faecale 
                 
                 
                   Actinoalloteichus 
                 
                 
                   Actinopolyspora mortivallis 
                 
                 
                   Amycolatopsis tolypomycina 
                 
                 
                   Arthrobacter 
                 
               
               
                 
                   Acetomicrobium flavidum 
                 
                 
                   cyanogriseus 
                 
                 
                   Actinosynnema 
                 
                 
                   Anabaena 
                 
                 
                   polychromogenes 
                 
               
               
                 
                   Acetonema 
                 
                 
                   Actinoalloteichus 
                 
                 
                   Actinosynnema mirum 
                 
                 
                   Anabaena cylindrica 
                 
                 
                   Atrhrobacter protophormiae 
                 
               
               
                 
                   Acetonema longum 
                 
                 
                   hymeniacidonis 
                 
                 
                   Actinotalea 
                 
                   Anabaena flos - aquae   
                 
                   Arthrobacter 
                 
               
               
                 
                   Acetothermus 
                 
                 
                   Actinoalloteichus spitiensis 
                 
                 
                   Actinotalea fermentans 
                 
                 
                   Anabaena variabilis 
                 
                 
                   psychrolactophilus 
                 
               
               
                 
                   Acetothermus paucivorans 
                 
                 
                   Actinobaccillus 
                 
                 
                   Aerococcus 
                 
                 
                   Anaeroarcus 
                 
                 
                   Arthrobacter ramosus 
                 
               
               
                 
                   Acholeplasma 
                 
                 
                   Actinobacillus capsulatus 
                 
                 
                   Aerococcus sanguinicola 
                 
                 
                   Anaeroarcus burkinensis 
                 
                 
                   Arthrobacter sulfonivorans 
                 
               
               
                 
                   Acholeplasma axanthum 
                 
                 
                   Actinobacillus delphinicola 
                 
                 
                   Aerococcus urinae 
                 
                 
                   Anaerobaculum 
                 
                 
                   Arthrobacter sulfureus 
                 
               
               
                 
                   Acholeplasma brassicae 
                 
                 
                   Actinobacillus hominis 
                 
                 
                   Aerococcus urinaeequi 
                 
                 
                   Anaerobaculum mobile 
                 
                 
                   Arthrobacter uratoxydans 
                 
               
               
                 
                   Acholeplasma cavigenitalium 
                 
                 
                   Actinobacillus indolicus 
                 
                 
                   Aerococcus urinaehominis 
                 
                 
                   Anaerobiospirillum 
                 
                 
                   Arthrobacter ureafaciens 
                 
               
               
                 
                   Acholeplasma equifetale 
                 
                 
                   Actinobacillus lignieresii 
                 
                 
                   Aerococcus viridans 
                 
                 
                   Anaerobiospirillum 
                 
                 
                   Arthrobacter viscosus 
                 
               
               
                 
                   Acholeplasma granularum 
                 
                 
                   Actinobacillus minor 
                 
                 
                   Aeromicrobium 
                 
                 
                   succiniciproducens 
                 
                 
                   Arthrobacter woluwensis 
                 
               
               
                 
                   Acholeplasma hippikon 
                 
                 
                   Actinobacillus muris 
                 
                 
                   Aeromicrobium erythreum 
                 
                 
                   Anaerobiospirillum thomasii 
                 
                 
                   Asaia 
                 
               
               
                 
                   Acholeplasma laidlawii 
                 
                 
                   Actinobacillus 
                 
                 
                   Aeromonas 
                 
                 
                   Anaerococcus 
                 
                 
                   Asaia bogorensis 
                 
               
               
                 
                   Acholeplasma modicum 
                 
                 
                   pleuropneumoniae 
                 
                 
                   Aeromonas 
                 
                 
                   Anaerococcus hydrogenalis 
                 
                 
                   Asanoa 
                 
               
               
                 
                   Acholeplasma morum 
                 
                 
                   Actinobacillus porcinus 
                 
                 
                   allosaccharophila 
                 
                 
                   Anaerococcus lactolyticus 
                 
                 
                   Asanoa ferruginea 
                 
               
               
                 
                   Acholeplasma multilocale 
                 
                 
                   Actinobacillus rossii 
                 
                 
                   Aeromonas bestiarum 
                 
                 
                   Anaerococcus prevotii 
                 
                 
                   Asticcacaulis 
                 
               
               
                 
                   Acholeplasma oculi 
                 
                 
                   Actinobacillus scotiae 
                 
                 
                   Aeromonas caviae 
                 
                 
                   Anaerococcus tetradius 
                 
                 
                   Asticcacaulis biprosthecium 
                 
               
               
                 
                   Acholeplasma palmae 
                 
                 
                   Actinobacillus seminis 
                 
                 
                   Aeromonas encheleia 
                 
                 
                   Anaerococcus vaginalis 
                 
                 
                   Asticcacaulis excentricus 
                 
               
               
                 
                   Acholeplasma parvum 
                 
                 
                   Actinobacillus succinogenes 
                 
                 
                   Aeromonas 
                 
                 
                   Anaerofustis 
                 
                 
                   Atopobacter 
                 
               
               
                 
                   Acholeplasma pleciae 
                 
                 
                   Actinobaccillus suis 
                 
                 
                   enteropelogenes 
                 
                 
                   Anaerofustis stercorihominis 
                 
                 
                   Atopobacter phocae 
                 
               
               
                 
                   Acholeplasma vituli 
                 
                 
                   Actinobacillus ureae 
                 
                 
                   Aeromonas eucrenophila 
                 
                 
                   Anaeromusa 
                 
                 
                   Atopobium 
                 
               
               
                 
                   Achromobacter 
                 
                 
                   Actinobaculum 
                 
                 
                   Aeromonas ichthiosmia 
                 
                 
                   Anaeromusa acidaminophila 
                 
                 
                   Atopobium fossor 
                 
               
               
                 
                   Achromobacter denitrificans 
                 
                 
                   Actinobaculum massiliense 
                 
                 
                   Aeromonas jandaei 
                 
                 
                   Anaeromyxobacter 
                 
                 
                   Atopobium minutum 
                 
               
               
                 
                   Achromobacter insolitus 
                 
                 
                   Actinobaculum schaalii 
                 
                 
                   Aeromonas media 
                 
                 
                   Anaeromyxobacter 
                 
                 
                   Atopobium parvulum 
                 
               
               
                 
                   Achromobacter piechaudii 
                 
                 
                   Actinobaculum suis 
                 
                 
                   Aeromonas popoffii 
                 
                 
                   dehalogenans 
                 
                 
                   Atopobium rimae 
                 
               
               
                 
                   Achromobacter ruhlandii 
                 
                 
                   Actinomyces urinale 
                 
                 
                   Aeromonas sobria 
                 
                 
                   Anaerorhabdus 
                 
                 
                   Atopobium vaginae 
                 
               
               
                 
                   Achromobacter spanius 
                 
                 
                   Actinocatenispora 
                 
                 
                   Aeromonas veronii 
                 
                 
                   Anaerorhabdus furcosa 
                 
                 
                   Aureobacterium 
                 
               
               
                 
                   Acidaminobacter 
                 
                 
                   Actinocatenispora rupis 
                 
                 
                   Agrobacterium 
                 
                 
                   Anaerosinus 
                 
                 
                   Aureobacterium barkeri 
                 
               
               
                 
                   Acidaminobacter 
                 
                 
                   Actinocatenispora 
                 
                 
                   Agrobacterium 
                 
                 
                   Anaerosinus glycerini 
                 
                 
                   Aurobacterium 
                 
               
               
                 
                   hydrogenoformans 
                 
                 
                   thailandica 
                 
                 
                   gelatinovorum 
                 
                 
                   Anaerovirgula 
                 
                 
                   Aurobacterium liquefaciens 
                 
               
               
                 
                   Acidaminococcus 
                 
                 
                   Actinocatenispora sera 
                 
                 
                   Agrococcus 
                 
                 
                   Anaerovirgula multivorans 
                 
                 
                   Avibacterium 
                 
               
               
                 
                   Acidaminococcus fermentans 
                 
                 
                   Actinocorallia 
                 
                 
                   Agrococcus citreus 
                 
                 
                   Ancalomicrobium 
                 
                 
                   Avibacterium avium 
                 
               
               
                 
                   Acidaminococcus intestini 
                 
                 
                   Actinocorallia aurantiaca 
                 
                 
                   Agrococcus jenensis 
                 
                 
                   Ancalomicrobium adetum 
                 
                 
                   Avibacterium gallinarum 
                 
               
               
                 
                   Acidicaldus 
                 
                 
                   Actinocorallia aurea 
                 
                 
                   Agromonas 
                 
                 
                   Ancylobacter 
                 
                 
                   Avibacterium paragallinarum 
                 
               
               
                 
                   Acidicaldus organivorans 
                 
                 
                   Actinocorallia cavernae 
                 
                 
                   Agromonas oligotrophica 
                 
                 
                   Ancylobacter aquaticus 
                 
                 
                   Avibacterium volantium 
                 
               
               
                 
                   Acidimicrobium 
                 
                 
                   Actinocorallia glomerata 
                 
                 
                   Agromyces 
                 
                 
                   Aneurinibacillus 
                 
                 
                   Azoarcus 
                 
               
               
                 
                   Acidimicrobium ferrooxidans 
                 
                 
                   Actinocorallia herbida 
                 
                 
                   Agromyces fucosus 
                 
                 
                   Aneurinibacillus aneurinilyticus 
                 
                 
                   Azoarcus indigens 
                 
               
               
                 
                   Acidiphilium 
                 
                 
                   Actinocorallia libanotica 
                 
                 
                   Agromyces hippuratus 
                 
                 
                   Aneurinibacillus migulanus 
                 
                 
                   Azoarcus tolulyticus 
                 
               
               
                 
                   Acidiphilium acidophilum 
                 
                 
                   Actinocorallia longicatena 
                 
                 
                   Agromyces luteolus 
                 
                 
                   Aneurinibacillus 
                 
                 
                   Azoarcus toluvorans 
                 
               
               
                 
                   Acidiphilium angustum 
                 
                 
                   Actinomadura 
                 
                 
                   Agromyces mediolanus 
                 
                 
                   thermoaerophilus 
                 
                 
                   Azohydromonas 
                 
               
               
                 
                   Acidiphilium cryptum 
                 
                 
                   Actinomadura alba 
                 
                 
                   Agromyces ramosus 
                 
                 
                   Angiococcus 
                 
                 
                   Azohydromonas australica 
                 
               
               
                 
                   Acidiphilium multivorum 
                 
                 
                   Actinomadura atramentaria 
                 
                 
                   Agromyces rhizospherae 
                 
                 
                   Angiococcus disciformis 
                 
                 
                   Azohydromonas lata 
                 
               
               
                 
                   Acidiphilium organovorum 
                 
                 
                   Actinomadura 
                 
                 
                   Akkermansia 
                 
                 
                   Angulomicrobium 
                 
                 
                   Azomonas 
                 
               
               
                 
                   Acidiphilium rubrum 
                 
                 
                   bangladeshensis 
                 
                 
                   Akkermansia muciniphila 
                 
                 
                   Angulomicrobium tetraedrale 
                 
                 
                   Azomonas agilis 
                 
               
               
                 
                   Acidisoma 
                 
                 
                   Actinomadura catellatispora 
                 
                 
                   Albidiferax 
                 
                 
                   Anoxybacillus 
                 
                 
                   Azomonas insignis 
                 
               
               
                 
                   Acidisoma sibiricum 
                 
                 
                   Actinomadura chibensis 
                 
                 
                   Albidiferax ferrireducens 
                 
                 
                   Anoxybacillus pushchinoensis 
                 
                 
                   Azomonas macrocytogenes 
                 
               
               
                 
                   Acidisoma tundrae 
                 
                 
                   Actinomadura chokoriensis 
                 
                 
                   Albidovulum 
                 
                 
                   Aquabacterium 
                 
                 
                   Azorhizobium 
                 
               
               
                 
                   Acidisphaera 
                 
                 
                   Actinomadura citrea 
                 
                 
                   Albidovulum inexpectatum 
                 
                 
                   Aquabacterium commune 
                 
                 
                   Azorhizobium caulinodans 
                 
               
               
                 
                   Acidisphaera rubrifaciens 
                 
                 
                   Actinomadura coerulea 
                 
                 
                   Alcaligenes 
                 
                 
                   Aquabacterium parvum 
                 
                 
                   Azorhizophilus 
                 
               
               
                 
                   Acidithiobacillus 
                 
                 
                   Actinomadura echinospora 
                 
                 
                   Alcaligenes denitrificans 
                 
                   
                 
                   Azorhizophilus paspali 
                 
               
               
                 
                   Acidithiobacillus albertensis 
                 
                 
                   Actinomadura fibrosa 
                 
                 
                   Alcaligenes faecalis 
                 
                   
                 
                   Azospirillum 
                 
               
               
                 
                   Acidithiobacillus caldus 
                 
                 
                   Actinomadura formosensis 
                 
                 
                   Alcanivorax 
                 
                   
                 
                   Azospirillum brasilense 
                 
               
               
                 
                   Acidithiobacillus ferrooxidans 
                 
                 
                   Actinomadura hibisca 
                 
                 
                   Alcanivorax borkumensis 
                 
                   
                 
                   Azospirillum halopraeferens 
                 
               
               
                 
                   Acidithiobacillus thiooxidans 
                 
                 
                   Actinomadura kijaniata 
                 
                 
                   Alcanivorax jadensis 
                 
                   
                 
                   Azospirillum irakense 
                 
               
               
                 
                   Acidobacterium 
                 
                 
                   Actinomadura latina 
                 
                 
                   Algicola 
                 
                   
                 
                   Azotobacter 
                 
               
               
                 
                   Acidobacterium capsulatum 
                 
                 
                   Actinomadura livida 
                 
                 
                   Algicola bacteriolytica 
                 
                   
                 
                   Azotobacter beijerinckii 
                 
               
               
                   
                 
                   Actinomadura 
                 
                 
                   Alicyclobacillus 
                 
                   
                 
                   Azotobacter chroococcum 
                 
               
               
                   
                 
                   luteofluorescens 
                 
                 
                   Alicyclobacillus 
                 
                   
                 
                   Azotobacter nigricans 
                 
               
               
                   
                 
                   Actinomadura macra 
                 
                 
                   disulfidooxidans 
                 
                   
                 
                   Azotobacter salinestris 
                 
               
               
                   
                 
                   Actinomadura madurae 
                 
                 
                   Alicyclobacillus 
                 
                   
                 
                   Azotobacter vinelandii 
                 
               
               
                   
                 
                   Actinomadura oligospora 
                 
                 
                   sendaiensis 
                 
               
               
                   
                 
                   Actinomadura pelletieri 
                 
                 
                   Alicyclobacillus vulcanalis 
                 
               
               
                   
                 
                   Actinomadura rubrobrunea 
                 
                 
                   Alishewanella 
                 
               
               
                   
                 
                   Actinomadura rugatobispora 
                 
                 
                   Alishewanella fetalis 
                 
               
               
                   
                 
                   Actinomadura umbrina 
                 
                 
                   Alkalibacillus 
                 
               
               
                   
                 
                   Actinomadura 
                 
                 
                   Alkalibacillus 
                 
               
               
                   
                 
                   verrucosospora 
                 
                 
                   haloalkaliphilus 
                 
               
               
                   
                 
                   Actinomadura vinacea 
                 
               
               
                   
                 
                   Actinomadura viridilutea 
                 
               
               
                   
                 
                   Actinomadura viridis 
                 
               
               
                   
                 
                   Actinomadura yumaensis 
                 
               
               
                 
                   Bacillus 
                 
                 
                   Bacteroides 
                 
                 
                   Bibersteinia 
                 
                 
                   Borrelia 
                 
                 
                   Brevinema 
                 
               
               
                 [see below] 
                 
                   Bacteroides caccae 
                 
                 
                   Bibersteinia trehalosi 
                 
                 
                   Borrelia afzelii 
                 
                 
                   Brevinema andersonii 
                 
               
               
                 
                   Bacteriovorax 
                 
                 
                   Bacteroides coagulans 
                 
                 
                   Bifidobacterium 
                 
                 
                   Borrelia americana 
                 
                 
                   Brevundimonas 
                 
               
               
                 
                   Bacteriovorax stolpii 
                 
                 
                   Bacteroides eggerthii 
                 
                 
                   Bifidobacterium adolescentis 
                 
                 
                   Borrelia burgdorferi 
                 
                 
                   Brevundimonas alba 
                 
               
               
                   
                 
                   Bacteroides fragilis 
                 
                 
                   Bifidobacterium angulatum 
                 
                 
                   Borrelia carolinensis 
                 
                 
                   Brevundimonas aurantiaca 
                 
               
               
                   
                 
                   Bacteroides galacturonicus 
                 
                 
                   Bifidobacterium animalis 
                 
                 
                   Borrelia coriaceae 
                 
                 
                   Brevundimonas diminuta 
                 
               
               
                   
                 
                   Bacteroides helcogenes 
                 
                 
                   Bifidobacterium asteroides 
                 
                 
                   Borrelia garinii 
                 
                 
                   Brevundimonas intermedia 
                 
               
               
                   
                 
                   Bacteroides ovatus 
                 
                 
                   Bifidobacterium bifidum 
                 
                 
                   Borrelia japonica 
                 
                 
                   Brevundimonas subvibrioides 
                 
               
               
                   
                 
                   Bacteroides pectinophilus 
                 
                 
                   Bifidobacterium boum 
                 
                 
                   Bosea 
                 
                 
                   Brevundimonas vancanneytii 
                 
               
               
                   
                 
                   Bacteroides pyogenes 
                 
                 
                   Bifidobacterium breve 
                 
                 
                   Bosea minatitlanensis 
                 
                 
                   Brevundimonas variabilis 
                 
               
               
                   
                 
                   Bacteroides salyersiae 
                 
                 
                   Bifidobacterium catenulatum 
                 
                 
                   Bosea thiooxidans 
                 
                 
                   Brevundimonas vesicularis 
                 
               
               
                   
                 
                   Bacteroides stercoris 
                 
                 
                   Bifidobacterium choerinum 
                 
                 
                   Brachybacterium 
                 
                 
                   Brochothrix 
                 
               
               
                   
                 
                   Bacteroides suis 
                 
                 
                   Bifidobacterium coryneforme 
                 
                 
                   Brachybacterium 
                 
                 
                   Brochothrix campestris 
                 
               
               
                   
                 
                   Bacteroides tectus 
                 
                 
                   Bifidobacterium cuniculi 
                 
                 
                   alimentarium 
                 
                 
                   Brochothrix thermosphacta 
                 
               
               
                   
                 
                   Bacteroides thetaiotaomicron 
                 
                 
                   Bifidobacterium dentium 
                 
                 
                   Brachybacterium faecium 
                 
                 
                   Brucella 
                 
               
               
                   
                 
                   Bacteroides uniformis 
                 
                 
                   Bifidobacterium gallicum 
                 
                 
                   Brachybacterium 
                 
                 
                   Brucella canis 
                 
               
               
                   
                 
                   Bacteroides ureolyticus 
                 
                 
                   Bifidobacterium gallinarum 
                 
                 
                   paraconglomeratum 
                 
                 
                   Brucella neotomae 
                 
               
               
                   
                 
                   Bacteroides vulgatus 
                 
                 
                   Bifidobacterium indicum 
                 
                 
                   Brachybacterium rhamnosum 
                 
                 
                   Bryobacter 
                 
               
               
                   
                 
                   Balnearium 
                 
                 
                   Bifidobacterium longum 
                 
                 
                   Brachybacterium 
                 
                 
                   Bryobacter aggregatus 
                 
               
               
                   
                 
                   Balnearium lithotrophicum 
                 
                 
                   Bifidobacterium 
                 
                 
                   tyrofermentans 
                 
                 
                   Burkholderia 
                 
               
               
                   
                 
                   Balneatrix 
                 
                 
                   magnumBifidobacterium 
                 
                 
                   Brachyspira 
                 
                 
                   Burkholderia ambifaria 
                 
               
               
                   
                 
                   Balneatrix alpica 
                 
                 
                   merycicum 
                 
                 
                   Brachyspira alvinipulli 
                 
                 
                   Burkholderia andropogonis 
                 
               
               
                   
                 
                   Balneola 
                 
                 
                   Bifidobacterium minimum 
                 
                 
                   Brachyspira hyodysenteriae 
                 
                 
                   Burkholderia anthina 
                 
               
               
                   
                 
                   Balneola vulgaris 
                 
                 
                   Bifidobacterium 
                 
                 
                   Brachyspira innocens 
                 
                 
                   Burkholderia caledonica 
                 
               
               
                   
                 
                   Barnesiella 
                 
                 
                   pseudocatenulatum 
                 
                 
                   Brachyspira murdochii 
                 
                 
                   Burkholderia caryophylli 
                 
               
               
                   
                 
                   Barnesiella viscericola 
                 
                 
                   Bifidobacterium 
                 
                 
                   Brachyspira 
                 
                 
                   Burkholderia cenocepacia 
                 
               
               
                   
                 
                   Bartonella 
                 
                 
                   pseudolongum 
                 
                 
                   pilosicoli 
                 
                 
                   Burkholderia cepacia 
                 
               
               
                   
                 
                   Bartonella alsatica 
                 
                 
                   Bifidobacterium pullorum 
                 
                 
                   Bradyrhizobium 
                 
                 
                   Burkholderia cocovenenans 
                 
               
               
                   
                 
                   Bartonella bacilliformis 
                 
                 
                   Bifidobacterium ruminantium 
                 
                 
                   Bradyrhizobium canariense 
                 
                 
                   Burkholderia dolosa 
                 
               
               
                   
                 
                   Bartonella clarridgeiae 
                 
                 
                   Bifidobacterium saeculare 
                 
                 
                   Bradyrhizobium elkanii 
                 
                 
                   Burkholderia fungorum 
                 
               
               
                   
                 
                   Bartonella doshiae 
                 
                 
                   Bifidobacterium subtile 
                 
                 
                   Bradyrhizobium japonicum 
                 
                 
                   Burkholderia glathei 
                 
               
               
                   
                 
                   Bartonella elizabethae 
                 
                 
                   Bifidobacterium 
                 
                 
                   Bradyrhizobium liaoningense 
                 
                 
                   Burkholderia glumae 
                 
               
               
                   
                 
                   Bartonella grahamii 
                 
                 
                   thermophilum 
                 
                 
                   Brenneria 
                 
                 
                   Burkholderia graminis 
                 
               
               
                   
                 
                   Bartonella henselae 
                 
                 
                   Bilophila 
                 
                 
                   Brenneria alni 
                 
                 
                   Burkholderia kururiensis 
                 
               
               
                   
                 
                   Bartonella rochalimae 
                 
                 
                   Bilophila wadsworthia 
                 
                 
                   Brenneria nigrifluens 
                 
                 
                   Burkholderia multivorans 
                 
               
               
                   
                 
                   Bartonella vinsonii 
                 
                 
                   Biostraticola 
                 
                 
                   Brenneria quercina 
                 
                 
                   Burkholderia phenazinium 
                 
               
               
                   
                 
