bluuebunny/crossref_metadata_2025 huggingface dataset:
API Listeria, a new and promising one-day system to identify Listeria isolates
Authors: Bille J , Catimel B , Bannerman E , Jacquet C , Yersin M N , Caniaux I , Monget D , Rocourt J
Published: January 2020

API Listeria is a new 10-test strip for 24-h biochemical identification of Listeria isolates. With this commercial system, 85% of 646 Listeria strains, including atypical isolates selected for this study, were recognized at the species and subspecies level without a complementary test. A new test differentiates Listeria monocytogenes from L. innocua on the basis of the absence of arylamidase from the former. With this system, 97.7% (252 of 258) of the L. monocytogenes strains tested were correctly identified and differentiated from 99.4% (175 of 176) of the L. innocua strains also tested. Gram-positive bacteria other than Listeria spp. gave quite different biochemical patterns. This system considerably reduced the time needed for conventional identification, since results were available within 18 to 24 h.
Hyperreiterated DNA regions are conserved among Bradyrhizobium japonicum serocluster 123 strains
Authors: Rodriguez-Quiñones F , Judd A K , Sadowsky M J , Liu R L , Cregan P B
Published: January 2020

We have identified and cloned two DNA regions which are highly reiterated in Bradyrhizobium japonicum serocluster 123 strains. While one of the reiterated DNA regions, pFR2503, is closely linked to the B. japonicum common and genotype-specific nodulation genes in strain USDA 424, the other, pMAP9, is located next to a Tn5 insertion site in a host-range extension mutant of B. japonicum USDA 438. The DNA cloned in pFR2503 and pMAP9 are reiterated 18 to 21 times, respectively, in the genomes of B. japonicum serocluster 123 strains. Gene probes from the reiterated regions share sequence homology, failed to hybridize (or hybridized poorly) to genomic DNA from other B. japonicum and Bradyrhizobium spp. strains, and did not hybridize to DNA from Rhizobium meliloti, Rhizobium fredii, Rhizobium leguminosarum biovars trifolii, phaseoli, and viceae, or Agrobacterium tumefacians. The restriction fragment length polymorphism hybridization profiles obtained by using these gene probes are useful for discriminating among serologically related B. japonicum serocluster 123 strains.
Genetic and Phenotypic Diversity of
Bacillus polymyxa
in Soil and in the Wheat Rhizosphere
Authors: Mavingui Patrick , Laguerre Gisèle , Berge Odile , Heulin Thierry
Published: January 2020

Diversity among 130 strains of
Bacillus polymyxa
was studied; the bacteria were isolated by immunotrapping from nonrhizosphere soil (32 strains), rhizosphere soil (38 strains), and the rhizoplane (60 strains) of wheat plantlets growing in a growth chamber. The strains were characterized phenotypically by 63 auxanographic (API 50 CHB and API 20B strips) and morphological features, serologically by an enzyme-linked immunosorbent assay, and genetically by restriction fragment length polymorphism (RFLP) profiles of total DNA in combination with hybridization patterns obtained with an rRNA gene probe. Cluster analysis of phenotypic characters by the unweighted pair group method with averages indicated four groups at a similarity level of 93%. Clustering of
B. polymyxa
strains from the various fractions showed that the strains isolated from nonrhizosphere soil fell into two groups (I and II), while the third group (III) mainly comprised strains isolated from rhizosphere soil. The last group (IV) included strains isolated exclusively from the rhizoplane. Strains belonging to a particular group exhibited a similarity level of 96%. Serological properties revealed a higher variability among strains isolated from nonrhizosphere and rhizosphere soil than among rhizoplane strains. RFLP patterns also revealed a greater genetic diversity among strains isolated from nonrhizosphere and rhizosphere soil and therefore could not be clearly grouped. The RFLP patterns of sorbitol-positive strains isolated from the rhizoplane were identical. These results indicate that diversity within populations of
B. polymyxa
isolated from nonrhizosphere and rhizosphere soil is higher than that of
B. polymyxa
isolated from the rhizoplane. It therefore appears that wheat roots may select a specific subpopulation from the soil
B. polymyxa
population.
Study on the epidemiology and control of Campylobacter jejuni in poultry broiler flocks
Authors: van de Giessen A , Mazurier S I , Jacobs-Reitsma W , Jansen W , Berkers P , Ritmeester W , Wernars K
Published: January 2020