                   Bavariicoccus 
                 
                 
                   Biostraticola tofi 
                 
                 
                   Brenneria quercina 
                 
                 
                   Burkholderia plantarii 
                 
               
               
                   
                 
                   Bavariicoccus seileri 
                 
                 
                   Bizionia 
                 
                 
                   Brenneria salicis 
                 
                 
                   Burkholderia pyrrocinia 
                 
               
               
                   
                 
                   Bdellovibrio 
                 
                 
                   Bizionia argentinensis 
                 
                 
                   Brevibacillus 
                 
                 
                   Burkholderia silvatlantica 
                 
               
               
                   
                 
                   Bdellovibrio bacteriovorus 
                 
                 
                   Blastobacter 
                 
                 
                   Brevibacillus agri 
                 
                 
                   Burkholderia stabilis 
                 
               
               
                   
                 
                   Bdellovibrio exovorus 
                 
                 
                   Blastobacter capsulatus 
                 
                 
                   Brevibacillus borstelensis 
                 
                 
                   Burkholderia thailandensis 
                 
               
               
                   
                 
                   Beggiatoa 
                 
                 
                   Blastobacter denitrificans 
                 
                 
                   Brevibacillus brevis 
                 
                 
                   Burkholderia tropica 
                 
               
               
                   
                 
                   Beggiatoa alba 
                 
                 
                   Blastococcus 
                 
                 
                   Brevibacillus centrosporus 
                 
                 
                   Burkholderia unamae 
                 
               
               
                   
                 
                   Beijerinckia 
                 
                 
                   Blastococcus aggregatus 
                 
                 
                   Brevibacillus choshinensis 
                 
                 
                   Burkholderia vietnamiensis 
                 
               
               
                   
                 
                   Beijerinckia derxii 
                 
                 
                   Blastococcus saxobsidens 
                 
                 
                   Brevibacillus invocatus 
                 
                 
                   Buttiauxella 
                 
               
               
                   
                 
                   Beijerinckia fluminensis 
                 
                 
                   Blastochloris 
                 
                 
                   Brevibacillus laterosporus 
                 
                 
                   Buttiauxella agrestis 
                 
               
               
                   
                 
                   Beijerinckia indica 
                 
                 
                   Blastochloris viridis 
                 
                 
                   Brevibacillus parabrevis 
                 
                 
                   Buttiauxella brennerae 
                 
               
               
                   
                 
                   Beijerinckia mobilis 
                 
                 
                   Blastomonas 
                 
                 
                   Brevibacillus reuszeri 
                 
                 
                   Buttiauxella ferragutiae 
                 
               
               
                   
                 
                   Belliella 
                 
                 
                   Blastomonas natatoria 
                 
                 
                   Brevibacterium 
                 
                 
                   Buttiauxella gaviniae 
                 
               
               
                   
                 
                   Belliella baltica 
                 
                 
                   Blastopirellula 
                 
                 
                   Brevibacterium abidum 
                 
                 
                   Buttiauxella izardii 
                 
               
               
                   
                 
                   Bellilinea 
                 
                 
                   Blastopirellula marina 
                 
                 
                   Brevibacterium album 
                 
                 
                   Buttiauxella noackiae 
                 
               
               
                   
                 
                   Bellilinea caldifistulae 
                 
                 
                   Blautia 
                 
                 
                   Brevibacterium aurantiacum 
                 
                 
                   Buttiauxella warmboldiae 
                 
               
               
                   
                 
                   Belnapia 
                 
                 
                   Blautia coccoides 
                 
                 
                   Brevibacterium celere 
                 
                 
                   Butyrivibrio 
                 
               
               
                   
                 
                   Belnapia moabensis 
                 
                 
                   Blautia hansenii 
                 
                 
                   Brevibacterium epidermidis 
                 
                 
                   Butyrivibrio fibrisolvens 
                 
               
               
                   
                 
                   Bergeriella 
                 
                 
                   Blautia producta 
                 
                 
                   Brevibacterium 
                 
                 
                   Butyrivibrio hungatei 
                 
               
               
                   
                 
                   Bergeriella denitrificans 
                 
                 
                   Blautia wexlerae 
                 
                 
                   frigoritolerans 
                 
                 
                   Butyrivibrio proteoclasticus 
                 
               
               
                   
                 
                   Beutenbergia 
                 
                 
                   Bogoriella 
                 
                 
                   Brevibacterium halotolerans 
                 
               
               
                   
                 
                   Beutenbergia cavernae 
                 
                 
                   Bogoriella caseilytica 
                 
                 
                   Brevibacterium iodinum 
                 
               
               
                   
                   
                 
                   Bordetella 
                 
                 
                   Brevibacterium linens 
                 
               
               
                   
                   
                 
                   Bordetella avium 
                 
                 
                   Brevibacterium lyticum 
                 
               
               
                   
                   
                 
                   Bordetella bronchiseptica 
                 
                 
                   Brevibacterium mcbrellneri 
                 
               
               
                   
                   
                 
                   Bordetella hinzii 
                 
                 
                   Brevibacterium otitidis 
                 
               
               
                   
                   
                 
                   Bordetella holmesii 
                 
                 
                   Brevibacterium oxydans 
                 
               
               
                   
                   
                 
                   Bordetella parapertussis 
                 
                 
                   Brevibacterium paucivorans 
                 
               
               
                   
                   
                 
                   Bordetella pertussis 
                 
                 
                   Brevibacterium stationis 
                 
               
               
                   
                   
                 
                   Bordetella petrii 
                 
               
               
                   
                   
                 
                   Bordetella trematum 
                 
               
               
                 
                   Bacillus 
                 
               
               
                 
                   B. acidiceler 
                 
                 
                   B. aminovorans 
                 
                 
                   B. glucanolyticus 
                 
                 
                   B. taeanensis 
                 
                 
                   B. lautus 
                 
               
               
                 
                   B. acidicola 
                 
                 
                   B. amylolyticus 
                 
                 
                   B. gordonae 
                 
                 
                   B. tequilensis 
                 
                 
                   B. lehensis 
                 
               
               
                 
                   B. acidiproducens 
                 
                 
                   B. andreesenii 
                 
                 
                   B. gottheilii 
                 
                 
                   B. thermantarcticus 
                 
                 
                   B. lentimorbus 
                 
               
               
                 
                   B. acidocaldarius 
                 
                 
                   B. aneurinilyticus 
                 
                 
                   B. graminis 
                 
                 
                   B. thermoaerophilus 
                 
                 
                   B. lentus 
                 
               
               
                 
                   B. acidoterrestris 
                 
                 
                   B. anthracis 
                 
                 
                   B. halmapalus 
                 
                 
                   B. thermoamylovorans 
                 
                 
                   B. licheniformis 
                 
               
               
                 
                   B. aeolius 
                 
                 
                   B. aquimaris 
                 
                 
                   B. haloalkaliphilus 
                 
                 
                   B. thermocatenulatus 
                 
                 
                   B. ligniniphilus 
                 
               
               
                 
                   B. aerius 
                 
                 
                   B. arenosi 
                 
                 
                   B. halochares 
                 
                 
                   B. thermocloacae 
                 
                 
                   B. litoralis 
                 
               
               
                 
                   B. aerophilus 
                 
                 
                   B. arseniciselenatis 
                 
                 
                   B. halodenitrificans 
                 
                 
                   B. thermocopriae 
                 
                 
                   B. locisalis 
                 
               
               
                 
                   B. agaradhaerens 
                 
                 
                   B. arsenicus 
                 
                 
                   B. halodurans 
                 
                 
                   B. thermodenitrificans 
                 
                 
                   B. luciferensis 
                 
               
               
                 
                   B. agri 
                 
                 
                   B. aurantiacus 
                 
                 
                   B. halophilus 
                 
                 
                   B. thermoglucosidasius 
                 
                 
                   B. luteolus 
                 
               
               
                 
                   B. aidingensis 
                 
                 
                   B. arvi 
                 
                 
                   B. halosaccharovorans 
                 
                 
                   B. thermolactis 
                 
                 
                   B. luteus 
                 
               
               
                 
                   B. akibai 
                 
                 
                   B. aryabhattai 
                 
                 
                   B. hemicellulosilyticus 
                 
                 
                   B. thermoleovorans 
                 
                 
                   B. macauensis 
                 
               
               
                 
                   B. alcalophilus 
                 
                 
                   B. asahii 
                 
                 
                   B. hemicentroti 
                 
                 
                   B. thermophilus 
                 
                 
                   B. macerans 
                 
               
               
                 
                   B. algicola 
                 
                 
                   B. atrophaeus 
                 
                 
                   B. herbersteinensis 
                 
                 
                   B. thermoruber 
                 
                 
                   B. macquariensis 
                 
               
               
                 
                   B. alginolyticus 
                 
                 
                   B. axarquiensis 
                 
                 
                   B. horikoshii 
                 
                 
                   B. thermosphaericus 
                 
                 
                   B. macyae 
                 
               
               
                 
                   B. alkalidiazotrophicus 
                 
                 
                   B. azotofixans 
                 
                 
                   B. horneckiae 
                 
                 
                   B. thiaminolyticus 
                 
                 
                   B. malacitensis 
                 
               
               
                 
                   B. alkalinitrilicus 
                 
                 
                   B. azotoformans 
                 
                 
                   B. horti 
                 
                 
                   B. thioparans 
                 
                 
                   B. mannanilyticus 
                 
               
               
                 
                   B. alkalisediminis 
                 
                 
                   B. badius 
                 
                 
                   B. huizhouensis 
                 
                 
                   B. thuringiensis 
                 
                 
                   B. marisflavi 
                 
               
               
                 
                   B. alkalitelluris 
                 
                 
                   B. barbaricus 
                 
                 
                   B. humi 
                 
                 
                   B. tianshenii 
                 
                 
                   B. marismortui 
                 
               
               
                 
                   B. altitudinis 
                 
                 
                   B. bataviensis 
                 
                 
                   B. hwajinpoensis 
                 
                 
                   B. trypoxylicola 
                 
                 
                   B. marmarensis 
                 
               
               
                 
                   B. alveayuensis 
                 
                 
                   B. beijingensis 
                 
                 
                   B. idriensis 
                 
                 
                   B. tusciae 
                 
                 
                   B. massiliensis 
                 
               
               
                 
                   B. alvei 
                 
                 
                   B. benzoevorans 
                 
                 
                   B. indicus 
                 
                 
                   B. validus 
                 
                 
                   B. megaterium 
                 
               
               
                 
                   B. amyloliquefaciens 
                 
                 
                   B. beringensis 
                 
                 
                   B. infantis 
                 
                 
                   B. vallismortis 
                 
                 
                   B. mesonae 
                 
               
               
                 B. 
                 
                   B. berkeleyi 
                 
                 
                   B. infernus 
                 
                 
                   B. vedderi 
                 
                 
                   B. methanolicus 
                 
               
               
                 a. subsp.  amyloliquefaciens   
                 
                   B. beveridgei 
                 
                 
                   B. insolitus 
                 
                 
                   B. velezensis 
                 
                 
                   B. methylotrophicus 
                 
               
               
                 B. a. subsp.  plantarum   
                 
                   B. bogoriensis 
                 
                 
                   B. invictae 
                 
                 
                   B. vietnamensis 
                 
                 
                   B. migulanus 
                 
               
               
                   
                 
                   B. boroniphilus 
                 
                 
                   B. iranensis 
                 
                 
                   B. vireti 
                 
                 
                   B. mojavensis 
                 
               
               
                 
                   B. dipsosauri 
                 
                 
                   B. borstelensis 
                 
                 
                   B. isabeliae 
                 
                 
                   B. vulcani 
                 
                 
                   B. mucilaginosus 
                 
               
               
                 
                   B. drentensis 
                 
                 
                   B. brevis Migula 
                 
                 
                   B. isronensis 
                 
                 
                   B. wakoensis 
                 
                 
                   B. muralis 
                 
               
               
                 
                   B. edaphicus 
                 
                 
                   B. butanolivorans 
                 
                 
                   B. jeotgali 
                 
                 
                   B. weihenstephanensis 
                 
                 
                   B. murimartini 
                 
               
               
                 
                   B. ehimensis 
                 
                 
                   B. canaveralius 
                 
                 
                   B. kaustophilus 
                 
                 
                   B. xiamenensis 
                 
                 
                   B. mycoides 
                 
               
               
                 
                   B. eiseniae 
                 
                 
                   B. carboniphilus 
                 
                 
                   B. kobensis 
                 
                 
                   B. xiaoxiensis 
                 
                 
                   B. naganoensis 
                 
               
               
                 
                   B. enclensis 
                 
                 
                   B. cecembensis 
                 
                 
                   B. kochii 
                 
                 
                   B. zhanjiangensis 
                 
                 
                   B. nanhaiensis 
                 
               
               
                 
                   B. endophyticus 
                 
                 
                   B. cellulosilyticus 
                 
                 
                   B. kokeshiiformis 
                 
                 
                   B. peoriae 
                 
                 
                   B. nanhaiisediminis 
                 
               
               
                 
                   B. endoradicis 
                 
                 
                   B. centrosporus 
                 
                 
                   B. koreensis 
                 
                 
                   B. persepolensis 
                 
                 
                   B. nealsonii 
                 
               
               
                 
                   B. farraginis 
                 
                 
                   B. cereus 
                 
                 
                   B. korlensis 
                 
                 
                   B. persicus 
                 
                 
                   B. neidei 
                 
               
               
                 
                   B. fastidiosus 
                 
                 
                   B. chagannorensis 
                 
                 
                   B. kribbensis 
                 
                 
                   B. pervagus 
                 
                 
                   B. neizhouensis 
                 
               
               
                 
                   B. fengqiuensis 
                 
                 
                   B. chitinolyticus 
                 
                 
                   B. krulwichiae 
                 
                 
                   B. plakortidis 
                 
                 
                   B. niabensis 
                 
               
               
                 
                   B. firmus 
                 
                 
                   B. chondroitinus 
                 
                 
                   B. laevolacticus 
                 
                 
                   B. pocheonensis 
                 
                 
                   B. niacini 
                 
               
               
                 
                   B. flexus 
                 
                 
                   B. choshinensis 
                 
                 
                   B. larvae 
                 
                 
                   B. polygoni 
                 
                 
                   B. novalis 
                 
               
               
                 
                   B. foraminis 
                 
                 
                   B. chungangensis 
                 
                 
                   B. laterosporus 
                 
                 
                   B. polymyxa 
                 
                 
                   B. oceanisediminis 
                 
               
               
                 
                   B. fordii 
                 
                 
                   B. cibi 
                 
                 
                   B. salexigens 
                 
                 
                   B. popilliae 
                 
                 
                   B. odysseyi 
                 
               
               
                 
                   B. formosus 
                 
                 
                   B. circulans 
                 
                 
                   B. saliphilus 
                 
                 
                   B. pseudalcalophilus 
                 
                 
                   B. okhensis 
                 
               
               
                 
                   B. fortis 
                 
                 
                   B. clarkii 
                 
                 
                   B. schlegelii 
                 
                 
                   B. pseudofirmus 
                 
                 
                   B. okuhidensis 
                 
               
               
                 
                   B. fumarioli 
                 
                 
                   B. clausii 
                 
                 
                   B. sediminis 
                 
                 
                   B. pseudomycoides 
                 
                 
                   B. oleronius 
                 
               
               
                 
                   B. funiculus 
                 
                 
                   B. coagulans 
                 
                 
                   B. selenatarsenatis 
                 
                 
                   B. psychrodurans 
                 
                 
                   B. oryzaecorticis 
                 
               
               
                 
                   B. fusiformis 
                 
                 
                   B. coahuilensis 
                 
                 
                   B. selenitireducens 
                 
                 
                   B. psychrophilus 
                 
                 
                   B. oshimensis 
                 
               
               
                 
                   B. galactophilus 
                 
                 
                   B. cohnii 
                 
                 
                   B. seohaeanensis 
                 
                 
                   B. psychrosaccharolyticus 
                 
                 
                   B. pabuli 
                 
               
               
                 
                   B. galactosidilyticus 
                 
                 
                   B. composti 
                 
                 
                   B. shacheensis 
                 
                 
                   B. psychrotolerans 
                 
                 
                   B. pakistanensis 
                 
               
               
                 
                   B. galliciensis 
                 
                 
                   B. curdlanolyticus 
                 
                 
                   B. shackletonii 
                 
                 
                   B. pulvifaciens 
                 
                 
                   B. pallidus 
                 
               
               
                 
                   B. gelatini 
                 
                 
                   B. cycloheptanicus 
                 
                 
                   B. siamensis 
                 
                 
                   B. pumilus 
                 
                 
                   B. pallidus 
                 
               
               
                 
                   B. gibsonii 
                 
                 
                   B. cytotoxicus 
                 
                 
                   B. silvestris 
                 
                 
                   B. purgationiresistens 
                 
                 
                   B. panacisoli 
                 
               
               
                 
                   B. ginsengi 
                 
                 
                   B. daliensis 
                 
                 
                   B. simplex 
                 
                 
                   B. pycnus 
                 
                 
                   B. panaciterrae 
                 
               
               
                 
                   B. ginsengihumi 
                 
                 
                   B. decisifrondis 
                 
                 
                   B. siralis 
                 
                 
                   B. qingdaonensis 
                 
                 
                   B. pantothenticus 
                 
               
               
                 
                   B. ginsengisoli 
                 
                 
                   B. decolorationis 
                 
                 
                   B. smithii 
                 
                 
                   B. qingshengii 
                 
                 
                   B. parabrevis 
                 
               
               
                   B. globisporus  (eg, B. 
                 
                   B. deserti 
                 
                 
                   B. soli 
                 
                 
                   B. reuszeri 
                 
                 
                   B. paraflexus 
                 
               
               
                 g. subsp.  Globisporus ; or B. 
                   
                 
                   B. solimangrovi 
                 
                 
                   B. rhizosphaerae 
                 
                 
                   B. pasteurii 
                 
               
               
                 g. subsp.  Marinus ) 
                   
                 
                   B. solisalsi 
                 
                 
                   B. rigui 
                 
                 
                   B. patagoniensis 
                 
               
               
                   
                   
                 
                   B. songklensis 
                 
                 
                   B. ruris 
                 
               
               
                   
                   
                 
                   B. sonorensis 
                 
                 
                   B. safensis 
                 
               
               
                   
                   
                 
                   B. sphaericus 
                 
                 
                   B. salarius 
                 
               
               
                   
                   
                 
                   B. sporothermodurans 
                 
               
               
                   
                   
                 
                   B. stearothermophilus 
                 
               
               
                   
                   
                 
                   B. stratosphericus 
                 
               
               
                   
                   
                 
                   B. subterraneus 
                 
               
               
                   
                   
                   B. subtilis  (eg, B. 
               
               
                   
                   
                 s. subsp.  Inaquosorum , or B. 
               
               
                   
                   
                 s. subsp.  Spizizenr , or B. 
               
               
                   
                   
                 s. subsp.  Subtilis ) 
               
               
                 
                   Caenimonas 
                 
                 
                   Campylobacter 
                 
                 
                   Cardiobacterium 
                 
                 
                   Catenuloplanes 
                 
                 
                   Curtobacterium 
                 
               
               
                 
                   Caenimonas koreensis 
                 
                 
                   Campylobacter coli 
                 
                 
                   Cardiobacterium hominis 
                 
                 
                   Catenuloplanes atrovinosus 
                 
                 
                   Curtobacterium albidum 
                 
               
               
                 
                   Caldalkalibacillus 
                 
                 
                   Campylobacter concisus 
                 
                 
                   Carnimonas 
                 
                 
                   Catenuloplanes castaneus 
                 
                 
                   Curtobacterium citreus 
                 
               
               
                 
                   Caldalkalibacillus uzonensis 
                 
                 
                   Campylobacter curvus 
                 
                 
                   Carnimonas nigrificans 
                 
                 
                   Catenuloplanes crispus 
                 
               
               
                 
                   Caldanaerobacter 
                 
                 
                   Campylobacter fetus 
                 
                 
                   Carnobacterium 
                 
                 
                   Catenuloplanes indicus 
                 
               
               
                 
                   Caldanaerobacter subterraneus 
                 
                 
                   Campylobacter gracilis 
                 
                 
                   Carnobacterium alterfunditum 
                 
                 
                   Catenuloplanes japonicus 
                 
               
               
                 
                   Caldanaerobius 
                 
                 
                   Campylobacter helveticus 
                 
                 
                   Carnobacterium divergens 
                 
                 
                   Catenuloplanes nepalensis 
                 
               
               
                 
                   Caldanaerobius fijiensis 
                 
                 
                   Campylobacter hominis 
                 
                 
                   Carnobacterium funditum 
                 
                 
                   Catenuloplanes niger 
                 
               
               
                 
                   Caldanaerobius 
                 
                 
                   Campylobacter hyointestinalis 
                 
                 
                   Carnobacterium gallinarum 
                 
                 
                   Chryseobacterium 
                 
               
               
                 
                   polysaccharolyticus 
                 
                 
                   Campylobacter jejuni 
                 
                 
                   Carnobacterium 
                 
                 
                   Chryseobacterium 
                 
               
               
                 
                   Caldanaerobius zeae 
                 
                 
                   Campylobacter lari 
                 
                 
                   maltaromaticum 
                 
                 
                   balustinum 
                 
               
               
                 
                   Caldanaerovirga 
                 
                 
                   Campylobacter mucosalis 
                 
                 
                   Carnobacterium mobile 
                 
                 
                   Citrobacter 
                 
               
               
                 
                   Caldanaerovirga acetigignens 
                 
                 
                   Campylobacter rectus 
                 
                 
                   Carnobacterium viridans 
                 
                 
                   C. amalonaticus 
                 
               
               
                 
                   Caldicellulosiruptor 
                 
                 
                   Campylobacter showae 
                 
                 
                   Caryophanon 
                 
                 
                   C. braakii 
                 
               
               
                 
                   Caldicellulosiruptor bescii 
                 
                 
                   Campylobacter sputorum 
                 
                 
                   Caryophanon latum 
                 
                 
                   C. diversus 
                 
               
               
                 
                   Caldicellulosiruptor kristjanssonii 
                 
                 
                   Campylobacter upsaliensis 
                 
                 
                   Caryophanon tenue 
                 
                 
                   C. farmeri 
                 
               
               
                 
                   Caldicellulosiruptor owensensis 
                 
                 
                   Capnocytophaga 
                 
                 
                   Catellatospora 
                 
                 
                   C. freundii 
                 
               
               
                   
                 
                   Capnocytophaga canimorsus 
                 
                 
                   Catellatospora citrea 
                 
                 
                   C. gillenii 
                 
               
               
                   
                 
                   Capnocytophaga cynodegmi 
                 
                 
                   Catellatospora 
                 
                 
                   C. koseri 
                 
               
               
                   
                 
                   Capnocytophaga gingivalis 
                 
                 
                   methionotrophica 
                 
                 
                   C. murliniae 
                 
               
               
                   
                 
                   Capnocytophaga granulosa 
                 
                 
                   Catenococcus 
                 
                 
                   C. pasteurii 
                   [1] 
                 
               
               
                   
                 
                   Capnocytophaga haemolytica 
                 
                 
                   Catenococcus thiocycli 
                 
                 
                   C. rodentium 
                 
               
               
                   
                 
                   Capnocytophaga ochracea 
                 
                   
                 
                   C. sedlakii 
                 
               
               
                   
                 
                   Capnocytophaga sputigena 
                 
                   
                 
                   C. werkmanii 
                 
               
               
                   
                   
                   
                 
                   C. youngae 
                 
               
               
                   
                   
                   
                 
                   Clostridium 
                 
               
               
                   
                   
                   
                 (see below) 
               
               
                   
                   
                   
                 
                   Coccochloris 
                 
               
               
                   
                   
                   
                 
                   Coccochloris elabens 
                 
               
               
                   
                   
                   
                 
                   Corynebacterium 
                 
               
               
                   
                   
                   
                 
                   Corynebacterium flavescens 
                 
               
               
                   
                   
                   
                 
                   Corynebacterium variabile 
                 
               
               
                 
                   Clostridium 
                 
               
            
           
           
               
            
               
                   Clostridium absonum ,  Clostridium aceticum ,  Clostridium acetireducens ,  Clostridium acetobutylicum ,  Clostridium acidisoli ,  Clostridium aciditolerans,    Clostridium acidurici ,  Clostridium aerotolerans ,  Clostridium   
               
               
                   aestuarii ,  Clostridium akagii ,  Clostridium aldenense ,  Clostridium aldrichii ,  Clostridium algidicarni ,  Clostridium algidixylanolyticum ,  Clostridium algifaecis ,  Clostridium algoriphilum ,  Clostridium alkalicellulosi , 
               
               
                   Clostridium aminophilum ,  Clostridium aminovalericum ,  Clostridium amygdalinum ,  Clostridium amylolyticum ,  Clostridium arbusti ,  Clostridium arcticum ,  Clostridium argentinense ,  Clostridium asparagiforme , 
               
               
                   Clostridium aurantibutyricum ,  Clostridium autoethanogenum ,  Clostridium baratii ,  Clostridium barkeri ,  Clostridium bartlettii ,  Clostridium beijerinckii ,  Clostridium bifermentans ,  Clostridium bolteae ,  Clostridium   
               
               
                   bornimense ,  Clostridium botulinum ,  Clostridium bowmanii ,  Clostridium bryantii ,  Clostridium butyricum ,  Clostridium cadaveris ,  Clostridium caenicola ,  Clostridium caminithermale ,  Clostridium carboxidivorans , 
               
               
                   Clostridium carnis ,  Clostridium cavendishii ,  Clostridium celatum ,  Clostridium celerecrescens ,  Clostridium cellobioparum ,  Clostridium cellulofermentans ,  Clostridium cellulolyticum ,  Clostridium cellulosi , 
               
               
                   Clostridium cellulovorans ,  Clostridium chartatabidum ,  Clostridium chauvoei ,  Clostridium chromiireducens ,  Clostridium citroniae ,  Clostridium clariflavum ,  Clostridium clostridioforme ,  Clostridium coccoides , 
               
               
                   Clostridium cochlearium ,  Clostridium colletant ,  Clostridium colicanis ,  Clostridium colinum ,  Clostridium collagenovorans ,  Clostridium cylindrosporum ,  Clostridium difficile ,  Clostridium diolis ,  Clostridium   
               
               
                   disporicum ,  Clostridium drakei ,  Clostridium durum ,  Clostridium estertheticum ,  Clostridium estertheticum estertheticum ,  Clostridium estertheticum laramiense ,  Clostridium fallax ,  Clostridium felsineum ,  Clostridium   
               
               
                   fervidum ,  Clostridium fimetarium ,  Clostridium formicaceticum ,  Clostridium frigidicarnis ,  Clostridium frigoris ,  Clostridium ganghwense ,  Clostridium gasigenes ,  Clostridium ghonii ,  Clostridium glycolicum , 
               
               
                   Clostridium glycyrrhizinilyticum ,  Clostridium grantii ,  Clostridium haemolyticum ,  Clostridium halophilum ,  Clostridium hastiforme ,  Clostridium hathewayi ,  Clostridium herbivorans ,  Clostridium hiranonis , 
               
               
                   Clostridium histolyticum ,  Clostridium homopropionicum ,  Clostridium huakuii ,  Clostridium hungatei ,  Clostridium hydrogeniformans ,  Clostridium hydroxybenzoicum ,  Clostridium hylemonae ,  Clostridium jejuense , 
               
               
                   Clostridium indolis ,  Clostridium innocuum ,  Clostridium intestinale ,  Clostridium irregulare ,  Clostridium isatidis ,  Clostridium josui ,  Clostridium kluyveri ,  Clostridium lactatifermentans ,  Clostridium lacusfryxellense , 
               
               
                   Clostridium laramiense ,  Clostridium lavalense ,  Clostridium lentocellum ,  Clostridium lentoputrescens ,  Clostridium leptum ,  Clostridium limosum ,  Clostridium litorale ,  Clostridium lituseburense ,  Clostridium ljungdahlii , 
               
               
                   Clostridium lortetii ,  Clostridium lundense ,  Clostridium magnum ,  Clostridium malenominatum ,  Clostridium mangenotii ,  Clostridium mayombei ,  Clostridium methoxybenzovorans ,  Clostridium methylpentosum , 
               
               
                   Clostridium neopropionicum ,  Clostridium nexile ,  Clostridium nitrophenolicum ,  Clostridium novyi ,  Clostridium oceanicum ,  Clostridium orbiscindens ,  Clostridium oroticum ,  Clostridium oxalicum ,  Clostridium   
               
               
                   papyrosolvens ,  Clostridium paradoxum ,  Clostridium paraperfringens  (Alias:  C. welchii ),  Clostridium paraputrificum ,  Clostridium pascui ,  Clostridium pasteurianum ,  Clostridium peptidivorans ,  Clostridium perenne , 
               
               
                   Clostridium perfringens ,  Clostridium pfennigii ,  Clostridium phytofermentans ,  Clostridium piliforme ,  Clostridium polysaccharolyticum ,  Clostridium populeti ,  Clostridium propionicum ,  Clostridium proteoclasticum , 
               
               
                   Clostridium proteolyticum ,  Clostridium psychrophilum ,  Clostridium puniceum ,  Clostridium purinilyticum ,  Clostridium putrefaciens ,  Clostridium putrificum ,  Clostridium quercicolum ,  Clostridium quinii , 
               
               
                   Clostridium ramosum ,  Clostridium rectum ,  Clostridium roseum ,  Clostridium saccharobutylicum ,  Clostridium saccharogumia ,  Clostridium saccharolyticum ,  Clostridium saccharoperbutylacetonicum ,  Clostridium   
               
               
                   sardiniense ,  Clostridium sartagoforme ,  Clostridium scatologenes ,  Clostridium schirmacherense ,  Clostridium scindens ,  Clostridium septicum ,  Clostridium sordellii ,  Clostridium sphenoides ,  Clostridium spiroforme , 
               
               
                   Clostridium sporogenes ,  Clostridium sporosphaeroides ,  Clostridium stercorarium ,  Clostridium stercorarium leptospartum ,  Clostridium stercorarium stercorarium ,  Clostridium stercorarium thermolacticum , 
               
               
                   Clostridium sticklandii ,  Clostridium straminisolvens ,  Clostridium subterminale ,  Clostridium sufflavum ,  Clostridium sulfidigenes ,  Clostridium symbiosum ,  Clostridium tagluense ,  Clostridium   
               
               
                   tepidiprofundi ,  Clostridium termitidis ,  Clostridium tertium ,  Clostridium tetani ,  Clostridium tetanomorphum ,  Clostridium thermaceticum ,  Clostridium thermautotrophicum ,  Clostridium thermoalcaliphilum , 
               
               
                   Clostridium thermobutyricum ,  Clostridium thermocellum ,  Clostridium thermocopriae ,  Clostridium thermohydrosulfuricum ,  Clostridium thermolacticum ,  Clostridium thermopalmarium , 
               
               
                   Clostridium thermopapyrolyticum ,  Clostridium thermosaccharolyticum ,  Clostridium thermosuccinogenes ,  Clostridium thermosulfurigenes ,  Clostridium thiosulfatireducens ,  Clostridium tyrobutyricum , 
               
               
                   Clostridium uliginosum ,  Clostridium ultunense ,  Clostridium villosum ,  Clostridium vincentii ,  Clostridium viride ,  Clostridium xylanolyticum ,  Clostridium xylanovorans   
               
            
           
           
               
               
               
               
               
            
               
                 
                   Dactylosporangium 
                 
                 
                   Deinococcus 
                 
                 
                   Delftia 
                 
                 
                   Echinicola 
                 
                   
               
               
                 
                   Dactylosporangium aurantiacum 
                 
                 
                   Deinococcus aerius 
                 
                 
                   Delftia acidovorans 
                 
                 
                   Echinicola pacifica 
                 
               
               
                 
                   Dactylosporangium fulvum 
                 
                 
                   Deinococcus apachensis 
                 
                 
                   Desulfovibrio 
                 
                 
                   Echinicola vietnamensis 
                 
               
               
                 
                   Dactylosporangium matsuzakiense 
                 
                 
                   Deinococcus aquaticus 
                 
                 
                   Desulfovibrio desulfuricans 
                 
               
               
                 
                   Dactylosporangium roseum 
                 
                 
                   Deinococcus aquatilis 
                 
                 
                   Diplococcus 
                 
               
               
                 
                   Dactylosporangium thailandense 
                 
                 
                   Deinococcus caeni 
                 
                 
                   Diplococcus pneumoniae 
                 
               
               
                 
                   Dactylosporangium vinaceum 
                 
                 
                   Deinococcus radiodurans 
                 
               
               
                   
                 
                   Deinococcus radiophilus 
                 
               
               
                 
                   Enterobacter 
                 
                 
                   Enterobacter kobei 
                 
                 
                   Faecalibacterium 
                 
                 
                   Flavobacterium 
                 
               
               
                 
                   E. aerogenes 
                 
                 
                   E. ludwigii 
                 
                 
                   Faecalibacterium prausnitzii 
                 
                 
                   Flavobacterium antarcticum 
                 
               
               
                 
                   E. amnigemis 
                 
                 
                   E. mori 
                 
                 
                   Fangia 
                 
                 
                   Flavobacterium aquatile 
                 
               
               
                 
                   E. agglomerans 
                 
                 
                   E. nimipressuralis 
                 
                 
                   Fangia hongkongensis 
                 
                 
                   Flavobacterium aquidurense 
                 
               
               
                 
                   E. arachidis 
                 
                 
                   E. oryzae 
                 
                 
                   Fastidiosipila 
                 
                 
                   Flavobacterium balustinum 
                 
               
               
                 
                   E. asburiae 
                 
                 
                   E. pulveris 
                 
                 
                   Fastidiosipila sanguinis 
                 
                 
                   Flavobacterium croceum 
                 
               
               
                 
                   E. cancerogenous 
                 
                 
                   E. pyrinus 
                 
                 
                   Fusobacterium 
                 
                 
                   Flavobacterium cucumis 
                 
               
               
                 
                   E. cloacae 
                 
                 
                   E. radicincitans 
                 
                 
                   Fusobacterium nucleatum 
                 
                 
                   Flavobacterium daejeonense 
                 
               
               
                 
                   E. cowanii 
                 
                 
                   E. taylorae 
                 
                   
                 
                   Flavobacterium defluvii 
                 
               
               
                 
                   E. dissolvens 
                 
                 
                   E. turicensis 
                 
                   
                 
                   Flavobacterium degerlachei 
                 
               
               
                 
                   E. gergoviae 
                 
                 
                   E. sakazakii Enterobacter soli 
                 
                   
                 
                   Flavobacterium 
                 
               
               
                 
                   E. helveticus 
                 
                 
                   Enterococcus 
                 
                   
                 
                   denitrificans 
                 
               
               
                 
                   E. hormaechei 
                 
                 
                   Enterococcus durans 
                 
                   
                 
                   Flavobacterium filum 
                 
               
               
                 
                   E. intermedins 
                 
                 
                   Enterococcus faecalis 
                 
                   
                 
                   Flavobacterium flevense 
                 
               
               
                   
                 
                   Enterococcus faecium 
                 
                   
                 
                   Flavobacterium frigidarium 
                 
               
               
                   
                 
                   Erwinia 
                 
                   
                 
                   Flavobacterium mizutaii 
                 
               
               
                   
                 
                   Erwinia hapontici 
                 
                   
                 
                   Flavobacterium 
                 
               
               
                   
                 
                   Escherichia 
                 
                   
                 
                   okeanokoites 
                 
               
               
                   
                 
                   Escherichia coli 
                 
               
               
                 
                   Gaetbulibacter 
                 
                 
                   Haemophilus 
                 
                 
                   Ideonella 
                 
                 
                   Janibacter 
                 
               
               
                 
                   Gaetbulibacter saemankumensis 
                 
                 
                   Haemophilus aegyptius 
                 
                 
                   Ideonella azotifigens 
                 
                 
                   Janibacter anophelis 
                 
               
               
                 
                   Gallibacterium 
                 
                 
                   Haemophilus aphrophilus 
                 
                 
                   Idiomarina 
                 
                 
                   Janibacter corallicola 
                 
               
               
                 
                   Gallibacterium anatis 
                 
                 
                   Haemophilus felis 
                 
                 
                   Idiomarina abyssalis 
                 
                 
                   Janibacter limosus 
                 
               
               
                 
                   Gallicola 
                 
                 
                   Haemophilus gallinarum 
                 
                 
                   Idiomarina baltica 
                 
                 
                   Janibacter melonis 
                 
               
               
                 
                   Gallicola barnesae 
                 
                 
                   Haemophilus haemolyticus 
                 
                 
                   Idiomarina fontislapidosi 
                 
                 
                   Janibacter terrae 
                 
               
               
                 
                   Garciella 
                 
                 
                   Haemophilus influenzae 
                 
                 
                   Idiomarina loihiensis 
                 
                 
                   Jannaschia 
                 
               
               
                 
                   Garciella nitratireducens 
                 
                 
                   Haemophilus paracuniculus 
                 
                 
                   Idiomarina ramblicola 
                 
                 
                   Jannaschia cystaugens 
                 
               
               
                 
                   Geobacillus 
                 
                 
                   Haemophilus parahaemolyticus 
                 
                 
                   Idiomarina seosinensis 
                 
                 
                   Jannaschia helgolandensis 
                 
               
               
                 
                   Geobacillus thermoglucosidasius 
                 
                 
                   Haemophilus parainfluenzae 
                 
                 
                   Idiomarina zobellii 
                 
                 
                   Jannaschia 
                 
               
               
                 
                   Geobacillus stearothermophilus 
                 
                 
                   Haemophilus 
                 
                 
                   Ignatzschineria 
                 
                 
                   pohangensis 
                 
               
               
                 
                   Geobacter 
                 
                 
                   paraphrohaemolyticus 
                 
                 
                   Ignatzschineria 
                 
                 
                   Jannaschia rubra 
                 
               
               
                 
                   Geobacter bemidjiensis 
                 
                 
                   Haemophilus parasuis 
                 
                 
                   larvae 
                 
                 
                   Janthinobacterium 
                 
               
               
                 
                   Geobacter bremensis 
                 
                 
                   Haemophilus pittmaniae 
                 
                 
                   Ignavigranum 
                 
                 
                   Janthinobacterium 
                 
               
               
                 
                   Geobacter chapellei 
                 
                 
                   Hafnia 
                 
                 
                   Ignavigranum ruoffiae 
                 
                 
                   agaricidamnosum 
                 
               
               
                 
                   Geobacter grbiciae 
                 
                 
                   Hafnia alvei 
                 
                 
                   Ilumatobacter 
                 
                 
                   Janthinobacterium lividum 
                 
               
               
                 
                   Geobacter hydrogenophilus 
                 
                 
                   Hahella 
                 
                 
                   Ilumatobacter fluminis 
                 
                 
                   Jejuia 
                 
               
               
                 
                   Geobacter lovleyi 
                 
                 
                   Hahella ganghwensis 
                 
                 
                   Ilyobacter 
                 
                 
                   Jejuia pallidilutea 
                 
               
               
                 
                   Geobacter metallireducens 
                 
                 
                   Halalkalibacillus 
                 
                 
                   Ilyobacter delafieldii 
                 
                 
                   Jeotgalibacillus 
                 
               
               
                 
                   Geobacter pelophilus 
                 
                 
                   Halalkalibacillus halophilus 
                 
                 
                   Ilyobacter insuetus 
                 
                 
                   Jeotgalibacillus 
                 
               
               
                 
                   Geobacter pickeringii 
                 
                 
                   Helicobacter 
                 
                 
                   Ilyobacter polytropus 
                 
                 
                   alimentarius 
                 
               
               
                 
                   Geobacter sulfurreducens 
                 
                 
                   Helicobacter pylori 
                 
                 
                   Ilyobacter tartaricus 
                 
                 
                   Jeotgalicoccus 
                 
               
               
                 
                   Geodermatophilus 
                 
                   
                   
                 
                   Jeotgalicoccus halotolerans 
                 
               
               
                 
                   Geodermatophilus obscurus 
                 
               
               
                 
                   Gluconacetobacter 
                 
               
               
                 
                   Gluconacetobacter xylinus 
                 
               
               
                 
                   Gordonia 
                 
               
               
                 
                   Gordonia rubripertincta 
                 
               
               
                 
                   Kaistia 
                 
                 
                   Labedella 
                 
                 
                   Listeria ivanovii 
                 
                 
                   Micrococcus 
                 
                 
                   Nesterenkonia 
                 
               
               
                 
                   Kaistia adipata 
                 
                 
                   Labedella gwakjiensis 
                 
                 
                   L. marthii 
                 
                 
                   Micrococcus luteus 
                 
                 
                   Nesterenkonia holobia 
                 
               
               
                 
                   Kaistia soli 
                 
                 
                   Labrenzia 
                 
                 
                   L. monocytogenes 
                 
                 
                   Micrococcus lylae 
                 
                 
                   Nocardia 
                 
               
               
                 
                   Kangiella 
                 
                 
                   Labrenzia aggregata 
                 
                 
                   L. newyorkensis 
                 
                 
                   Moraxella 
                 
                 
                   Nocardia argentinensis 
                 
               
               
                 
                   Kangiella aquimarina 
                 
                 
                   Labrenzia alba 
                 
                 
                   L. riparia 
                 
                 
                   Moraxella bovis 
                 
                 
                   Nocardia corallina 
                 
               
               
                 
                   Kangiella 
                 
                 
                   Labrenzia alexandrii 
                 
                 
                   L. rocourtiae 
                 
                 
                   Moraxella nonliquefaciens 
                 
                 
                   Nocardia 
                 
               
               
                 
                   koreensis 
                 
                 
                   Labrenzia marina 
                 
                 
                   L. seeligeri 
                 
                 
                   Moraxella osloensis 
                 
                 
                   otitidiscaviarum 
                 
               
               
                 
                   Kerstersia 
                 
                 
                   Labrys 
                 
                 
                   L. weihenstephanensis 
                 
                 
                   Nakamurella 
                 
               
               
                 
                   Kerstersia gyiorum 
                 
                 
                   Labrys methylaminiphilus 
                 
                 
                   L. welshimeri 
                 
                 
                   Nakamurella multipartita 
                 
               
               
                 
                   Kiloniella 
                 
                 
                   Labrys miyagiensis 
                 
                 
                   Listonella 
                 
                 
                   Nannocystis 
                 
               
               
                 
                   Kiloniella laminariae 
                 
                 
                   Labrys monachus 
                 
                 
                   Listonella anguillarum 
                 
                 
                   Nannocystis pusilia 
                 
               
               
                 
                   Klebsiella 
                 
                 
                   Labrys okinawensis 
                 
                 
                   Macrococcus 
                 
                 
                   Natranaerobius 
                 
               
               
                 
                   K. gramilomatis 
                 
                 
                   Labrys 
                 
                 
                   Macrococcus bovicus 
                 
                 
                   Natranaerobius 
                 
               
               
                 
                   K. oxytoca 
                 
                 
                   portucalensis 
                 
                 
                   Marinobacter 
                 
                 
                   thermophilus 
                 
               
               
                 
                   K. pneumoniae 
                 
                 
                   Lactobacillus 
                 
                 
                   Marinobacter algicola 
                 
                 
                   Natranaerobius trueperi 
                 
               
               
                 
                   K. terrigena 
                 
                 [see below] 
                 
                   Marinobacter bryozoorum 
                 
                 
                   Naxibacter 
                 
               
               
                 
                   K. variicola 
                 
                 
                   Laceyella 
                 
                 
                   Marinobacter flavimaris 
                 
                 
                   Naxibacter alkalitolerans 
                 
               
               
                 
                   Kluyvera 
                 
                 
                   Laceyella putida 
                 
                 
                   Meiothermus 
                 
                 
                   Neisseria 
                 
               
               
                 
                   Kluyvera ascorbata 
                 
                 
                   Lechevalieria 
                 
                 
                   Meiothermus ruber 
                 
                 
                   Neisseria cinerea 
                 
               
               
                 
                   Kocuria 
                 
                 
                   Lechevalieria aerocolonigenes 
                 
                 
                   Methylophilus 
                 
                 
                   Neisseria denitrificans 
                 
               
               
                 
                   Kocuria roasea 
                 
                 
                   Legionella 
                 
                 
                   Methylophilus methylotrophus 
                 
                 
                   Neisseria gonorrhoeae 
                 
               
               
                 
                   Kocuria varians 
                 
                 [see below] 
                 
                   Microbacterium 
                 
                 
                   Neisseria lactamica 
                 
               
               
                 
                   Kurthia 
                 
                 
                   Listeria 
                 
                 
                   Microbacterium 
                 
                 
                   Neisseria mucosa 
                 
               
               
                 
                   Kurthia zopfii 
                 
                 
                   L. aquatica 
                 
                 
                   ammoniaphilum 
                 
                 
                   Neisseria sicca 
                 
               
               
                   
                 
                   L. booriae 
                 
                 
                   Microbacterium arborescens 
                 
                 
                   Neisseria subflava 
                 
               
               
                   
                 
                   L. cornellensis 
                 
                 
                   Microbacterium liquefaciens 
                 
                 
                   Neptunomonas 
                 
               
               
                   
                 
                   L. fleischmannii 
                 
                 
                   Microbacterium oxydans 
                 
                 
                   Neptunomonas japonica 
                 
               
               
                   
                 
                   L. floridensis 
                 
               
               
                   
                 
                   L. grandensis 
                 
               
               
                   
                 
                   L. grayi 
                 
               
               
                   
                 
                   L. innocua 
                 
               
               
                 
                   Lactobacillus 
                 
               
               
                 
                   L. acetotolerans 
                 
                 
                   L. catenaformis 
                 
                 
                   L. mali 
                 
                 
                   L. parakefiri 
                 
                 
                   L. sakei 
                 
               
               
                 
                   L. acidifarinae 
                 
                 
                   L. ceti 
                 
                 
                   L. manihotivorans 
                 
                 
                   L. paralimentarius 
                 
                 
                   L. salivarius 
                 
               
               