Broiler flocks are frequently infected with Campylobacter jejuni. The origin of the infection is still unclear. The question of whether colonization of flocks results from transmission of C. jejuni from breeder flocks to progeny (vertical transmission) or from environmental sources (horizontal transmission) remains to be answered. Therefore, in this study samples were taken from successive broiler flocks in two broiler houses (house A on farm A and house B1 on farm B) as well as from the environment of the houses. All C. jejuni isolates were typed by using the Penner serotyping system, and part of the isolates from farm B were typed by using a randomly amplified polymorphic DNA-typing system. In poultry house A, C. jejuni was isolated from the first flock but not from subsequent flocks. In poultry house B1, C. jejuni strains of the same Penner serotypes and exhibiting identical DNA profiles were isolated from successive flocks. Infection of the flocks from a common source via horizontal pathways is suspected, while a vertical route of infection is not likely to exist. Application of measures to control horizontal transmission of C. jejuni on farm B was successful.
Transformation of Bacillus subtilis by DNA bound on montmorillonite and effect of DNase on the transforming ability of bound DNA
Authors: Khanna M , Stotzky G
Published: January 2020

The equilibrium adsorption and binding of DNA from Bacillus subtilis on the clay mineral montmorillonite, the ability of bound DNA to transform competent cells, and the resistance of bound DNA to degradation by DNase I are reported. Maximum adsorption of DNA on the clay occurred after 90 min of contact and was followed by a plateau. Adsorption was pH dependent and was greatest at pH 1.0 (19.9 micrograms of DNA mg of clay-1) and least at pH 9.0 (10.7 micrograms of DNA mg of clay-1). The transformation frequency increased as the pH at which the clay-DNA complexes were prepared increased, and there was no transformation by clay-DNA complexes prepared at pH 1. After extensive washing with deionized distilled water (pH 5.5) or DNA buffer (pH 7.5), 21 and 28%, respectively, of the DNA remained bound. Bound DNA was capable of transforming competent cells (as was the desorbed DNA), indicating that adsorption, desorption, and binding did not alter the transforming ability of the DNA. Maximum transformation by bound DNA occurred at 37 degrees C (the other temperatures evaluated were 0, 25, and 45 degrees C). DNA bound on montmorillonite was protected against degradation by DNase, supporting the concept that "cryptic genes" may persist in the environment when bound on particulates. The concentration of DNase required to inhibit transformation by bound DNA was higher than that required to inhibit transformation by comparable amounts of free DNA, and considerably more bound than free DNase was required to inhibit transformation by the same amount of free DNA. Similarly, when DNA and DNase were bound on the same or separate samples of montmorillonite, the bound DNA was protected from the activity of DNase.
An enzyme immunoassay of phaseolinone and its application in estimation of the amount of toxin in Macrophomina phaseolina-infected seeds
Authors: Bhattacharya D , Dhar T K , Ali E
Published: January 2020

A microtiter plate-based enzyme immunoassay has been developed for phaseolinone, a phytotoxin isolated from the culture filtrate of the plant-pathogenic fungus Macrophomina phaseolina (Tassi) Goid. The smallest amount of phaseolinone detectable by the method is 5 pg per well. The method is validated by comparison with high-performance liquid chromatography and used to confirm and estimate phaseolinone production in seeds infected with the fungus. The degree of seed inhibition correlated well with the amount of toxin produced in infected seeds, 50% inhibition being observed at a toxin concentration of 0.60 micrograms/g of wet tissue.
Biological suppression of potato ring rot by fluorescent pseudomonads
Authors: de la Cruz A R , Poplawsky A R , Wiese M V
Published: January 2020

Three strains of fluorescent pseudomonads (IS-1, IS-2, and IS-3) isolated from potato underground stems with roots showed in vitro antibiosis against 30 strains of the ring rot bacterium Clavibacter michiganensis subsp. sepedonicus. On the basis of morphological and biochemical tests and fatty acid analysis, IS-1 and IS-2 were identified as Pseudomonas aureofaciens and IS-3 was identified as P. fluorescens biovar III. IS-1 was the most inhibitory to C. michiganensis subsp. sepedonicus strains in vitro, followed by IS-3 and IS-2. Suppression of ring rot by these antagonists was demonstrated in greenhouse trials with stem-cultured potato (cv. Russet Burbank) seedlings. Although each antagonist significantly reduced C. michiganensis subsp. sepedonicus populations, only IS-1 reduced infection by C. michiganensis subsp. sepedonicus. In a second experiment, treatment with IS-1 (10(9) CFU/ml) significantly reduced ring rot infection by 23.4 to 26.7% after 5 to 8 weeks. The average C. michiganensis subsp. sepedonicus population was also significantly reduced by 50 to 52%. Application of different combinations of antagonist strains was not more effective than single-strain treatment.
Incubation of water samples containing amoebae improves detection of legionellae by the culture method
Authors: Sanden G N , Morrill W E , Fields B S , Breiman R F , Barbaree J M
Published: January 2020