                 
                   L. acidipiscis 
                 
                 
                   L. coleohominis 
                 
                 
                   L. mindensis 
                 
                 
                   L. paraplantarum 
                 
                 
                   L. sanfranciscensis 
                 
               
               
                 
                   L. acidophilus 
                 
                 
                   L. collinoides 
                 
                 
                   L. mucosae 
                 
                 
                   L. pentosus 
                 
                 
                   L. satsumensis 
                 
               
               
                 
                   Lactobacillus agilis 
                 
                 
                   L. composti 
                 
                 
                   L. murinus 
                 
                 
                   L. perolens 
                 
                 
                   L. secaliphilus 
                 
               
               
                 
                   L. algidus 
                 
                 
                   L. concavus 
                 
                 
                   L. nagelii 
                 
                 
                   L. plantarum 
                 
                 
                   L. sharpeae 
                 
               
               
                 
                   L. alimentarius 
                 
                 
                   L. coryniformis 
                 
                 
                   L. namurensis 
                 
                 
                   L. pontis 
                 
                 
                   L. siliginis 
                 
               
               
                 
                   L. amylolyticus 
                 
                 
                   L. crispatus 
                 
                 
                   L. nantensis 
                 
                 
                   L. protectus 
                 
                 
                   L. spicheri 
                 
               
               
                 
                   L. amylophilus 
                 
                 
                   L. crustorum 
                 
                 
                   L. oligofermentans 
                 
                 
                   L. psittaci 
                 
                 
                   L. suebicus 
                 
               
               
                 
                   L. amylotrophicus 
                 
                 
                   L. curvatus 
                 
                 
                   L. oris 
                 
                 
                   L. rennini 
                 
                 
                   L. thailandensis 
                 
               
               
                 
                   L. amylovorus 
                 
                   L. delbrueckii  subsp.  bulgaricus   
                 
                   L. panis 
                 
                 
                   L. reuteri 
                 
                 
                   L. ultunensis 
                 
               
               
                 
                   L. animalis 
                 
                   L. delbrueckii  subsp. 
                 
                   L. pantheris 
                 
                 
                   L. rhamnosus 
                 
                 
                   L. vaccinostercus 
                 
               
               
                 
                   L. antri 
                 
                 
                   delbrueckii 
                 
                 
                   L. parabrevis 
                 
                 
                   L. rimae 
                 
                 
                   L. vaginalis 
                 
               
               
                 
                   L. apodemi 
                 
                   L. delbrueckii  subsp.  lactis   
                 
                   L. parabuchneri 
                 
                 
                   L. rogosae 
                 
                 
                   L. versmoldensis 
                 
               
               
                 
                   L. aviarius 
                 
                 
                   L. dextrinicus 
                 
                 
                   L. paracasei 
                 
                 
                   L. rossiae 
                 
                 
                   L. vini 
                 
               
               
                 
                   L. bifermentans 
                 
                 
                   L. diolivorans 
                 
                 
                   L. paracollinoides 
                 
                 
                   L. ruminis 
                 
                 
                   L. vitulinus 
                 
               
               
                 
                   L. brevis 
                 
                 
                   L. equi 
                 
                 
                   L. parafarraginis 
                 
                 
                   L. saerimneri 
                 
                 
                   L. zeae 
                 
               
               
                 
                   L. buchneri 
                 
                 
                   L. equigenerosi 
                 
                 
                   L. homohiochii 
                 
                 
                   L. jensenii 
                 
                 
                   L. zymae 
                 
               
               
                 
                   L. camelliae 
                 
                 
                   L. farraginis 
                 
                 
                   L. iners 
                 
                 
                   L. johnsonii 
                 
                 
                   L. gastricus 
                 
               
               
                 
                   L. casei 
                 
                 
                   L. farciminis 
                 
                 
                   L. ingluviei 
                 
                 
                   L. kalixensis 
                 
                 
                   L. ghanensis 
                 
               
               
                 
                   L. kitasatonis 
                 
                 
                   L. fermentum 
                 
                 
                   L. intestinalis 
                 
                 
                   L. kefiranofaciens 
                 
                 
                   L. graminis 
                 
               
               
                 
                   L. kunkeei 
                 
                 
                   L. fornicalis 
                 
                 
                   L. fuchuensis 
                 
                 
                   L. kefiri 
                 
                 
                   L. hammesii 
                 
               
               
                 
                   L. leichmannii 
                 
                 
                   L. fructivorans 
                 
                 
                   L. gallinarum 
                 
                 
                   L. kimchii 
                 
                 
                   L. hamsteri 
                 
               
               
                 
                   L. lindneri 
                 
                 
                   L. frumenti 
                 
                 
                   L. gasseri 
                 
                 
                   L. helveticus 
                 
                 
                   L. harbinensis 
                 
               
               
                 
                   L. malefermentans 
                 
                   
                   
                 
                   L. hilgardii 
                 
                 
                   L. hayakitensis 
                 
               
               
                 
                   Legionella 
                 
               
               
                 
                   Legionella adelaidensis 
                 
                 
                   Legionella drancourtii 
                 
                 
                   Candidatus Legionella jeonii 
                 
                 
                   Legionella quinlivanii 
                 
               
               
                 
                   Legionella anisa 
                 
                 
                   Legionella dresdenensis 
                 
                 
                   Legionella jordanis 
                 
                 
                   Legionella rowbothamii 
                 
               
               
                 
                   Legionella beliardensis 
                 
                 
                   Legionella drozanskii 
                 
                 
                   Legionella lansingensis 
                 
                 
                   Legionella rubrilucens 
                 
               
               
                 
                   Legionella birminghamensis 
                 
                 
                   Legionella dumoffii 
                 
                 
                   Legionella londiniensis 
                 
                 
                   Legionella sainthelensi 
                 
               
               
                 
                   Legionella bozemanae 
                 
                 
                   Legionella erythra 
                 
                 
                   Legionella longbeachae 
                 
                 
                   Legionella santicrucis 
                 
               
               
                 
                   Legionella brunensis 
                 
                 
                   Legionella fairfieldensis 
                 
                 
                   Legionella lytica 
                 
                 
                   Legionella shakespearei 
                 
               
               
                 
                   Legionella busanensis 
                 
                 
                   Legionella fallonii 
                 
                 
                   Legionella maceachernii 
                 
                 
                   Legionella spiritensis 
                 
               
               
                 
                   Legionella cardiaca 
                 
                 
                   Legionella feeleii 
                 
                 
                   Legionella massiliensis 
                 
                 
                   Legionella steelei 
                 
               
               
                 
                   Legionella cherrii 
                 
                 
                   Legionella geestiana 
                 
                 
                   Legionella micdadei 
                 
                 
                   Legionella steigerwaltii 
                 
               
               
                 
                   Legionella cincinnatiensis 
                 
                 
                   Legionella genomospecies 
                 
                 
                   Legionella monrovica 
                 
                 
                   Legionella taurinensis 
                 
               
               
                 
                   Legionella clemsonensis 
                 
                 
                   Legionella gormanii 
                 
                 
                   Legionella moravica 
                 
                 
                   Legionella tucsonensis 
                 
               
               
                 
                   Legionella donaldsonii 
                 
                 
                   Legionella gratiana 
                 
                 
                   Legionella nagasakiensis 
                 
                 
                   Legionella tunisiensis 
                 
               
               
                   
                 
                   Legionella gresilensis 
                 
                 
                   Legionella nautarum 
                 
                 
                   Legionella wadsworthii 
                 
               
               
                   
                 
                   Legionella hackeliae 
                 
                 
                   Legionella norrlandica 
                 
                 
                   Legionella waltersii 
                 
               
               
                   
                 
                   Legionella impletisoli 
                 
                 
                   Legionella oakridgensis 
                 
                 
                   Legionella worsleiensis 
                 
               
               
                   
                 
                   Legionella israelensis 
                 
                 
                   Legionella parisiensis 
                 
                 
                   Legionella yabuuchiae 
                 
               
               
                   
                 
                   Legionella jamestowniensis 
                 
                 
                   Legionella pittsburghensis 
                 
               
               
                   
                   
                 
                   Legionella pneumophila 
                 
               
               
                   
                   
                 
                   Legionella quateirensis 
                 
               
               
                 
                   Oceanibulbus 
                 
                 
                   Paenibacillus 
                 
                 
                   Prevotella 
                 
                 
                   Quadrisphaera 
                 
               
               
                 
                   Oceanibulbus indolifex 
                 
                 
                   Paenibacillus thiaminolyticus 
                 
                 
                   Prevotella albensis 
                 
                 
                   Quadrisphaera 
                 
               
               
                 
                   Oceanicaulis 
                 
                 
                   Pantoea 
                 
                 
                   Prevotella amnii 
                 
                 
                   granulorum 
                 
               
               
                 
                   Oceanicaulis alexandrii 
                 
                 
                   Pantoea 
                 
                 
                   Prevotella bergensis 
                 
                 
                   Quatrionicoccus 
                 
               
               
                 
                   Oceanicola 
                 
                 
                   agglomerans 
                 
                 
                   Prevotella bivia 
                 
                 
                   Quatrionicoccus 
                 
               
               
                 
                   Oceanicola batsensis 
                 
                 
                   Paracoccus 
                 
                 
                   Prevotella brevis 
                 
                 
                   australiensis 
                 
               
               
                 
                   Oceanicola granulosus 
                 
                 
                   Paracoccus alcaliphilus 
                 
                 
                   Prevotella bryantii 
                 
                 
                   Quinella 
                 
               
               
                 
                   Oceanicola nanhaiensis 
                 
                 
                   Paucimonas 
                 
                 
                   Prevotella buccae 
                 
                 
                   Quinella 
                 
               
               
                 
                   Oceanimonas 
                 
                 
                   Paucimonas lemoignei 
                 
                 
                   Prevotella buccalis 
                 
                 
                   ovalis 
                 
               
               
                 
                   Oceanimonas baumannii 
                 
                 
                   Pectobacterium 
                 
                 
                   Prevotella copri 
                 
                 
                   Ralstonia 
                 
               
               
                 
                   Oceaniserpentilla 
                 
                 
                   Pectobacterium aroidearum 
                 
                 
                   Prevotella dentalis 
                 
                 
                   Ralstonia eutropha 
                 
               
               
                 
                   Oceaniserpentilla haliotis 
                 
                 
                   Pectobacterium atrosepticum 
                 
                 
                   Prevotella denticola 
                 
                 
                   Ralstonia insidiosa 
                 
               
               
                 
                   Oceanisphaera 
                 
                 
                   Pectobacterium betavasculorum 
                 
                 
                   Prevotella disiens 
                 
                 
                   Ralstonia mannitolilytica 
                 
               
               
                 
                   Oceanisphaera donghaensis 
                 
                 
                   Pectobacterium cacticida 
                 
                 
                   Prevotella histicola 
                 
                 
                   Ralstonia pickettii 
                 
               
               
                 
                   Oceanisphaera litoralis 
                 
                 
                   Pectobacterium carnegieana 
                 
                 
                   Prevotella intermedia 
                 
                 
                   Ralstonia 
                 
               
               
                 
                   Oceanithermus 
                 
                 
                   Pectobacterium carotovorum 
                 
                 
                   Prevotella maculosa 
                 
                 
                   pseudosolanacearum 
                 
               
               
                 
                   Oceanithermus desulfurans 
                 
                 
                   Pectobacterium chrysanthemi 
                 
                 
                   Prevotella marshii 
                 
                 
                   Ralstonia syzygii 
                 
               
               
                 
                   Oceanithermus profundus 
                 
                 
                   Pectobacterium cypripedii 
                 
                 
                   Prevotella melaninogenica 
                 
                 
                   Ralstonia solanacearum 
                 
               
               
                 
                   Oceanobacillus 
                 
                 
                   Pectobacterium rhapontici 
                 
                 
                   Prevotella micans 
                 
                 
                   Ramlibacter 
                 
               
               
                 
                   Oceanobacillus caeni 
                 
                 
                   Pectobacterium wasabiae 
                 
                 
                   Prevotella multiformis 
                 
                 
                   Ramlibacter henchirensis 
                 
               
               
                 
                   Oceanospirillum 
                 
                 
                   Planococcus 
                 
                 
                   Prevotella nigrescens 
                 
                 
                   Ramlibacter 
                 
               
               
                 
                   Oceanospirillum linum 
                 
                 
                   Planococcus citreus 
                 
                 
                   Prevotella oralis 
                 
                 
                   tataouinensis 
                 
               
               
                   
                 
                   Planomicrobium 
                 
                 
                   Prevotella oris 
                 
                 
                   Raoultella 
                 
               
               
                   
                 
                   Planomicrobium okeanokoites 
                 
                 
                   Prevotella oulorum 
                 
                 
                   Raoultella ornithinolytica 
                 
               
               
                   
                 
                   Plesiomonas 
                 
                 
                   Prevotella pallens 
                 
                 
                   Raoultella planticola 
                 
               
               
                   
                 
                   Plesiomonas shigelloides 
                 
                 
                   Prevotella salivae 
                 
                 
                   Raoultella terrigena 
                 
               
               
                   
                 
                   Proteus 
                 
                 
                   Prevotella stercorea 
                 
                 
                   Rathayibacter 
                 
               
               
                   
                 
                   Proteus vulgaris 
                 
                 
                   Prevotella tannerae 
                 
                 
                   Rathayibacter caricis 
                 
               
               
                   
                   
                 
                   Prevotella timonensis 
                 
                 
                   Rathayibacter festucae 
                 
               
               
                   
                   
                 
                   Prevotella veroralis 
                 
                 
                   Rathayibacter iranicus 
                 
               
               
                   
                   
                 
                   Providencia 
                 
                 
                   Rathayibacter rathayi 
                 
               
               
                   
                   
                 
                   Providencia stuartii 
                 
                 
                   Rathayibacter toxicus 
                 
               
               
                   
                   
                 
                   Pseudomonas 
                 
                 
                   Rathayibacter tritici 
                 
               
               
                   
                   
                 
                   Pseudomonas aeruginosa 
                 
                 
                   Rhodobacter 
                 
               
               
                   
                   
                 
                   Pseudomonas alcaligenes 
                 
                 
                   Rhodobacter sphaeroides 
                 
               
               
                   
                   
                 
                   Pseudomonas anguillispetica 
                 
                 
                   Ruegeria 
                 
               
               
                   
                   
                 
                   Pseudomonas fluorescens 
                 
                 
                   Ruegeria gelatinovorans 
                 
               
               
                   
                   
                 
                   Pseudoalteromonas 
                 
               
               
                   
                   
                 
                   haloplanktis 
                 
               
               
                   
                   
                 
                   Pseudomonas mendocina 
                 
               
               
                   
                   
                 
                   Pseudomonas 
                 
               
               
                   
                   
                 
                   pseudoalcaligenes 
                 
               
               
                   
                   
                 
                   Pseudomonas putida 
                 
               
               
                   
                   
                 
                   Pseudomonas tutzeri 
                 
               
               
                   
                   
                 
                   Pseudomonas syringae 
                 
               
               
                   
                   
                 
                   Psychrobacter 
                 
               
               
                   
                   
                 
                   Psychrobacter faecalis 
                 
               
               
                   
                   
                 
                   Psychrobacter 
                 
               
               
                   
                   
                 
                   phenylpyruvicus 
                 
               
               
                 
                   Saccharococcus 
                 
                 
                   Sagittula 
                 
                 
                   Sanguibacter 
                 
                 
                   Stenotrophomonas 
                 
                 
                   Tatlockia 
                 
               
               
                 
                   Saccharococcus thermophilus 
                 
                 
                   Sagittula stellata 
                 
                 
                   Sanguibacter keddieii 
                 
                 
                   Stenotrophomonas 
                 
                 
                   Tatlockia maceachernii 
                 
               
               
                 
                   Saccharomonospora 
                 
                 
                   Salegentibacter 
                 
                 
                   Sanguibacter suarezii 
                 
                 
                   maltophilia 
                 
                 
                   Tatlockia micdadei 
                 
               
               
                 
                   Saccharomonospora azurea 
                 
                 
                   Salegentibacter salegens 
                 
                 
                   Saprospira 
                 
                 
                   Streptococcus 
                 
                 
                   Tenacibaculum 
                 
               
               
                 
                   Saccharomonospora cyanea 
                 
                 
                   Salimicrobium 
                 
                 
                   Saprospira grandis 
                 
                 [also see below] 
                 
                   Tenacibaculum 
                 
               
               
                 
                   Saccharomonospora viridis 
                 
                 
                   Salimicrobium album 
                 
                 
                   Sarcina 
                 
                 
                   Streptomyces 
                 
                 
                   amylolyticum 
                 
               
               
                 
                   Saccharophagus 
                 
                 
                   Salinibacter 
                 
                 
                   Sarcina maxima 
                 
                 
                   Streptomyces 
                 
                 
                   Tenacibaculum discolor 
                 
               
               
                 
                   Saccharophagus degradans 
                 
                 
                   Salinibacter ruber 
                 
                 
                   Sarcina ventriculi 
                 
                 
                   achromogenes 
                 
                 
                   Tenacibaculum 
                 
               
               
                 
                   Saccharopolyspora 
                 
                 
                   Salinicoccus 
                 
                 
                   Sebaldella 
                 
                 
                   Streptomyces 
                 
                 
                   gallaicum 
                 
               
               
                 
                   Saccharopolyspora erythraea 
                 
                 
                   Salinicoccus alkaliphilus 
                 
                 
                   Sebaldella 
                 
                 
                   cesalbus 
                 
                 
                   Tenacibaculum 
                 
               
               
                 
                   Saccharopolyspora gregorii 
                 
                 
                   Salinicoccus hispanicus 
                 
                 
                   termitidis 
                 
                 
                   Streptomyces cescaepitosus 
                 
                 
                   lutimaris 
                 
               
               
                 
                   Saccharopolyspora hirsuta 
                 
                 
                   Salinicoccus roseus 
                 
                 
                   Serratia 
                 
                 
                   Streptomyces cesdiastaticus 
                 
                 
                   Tenacibaculum 
                 
               
               
                 
                   Saccharopolyspora hordei 
                 
                 
                   Salinispora 
                 
                 
                   Serratia fonticola 
                 
                 
                   Streptomyces cesexfoliatus 
                 
                 
                   mesophilum 
                 
               
               
                 
                   Saccharopolyspora rectivirgula 
                 
                 
                   Salinispora arenicola 
                 
                 
                   Serratia marcescens 
                 
                 
                   Streptomyces fimbriatus 
                 
                 
                   Tenacibaculum 
                 
               
               
                 
                   Saccharopolyspora spinosa 
                 
                 
                   Salinispora tropica 
                 
                 
                   Sphaerotilus 
                 
                 
                   Streptomyces fradiae 
                 
                 
                   skagerrakense 
                 
               
               
                 
                   Saccharopolyspora taberi 
                 
                 
                   Salinivibrio 
                 
                 
                   Sphaerotilus natans 
                 
                 
                   Streptomyces fulvissimus 
                 
                 
                   Tepidanaerobacter 
                 
               
               
                 
                   Saccharothrix 
                 
                 
                   Salinivibrio costicola 
                 
                 
                   Sphingobacterium 
                 
                 
                   Streptomyces griseoruber 
                 
                 
                   Tepidanaerobacter 
                 
               
               
                 
                   Saccharothrix australiensis 
                 
                 
                   Salmonella 
                 
                 
                   Sphingobacterium multivorum 
                 
                 
                   Streptomyces griseus 
                 
                 
                   syntrophicus 
                 
               
               
                 
                   Saccharothrix coeruleofusca 
                 
                 
                   Salmonella bongori 
                 
                 
                   Staphylococcus 
                 
                 
                   Streptomyces lavendulae 
                 
                 
                   Tepidibacter 
                 
               
               
                 
                   Saccharothrix espanaensis 
                 
                 
                   Salmonella enterica 
                 
                 [see below] 
                 
                   Streptomyces 
                 
                 
                   Tepidibacter 
                 
               
               
                 
                   Saccharothrix longispora 
                 
                 
                   Salmonella subterranea 
                 
                   
                 
                   phaeochromogenes 
                 
                 
                   formicigenes 
                 
               
               
                 
                   Saccharothrix mutabilis 
                 
                 
                   Salmonella typhi 
                 
                   
                 
                   Streptomyces 
                 
                 
                   Tepidibacter thalassicus 
                 
               
               
                 
                   Saccharothrix syringae 
                 
                   
                   
                 
                   thermodiastaticus 
                 
                 
                   Thermus 
                 
               
               
                 
                   Saccharothrix tangerinus 
                 
                   
                   
                 
                   Streptomyces tubercidicus 
                 
                 
                   Thermus aquaticus 
                 
               
               
                 
                   Saccharothrix texasensis 
                 
                   
                   
                   
                 
                   Thermus filiformis 
                 
               
               
                   
                   
                   
                   
                 
                   Thermus thermophilus 
                 
               
               
                 
                   Staphylococcus 
                 
               
               
                 
                   S. arlettae 
                 
                 
                   S. equorum 
                 
                 
                   S. microti 
                 
                 
                   S. schleiferi 
                 
               
               
                 
                   S. agnetis 
                 
                 
                   S. felis 
                 
                 
                   S. muscae 
                 
                 
                   S. sciuri 
                 
               
               
                 
                   S. aureus 
                 
                 
                   S. fleurettii 
                 
                 
                   S. nepalensis 
                 
                 
                   S. simiae 
                 
               
               
                 
                   S. auricularis 
                 
                 
                   S. gallinarum 
                 
                 
                   S. pasteuri 
                 
                 
                   S. simulans 
                 
               
               
                 
                   S. capitis 
                 
                 
                   S. haemolyticus 
                 
                 
                   S. petrasii 
                 
                 
                   S. stepanovicii 
                 
               
               
                 
                   S. caprae 
                 
                 
                   S. hominis 
                 
                 
                   S. pettenkoferi 
                 
                 
                   S. succinus 
                 
               
               
                 
                   S. carnosus 
                 
                 
                   S. hyicus 
                 
                 
                   S. piscifermentans 
                 
                 
                   S. vitulinus 
                 
               
               
                 
                   S. caseolyticus 
                 
                 
                   S. intermedius 
                 
                 
                   S. pseudintermedius 
                 
                 
                   S. warneri 
                 
               
               
                 
                   S. chromogenes 
                 
                 
                   S. kloosii 
                 
                 
                   S. pseudolugdunensis 
                 
                 
                   S. xylosus 
                 
               
               
                 
                   S. cohnii 
                 
                 
                   S. leei 
                 
                 
                   S. pulvereri 
                 
               
               
                 
                   S. condimenti 
                 
                 
                   S. lentus 
                 
                 
                   S. rostri 
                 
               
               
                 
                   S. delphini 
                 
                 
                   S. lugdunensis 
                 
                 
                   S. saccharolyticus 
                 
               
               
                 
                   S. devriesei 
                 
                 
                   S. lutrae 
                 
                 
                   S. saprophyticus 
                 
               
               
                 
                   S. epidermidis 
                 
                 
                   S. lyticans 
                 
               
               
                   
                 
                   S. massiliensis 
                 
               
               
                 
                   Streptococcus 
                 
               
               