Some protozoans isolated from aquatic habitats, including domestic water supplies, can support the intracellular replication of autochthonous legionellae in vitro. We studied the effect of incubating water samples containing amoebae on the sensitivity of culture for legionellae. Samples collected during investigations of legionellosis epidemics and shown by conventional culture procedures to contain amoebae, but not legionellae, were incubated at 35 degrees C and replated. Legionellae were recovered from 59 of 144 such samples. Species isolated included L. pneumophila, L. anisa, L. bozemanii, L. gormanii, L. micdadei, L. rubrilucens, L. sainthelensi, L. steigerwaltii, and an unnamed species. Acanthamoeba polyphaga, Acanthamoeba hatchetti, a Rosculus sp., Hartmannella vermiformis, and Vahlkampfia spp. were among the autochthonous amoebae identified. Legionellae were recovered by this procedure from only 3 of 63 samples that were negative for amoebae by conventional culture procedures. These results show that water samples negative for legionellae, but positive for amoebae, by standard culture techniques should be incubated and replated to maximize the sensitivity of culture for legionellae.
Differentiation of Gram-Negative, Nonfermentative Bacteria Isolated from Biofilters on the Basis of Fatty Acid Composition, Quinone System, and Physiological Reaction Profiles
Authors: Lipski André , Klatte Stefan , Bendinger Bernd , Altendorf Karlheinz
Published: January 2020

Gram-negative, nonfermentative bacteria isolated from biofilters for off-gas treatment of animal-rendering-plant emissions were differentiated by whole-cell fatty acid analysis, quinone analysis, and numerical taxonomy based on their physiological reaction profiles. The last system consisted of 60 physiological tests and was arranged as a microtest system on microtitration plates. Based on fatty acid analyses, 31 isolates were separated into six clusters and five single-member clusters. The isolates of two clusters were identified as
Alcaligenes faecalis
and
Pseudomonas diminuta.
The remaining nine clusters were characterized by their fatty acid profiles, quinone systems, and physiological reaction profiles. Clusters resulting from fatty acid analyses were compared with those resulting from physiological reaction profiles. Six clusters could be confirmed this way. The efficiency of the physiological test system was increased by the prearrangement of the isolates according to their quinone type.
Acid adaptation promotes survival of Salmonella spp. in cheese
Authors: Leyer G J , Johnson E A
Published: January 2020

Salmonella typhimurium was adapted to acid by exposure to hydrochloric acid at pH 5.8 for one to two doublings. Acid-adapted cells had increased resistance to inactivation by organic acids commonly present in cheese, including lactic, propionic, and acetic acids. Recovery of cells during the treatment with organic acids was increased 1,000-fold by inclusion of 0.1% sodium pyruvate in the recovery medium. Acid-adapted S. typhimurium cells survived better than nonadapted cells during a milk fermentation by a lactic acid culture. Acid-adapted cells also showed enhanced survival over a period of two months in cheddar, Swiss, and mozzarella cheeses kept at 5 degrees C. Acid adaptation was found in Salmonella spp., including Salmonella enteritidis, Salmonella choleraesuis subsp. choleraesuis serotype heidelberg, and Salmonella choleraesuis subsp. choleraesuis serotype javiana, associated with food poisoning. These observations support the theory that acid adaptation is an important survival mechanism enabling Salmonella spp. to persist in fermented dairy products and possibly other acidic food products.
Acquisition of a sucrose utilization system in Escherichia coli K-12 derivatives and its application to industry
Authors: Tsunekawa H , Azuma S , Okabe M , Okamoto R , Aiba S
Published: January 2020