                 
                   Streptococcus agalactiae 
                 
                 
                   Streptococcus infantarius 
                 
                 
                   Streptococcus orisratti 
                 
                 
                   Streptococcus thermophilus 
                 
               
               
                 
                   Streptococcus anginosus 
                 
                 
                   Streptococcus iniae 
                 
                 
                   Streptococcus parasanguinis 
                 
                 
                   Streptococcus sanguinis 
                 
               
               
                 
                   Streptococcus bovis 
                 
                 
                   Streptococcus intermedius 
                 
                 
                   Streptococcus peroris 
                 
                 
                   Streptococcus sobrinus 
                 
               
               
                 
                   Streptococcus canis 
                 
                 
                   Streptococcus lactarius 
                 
                 
                   Streptococcus pneumoniae 
                 
                 
                   Streptococcus suis 
                 
               
               
                 
                   Streptococcus constellatus 
                 
                 
                   Streptococcus milleri 
                 
                 
                   Streptococcus 
                 
                 
                   Streptococcus uberis 
                 
               
               
                 
                   Streptococcus downei 
                 
                 
                   Streptococcus mitis 
                 
                 
                   pseudopneumoniae 
                 
                 
                   Streptococcus vestibularis 
                 
               
               
                 
                   Streptococcus dysgalactiae 
                 
                 
                   Streptococcus mutans 
                 
                 
                   Streptococcus pyogenes 
                 
                 
                   Streptococcus viridans 
                 
               
               
                 
                   Streptococcus equines 
                 
                 
                   Streptococcus oralis 
                 
                 
                   Streptococcus ratti 
                 
                 
                   Streptococcus 
                 
               
               
                 
                   Streptococcus faecalis 
                 
                 
                   Streptococcus tigurinus 
                 
                 
                   Streptococcus salivariu 
                 
                 
                   zooepidemicus 
                 
               
               
                 
                   Streptococcus ferus 
                 
               
               
                 
                   Uliginosibacterium 
                 
                 
                   Vagococcus 
                 
                 
                   Vibrio 
                 
                 
                   Virgibacillus 
                 
                 
                   Xanthobacter 
                 
               
               
                 
                   Uliginosibacterium 
                 
                 
                   Vagococcus carniphilus 
                 
                 
                   Vibrio aerogenes 
                 
                 
                   Virgibacillus 
                 
                 
                   Xanthobacter agilis 
                 
               
               
                 
                   gangwonense 
                 
                 
                   Vagococcus elongatus 
                 
                 
                   Vibrio aestuarianus 
                 
                 
                   halodenitrificans 
                 
                 
                   Xanthobacter 
                 
               
               
                 
                   Ulvibacter 
                 
                 
                   Vagococcus fessus 
                 
                 
                   Vibrio albensis 
                 
                 
                   Virgibacillus 
                 
                 
                   aminoxidans 
                 
               
               
                 
                   Ulvibacter litoralis 
                 
                 
                   Vagococcus fluvialis 
                 
                 
                   Vibrio alginolyticus 
                 
                 
                   pantothenticus 
                 
                 
                   Xanthobacter 
                 
               
               
                 
                   Umezawaea 
                 
                 
                   Vagococcus lutrae 
                 
                 
                   Vibrio campbellii 
                 
                 
                   Weissella 
                 
                 
                   autotrophicus 
                 
               
               
                 
                   Umezawaea tangerina 
                 
                 
                   Vagococcus salmoninarum 
                 
                 
                   Vibrio cholerae 
                 
                 
                   Weissella cibaria 
                 
                 
                   Xanthobacter flavus 
                 
               
               
                 
                   Undibacterium 
                 
                 
                   Variovorax 
                 
                 
                   Vibrio cincinnatiensis 
                 
                 
                   Weissella confusa 
                 
                 
                   Xanthobacter tagetidis 
                 
               
               
                 
                   Undibacterium pigrum 
                 
                 
                   Variovorax boronicumulans 
                 
                 
                   Vibrio coralliilyticus 
                 
                 
                   Weissella halotolerans 
                 
                 
                   Xanthobacter viscosus 
                 
               
               
                 
                   Ureaplasma 
                 
                 
                   Variovorax dokdonensis 
                 
                 
                   Vibrio cyclitrophicus 
                 
                 
                   Weissella hellenica 
                 
                 
                   Xanthomonas 
                 
               
               
                 
                   Ureaplasma 
                 
                 
                   Variovorax paradoxus 
                 
                 
                   Vibrio diazotrophicus 
                 
                 
                   Weissella kandleri 
                 
                 
                   Xanthomonas 
                 
               
               
                 
                   urealyticum 
                 
                 
                   Variovorax soli 
                 
                 
                   Vibrio fluvialis 
                 
                 
                   Weissella koreensis 
                 
                 
                   albilineans 
                 
               
               
                 
                   Ureibacillus 
                 
                 
                   Veillonella 
                 
                 
                   Vibrio furnissii 
                 
                 
                   Weissella minor 
                 
                 
                   Xanthomonas alfalfae 
                 
               
               
                 
                   Ureibacillus composti 
                 
                 
                   Veillonella atypica 
                 
                 
                   Vibrio gazogenes 
                 
                 
                   Weissella 
                 
                 
                   Xanthomonas 
                 
               
               
                 
                   Ureibacillus suwonensis 
                 
                 
                   Veillonella caviae 
                 
                 
                   Vibrio halioticoli 
                 
                 
                   paramesenteroides 
                 
                 
                   arboricola 
                 
               
               
                 
                   Ureibacillus terrenus 
                 
                 
                   Veillonella criceti 
                 
                 
                   Vibrio harveyi 
                 
                 
                   Weissella soli 
                 
                 
                   Xanthomonas 
                 
               
               
                 
                   Ureibacillus thermophilus 
                 
                 
                   Veillonella dispar 
                 
                 
                   Vibrio ichthyoenteri 
                 
                 
                   Weissella thailandensis 
                 
                 
                   axonopodis 
                 
               
               
                 
                   Ureibacillus thermosphaericus 
                 
                 
                   Veillonella montpellierensis 
                 
                 
                   Vibrio mediterranei 
                 
                 
                   Weissella viridescens 
                 
                 
                   Xanthomonas 
                 
               
               
                   
                 
                   Veillonella parvula 
                 
                 
                   Vibrio metschnikovii 
                 
                 
                   Williamsia 
                 
                 
                   campestris 
                 
               
               
                   
                 
                   Veillonella ratti 
                 
                 
                   Vibrio mytili 
                 
                 
                   Williamsia marianensis 
                 
                 
                   Xanthomonas citri 
                 
               
               
                   
                 
                   Veillonella rodentium 
                 
                 
                   Vibrio natriegens 
                 
                 
                   Williamsia maris 
                 
                 
                   Xanthomonas codiaei 
                 
               
               
                   
                 
                   Venenivibrio 
                 
                 
                   Vibrio navarrensis 
                 
                 
                   Williamsia serinedens 
                 
                 
                   Xanthomonas 
                 
               
               
                   
                 
                   Venenivibrio stagnispumantis 
                 
                 
                   Vibrio nereis 
                 
                 
                   Winogradskyella 
                 
                 
                   cucurbitae 
                 
               
               
                   
                 
                   Verminephrobacter 
                 
                 
                   Vibrio nigripulchritudo 
                 
                 
                   Winogradskyella 
                 
                 
                   Xanthomonas 
                 
               
               
                   
                 
                   Verminephrobacter eiseniae 
                 
                 
                   Vibrio ordalii 
                 
                 
                   thalassocola 
                 
                 
                   euvesicatoria 
                 
               
               
                   
                 
                   Verrucomicrobium 
                 
                 
                   Vibrio orientalis 
                 
                 
                   Wolbachia 
                 
                 
                   Xanthomonas fragariae 
                 
               
               
                   
                 
                   Verrucomicrobium spinosum 
                 
                 
                   Vibrio parahaemolyticus 
                 
                 
                   Wolbachia persica 
                 
                 
                   Xanthomonas fuscans 
                 
               
               
                   
                   
                 
                   Vibrio pectenicida 
                 
                 
                   Wolinella 
                 
                 
                   Xanthomonas gardneri 
                 
               
               
                   
                   
                 
                   Vibrio penaeicida 
                 
                 
                   Wolinella succinogenes 
                 
                 
                   Xanthomonas hortorum 
                 
               
               
                   
                   
                 
                   Vibrio proteolyticus 
                 
                 
                   Zobellia 
                 
                 
                   Xanthomonas hyacinthi 
                 
               
               
                   
                   
                 
                   Vibrio shilonii 
                 
                 
                   Zobellia galactanivorans 
                 
                 
                   Xanthomonas perforans 
                 
               
               
                   
                   
                 
                   Vibrio splendidus 
                 
                 
                   Zobellia uliginosa 
                 
                 
                   Xanthomonas phaseoli 
                 
               
               
                   
                   
                 
                   Vibrio tubiashii 
                 
                 
                   Zoogloea 
                 
                 
                   Xanthomonas pisi 
                 
               
               
                   
                   
                 
                   Vibrio vulnificus 
                 
                 
                   Zoogloea ramigera 
                 
                 
                   Xanthomonas populi 
                 
               
               
                   
                   
                   
                 
                   Zoogloea resiniphila 
                 
                 
                   Xanthomonas theicola 
                 
               
               
                   
                   
                   
                   
                 
                   Xanthomonas 
                 
               
               
                   
                   
                   
                   
                 
                   translucens 
                 
               
               
                   
                   
                   
                   
                 
                   Xanthomonas 
                 
               
               
                   
                   
                   
                   
                 
                   vesicatoria 
                 
               
               
                   
                   
                   
                   
                 
                   Xylella 
                 
               
               
                   
                   
                   
                   
                 
                   Xylella fastidiosa 
                 
               
               
                   
                   
                   
                   
                 
                   Xylophilus 
                 
               
               
                   
                   
                   
                   
                 
                   Xylophilus ampelinus 
                 
               
               
                 
                   Xenophilus 
                 
                 
                   Yangia 
                 
                 
                   Yersinia mollaretii 
                 
                 
                   Zooshikella 
                 
                 
                   Zobellella 
                 
               
               
                 
                   Xenophilus azovorans 
                 
                 
                   Yangia pacifica 
                 
                 
                   Yersinia philomiragia 
                 
                 
                   Zooshikella ganghwensis 
                 
                 
                   Zobellella denitrificans 
                 
               
               
                 
                   Xenorhabdus 
                 
                 
                   Yaniella 
                 
                 
                   Yersinia pestis 
                 
                 
                   Zunongwangia 
                 
                 
                   Zobellella taiwanensis 
                 
               
               
                 
                   Xenorhabdus beddingii 
                 
                 
                   Yaniella flava 
                 
                 
                   Yersinia pseudotuberculosis 
                 
                 
                   Zunongwangia profunda 
                 
                 
                   Zeaxanthinibacter 
                 
               
               
                 
                   Xenorhabdus bovienii 
                 
                 
                   Yaniella halotolerans 
                 
                 
                   Yersinia rohdei 
                 
                 
                   Zymobacter 
                 
                 
                   Zeaxanthinibacter 
                 
               
               
                 
                   Xenorhabdus cabanillasii 
                 
                 
                   Yeosuana 
                 
                 
                   Yersinia ruckeri 
                 
                 
                   Zymobacter palmae 
                 
                 
                   enoshimensis 
                 
               
               
                 
                   Xenorhabdus doucetiae 
                 
                 
                   Yeosuana aromativorans 
                 
                 
                   Yokenella 
                 
                 
                   Zymomonas 
                 
                 
                   Zhihengliuella 
                 
               
               
                 
                   Xenorhabdus griffiniae 
                 
                 
                   Yersinia 
                 
                 
                   Yokenella regensburgei 
                 
                 
                   Zymomonas mobilis 
                 
                 
                   Zhihengliuella 
                 
               
               
                 
                   Xenorhabdus hominickii 
                 
                 
                   Yersinia aldovae 
                 
                 
                   Yonghaparkia 
                 
                 
                   Zymophilus 
                 
                 
                   halotolerans 
                 
               
               
                 
                   Xenorhabdus koppenhoeferi 
                 
                 
                   Yersinia bercovieri 
                 
                 
                   Yonghaparkia alkaliphila 
                 
                 
                   Zymophilus paucivorans 
                 
                 
                   Xylanibacterium 
                 
               
               
                 
                   Xenorhabdus nematophila 
                 
                 
                   Yersinia enterocolitica 
                 
                 
                   Zavarzinia 
                 
                 
                   Zymophilus raffinosivorans 
                 
                 
                   Xylanibacterium ulmi 
                 
               
               
                 
                   Xenorhabdus poinarii 
                 
                 
                   Yersinia entomophaga 
                 
                 
                   Zavarzinia compransoris 
                 
               
               
                 
                   Xylanibacter 
                 
                 
                   Yersinia frederiksenii 
                 
               
               
                 
                   Xylanibacter oryzae 
                 
                 
                   Yersinia intermedia 
                 
               
               
                   
                 
                   Yersinia kristensenii 
                 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 6 
               
               
                   
               
               
                 Sequences: 
               
               
                 SEQ 
               
               
                 ID 
               
               
                 NO: 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
            
               
                  1 
                 Salcr7-1 
               
               
                   
                 Sequence written 5′ to 3′. 
               
               
                   
                 Underlined: direct repeat 
               
               
                   
                 Between direct repeats: spacer region corresponding (in 5′ to 3′ direction) to selected target sequences  
               
               
                   
                 TGAAACACGCATTGATTTGAGTCAGCTAGGAGGTGACTGAAGTATATTTTAGATGAAGATTATTTCTTAATAACTAAAAATATGGTATAATACTCTTAATAAATGCAGTAATACAGGGGCTTTTCAAGACTGAAGT 
               
               
                   
                 CTAGCTGAGACAAATAGTGCGATTACGAAATTTTTTAGACAAAAATAGTCTACGAG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC CGCTTTGGGTATACGCATTTTGAAGTACGG GTTTTAGAGCTATG   
               
               
                   
                   CTGTTTTGAATGGTCCCAAAAC TGTCAACGGGTGTACTATATGTCTGTCATG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC GCAAACCAGTACCGAAGAAGAGGCGCTCAC GTTTTAGAGCTATGCTGT   
               
               
                   
                   TTTGAATGGTCCCAAAAC TGCCGTTCTGGTCATCCTGCTCGAAGCCGC GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC CCAGAAATGAATCGCCTGGCTTCATTATCG GTTTTAGAGCTATGCTGTTTTG   
               
               
                   
                   AATGGTCCCAAAAC ATCAGCAGGAAGCGCTCAAAAACATACTGC GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC AAGTGCGCCTGGTAGTCTTCCGGATAGCGG GTTTTAGAGCTATGCTGTTTTGAATG   
               
               
                   
                   GACTCCATTC TTCAGCACACTGAGACTTGTTGAGTTCCATGTTTTAGAGCTATGCTGTTTTGAATGGACTCCATTCAACATTGCCGATGATAACTTGAGAAAGAGGGTTAATACCAGCAGTCGGATACCTTCCTAT 
               
               
                   
                 TCTTTCTGTTAAAGCGTTTTCATGTTATAATAGGCAAAAGAAGAGTAGTGTGAT 
               
               
                  2 
                 Salcr7-2 
               
               
                   
                 Sequence written 5′ to 3′. 
               
               
                   
                 Underlined: direct repeat 
               
               
                   
                 Between direct repeats: spacer region corresponding (in 5′ to 3′ direction) to selected target sequences of pipA, pipB, pipC, hilA, marT,  
               
               
                   
                 sicP and sopB. 
               
               
                   
                 TGAAACACGCATTGATTTGAGTCAGCTAGGAGGTGACTGAAGTATATTTTAGATGAAGATTATTTCTTAATAACTAAAAATATGGTATAATACTCTTAATAAATGCAGTAATACAGGGGCTTTTCAAGACTGAAGT 
               
               
                   
                 CTAGCTGAGACAAATAGTGCGATTACGAAATTTTTTAGACAAAAATAGTCTACGAG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TCTTTTTCATATGCGTAATTCATCAGTCTG GTTTTAGAGCTATG   
               
               
                   
                   CTGTTTTGAATGGTCCCAAAAC TATAACCGAGGATGGTTTTCTGAACCTGCG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TATTAAATCGTTTATATGACGCGTTAGGCC GTTTTAGAGCTATGCTGT   
               
               
                   
                   TTTGAATGGTCCCAAAAC CACAATAATCCACAAGCTTTAGGATTACTG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TCTATTTCCCTTCCAGCAGTCGCACCCCAG GTTTTAGAGCTATGCTGTTTTG   
               
               
                   
                   AATGGTCCCAAAAC TTATCGCCTGTTTGTGGCGATTCTATCTGG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TACTCACCGCGTCGAATATTTTCGGCAAAG GTTTTAGAGCTATGCTGTTTTGAATG   
               
               
                   
                   GACTCCATTC TTCAGCACACTGAGACTTGTTGAGTTCCATGTTTTAGAGCTATGCTGTTTTGAATGGACTCCATTCAACATTGCCGATGATAACTTGAGAAAGAGGGTTAATACCAGCAGTCGGATACCTTCCTAT 
               
               
                   
                 TCTTTCTGTTAAAGCGTTTTCATGTTATAATAGGCAAAAGAAGAGTAGTGTGAT 
               
               
                   
               
               
                  3 
                 Salcr7-3 
               
               
                   
                 Sequence written 5′ to 3′. 
               
               
                   
                 Underlined: direct repeat 
               
               
                   
                 Between direct repeats: spacer region corresponding (in 5′ to 3′ direction) to selected target sequences of pipA, pipB, pipC, hilA, marT,   
               
               
                   
                 sicP and sopB. 
               
               
                   
                 TGAAACACGCATTGATTTGAGTCAGCTAGGAGGTGACTGAAGTATATTTTAGATGAAGATTATTTCTTAATAACTAAAAATATGGTATAATACTCTTAATAAATGCAGTAATACAGGGGCTTTTCAAGACTGAAGT 
               
               
                   
                 CTAGCTGAGACAAATAGTGCGATTACGAAATTTTTTAGACAAAAATAGTCTACGAG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC CTATTTATTGAAGATGTAGACCATTCTGGG GTTTTAGAGCTATG   
               
               
                   
                   CTGTTTTGAATGGTCCCAAAAC AGAAGCAATGAAAAGTGCAACTTCACCACG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC ATCGCTTGCCGCAAACCAGTACCGAAGAAG GTTTTAGAGCTATGCTGT   
               
               
                   
                   TTTGAATGGTCCCAAAAC TTCGATTGACAGTACTATGGTTTACTTACG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC ATTGATTTGTCAGCAACCTTATAAAACGCG GTTTTAGAGCTATGCTGTTTTG   
               
               
                   
                   AATGGTCCCAAAAC GAAGCAATCTTATAAAAGATATAATTCAAG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TAACCTGGAGATTCAGAAACAAAATACGGG GTTTTAGAGCTATGCTGTTTTGAATG   
               
               
                   
                   GACTCCATTC TTCAGCACACTGAGACTTGTTGAGTTCCATGTTTTAGAGCTATGCTGTTTTGAATGGACTCCATTCAACATTGCCGATGATAACTTGAGAAAGAGGGTTAATACCAGCAGTCGGATACCTTCCTAT 
               
               
                   
                 TCTTTCTGTTAAAGCGTTTTCATGTTATAATAGGCAAAAGAAGAGTAGTGTGAT 
               
               
                   
               
               
                  4 
                 tracrRNA-Cas9 fragment sequence based on pCas9 
               
               
                   
                 Bold: tracrRNA 
               
               
                   
                 Underlined: Cas9 coding sequence 
               
               
                   
                 CGAAATCATCCTGTGGAGCTTAGTAGGTTTAGCAAGATGGCAGCGCCTAAATGTAGAATGATAAAAGGATTAAGAGATTAATTTCCCTAAAAATGATAAAACAAGCGTTTTGAAAGCGCTTTTTTTGGTTTGCAGT 
               
               
                   
                 
                   
                     
                     
                         
                         
                     
                   
                 
               
               
                   
                 
                   
                     
                     
                         
                         
                     
                   
                 
               
               
                   
                 TGAAGAGATATTTTGAAAAAGAAAAATTAAAGCATATTAAACTAATTTCGGAGGTCATTAAAACTATTATTGAAATCATCAAACTCATTATGGATTTAATTTAAACTTTTTATTTTAGGAGGCAAAA ATGGATAAG   
               
               
                   
                 
                   AAATACTCAATAGGCTTAGATATCGGCACAAATAGCGTCGGATGGGCGGTGATCACTGATGAATATAAGGTTCCGTCTAAAAAGTTCAAGGTTCTGGGAAATACAGACCGCCACAGTATCAAAAAAAATCTTATAG 
                 
               
               
                   
                 
                   GGGCTCTTTTATTTGACAGTGGAGAGACAGCGGAAGCGACTCGTCTCAAACGGACAGCTCGTAGAAGGTATACACGTCGGAAGAATCGTATTTGTTATCTACAGGAGATTTTTTCAAATGAGATGGCGAAAGTAGA 
                 
               
               
                   
                 
                   TGATAGTTTCTTTCATCGACTTGAAGAGTCTTTTTTGGTGGAAGAAGACAAGAAGCATGAACGTCATCCTATTTTTGGAAATATAGTAGATGAAGTTGCTTATCATGAGAAATATCCAACTATCTATCATCTGCGA 
                 
               
               
                   
                 
                   AAAAAATTGGTAGATTCTACTGATAAAGCGGATTTGCGCTTAATCTATTTGGCCTTAGCGCATATGATTAAGTTTCGTGGTCATTTTTTGATTGAGGGAGATTTAAATCCTGATAATAGTGATGTGGACAAACTAT 
                 
               
               
                   
                 
                   TTATCCAGTTGGTACAAACCTACAATCAATTATTTGAAGAAAACCCTATTAACGCAAGTGGAGTAGATGCTAAAGCGATTCTTTCTGCACGATTGAGTAAATCAAGACGATTAGAAAATCTCATTGCTCAGCTCCC 
                 
               
               
                   
                 
                   CGGTGAGAAGAAAAATGGCTTATTTGGGAATCTCATTGCTTTGTCATTGGGTTTGACCCCTAATTTTAAATCAAATTTTGATTTGGCAGAAGATGCTAAATTACAGCTTTCAAAAGATACTTACGATGATGATTTA 
                 
               
               
                   
                 
                   GATAATTTATTGGCGCAAATTGGAGATCAATATGCTGATTTGTTTTTGGCAGCTAAGAATTTATCAGATGCTATTTTACTTTCAGATATCCTAAGAGTAAATACTGAAATAACTAAGGCTCCCCTATCAGCTTCAA 
                 
               
               
                   
                 
                   TGATTAAACGCTACGATGAACATCATCAAGACTTGACTCTTTTAAAAGCTTTAGTTCGACAACAACTTCCAGAAAAGTATAAAGAAATCTTTTTTGATCAATCAAAAAACGGATATGCAGGTTATATTGATGGGGG 
                 
               
               
                   
                 
                   AGCTAGCCAAGAAGAATTTTATAAATTTATCAAACCAATTTTAGAAAAAATGGATGGTACTGAGGAATTATTGGTGAAACTAAATCGTGAAGATTTGCTGCGCAAGCAACGGACCTTTGACAACGGCTCTATTCCC 
                 
               
               
                   
                 
                   CATCAAATTCACTTGGGTGAGCTGCATGCTATTTTGAGAAGACAAGAAGACTTTTATCCATTTTTAAAAGACAATCGTGAGAAGATTGAAAAAATCTTGACTTTTCGAATTCCTTATTATGTTGGTCCATTGGCGC 
                 
               
               
                   
                 
                   GTGGCAATAGTCGTTTTGCATGGATGACTCGGAAGTCTGAAGAAACAATTACCCCATGGAATTTTGAAGAAGTTGTCGATAAAGGTGCTTCAGCTCAATCATTTATTGAACGCATGACAAACTTTGATAAAAATCT 
                 
               
               
                   
                 
                   TCCAAATGAAAAAGTACTACCAAAACATAGTTTGCTTTATGAGTATTTTACGGTTTATAACGAATTGACAAAGGTCAAATATGTTACTGAAGGAATGCGAAAACCAGCATTTCTTTCAGGTGAACAGAAGAAAGCC 
                 
               
               
                   
                 
                   ATTGTTGATTTACTCTTCAAAACAAATCGAAAAGTAACCGTTAAGCAATTAAAAGAAGATTATTTCAAAAAAATAGAATGTTTTGATAGTGTTGAAATTTCAGGAGTTGAAGATAGATTTAATGCTTCATTAGGTA 
                 
               
               
                   
                 
                   CCTACCATGATTTGCTAAAAATTATTAAAGATAAAGATTTTTTGGATAATGAAGAAAATGAAGATATCTTAGAGGATATTGTTTTAACATTGACCTTATTTGAAGATAGGGAGATGATTGAGGAAAGACTTAAAAC 
                 
               
               
                   
                 
                   ATATGCTCACCTCTTTGATGATAAGGTGATGAAACAGCTTAAACGTCGCCGTTATACTGGTTGGGGACGTTTGTCTCGAAAATTGATTAATGGTATTAGGGATAAGCAATCTGGCAAAACAATATTAGATTTTTTG 
                 
               
               
                   
                 
                   AAATCAGATGGTTTTGCCAATCGCAATTTTATGCAGCTGATCCATGATGATAGTTTGACATTTAAAGAAGACATTCAAAAAGCACAAGTGTCTGGACAAGGCGATAGTTTACATGAACATATTGCAAATTTAGCTG 
                 
               
               
                   
                 
                   GTAGCCCTGCTATTAAAAAAGGTATTTTACAGACTGTAAAAGTTGTTGATGAATTGGTCAAAGTAATGGGGCGGCATAAGCCAGAAAATATCGTTATTGAAATGGCACGTGAAAATCAGACAACTCAAAAGGGCCA 
                 
               
               
                   
                 
                   GAAAAATTCGCGAGAGCGTATGAAACGAATCGAAGAAGGTATCAAAGAATTAGGAAGTCAGATTCTTAAAGAGCATCCTGTTGAAAATACTCAATTGCAAAATGAAAAGCTCTATCTCTATTATCTCCAAAATGGA 
                 
               
               
                   
                 
                   AGAGACATGTATGTGGACCAAGAATTAGATATTAATCGTTTAAGTGATTATGATGTCGATCACATTGTTCCACAAAGTTTCCTTAAAGACGATTCAATAGACAATAAGGTCTTAACGCGTTCTGATAAAAATCGTG 
                 
               
               
                   
                 
                   GTAAATCGGATAACGTTCCAAGTGAAGAAGTAGTCAAAAAGATGAAAAACTATTGGAGACAACTTCTAAACGCCAAGTTAATCACTCAACGTAAGTTTGATAATTTAACGAAAGCTGAACGTGGAGGTTTGAGTGA 
                 
               
               
                   
                 
                   ACTTGATAAAGCTGGTTTTATCAAACGCCAATTGGTTGAAACTCGCCAAATCACTAAGCATGTGGCACAAATTTTGGATAGTCGCATGAATACTAAATACGATGAAAATGATAAACTTATTCGAGAGGTTAAAGTG 
                 
               
               
                   
                 
                   ATTACCTTAAAATCTAAATTAGTTTCTGACTTCCGAAAAGATTTCCAATTCTATAAAGTACGTGAGATTAACAATTACCATCATGCCCATGATGCGTATCTAAATGCCGTCGTTGGAACTGCTTTGATTAAGAAAT 
                 
               
               
                   
                 
                   ATCCAAAACTTGAATCGGAGTTTGTCTATGGTGATTATAAAGTTTATGATGTTCGTAAAATGATTGCTAAGTCTGAGCAAGAAATAGGCAAAGCAACCGCAAAATATTTCTTTTACTCTAATATCATGAACTTCTT 
                 
               
               
                   
                 
                   CAAAACAGAAATTACACTTGCAAATGGAGAGATTCGCAAACGCCCTCTAATCGAAACTAATGGGGAAACTGGAGAAATTGTCTGGGATAAAGGGCGAGATTTTGCCACAGTGCGCAAAGTATTGTCCATGCCCCAA 
                 
               
               
                   
                 
                   GTCAATATTGTCAAGAAAACAGAAGTACAGACAGGCGGATTCTCCAAGGAGTCAATTTTACCAAAAAGAAATTCGGACAAGCTTATTGCTCGTAAAAAAGACTGGGATCCAAAAAAATATGGTGGTTTTGATAGTC 
                 
               
               
                   
                 
                   CAACGGTAGCTTATTCAGTCCTAGTGGTTGCTAAGGTGGAAAAAGGGAAATCGAAGAAGTTAAAATCCGTTAAAGAGTTACTAGGGATCACAATTATGGAAAGAAGTTCCTTTGAAAAAAATCCGATTGACTTTTT 
                 
               
               
                   
                 
                   AGAAGCTAAAGGATATAAGGAAGTTAAAAAAGACTTAATCATTAAACTACCTAAATATAGTCTTTTTGAGTTAGAAAACGGTCGTAAACGGATGCTGGCTAGTGCCGGAGAATTACAAAAAGGAAATGAGCTGGCT 
                 
               
               
                   
                 
                   CTGCCAAGCAAATATGTGAATTTTTTATATTTAGCTAGTCATTATGAAAAGTTGAAGGGTAGTCCAGAAGATAACGAACAAAAACAATTGTTTGTGGAGCAGCATAAGCATTATTTAGATGAGATTATTGAGCAAA 
                 
               
               
                   
                 
                   TCAGTGAATTTTCTAAGCGTGTTATTTTAGCAGATGCCAATTTAGATAAAGTTCTTAGTGCATATAACAAACATAGAGACAAACCAATACGTGAACAAGCAGAAAATATTATTCATTTATTTACGTTGACGAATCT 
                 
               
               
                   
                 
                   TGGAGCTCCCGCTGCTTTTAAATATTTTGATACAACAATTGATCGTAAACGATATACGTCTACAAAAGAAGTTTTAGATGCCACTCTTATCCATCAATCCATCACTGGTCTTTATGAAACACGCATTGATTTGAGT 
                 
               
               
                   
                 
                   CAGCTAGGAGGTGACTGA 
                 
               
               
                   
               
               
                  5 
                 Salcr3.2 (crRNA-pipC-hilA-marT) 
               
               
                   
                 Sequence written 5′ to 3′. 
               
               
                   
                 Underlined: direct repeat 
               
               
                   
                 Between direct repeats: spacer region corresponding (in 5′ to 3′ direction) to selected target sequences of pipC-hilA-marT  
               
               
                   
                 TGTCGACGGTATATTTTAGATGAAGATTATTTCTTAATAACTAAAAATATGGTATAATACTCTTAATAAATGCAGTAATACAGGGGCTTTTCAAGACTGAAGTCTAGCTGAGACAAATAGTGCGATTACGAAATTT 
               
               
                   
                 TTTAGACAAAAATAGTCTACGAG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC ATCGCTTGCCGCAAACCAGTACCGAAGAAG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TTCGATTGACA 
               
               
                   
                 GTACTATGGTTTACTTACG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC ATTGATTTGTCAGCAACCTTATAAAACGCG GTTTTAGAGCTATGCTGTTTTGAATGGTCCCAAAAC TTCAGCACACTGAGA 
               
               
                   
                 CTTGTTGAGTTCCATGTTTTAGAGCTATGCTGTTTTGAATGGTCTCCATTCAACATTGCCGATGATAACTTGAGAAAGAGGGTTAATACCAGCAGTCGGATACCTTCCTATTCTTTCTGTTAAAGCGTTTTCATGT 
               
               
                   
                 TATAATAGGCAAAAGAAGAGTAGTGTGAT 
               
               
                   
               
               
                  6 
                 mob (oriT) 
               
               
                   
                 TTGAGCACCGCCAGGTGCGAATAAGGGACAGTGAAGAAGGAACACCCGCTCGCGGGTGGGCCTACTTCACCTATCCTGCCCGGCTGACGCCGTTGGATACACCAAGGAAAGTCTACACGAA 
               
               
                   
               
               
                  7 
                 pZA31MCS 
               
               
                   
                 ATCCCATGGTACGCGTGCTAGAGGCATCAAATAAAACGAAAGGCTCAGTCGAAAGACTGGGCCTTTCGTTTTATCTGTTGTTTGTCGGTGAACGCTCTCCTGAGTAGGACAAATCCGCCGCCCTAGACCTAGGGAT 
               
               
                   
                 ATATTCCGCTTCCTCGCTCACTGACTCGCTACGCTCGGTCGTTCGACTGCGGCGAGCGGAAATGGCTTACGAACGGGGCGGAGATTTCCTGGAAGATGCCAGGAAGATACTTAACAGGGAAGTGAGAGGGCCGCGG 
               
               
                   
                 CAAAGCCGTTTTTCCATAGGCTCCGCCCCCCTGACAAGCATCACGAAATCTGACGCTCAAATCAGTGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGCGGCTCCCTCGTGCGCTCTCC 
               
               
                   
                 TGTTCCTGCCTTTCGGTTTACCGGTGTCATTCCGCTGTTATGGCCGCGTTTGTCTCATTCCACGCCTGACACTCAGTTCCGGGTAGGCAGTTCGCTCCAAGCTGGACTGTATGCACGAACCCCCCGTTCAGTCCGA 
               
               
                   
                 CCGCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCCAACCCGGAAAGACATGCAAAAGCACCACTGGCAGCAGCCACTGGTAATTGATTTAGAGGAGTTAGTCTTGAAGTCATGCGCCGGTTAAGGCTAAACTGA 
               
               
                   
                 AAGGACAAGTTTTGGTGACTGCGCTCCTCCAAGCCAGTTACCTCGGTTCAAAGAGTTGGTAGCTCAGAGAACCTTCGAAAAACCGCCCTGCAAGGCGGTTTTTTCGTTTTCAGAGCAAGAGATTACGCGCAGACCA 
               
               
                   
                 AAACGATCTCAAGAAGATCATCTTATTAATCAGATAAAATATTACTAGATTTCAGTGCAATTTATCTCTTCAAATGTAGCACCTGAAGTCAGCCCCATACGATATAAGTTGTTACTAGTGCTTGGATTCTCACCAA 
               
               
                   
                 TAAAAAACGCCCGGCGGCAACCGAGCGTTCTGAACAAATCCAGATGGAGTTCTGAGGTCATTACTGGATCTATCAACAGGAGTCCAAGCGAGCTCGATATCAAATTACGCCCCGCCCTGCCACTCATCGCAGTACT 
               
               
                   
                 GTTGTAATTCATTAAGCATTCTGCCGACATGGAAGCCATCACAGACGGCATGATGAACCTGAATCGCCAGCGGCATCAGCACCTTGTCGCCTTGCGTATAATATTTGCCCATGGTGAAAACGGGGGCGAAGAAGTT 
               
               
                   
                 GTCCATATTGGCCACGTTTAAATCAAAACTGGTGAAACTCACCCAGGGATTGGCTGAGACGAAAAACATATTCTCAATAAACCCTTTAGGGAAATAGGCCAGGTTTTCACCGTAACACGCCACATCTTGCGAATAT 
               
               
                   
                 ATGTGTAGAAACTGCCGGAAATCGTCGTGGTATTCACTCCAGAGCGATGAAAACGTTTCAGTTTGCTCATGGAAAACGGTGTAACAAGGGTGAACACTATCCCATATCACCAGCTCACCGTCTTTCATTGCCATAC 
               
               
                   
                 GAAATTCCGGATGAGCATTCATCAGGCGGGCAAGAATGTGAATAAAGGCCGGATAAAACTTGTGCTTATTTTTCTTTACGGTCTTTAAAAAGGCCGTAATATCCAGCTGAACGGTCTGGTTATAGGTACATTGAGC 
               
               
                   
                 AACTGACTGAAATGCCTCAAAATGTTCTTTACGATGCCATTGGGATATATCAACGGTGGTATATCCAGTGATTTTTTTCTCCATTTTAGCTTCCTTAGCTCCTGAAAATCTCGATAACTCAAAAAATACGCCCGGT 
               
               
                   
                 AGTGATCTTATTTCATTATGGTGAAAGTTGGAACCTCTTACGTGCCGATCAACGTCTCATTTTCGCCAGATATCGACGTCTAAGAAACCATTATTATCATGACATTAACCTATAAAAATAGGCGTATCACGAGGCC 
               
               
                   
                 CTTTCGTCTTCAC 
               
               
                   
               
               
                  8 
                 spCas9-6 forward primer 
               
               
                   
                 ATTGTTTGTGGAGCAGCATAAGC 
               
               
                   
               
               
                  9 
                 mob2 reverse primer 
               
               
                   
                 GCCTCTAGCACGCGTACCATGGGAT 
               
               
                   
               
               
                 10 
                 trfA module for genomic insertion DH10B (written 5′ to 3&#39;) 
               
               
                   
                 Bracketed sequence: encodes trfA mRNA (complement strand) 
               
               
                   
                 Capital letters at 5′: bi-directional synthetic rho-independent terminator to prevent read-through from the trfA gene as well as upstream 
               
               
                   
                 gene on the chromosome 
               
               
                   
                 Lower case letters at 3′: J23105 promoter, a weak to intermediate strength promoter (capital T indicates transcription start site) 
               
               
                   
                 CTTAATAAAAACCCGCTTGGGCAAGCGGGTTTTATTGTGCAATTAGGCC[ CTA GCGTTTGCAATGCACCAGGTCATCATTGACCCAGGCGTGTTCCACCAGGCCGCTGCCTCGCAACTCTTCGCAGGCTTCGCCGA 
               
               
                   
                 CCTGCTCGCGCCACTTCTTCACGCGGGTGGAATCCGATCCGCACATGAGGCGGAAGGTTTCCAGCTTGAGCGGGTACGGCTCCCGGTGCGAGCTGAAATAGTCGAACATCCGTCGGGCCGTCGGCGACAGCTTGCG 
               
               
                   
                 GTACTTCTCCCATATGAATTTCGTGTAGTGGTCGCCAGCAAACAGCACGACGATTTCCTCGTCGATCAGGACCTGGCAACGGGACGTTTTCTTGCCACGGTCCAGGACGCGGAAGCGGTGCAGCAGCGACACCGAT 
               
               
                   
                 TCCAGGTGCCCAACGCGGTCGGACGTGAAGTCCATCGCCGTCGCCTGTAGGCGCGACAGGCATTCCTCGGCCTTCGTGTAATACCGGCCATTGATCGACCAGCCCAGGTCCTGGCAAAGCTCGTAGAACGTGAAGG 
               
               
                   
                 TGATCGGCTCGCCGATAGGGGTGCGCTTCGCGTACTCCAACACCTGCTGCCACACCAGTTCGTCATCGTCGGCCCGCAGCTCGACGCCGGTGTAGGTGATCTTCACGTCCTTGTTGACGTGGAAAATGACCTTGTT 
               
               
                   
                 TTGCAGCGCCTCGCGCGGGATTTTCTTGTTGCGCGTGGTGAACAGGGCAGAGCGGGCCGTGTCGTTTGGCATCGCTCGCATCGTGTCCGGCCACGGCGCAATATCGAACAAGGAAAGCTGCATTTCCTTGATCTGC 
               
               
                   
                 TGCTTCGTGTGTTTCAGCAACGCGGCCTGCTTGGCCTCGCTGACCTGTTTTGCCAGGTCCTCGCCGGCGGTTTTTCGCTTCTTGGTCGTCATAGTTCCTCGCGTGTCGATGGTCATCGACTTCGCCAAACCTGCCG 
               
               
                   
                 CCTCCTGTTCGAGACGACGCGAACGCTCCACGGCGGCCGATGGCGCGGGCAGGGCAGGGGGAGCCAGTTGCACGCTGTCGCGCTCGATCTTGGCCGTAGCTTGCTGGACCATCGAGCCGACGGACTGGAAGGTTTC 
               
               
                   
                 GCGGGGCGCACGCATGACGGTGCGGCTTGCGATGGTTTCGGCATCCTCGGCGGAAAACCCCGCGTCGATCAGTTCTTGCCTGTATGCCTTCCGGTCAAACGTCCGATT CAT TCACCCTCCTTGCGGGATTGCCCCG 
               
               
                   
                 ACTCACGCCGGGGCAATGTGCCCTT]gTagctagcatagtacctaggactgagctagccgtaaa 
               
               
                   
               
               
                 11 
                 aroA coding sequence 
               
               
                   
                 ATGGAATCCCTGACGTTACAACCCATCGCTCGTGTCGATGGCACTATTAATCTGCCCGGTTCCAAGAGCGTTTCTAACCGCGCTTTATTGCTGGCGGCATTAGCACACGGCAAAACAGTATTAACCAATCTGCTGG 
               
               
                   
                 ATAGCGATGACGTGCGCCATATGCTGAATGCATTAACAGCGTTAGGGGTAAGCTATACGCTTTCAGCCGATCGTACGCGTTGCGAAATTATCGGTAACGGCGGTCCATTACACGCAGAAGGTGCCCTGGAGTTGTT 
               
               
                   
                 CCTCGGTAACGCCGGAACGGCAATGCGTCCGCTGGCGGCAGCTCTTTGTCTGGGTAGCAATGATATTGTGCTGACCGGTGAGCCGCGTATGAAAGAACGCCCGATTGGTCATCTGGTGGATGCGCTGCGCCTGGGC 
               
               
                   
                 GGGGCGAAGATCACTTACCTGGAACAAGAAAATTATCCGCCGTTGCGTTTACAGGGCGGCTTTACTGGCGGCAACGTTGACGTTGATGGCTCCGTTTCCAGCCAATTCCTCACCGCACTGTTAATGACTGCGCCTC 
               
               
                   
                 TTGCGCCGGAAGATACGGTGATTCGTATTAAAGGCGATCTGGTTTCTAAACCTTATATCGACATCACACTCAATCTGATGAAGACGTTTGGTGTTGAAATTGAAAATCAGCACTATCAACAATTTGTCGTAAAAGG 
               
               
                   
                 CGGGCAGTCTTATCAGTCTCCGGGTACTTATTTGGTCGAAGGCGATGCATCTTCGGCTTCTTACTTTCTGGCAGCAGCAGCAATCAAAGGCGGCACTGTAAAAGTGACCGGTATTGGACGTAACAGTATGCAGGGT 
               
               
                   
                 GATATTCGCTTTGCTGATGTGCTGGAAAAAATGGGCGCGACCATTTGCTGGGGCGATGATTATATTTCCTGCACGCGTGGTGAACTGAACGCTATTGATATGGATATGAACCATATTCCTGATGCGGCGATGACCA 
               
               
                   
                 TTGCCACGGCGGCGTTATTTGCAAAAGGCACCACCACGCTGCGCAATATCTATAACTGGCGTGTTAAAGAGACCGATCGCCTGTTTGCGATGGCAACAGAACTGCGTAAAGTCGGCGCGGAAGTGGAAGAGGGGCA 
               
               
                   
                 CGATTACATTCGTATCACTCCTCCGGAAAAACTGAACTTTGCCGAGATCGCGACATACAATGATCACCGGATGGCGATGTGTTTCTCGCTGGTGGCGTTGTCAGATACACCAGTGACGATTCTTGATCCCAAATGC 
               
               
                   
                 ACGGCCAAAACATTTCCGGATTATTTCGAGCAGCTGGCGCGGATTAGCCAGGCAGCCTGA 
               
               
                   
               
               
                 12 
                 oriV RK2 
               
               
                   
                 Tgacacttgaggggcgtttagagcgagccaggaaagccgaccccctccttggagtaaaaacccttgcggcgttgcagccggcacggatcttccgatcgggcgcggtggtggccgcgtctgtgacctaaaaaggg 
               
               
                   
                 gggagtccagaggggcgcagcccctttgggcatagcgcagcgtaatcggagacgtaattgagcatttccaggcgcttgcgcctggtcaacgaaagagtcagcgccgtaggcgctgccatttttggggtgaggccg 
               
               
                   
                 ttcgcggccgaggggcgcagcccctggggggatgggaggcccgcgttagcgggccgggagggttcgagaagggggggcaccccccttcggcgtgcgcggtcacgcgccagggcgcagccctggttaaaaa 
               
               
                   
                 caaggtttataaatattggtttaaaagcaggttaaaagacaggttagcggtggccgaaaaacgggcggaaacccttgcaaatgctggattttctgcctgtggacagcccctcaaatgtcaataggtgcgcccctcatctg 
               
               
                   
                 tcatcactctgcccctcaagtgtcaaggatcgcgcccctcatctgtcagtagtcgcgcccctcaagtgtcaataccgcagggcacttatccccaggcttgtccacatcatctgtgggaaactcgcgtaaaatcaggcgtt 
               
               
                   
                 ttcgccgatttgcgaggctggccagctccacgtcgccggccgaaatcgagcctgcccctcatctgtcaacgccgcgccgggtgagtcggcccctcaagtgtcaacgtccgcccctcatctgtcagtgagggccaag 
               
               
                   
                 ttttccgcgtggtatccacaacgccggcggccgcggtgtctcgcacacggcttcgacggcgtttctggcgcgtttgcagggccatagacggccgccagcccagcggcgagggcaaccagcccggtgagcgtcgg 
               
               
                   
                 aaaggcgctggaagccccgtagcgacgcggagaggggcgagacaagccaagggcgcaggctcgatgcgcagcacgacatagccggttctcgcaaggacgagaatttccctgcggtgcccctcaagtgtcaa 
               
               
                   
               
               
                 13 
                 tra2 module trbB-L RK2 
               
               
                   
                 ttgcaggctaaacactttcggtatatcgtttgcctgtgcgataatgttgctaatgatttgttgcgtaggggttactgaaaagtgagcgggaaagaagagtttcagaccatcaaggagcgggccaagcgcaagctggaac 
               
               
                   
                 gcgacatgggtgcggacctgttggccgcgctcaacgacccgaaaaccgttgaagtcatgctcaacgcggacggcaaggtgtggcacgaacgccttggcgagccgatgcggtacatctgcgacatgcggcccag 
               