An Escherichia coli strain, B-62, that was isolated from a clinical source and was epidemiologically unrelated to E. coli K-12 was the source of chromosomal DNA for a sucrose utilization system (Scr+) in the construction of a plasmid, pST621. The cloned insert of a gene encoding Scr+ in pST621 conferred a sucrose-positive phenotype onto transformed cells of E. coli K-12 derivatives. Sucrase activity of the transformants was as high as that which would correspond to a "gene dosage effect" of a vector plasmid pBR322, whereas the transformants' sucrose uptake activity was always lower than that of E. coli B-62. A region within an XhoI-SacI fragment (3.2 kb) of pBR322-glyA was replaced in the construction of another plasmid, pST5R7, by a fragment (about 2.6 kb) of pST622 containing the gene encoding Scr+. A genetically stable Scr+ derivative of E. coli K-12 was obtained by introducing the gene encoding Scr+ onto E. coli chromosome via homologous recombination between pST5R7 and the chromosome and subsequent plasmid segregation. The use of low-copy-number plasmid RP4 as a cloning vector was also effective for enhancing the stability of Scr+. Tryptophan producers E. coli SGIII1032S, in which the gene encoding Scr+ was cloned onto the chromosome, and E. coli SGIII1032, which carried Scr+ plasmid RP4.5R7, produced from 6% sucrose in shake flasks (33 degrees C, 96 h) 2.3 and 5.7 g of tryptophan per liter, respectively.
Assessment of genetic diversity and population structure of Xanthomonas oryzae pv. oryzae with a repetitive DNA element
Authors: Leach J E , Rhoads M L , Vera Cruz C M , White F F , Mew T W , Leung H
Published: January 2020

A repetitive DNA element cloned from Xanthomonas oryzae pv. oryzae was used to assess the population structure and genetic diversity of 98 strains of X. oryzae pv. oryzae collected between 1972 and 1988 from the Philippine Islands. Genomic DNA from X. oryzae pv. oryzae was digested with EcoRI and analyzed for restriction fragment length polymorphisms (RFLPs) with repetitive DNA element as a probe. Twenty-seven RFLP types were identified; there was no overlap of RFLP types among the six races from the Philippines. Most variability (20 RFLP types) was found in strains of races 1, 2, and 3, which were isolated from tropical lowland areas. Four RFLP types (all race 5) were found among strains isolated from cultivars grown in the temperate highlands. The genetic diversity of the total population of X. oryzae pv. oryzae was 0.93, of which 42% was due to genetic differentiation between races. The genetic diversities of strains collected in 1972 to 1976, 1977 to 1981, and 1982 to 1986, were 0.89, 0.90, and 0.92, respectively, suggesting a consistently high level of variability in the pathogen population over the past 15 years. Cluster analysis based on RFLP banding patterns showed five groupings at 85% similarity. The majority of strains from a given race were contained within one cluster, except for race 3 strains, which were distributed in three of the five clusters.
Influence of ecosystematic factors on survival of Escherichia coli after large-scale release into lake water mesocosms
Authors: Brettar I , Höfle M G
Published: January 2020

Mass cultures of an Escherichia coli K-12 strain were released into exposed mesocosms in a eutrophic lake. The release was performed with and without additional input of the E. coli culture medium to stimulate the scenario of leakage of a production fermenter on one hand and to compare the influence of the added organic nutrients with that of the added strain on the other hand. The survival of the introduced strain and the influence on ecological processes in the mesocosms were monitored for 10 weeks after release. For comparison, survival of the strain in microcosms with sterile lake water was also monitored. Survival of the strain was determined by means of immunofluorescence and growth on selective agar medium. In lake mesocosms, E. coli showed a rapid and constant dieback during the first week. After 4 days, cells were mostly restricted to particles, which seemed to provide niches for survival. From the second week onward, survival was improved in mesocosms with culture medium added. In microcosms with sterile lake water, plate counts of E. coli showed a strong decrease within 2 weeks, while total cell numbers remained approximately the same. The rapid elimination of E. coli from the free-water phase of the mesocosms was probably due to the combined effect of the inability to grow in lake water and grazing. The better survival of E. coli (mainly on particles) in mesocosms with added medium was attributed to the medium-induced enhancement of primary production, which was the source of a large quantity of particles. These particles, in turn, may have functioned as niches for prolonged survival as well as transport vehicles for sedimentation of the E. coli cells.
Applications of a colorimetric plate assay for soluble methane monooxygenase activity
Authors: Graham D W , Korich D G , LeBlanc R P , Sinclair N A , Arnold R G
Published: January 2020