               
                   
                 ccagtcgcaggcgattatagaaacggtggccggattccacggcaaagaggtcacgcggcattcgcccatcctggaaggcgagttccccttggatggcagccgctttgccggccaattgccgccggtcgtggccgc 
               
               
                   
                 gccaacctttgcgatccgcaagcgcgcggtcgccatcttcacgctggaacagtacgtcgaggcgggcatcatgacccgcgagcaatacgaggtcattaaaagcgccgtcgcggcgcatcgaaacatcctcgtcatt 
               
               
                   
                 ggcggtactggctcgggcaagaccacgctcgtcaacgcgatcatcaatgaaatggtcgccttcaacccgtctgagcgcgtcgtcatcatcgaggacaccggcgaaatccagtgcgccgcagagaacgccgtccaa 
               
               
                   
                 taccacaccagcatcgacgtctcgatgacgctgctgctcaagacaacgctgcgtatgcgccccgaccgcatcctggtcggtgaggtacgtggccccgaagcccttgatctgttgatggcctggaacaccgggcatg 
               
               
                   
                 aaggaggtgccgccaccctgcacgcaaacaaccccaaagcgggcctgagccggctcgccatgcttatcagcatgcacccggattcaccgaaacccattgagccgctgattggcgaggcggttcatgtggtcgtcc 
               
               
                   
                 atatcgccaggacccctagcggccgtcgagtgcaagaaattctcgaagttcttggttacgagaacggccagtacatcaccaaaaccctgtaaggagtatttccaatgacaacggctgttccgttccgtctgaccatgaa 
               
               
                   
                 tcgcggcattttgttctaccttgccgtgttcttcgttctcgctctcgcgttatccgcgcatccggcgatggcctcggaaggcaccggcggcagcttgccatatgagagctggctgacgaacctgcgcaactccgtaaccg 
               
               
                   
                 gcccggtggccttcgcgctgtccatcatcggcatcgtcgtcgccggcggcgtgctgatcttcggcggcgaactcaacgccttcttccgaaccctgatcttcctggttctggtgatggcgctgctggtcggcgcgcaga 
               
               
                   
                 acgtgatgagcaccttcttcggtcgtggtgccgaaatcgcggccctcggcaacggggcgctgcaccaggtgcaagtcgcggcggcggatgccgtgcgtgcggtagcggctggacggctcgcctaatcatggctc 
               
               
                   
                 tgcgcacgatccccatccgtcgcgcaggcaaccgagaaaacctgttcatgggtggtgatcgtgaactggtgatgttctcgggcctgatggcgtttgcgctgattttcagcgcccaagagctgcgggccaccgtggtc 
               
               
                   
                 ggtctgatcctgtggttcggggcgctctatgcgttccgaatcatggcgaaggccgatccgaagatgcggttcgtgtacctgcgtcaccgccggtacaagccgtattacccggcccgctcgaccccgttccgcgagaa 
               
               
                   
                 caccaatagccaagggaagcaataccgatgatccaagcaattgcgattgcaatcgcgggcctcggcgcgcttctgttgttcatcctctttgcccgcatccgcgcggtcgatgccgaactgaaactgaaaaagcatcgt 
               
               
                   
                 tccaaggacgccggcctggccgatctgctcaactacgccgctgtcgtcgatgacggcgtaatcgtgggcaagaacggcagctttatggctgcctggctgtacaagggcgatgacaacgcaagcagcaccgacca 
               
               
                   
                 gcagcgcgaagtagtgtccgcccgcatcaaccaggccctcgcgggcctgggaagtgggtggatgatccatgtggacgccgtgcggcgtcctgctccgaactacgcggagcggggcctgtcggcgttccctgac 
               
               
                   
                 cgtctgacggcagcgattgaagaagagcgccggcggcatttcgagagcctgggaacgatgtacgagggctatttcgtcctcaccttgacctggttcccgccgctgctcgcccagcgcaagttcgtcgagctgatgttt 
               
               
                   
                 gacgacgacgcgaccgcaccggatcgcaaggcgcgcacgcggggcctcatcgaccaattcaagcgtgacgtgcgcagcatcgagtcgcgcctgtcgtcggccgtgtcgctcactcgcttgaaggggcacaaga 
               
               
                   
                 tcgtcaacgaggacggcacgaccgtcacgcatgacgacttcctgcgctggctgcaattctgcgtgacgggcctgcaccatccggtgcagctccccagcaacccgatgtacctggacgccctggtcggcggacagg 
               
               
                   
                 aaatgtggggcggggtagtgcccaaggtcggccgcaagttcgtccaggtggtcgctctcgaaggcttccccttggagtcctatcccggcatcctgacggcgctcggcgagctgccctgcgagtatcggtggtcgag 
               
               
                   
                 ccggttcatcttcatggaccagcacgaagccgtgaagcacctcgacaagttccgcaagaagtggcggcagaagattcgcggcttcttcgaccaggtgttcaacacgaacaccggcccggtcgatcaggacgcgctt 
               
               
                   
                 tcgatggtggccgatgctgaggcggccattgccgaagtcaacagcggcatcgtggccgtgggctactacaccagcgtcgtcgtgctgatggatgaggaccgcacgcgcctggaagctgcggcccgcgatgttga 
               
               
                   
                 aaaggccgtcaaccggttgggctttgccgcgcgcatcgagtccatcaacaccctggacgccttccttggtagtttgccgggccacggcgtggaaaacgtccgccggccgctcatcaacacgatgaacctggccgac 
               
               
                   
                 ctgctgccgaccagcaccatctggaccggcaacgcgaacgcgccatgcccgatgtacccgccgctgtcgccggcgctcatgcactgcgtcacgcaaggatcaacgccgttccggctgaacctgcacgtgcgcga 
               
               
                   
                 cctcggccacacctttatgttcgggccgaccggcgcaggtaaatcgacgcacctggcgatcctcgccgcgcagctccgtcgctatgccggcatgtcgatcttcgcctttgacaagggcatgtcgatgtacccgctgg 
               
               
                   
                 ccgccggcatccgtgcggccacgaagggcaccagcggcctgcacttcaccgtggcggccgacgacgaacgcctggcgttctgcccgttgcagttcctgagcaccaagggcgaccgtgcttgggcgatggagtg 
               
               
                   
                 gatcgacaccatcctggcgttgaacggcgtcgaaacgaccccggcccagcgcaacgaaatcggcaacgcgatcatgagcatgcacgccagcggcgcgcgcacgctctccgagttcagcgtgacgattcaggat 
               
               
                   
                 gaggcgatccgcgaggcgatccgccagtacaccgtcgatggcgcaatgggccatctgctcgacgccgaagaggacggcttggcgctgtccgactttacagtgttcgagatcgaagagctgatgaacctcggcga 
               
               
                   
                 gaaattcgccctgcctgtgttgctctacctgttccgccgtatcgagcgcgccctgacgggccagccggccgtcatcatcctggacgaagcctggttgatgctcggccacccggcattccgcgcgaagatcagggaat 
               
               
                   
                 ggctcaaggtgctgcgtaaggccaactgccttgtgctgatggcaacgcagagcctgtccgacgccgccaacagcggcatcctggacgtgatcgtggaatcgaccgcgaccaagattttcctgccgaatatttacgcc 
               
               
                   
                 agggatgaggacacggcggccctgtaccgccgcatgggcctgaacgctcgccagatcgagattctggcccaggccgttcccaagcgtcagtactactacgtgtcggaaaacggccgccgtctctacgacctggca 
               
               
                   
                 cttggcccgctcgcgctcgcgttcgtcggcgcatccgacaaggaatccgtcgccatcatcaagaacctggaagccaagttcggcgaccagtgggtggatgaatggctgcgtggccggggcctcgcccttgatgaat 
               
               
                   
                 acctggaggcagcatgagttttgcagacacgatcaagggcttgatcttcaagaagaagcccgcaacggccgcagcagcggcgacgccggccgcgaccggcccgcaaaccgacaacccgtacctgacggcgcg 
               
               
                   
                 gcgcacctggaacgaccacgttggttccgttgtgtcgcaaaagcagacctggcaggttgtcggcatcctttcgctgatgatcgtcctcgcggcggtcggcggcatcatccacatcggcagccagtcgaagttcgtgc 
               
               
                   
                 cctatgtctacgaggtagacaagctcgggcagacggccgccgtggggccgatgaccagggcgtcgaaagccgatccgcgtgtcattcacgcctcggtggctgagttcgtcggcgatgctcgcctggtgacgccg 
               
               
                   
                 gacgtagctttgcagcgcaaggccgtctaccgcctctatgccaagctcgggccgaatgacccggccaccgccaagatgaacgaatggctcaacggcaccgccgacgccagcccgttcgctcgcgcggccgtcg 
               
               
                   
                 aaacggtcagcaccgaaatcacttccgtaatcccgcagacgcccgacacctggcaggtcgattgggtcgagacgacgcgcgacaggcaaggcgtggtgaaaggccagcccgtgcgcatgcgggccttggtgac 
               
               
                   
                 ggtctacgtcgtcgagccgacggcggacaccaaggaagaacaactgcgaaacaacccggccgggatctacgtccgggacttctcctggtcgagacttctgtgaggcactgaattatgaaaaaggaactgtttgcttt 
               
               
                   
                 ggtcctggccgcgtccgttagcgtgcctgcatttgccgccgatcccggcgcggacctgactgacctctatttttccggcaagaacccggagctgaccgcgcaagagcgggcggccatcgccatcgccaagaagtg 
               
               
                   
                 ggaggcgggtaccgccggcatgcggccggtggccggccccggtggttcggtgcgcttcctgttcggcgcgcagcagccgagcatcgtatgcgccgtgctgcaagtgtgcgacgtggccctgcaacccggcga 
               
               
                   
                 gcaagtcaactcgatcaacctgggcgacaccgcccgttggacggtcgagccggccattaccggcagcggcgcgaacgaaacccagcacctcatcatcaagccgatggatgtgggcctggaaaccagcctggtc 
               
               
                   
                 gtgaccacggaccgccgcagctaccacatgcgcctgcgctcgcatcgcacgcagtacatgccgcaggtgtcgttcacctacccggaagatgcccttgcgaagtgggacgccatcaagaaccgcgaacagcggg 
               
               
                   
                 atcgcgtcgagaaaaccattccgcagaccggcgagtacctgggcaacctgagcttcaactactccgtcagcgggtccacgtcgtggaagccggtgcgcgtctacaacgacggcaagaaaaccatcatccagatgc 
               
               
                   
                 cgcactcgatggaacagaccgaagcgccgacgctcctggtcgttcgcagggagggcggcctgttctccgacgatgaaacggtgatggtcaactaccgggtccagggcgaccgctacatcgtcgatacgattttcg 
               
               
                   
                 acaaggccatcctcatcgcgggcgtgggcagcagccaggaccgcgtgaccatttcaagggggaactaaaccatgcgtaagattctgaccgtcatcgcactcgcggccacgttggccggctgcgcgacctccaagt 
               
               
                   
                 acggcagcttcgtccaggacgcgccggccgcctacaaccagaccattgcgaccgacgcggtgaagcagctcgtcaagctctacccgccggcgcaaaccaagctggaattgcagcaggctacgcccgatccgttc 
               
               
                   
                 ggcattgccctggtcactgaccttcgcgcccagggctatgctgtcatggagtacaagcccgacggcaacgcggccgcagctccggctgctgcgtcctcggccgctgcgaagccggcaacgccgcaagcccagg 
               
               
                   
                 gcggctatccgctgcgctacgtgctggaccaattcagcgacagcaacctgtatcgcctgaccgtcatggtcggctctcaatcgctcacgcgcgcctacctcgcccaaaacaacacgatggtcccggccggcgcatg 
               
               
                   
                 ggttcggaaggagtaagccaatgagcgaagatcaaatggcaccggacgcatcgccagatgcggtcaagccgaaaagcggggttcgccgcgtcaacaacatgccgatgtacctcatcggcggtgtgctcggcatc 
               
               
                   
                 ttcctgctggtgatggccctggttgctgcggatcgcgctgcgcagcagaaccagccgggagctgcgaaggctgagaaggccggcagcaccagcatgtttgccgacgaaattgccggcaaacagcaggacggca 
               
               
                   
                 tcatcaaggccaagccgctggagattccgccggaacaaaccgcccagcaaccgacgacggagctgacgccagccccggcgcagggaacgactatcacggtcgcacggcccgagaacctggaccagccccc 
               
               
                   
                 gacgccgccgcagggtgcgcgcaacgaggacctggaccgcatccgcatggcgaagttgcagatgctggaagaggcgatcaaggccaagacgacggtgcgcatcgacgcgccgcgcagccagggcagcgc 
               
               
                   
                 cggcggcggtgctccgcagggccgcgaggaaacccttgcgcgcatccaggagctgcgtcggcaggctgagaacgcccgcgccaccgatccgaccgccgcctatcaggccgcgcttgcgcaggctcgcacga 
               
               
                   
                 tgggcggcgcggcagggggtggcggtatgggcggctcgggtgcgccgaccctcgtgcagacctcgaaccgcagtggtggcggcgctggctatgggtcgttcgacaaccgcagcgagggcgaccgttggcgg 
               
               
                   
                 ctcgactcccagccggaagcacctgcaacgccctatgtgctgcgcgctggcttcgtcgttccggctacgcttatctcgggcatcaactccgatctgccaggccaaatcatggcccaggtatcgcagtcggtgtacgac 
               
               
                   
                 acggcgaccggcaagcacatgctcatcccccaaggctcgcgcctggtgggcagctactcgaacgatgtggcctacgggcagaagcgcgttctggtggcatggcagcgcatcatcttccccgacggcaaggcaat 
               
               
                   
                 ggacattggggccatgccgggcggcgatagcgctgggtatgcaggcttcaacgacaaggtcaacaaccactacttccgcaccttcgcatcggcattcctcatgtcgggcgtcgttgcgggcatcagcttgagtcag 
               
               
                   
                 gaccgtggcaacagcaacagcggttacggacgacaagacgcgggttccgcgatgagtgaagcgttgggtcaacagctcggccaagtaacggcgcagatgatcgccaaaaacttgaatatcgcgccgacgctgg 
               
               
                   
                 aaatccgtccgggctatcgcttcaacgtcattgtcacgaaagacatgacgttttctaagccctaccaggcgtttgactattaactccaaggagtaacttatgaagaagctcgctaagaatgttttagccgctaaagtag 
               
               
                   
                 ctctggtgctggccctctcggtcggcaccttggcggtcacgcctgcgcaagcgggcattccggtcatcgacggcaccaacctgtcacaaaccactgtcaccgcgattcagcaggttgcgcaggtccagaagcaaatcga 
               
               
                   
                 ggaataccggacgcagttgcagcagtacgaaaacatgctgcaaaacacggtggccccggccgcctacgtgtgggaccaggcgcagtccaccatcaacggcctgatgagcgccgttgataccctgaactactaca 
               
               
                   
                 agaaccaggcgggcagcatcgacgcttacctgggcaagttcaaggacgtgtcctactacaaggggtcgccgtgcttctccctgtcgggctgctcggaaagcgagcgcaaggcgatggaagagaaccgccgcctg 
               
               
                   
                 gcgtccgaatcgcagaaaaaggccaacgatgcgctgttccgtggcctcgatcagcagcagagcaacctcaagtccgacgccgccacgctggagcaattgaagggcaaggcgacgacggcgcagggccagttg 
               
               
                   
                 gaagccctcggctacgccaaccagttcgccagccagcaggccaaccagctcatgcaaatccgtggccttctgcttgcgcagcagaacgccatcgccacgcagatgcaggcccagcaggaccggcaggcccagc 
               
               
                   
                 aggacgctgcgggcgcgaagctgcgcgagggttcgtaccgcgcaagcccgtctaagacctggtgaggggaggcgcgatgaagaaatccaacttcatcgcagttgccgcgctggccgccgtcatggcggccag 
               
               
                   
                 cctggcaggctgcgacaacaagcccgacaccgacaagctgacctgcgccgatctgccgaaggtcacggatgccgctcaacgcgcggagctgttgaagaagtgcccgcgcggagaaccgggaggcttcaagcc 
               
               
                   
                 cagcgaaaagaaagagtggtgatgacgtatgaaaatccagactagagctgccgcgctcgcggtcctgatgctggccttgatgccggtagcggcatacgcccaaatcgacaattcgggcatcctcgacaacgtattg 
               
               
                   
                 cagcgctaccagaacgccgcgagcggctgggccactgtcgtccagaacgccgcaacctggctgttctggaccttgaccgtgattagcatggtctggaccttcggcatgatggcactgcgcaaggccgacattggc 
               
               
                   
                 gagttcttcgccgagttcgtgcggttcaccatcttcaccggcttcttctggtggctgctgaccaacggcccgaatttcgcgtcgtccatctatgcgtccctgcggcagattgcaggccaggcaacggggttggggcagg 
               
               
                   
                 ggctttcgccgtccggcatcgtcgatgttggcttcgagattttcttcaaggtgatggacgaaacctcgtactggtcgccggtcgatagcttcgtcggtgcctcgttggcggccgccatcctctgcatcctggccctggtcg 
               
               
                   
                 gcgtgaatatgcttctgctcctggcgtccggatggattcttgcctacggcggtgtgttcttcctgggcttcggcggctcgcgctggacctcggacatggcgatcaactactacaagaccgtcctcggggtcgccgcgca 
               
               
                   
                 gctcttcgcaatggtgctgctcgtaggcatcggcaagaccttcctcgatgactactacagccgcatgagcgaaggcatcaacttcaaggaacttggagtgatgctgatcgtcggcctgatcctgctcgttctggtcaaca 
               
               
                   
                 aggtgccgcagctcatcgccggcatcatcaccggcgcgagcgtcggcggtgctggtatcggccagttcggcgctggcacgctcgtcggtgcggccgcgacggccggcgcggcaatcgcaactggcggcgcat 
               
               
                   
                 ctatcgcggccggcgctgcggcggcggccggtggcgcgcaggccatcatggcggccgcgtcgaaggccagcgataacgtctctgccggcactgacattctgtcgagcatgatgggcggcggcggtggcggcg 
               
               
                   
                 gcggtggtagcgccggcaccagcggcggcgacggcggcggctcgggtggcggcggtggctcgggcggcggtgaaaccccgatggcctcggccgccggcgacaacagcagcggcgcacgcggcggcagt 
               
               
                   
                 tcgggcggcggctcgggtggtggccgttcgtctggcggtatcggtgccacggcggccaagggcggccggatcgcggccgataccgtcgccaacctggcgaaaggtgccggctcgattgccaaggccaaggcc 
               
               
                   
                 ggcgaaatgcgcgcatcggcccaggaacgcatcggcgataccgtaggcggcaagatcgcgcaggcaattcgcggcgcgggtgcggcggcgcagaccgctgcaaccgtcgccgatagcaacagccaggcgc 
               
               
                   
                 aggaacaacctgcaccggcacccgcaccgtcgttcgacgacaacagcctttccgcaagcaacaacagggaagcggccgccgacgcggattccgaagtggcgagcttcgtcaacaagcccgcccaatcctga 
               
               
                   
               
               
                 14 
                 tra1 module traFG-traJXIH-traKLM RK2 
               
               
                   
                 tcaccaggtcagaaccggcctgatgacggtgatgatttgcgaacgattgacaggcccgaagtagcggccgtcgaaagacgtgtcgcttacgtcggacataagcagaacctcggcggtccccagggtgtagctgtc 
               
               
                   
                 ggactgataacgaggcagcggccgtcctgatggatcggccttgatgagcgcgctgtgaggcagcagcccgccattcacgcgcacgccggcgtcggtgatggcaacctcgtcgcctttagcggctaaaactcgctt 
               
               
                   
                 catcatgtagccgtagtcgccggggcagaaaccgccggcgatgtagccccgctccttggcgtccgaaaacacgccgacttgcggcgggcagaacatgacgtaagcccccttctccaccggcgcattcgatttcca 
               
               
                   
                 gtacaggccgaccggaatgcttttggtggtgttgaccttcgcgccggcgagataggccgcgccggcgagcaacaaggccgcgccgcctccgatggcgacgtacttggtgaggcgctggaagcggctcatatcgt 
               
               
                   
                 gatcccctccccttcctcgacggtggccgtctggatcagcttgtcgctgaccttcggagccggtacggccgcgcgggcctggaatatcgggtctttgaagtagagcggctgcttgccgtagatcgcgggatagccgg 
               
               
                   
                 cgacgtacacaaccatgtcgcccgcctcttcaatgctgccgtcggcgctcttcttcggccccggcatgcgcaggcattcatcgggggtcagcaatggccgctgcacttcctggaaggtccgcgagacgttgcccaac 
               
               
                   
                 agcgccgacgtgcggcggccgctcgtcgtgatctgctccttcacgatggtcgtggtgcctgtcagttttgacaggtgctcggccgtctccacgcggttcggcgggtaggcgttctgcacgtggcagttcgacgtgatg 
               
               
                   
                 ctttcgtcgtggccgtagccggtttcgcggctcttgagctggttaatgtcctggcagatgaggtagcacttgatgccgtagccggcgacgaaggcaagggactcttgcaggatttcgagcttgcccaggctggggaac 
               
               
                   
                 tcgtcgagcatcatcagcagacgatgcttgtagtgcgcgacaggacggccgttctcgaagtccatcttgtcggccagcagccggacgatcatgttgaccatgacgcgcaccagaggccgcagacgggccttgtcgt 
               
               
                   
                 tgggctgcgtcacgatgaacaggcttaccgggtcgtcgtggtgcatcagttgcttgatgcggaagtcggacttgctgacgttgcgggccacaaccgggtcgcggtacagggccaggtaggacttggcggtggaca 
               
               
                   
                 gcacggaaccggattcttcctccgggcggtccatcatgtcgcgggccgcagagccgaccgcagggtggttctgcccgtcaacgtggccgtaggtggtcatttccatccaaagctcgcccacgtcgcggttcgggtc 
               
               
                   
                 ggcaagcatgccgtccaccgacggcagggtggccggcgtaccctcgttcttagccttgtagagcgcgtgcaggatgacgccgacaagcagcgcctggctggttttctgccagtgcgattccaggcccttgccgtcc 
               
               
                   
                 ggatcgacgatcagggtggcaaggttctgcacgtcgccaacctcgtactcggtccccaagcggatttcatcgagcgggttccagcacgcgctaccctgcgcggatgccggctcaaagcgcacgaccttgttgcgg 
               
               
                   
                 gcatgcttcttccgccagccggcggtcagcgcccacaactcgcctttcaggtcggtgatgacggcgctgtgcgcccaggaaagcagcgtcggaacgaccaggccgacgcccttgccggagcgcgtcggcgcgt 
               
               
                   
                 aggtcaagacgtgctcggggccgttgtgccgcaggtagtggaacttgccgtccttgtcctgccagccgcccacatagacgccgctggaagtgggcgggtgtttgcctgacaccagctcgacgacggtgcgcggcc 
               
               
                   
                 ggggcagcaggccggcggcctgtatgtccttcttgtcggcccagcgggccgaaccgtgcagatagtcgttcgccttgccggtgttcgccttgaccatctgcgtgacggccgtgcccagcaggcccacggtcgaaa 
               