A straightforward method is described for screening methanotrophic colonies for soluble methane monooxygenase (sMMO) activity on solid media. Such activity results in the development of a colored complex between 1-naphthol, which is formed when sMMO reacts with naphthalene, and o-dianisidine (tetrazotized). Methanotrophic colonies expressing sMMO turned deep purple when exposed successively to naphthalene and o-dianisidine. The method was evaluated within the contexts of two potential applications. The first was for the enumeration of Methylosinus trichosporium OB3b in a methane-amended, unsaturated soil column dedicated to vinyl chloride treatment. The second application was for the isolation and enumeration of sMMO-bearing methanotrophs from sanitary landfill soils. The technique was effective in both applications.
The Composition and Attributes of
Colletotrichum truncatum
Spores Are Altered by the Nutritional Environment
Authors: Jackson Mark A. , Schisler David A.
Published: January 2020

Previous sporulation studies with
Colletotrichum truncatum
NRRL 13737, a fungal pathogen of the noxious weed
Sesbania exaltata
, showed that the carbon-to-nitrogen (CN) ratio of the conidiation medium influenced spore yield, morphology, and efficacy in inciting disease in
S. exaltata
. Spores produced in a medium with a CN ratio of 10:1 were more effective than were spores produced in a 30:1 or 80:1 ratio in causing disease in
S. exaltata
. With a basal salts medium supplemented with glucose and Casamino Acids, substrate utilization, spore production, biomass accumulation, and biomass and spore composition were compared in submerged cultures of
C. truncatum
grown in media with CN ratios of 80:1, 30:1, and 10:1. All cultures were sporulating by day 2, and spore concentrations in 5-day-old cultures were significantly different: 30:1 > 10:1 > 80:1. Amino acid and glucose utilization was balanced in cultures grown in media with a CN ratio of 10:1, whereas cultures grown in media with a CN ratio of 30:1 or 80:1 depleted amino acids prior to glucose. Conidia produced in media with a CN ratio of 10:1 contained significantly more protein (32% of dry weight) and less lipid (17% of dry weight) than conidia produced in media with a CN ratio of either 30:1 (15% protein, 33% lipid) or 80:1 (12% protein, 37% lipid). The higher lipid content of spores produced in media with a CN ratio of 30:1 or 80:1 was associated with the presence of increased numbers of lipid droplets. Optimization studies on conidia produced in media with CN ratios between 30:1 and 10:1 which compared yield, attributes, and efficacy in inciting disease in
S. exaltata
suggest that media with a CN ratio of 15:1 to 20:1 may be optimal for conidium production.
Comparative acid tolerances and inhibitor sensitivities of isolated F-ATPases of oral lactic acid bacteria
Authors: Sturr M G , Marquis R E
Published: January 2020

pH activity profiles and inhibitor sensitivities were compared for membrane ATPases isolated from three oral lactic acid bacteria, Lactobacillus casei ATCC 4646, Streptococcus mutans GS-5, and Streptococcus sanguis NCTC 10904, with, respectively, high, moderate, and low levels of acid tolerance. Membranes containing F1F0 ATPases were isolated by means of salt lysis of cells treated with muralytic enzymes. Membrane-free F1F0 complexes were then isolated from membranes by detergent extraction with Triton X-100 or octylglucoside. Finally, F1 complexes free of the proton-conducting F0 sector were obtained by washing membranes with buffers of low ionic strength. The pH activity profiles of the membrane-associated enzymes reflected the general acid tolerances of the organisms from which they were isolated; for example, pH optima were approximately 5.5, 6.0, and 7.0, respectively, for enzymes from L. casei, S. mutans, and S. sanguis. Roughly similar profiles were found for membrane-free F1F0 complexes, which were stabilized by phospholipids against loss of activity during storage. However, profiles for F1 enzymes were distinctly narrower, indicating that association with F0 and possibly other membrane components enhanced tolerance to both acid and alkaline media. All of the enzymes were found to have similar sensitivities to Al-F complexes, but only F1F0 enzymes were highly sensitive to dicyclohexylcarbodiimide. The procedures described for isolation of membrane-free F1F0 forms of the enzymes from oral lactic acid bacteria will be of use in future studies of the characteristics of the enzymes, especially in studies with liposomes.
Combined subtraction hybridization and polymerase chain reaction amplification procedure for isolation of strain-specific Rhizobium DNA sequences
Authors: Bjourson A J , Stone C E , Cooper J E
Published: January 2020