               
                   
                 cgaccatacccatgctggccgcgcgcatgaaatcgtcgggatattggccgtaccacttgccggcccattgaaggatcgaccagggcgtgtagacgtggttgatattccagccaagtccggcctgatactggaagga 
               
               
                   
                 atgggcgaaatattgcgtcgcggtctgcaagcctgccccaagggacaggccggcgaggatgggaacggtcttgctggccttcggttttttcgcccgtatctgtggccccacggcgttgtttcggttcttcatctactcct 
               
               
                   
                 acctcgggtagttttaagggagcctcgcggggtcacggtgacgggatcaccgatggcgaggcgcttcatgcgttgcaccgtggccttatcgacgggcagcaccagaatctcgtcgttttctttcctcaacagggcca 
               
               
                   
                 gcgcctggtcctcgacgttccgggtgcctgcataggacagcgcaccaacataatcagtatatcgtgcatgcttcggtatatcgaagccgtttagccgcttttgctcgcgctcggcaacatatttctcggccgccgcgatc 
               
               
                   
                 tgttcgggctttagccctcttcctggcccagaaactccccgtcgcagtgcgtgagctggttcggctccttgctgctccacgtgaccaggaacatcacgcggcaatagcatttcagctccgccggcgatgcgaaccaca 
               
               
                   
                 ccgagttgggacagcgctcgcaaacggttttggctttggggcggcggctttcgtccaatgcgtccaacgttgggcttgcggagtgcgacggttccgccggcgctgacggcgcgagcgtcccgtcggtcgccgtcg 
               
               
                   
                 ccgcctgtggcgttgagggtggttctggctgcggcaggtcgaatgcctccatcgccgccgcgatctcttcgtccgtcatttcgttcgggttgctcatgtgcttgctccttcgtcagtagttcttgacggcggcgctcaagg 
               
               
                   
                 gcggcgtcgtcaaaggtgattgccagacggccagcggcggccgcctgcgcgatccgctccttgaactctgctgtgccgttgacggtgatccggtcgccgaagcgctccattgccaggcgcagggcggcgtccag 
               
               
                   
                 gccgtccgtggtggcctcgcgcgagacttgcaggcggtcgccgtcgtcgcggacggcgctgctgccgacgcgatagatgatggttcccttcttcgtgatgttgtccgtcacggccgcatggcccggcttggcctcg 
               
               
                   
                 ccgctgccctggatggtgttgcccttgaggtcgctgcggccctcgcgtgcgcgcagcgcggccagggccttgtcgtcgcccttcatcgcctcggccttgagccagtcggcccacgcgcggcgctgcgtgcgctcct 
               
               
                   
                 ggaccgcctgacggccctgccggtactcgcggttgatcttgtccaggtcggcgcgcagagccttgtgcgcctgcgcgtacatcagtcgctttgcaatgcgcccctcgcccagcagcttgatagcggcgcggcgca 
               
               
                   
                 gccggttgctgcgcatcgcggcttcaatcaggcggtcacgacgccggcgcagcgtgtccagctcgcccttgcgcacggcccccatttcctggcgttcagactgataccgggcgtatagctcggtggtgtcgatgcg 
               
               
                   
                 ggtcttgagcggcttcgctcgatactcccgccgccggggggcttcgccgccctcggctggcgtgaatgccccgaatcgggcttcgagcttcggcttggacaggtcgcgcgaaacggtgctggccttgaccgtcgtg 
               
               
                   
                 ccgtcgccggcctcgaagatgaagccgtttccgcgctcgcgcagcttaagcccgttttcccgcaggacgcggtgcaggtcctcccaggattgcgccgcttgcagctccggcaggcattcgcgcttgatccagccga 
               
               
                   
                 ccaggctttccacgcccgcgtgccgctccatgtcgttcgcgcggttctcggaaacgcgctgccgcgtttcgtgattgtcacgctcaagcccgtagtcccgttcgagcgtcgcgcagaggtcagcgagggcgcggta 
               
               
                   
                 ggcccgatacggctcatggatggtgtttcgggtcgggtgaatcttgttgatggcgatatggatgtgcaggttgtcggtgtcgtgatgcacggcactgacgcgctgatgctcggcgaagccaagcccagcgcagatgc 
               
               
                   
                 ggtcctcaatcgcgcgcaacgtctccgcgtcgggcttctctcccgcgcggaagctaaccagcaggtgataggtcttgtcggcctcggaacgggtgttgccgtgctgggtcgccatcacctcggccatgacagcggg 
               
               
                   
                 cagggtgtttgcctcgcagttcgtgacgcgcacgtgacccaggcgctcggtcttgccttgctcgtcggtgatgtacttcaccagctccgcgaagtcgctcttcttgatggagcgcatggggacgtgcttggcaatcacg 
               
               
                   
                 cgcaccccccggccgttttagcggctaaaaaagtcatggctctgccctcgggcggaccacgcccatcatgaccttgccaagctcgtcctgcttctcttcgatcttcgccagcagggcgaggatcgtggcatcaccgaa 
               
               
                   
                 ccgcgccgtgcgcgggtcgtcggtgagccagagtttcagcaggccgcccaggcggcccaggtcgccattgatgcgggccagctcgcggacgtgctcatagtccacgacgcccgtgattttgtagccctggccga 
               
               
                   
                 cggccagcaggtaggccgacaggctcatgccggccgccgccgccttttcctcaatcgctcttcgttcgtctggaaggcagtacaccttgataggtgggctgcccttcctggttggcttggtttcatcagccatccgcttg 
               
               
                   
                 ccctcatctgttacgccggcggtagccggccagcctcgcagagcaggattcccgttgagcaccgccaggtgcgaataagggacagtgaagaaggaacacccgctcgcgggtgggcctacttcacctatcctgccc 
               
               
                   
                 ggctgacgccgttggatacaccaaggaaagtctacacgaaccctttggcaaaatcctgtatatcgtgcgaaaaaggatggatataccgaaaaaatcgctataatgaccccgaagcagggttatgcagcggaaaagc 
               
               
                   
                 gctgcttccctgctgttttgtggaatatctaccgactggaaacaggcaaatgcaggaaattactgaactgaggggacaggcgagagacgatgccaaagagctacaccgacgagctggccgagtgggttgaatcccg 
               
               
                   
                 cgcggccaagaagcgccggcgtgatgaggctgcggttgcgttcctggcggtgagggcggatgtcgaggcggcgttagcgtccggctatgcgctcgtcaccatttgggagcacatgcgggaaacggggaaggtc 
               
               
                   
                 aagttctcctacgagacgttccgctcgcacgccaggcggcacatcaaggccaagcccgccgatgtgcccgcaccgcaggccaaggctgcggaacccgcgccggcacccaagacgccggagccacggcggcc 
               
               
                   
                 gaagcaggggggcaaggctgaaaagccggcccccgctgcggccccgaccggcttcaccttcaacccaacaccggacaaaaaggatctactgtaatggcgaaaattcacatggttttgcagggcaagggcgggg 
               
               
                   
                 tcggcaagtcggccatcgccgcgatcattgcgcagtacaagatggacaaggggcagacacccttgtgcatcgacaccgacccggtgaacgcgacgttcgagggctacaaggccctgaacgtccgccggctgaa 
               
               
                   
                 catcatggccggcgacgaaattaactcgcgcaacttcgacaccctggtcgagctgattgcgccgaccaaggatgacgtggtgatcgacaacggtgccagctcgttcgtgcctctgtcgcattacctcatcagcaacca 
               
               
                   
                 ggtgccggctctgctgcaagaaatggggcatgagctggtcatccataccgtcgtcaccggcggccaggctctcctggacacggtgagcggcttcgcccagctcgccagccagttcccggccgaagcgcttttcgtg 
               
               
                   
                 gtctggctgaacccgtattgggggcctatcgagcatgagggcaagagctttgagcagatgaaggcgtacacggccaacaaggcccgcgtgtcgtccatcatccagattccggccctcaaggaagaaacctacggc 
               
               
                   
                 cgcgatttcagcgacatgctgcaagagcggctgacgttcgaccaggcgctggccgatgaatcgctcacgatcatgacgcggcaacgcctcaagatcgtgcggcgcggcctgtttgaacagctcgacgcggcggc 
               
               
                   
                 cgtgctatgagcgaccagattgaagagctgatccgggagattgcggccaagcacggcatcgccgtcggccgcgacgacccggtgctgatcctgcataccatcaacgcccggctcatggccgacagtgcggccaa 
               
               
                   
                 gcaagaggaaatccttgccgcgttcaaggaagagctggaagggatcgcccatcgttggggcgaggacgccaaggccaaagcggagcggatgctgaacgcggccctggcggccagcaaggacgcaatggcg 
               
               
                   
                 aaggtaatgaaggacagcgccgcgcaggcggccgaagcgatccgcagggaaatcgacgacggccttggccgccagctcgcggccaaggtcgcggacgcgcggcgcgtggcgatgatgaacatgatcgccg 
               
               
                   
                 gcggcatggtgttgttcgcggccgccctggtggtgtgggcctcgttatgaatcgcagaggcgcagatgaaaaagcccggcgttgccgggcttgttttt 
               
               
                   
               
               
                 15 
                 Sal-crRNA Array 1 (522bp) 
               
               
                   
                 TTTGAAAACAAAGAATTAGCTGATCTTTAATAATAAGGAAA TGTTACATT AAGGTTGGTGGGTTGTTTTTATGGGAAAAAATGCTTTAAGAACAAATGTATACTTTTAGA GAGTTCCCCGCGCCAGCGGGGATAAACCG   
               
               
                   
                 
                   GCTTTAAGCGTTAGCTCCCCATTCTGCTCCCC 
                   GAGTTCCCCGCGCCAGCGGGGATAAACCG 
                   GTACGTTAGCGTATATTGATGCCGCAGAGACG 
                   GAGTTCCCCGCGCCAGCGGGGATAAACCG 
                   GCGATAACTGGACAG 
                 
               
               
                   
                   TTTTATCCGCCGAGCAT   GAGTTCCCCGCGCCAGCGGGGATAAACCG GGCGCACTGGATGCGATGATGGATATCACTTGGAGTTCCCCCGCCTCTGCGGTAGAACTCCCAGCTCCCATTTTCAAACCCATCAAGACGCC 
               
               
                   
                 TTCGCCAACTCCTTCACCAGAGGTAGCATTATCCGCATAACGTCACGGCAGCGACGTTCTATTCTTCCAGGAAGAGCCTTATCAATATGTTGGTGATTATCCAGTCTT 
               
               
                   
               
               
                 16 
                 Repeat Sequence 
               
               
                   
                 GAGTTCCCCGCGCCAGCGGGGATAAACCG 
               
               
                   
               
               
                 17 
                 Spacer Sequence for targeting the  Salmonella  invB gene 
               
               
                   
                 GCTTTAAGCGTTAGCTCCCCATTCTGCTCCCC 
               
               
                   
               
               
                 18 
                 Spacer Sequence for targeting the  Salmonella  sicP gene 
               
               
                   
                 GTACGTTAGCGTATATTGATGCCGCAGAGACG 
               
               
                   
               
               
                 19 
                 Spacer Sequence for targeting the  Salmonella  seeE gene 
               
               
                   
                 GCGATAACTGGACAGTTTTATCCGCCGAGCAT 
               
               
                   
               
               
                 20 
                 invB from  Salmonella enterica  subsp.  enterica  serovar Typhimurium str. LT2; spacer sequence in bold 
               
               
                   
                 atgcaacatttggatatcgctgaattagttcgttccgcactggaagtaagtggttgcgatccttcattaattggaggaatagatagccattcaacaattgttctggatttatttgcattgccaagtatctgtat 
               
               
                   
                 cagcgtcaaggacgatgatgtatggatctgggcgcaattgggtgctgacagcatggtggtattacaacagcgggcttatgaaatcttaatgaccataatggaaggatgccattttgcccgcggcggg 
               
               
                   
                 caattactactg ggggagcagaatggggagctaacgcttaaagc   ctt agtgcatccggattttttatctgacggtgaaaagttctctactgccttgaatgggttttacaactatctggaagtttttagtc 
               
               
                   
                 ggtcgctaatgagatga 
               
               
                   
               
               
                 21 
                 sicP from  Salmonella enterica  subsp.  enterica  serovar Typhimurium str. LT2; spacer sequence in bold 
               
               
                   
                 ttgcaagcacaccaggatattatcgctaatattggtgagaaattgggtttaccgctcacttttgacgacaacaatcagtgcttattattactcgatagcgatatttttacgtctattgaagctaaagatgatat 
               
               
                   
                 ctggttattgaacggtatgattataccgttatcgcctgtttgtggcgattctatctggcggcagattatggtgattaatggtgaactggctgcgaataatgaaggtacgttagcgtatattgatgccgca 
               
               
                   
                   gagacg ttgttgatatacatgcaattaccgatagacaaatacttaccatattatatcgcagcttgagtcatttgtgaatcagcaggaagcgctcaaaaacatactgcaggaatatgctaaagtatga 
               
               
                   
               
               
                 22 
                 seeE from  Salmonella enterica  subsp.  enterica  serovar Typhimurium str. LT2; spacer sequence in bold 
               
               
                   
                 atggtgcaagaaatagagcaatggttacgtcggcatcaggtgtttactgagcctgcatatttaggggagaccgccatattacttgggcagcagtttatattatcgccttacctggtgatctatcgtattga 
               
               
                   
                 ggcaaaagaaatgattatttgtgagttcaggcgcctgacgcccgggcaacctcgaccacagcaattgtttcacttactgggacttttacgcgggatatttgtgcatcacccgcagttaacatgtttaaag 
               
               
                   
                 atgttgataatcaccgacgttctggatgaaaaaaaagccatgctacgcaggaaattattgcgcatcctgacagtaatgggagcgacctttacacagcttg atg   gcgataactggacagttttatccg   
               
               
                   
                   ccgagcat cttatccagcgacgtttttaa 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 7 
               
             
            
               
                   
               
               
                 Function and origin of each genetic element of the pFS-Sal- 
               
               
                 09-proAB-rm plasmid containing a Type I CRISPR/Cas system. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                 Size 
                   
                   
               
               
                 Component 
                 Location 
                 (bp) 
                 Origin 
                 Function 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 J23114 
                  1-29 
                 29 
                 Synthetic 
                 Initiates transcription of Cas3 
               
               
                 promoter 
                   
                   
                 promoter 
                 and Cas3 cascade 
               
               
                 Cas3 cascade 
                  105-4455 
                 4351 
                   E. coli  K12 
                 Recruits Cas3 and recognition 
               
               
                 (casABCDE) 
                   
                   
                   
                 of PAM 
               
               
                 Cas3 
                 4472-7138 
                 2667 
                   E. coli  K12 
                 Nuclease activity; cleavage of 
               
               
                   
                   
                   
                   
                 DNA 
               
               
                 Promoter- 
                 7182-7210 
                 29 
                   E. coli  K12 
                 Initiates transcription of pre- 
               
               
                 crRNA array 
                   
                   
                   
                 crRNA array to mature 
               
               
                   
                   
                   
                   
                 crRNA array 
               
               
                 invB-cr4 
                 7278-7309 
                 32 
                 Synthetic 
                 Spacer sequence of guide 
               
               
                   
                   
                   
                 sequence 
                 RNA. Binds to the 
               
               
                   
                   
                   
                   
                 complementary target DNA 
               
               
                   
                   
                   
                   
                 sequence of invB gene in 
               
               
                   
                   
                   
                   
                 
                   Salmonella enterica 
                 
               
               
                 sicP-cr1 
                 7339-7370 
                 32 
                 Synthetic 
                 Spacer sequence of guide 
               
               
                   
                   
                   
                 sequence 
                 RNA. Binds to the 
               
               
                   
                   
                   
                   
                 complementary target DNA 
               
               
                   
                   
                   
                   
                 sequence of sicP gene in 
               
               
                   
                   
                   
                   
                 
                   Salmonella enterica 
                 
               
               
                 sseE-cr2 
                 7400-7431 
                 32 
                 Synthetic 
                 Spacer sequence of guide 
               
               
                   
                   
                   
                 sequence 
                 RNA. Binds to the 
               
               
                   
                   
                   
                   
                 complementary target DNA 
               
               
                   
                   
                   
                   
                 sequence of sseE gene in 
               
               
                   
                   
                   
                   
                 
                   Salmonella enterica 
                 
               
               
                 oriT 
                 7661-7781 
                 121 
                 RP4 plasmid 
                 Origin of transfer. Transfer of 
               
               
                   
                   
                   
                   
                 CRISPR/CAS3 system to 
               
               
                   
                   
                   
                   
                 recipient 
               
               
                 rrnB T1 
                 7809-7895 
                 87 
                   E. coli  rrnB gene 
                 Transcription termination 
               
               
                 terminator 
               
               
                 p15A ori 
                 8064-8609 
                 546 
                 pZA31MCS 
                 Plasmid replication 
               
               
                   
                   
                   
                 vector 
               
               
                 Lambda T0 
                 8723-8817 
                 95 
                 Phage lambda 
                 Transcription termination 
               
               
                 terminator 
               
               
                 proA 
                  8837-10090 
                 1254 
                   E. coli  K12 
                 Proline biosynthesis 
               
               
                 proB 
                 10102-11205 
                 1104 
                   E. coli  K12 
                 Proline biosynthesis 
               
               
                 proAB promoter 
                 11250-11278 
                 29 
                   E. coli  K12 
                 Initiates transcription of 
               
               
                   
                   
                   
                   
                 proAB 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 8 
               
             
            
               
                   
               
               
                 Conjugation results in JM109. Conjugation from S17-1::pFS-EcoCas3-01- 
               
               
                 rm, S17-1::pFS-Sal-09-rm, S17-1::pFS-EcoCas3-03-rm and S17-1::pFS- 
               
               
                 Sal-08-rm, in  E. coli  JM109 strain showed a comparable efficiency 
               
               
                 between the two constructs and the respective control plasmids. 
               
               
                 Conjugation efficiency is calculated dividing the number (CFU/mL) 
               
               
                 of transconjugants by the number of recipients. 
               
            
           
           
               
               
               
               
            
               
                   
                   
                   
                 Conjugation 
               
               
                 Donor 
                 Recipient 
                 CFU/mL 
                 Efficiency 
               
               
                   
               
               
                 S17-1::pFS-EcoCas3-01-rm 
                 JM109 
                 1.2 × 107 
                 1.4 × 10−2 
               
               
                 S17-1::pFS-Sal-09-rm 
                 JM109 
                 7.0 × 106 
                 7.4 × 10−3 
               
               
                 S17-1::pFS-EcoCas3-03-rm 
                 JM109 
                 8.1 × 106 
                 3.4 × 10−2 
               
               
                 S17-1::pFS-Sal-08-rm 
                 JM109 
                 1.5 × 107 
                 2.9 × 10−2 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 9 
               
             
            
               
                   
               
               
                 Conjugation results in FS26. Conjugation from S17-1::pFS-EcoCas3-01-rm, 
               
               
                 S17-1::pFS-Sal-09-rm, S17-1::pFS-EcoCas3-03-rm and S17-1::pFS-Sal-08-rm in 
               
               
                   S. Enteritidis  strain FS26 were carried out in two biological replicates. 
               
               
                 Results showed significant reduction in  S. Enteritidis  strain FS26 
               
               
                 by conjugation with the active constructs, when compared to the respective 
               
               
                 control plasmids. Conjugation efficiency is calculated dividing the number 
               
               
                 (CFU/mL) of transconjugants by the number of recipients. 
               
            
           
           
               
               
               
               
               
            
               
                   
                   
                   
                 Transconjugants 
                 Conjugation 
               
               
                 Replicate 
                 Donor 
                 Recipient 
                 (CFU/mL) 
                 Efficiency 
               
               
                   
               
               
                 1st 
                 S17-1::pFS-EcoCas3-01-rm 
                 FS26 
                 7.6 × 106 
                 1.2 × 10−2 
               
               
                   
                 S17-1::pFS-Sal-09-rm 
                 FS26 
                 7.0 × 102 
                 2.7 × 10−6 
               
               
                 2nd 
                 S17-1::pFS-EcoCas3-01-rm 
                 FS26 
                 1.3 × 107 
                 1.0 × 10−2 
               
               
                   
                 S17-1::pFS-Sal-09-rm 
                 FS26 
                 1.0 × 103 
                 1.2 × 10−6 
               
               
                 1st 
                 S17-1::pFS-EcoCas3-03-rm 
                 FS26 
                 3.0 × 106 
                 1.0 × 10−2 
               
               
                   
                 S17-1::pFS-Sal-08-rm 
                 FS26 
                 3.7 × 102 
                 2.7 × 10−6 
               
               
                 2nd 
                 S17-1::pFS-EcoCas3-03-rm 
                 FS26 
                 1.0 × 107 
                 2.9 × 10−2 
               
               
                   
                 S17-1::pFS-Sal-08-rm 
                 FS26 
                 2.0 × 103 
                 5.6 × 10−6 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 10 
               
             
            
               
                   
               
               
                 Selected spacer conservation in  Salmonella   
               
            
           
           
               
               
               
               
            
               
                   
                   
                 conservation level 
                   
               
               
                   
                 # hits in 
                 in  Salmonella   
                 non- Salmonella   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 Spacer 
                 PAM 
                 
                   S. enterica 
                 
                 100 
                 ≥95 
                 ≥90 
                 &lt;90 
                 hits 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 invB 
                 AAG 
                 10613 
                 96 
                 2 
                 2 
                   
                 23 
               
               
                 sicP 
                 AAG 
                 10610 
                 74.5 
                 20.7 
                 4.7 
                 &lt;1 
                 46 
               
               
                 sseE 
                 ATG 
                 10604 
                 68.1 
                 30 
                 1.9 
                   
                 52 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
               
               
               
             
               
                 TABLE 11 
               
               
                   
               
               
                   
                 Under 
                 Ambient temperature 
                 Relative 
               
               
                 Age (days) 
                 lamp (° C.) 
                 in room (° C.) 
                 humidity (%) 
               
               
                   
               
             
            
               
                 Up to 1 
                 35 
                 25-30 
                 60-80 
               
               
                 1-7 
                 32 
                 22-27 
                 60-80 
               
               
                  7-14 
                 29 
                 19-25 
                 40-80 
               
               
                 14-21 
                 26 
                 18-25 
                 40-80 
               
               
                 21-28 
                 24 
                 18-25 
                 40-80 
               
               
                 28-35 
                 — 
                 18-25 
                 40-80 
               
               
                 Over 35 
                 — 
                 15-25 
                 40-80 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 12 
               
             
            
               
                   
               
               
                 Plating protocol 
               
            
           
           
               
               
               
            
               
                   
                 Plating for  Salmonella   
                 Enrichment for  Salmonella   
               
               
                   
                   
               
               
                   
                 XLD + Nalidixic acid 
                 Selenite broth, Plate onto XLD + 
               
               
                   
                 (25 μg/mL) 
                 Nalidixic acid (25 μg/mL) 
               
               
                   
                 Direct counts -  Salmonella   
                 Enriched counts -  Salmonella