A novel subtraction hybridization procedure, incorporating a combination of four separation strategies, was developed to isolate unique DNA sequences from a strain of Rhizobium leguminosarum bv. trifolii. Sau3A-digested DNA from this strain, i.e., the probe strain, was ligated to a linker and hybridized in solution with an excess of pooled subtracter DNA from seven other strains of the same biovar which had been restricted, ligated to a different, biotinylated, subtracter-specific linker, and amplified by polymerase chain reaction to incorporate dUTP. Subtracter DNA and subtracter-probe hybrids were removed by phenol-chloroform extraction of a streptavidin-biotin-DNA complex. NENSORB chromatography of the sequences remaining in the aqueous layer captured biotinylated subtracter DNA which may have escaped removal by phenol-chloroform treatment. Any traces of contaminating subtracter DNA were removed by digestion with uracil DNA glycosylase. Finally, remaining sequences were amplified by polymerase chain reaction with a probe strain-specific primer, labelled with 32P, and tested for specificity in dot blot hybridizations against total genomic target DNA from each strain in the subtracter pool. Two rounds of subtraction-amplification were sufficient to remove cross-hybridizing sequences and to give a probe which hybridized only with homologous target DNA. The method is applicable to the isolation of DNA and RNA sequences from both procaryotic and eucaryotic cells.
Localization of methanol dehydrogenase in two strains of methylotrophic bacteria detected by immunogold labeling
Authors: Fassel T A , Buchholz L A , Collins M L , Remsen C C
Published: January 2020

Antibodies to methanol dehydrogenase purified from Methylobacterium sp. strain AM1 and Methylomonas sp. strain A4 were raised. The antibody preparations were used in indirect immunogold labeling studies. With this approach, methanol dehydrogenase was found to be preferentially localized to the periplasmic region of the methylotroph Methylobacterium sp. strain AM1 and to the intracytoplasmic membrane of the methanotroph Methylomonas sp. strain A4. Antibody cross-reactivity to other methylotrophic bacteria was detected.
Monoclonal antibody-colony immunoblot method specific for isolation of Pediococcus acidilactici from foods and correlation with pediocin (bacteriocin) production
Authors: Bhunia A K , Johnson M G
Published: January 2020

BALB/c mice were immunized with broken, heat-killed cells of Pediococcus acidilactici H. After murine cell fusions, one monoclonal antibody (MAb), Ped-2B2, was selected on the basis of its positive reaction with seven of seven strains tested in an enzyme-linked immunosorbent assay with whole cells of P. acidilactici. The MAb Ped-2B2 did not show any cross-reactions with other lactic-acid bacteria or other gram-positive or gram-negative organisms. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot (immunoblot) analysis of surface proteins of P. acidilactici indicated that Ped-2B2 reacted with a protein of 116 kDa. MAb Ped-2B2 was used as a probe to isolate Pediococcus species from fermented-meat products by colony immunoblotting. A total of 18 Ped-2B2-reactive Pediococcus spp. isolates were isolated from eight food samples and assayed for bacteriocin production. All of the isolates produced bacteriocins which were heat stable, proteinaceous, and inhibitory to Lactobacillus plantarum NCDO 955. Biochemical characterization of these isolates indicated that they were all P. acidilactici.
Changes in fatty acid branching and unsaturation of Streptomyces griseus and Brevibacterium fermentans as a response to growth temperature
Authors: Suutari M , Laakso S
Published: January 2020

Streptomyces griseus showed three different modes of changing fatty acids in response to temperature change. In Brevibacterium fermentans, two such responses were found. The responses involved changes in fatty acid branching, unsaturation, or chain length, depending on growth temperature range. Changes in unsaturation of branched-chain acids were characteristic at low growth temperatures.
Effect of Starvation on Induction of Quinoline Degradation for a Subsurface Bacterium in a Continuous-Flow Column
Authors: Truex Michael J. , Brockman Fred J. , Johnstone Donald L. , Fredrickson James K.
Published: January 2020

Differences in the induction response and the initial two reactions of quinoline degradation between short-term (2 days)- and long-term (60 to 80 days)-starved cells of a subsurface
Pseudomonas cepacia
strain were examined by using continuous-flow columns. The ability of bacteria that are indigenous to oligotrophic environments to respond to a contaminant was assessed by using long-term starvation to induce a cell physiology that simulates the in situ physiology of the bacteria. With quinoline concentrations of 39 and 155 μM, long-term-starved cells converted quinoline to degradation products more efficiently than did short-term-starved cells. Quinoline concentrations of 155 μM and, to a greater extent, 775 μM had an inhibitory effect on induction in long-term-starved cells. However, only the length of the induction process was affected with these quinoline concentrations; degradation of quinoline at the steady state for long-term-starved cells was equal to or better than that for short-term-starved cells. The induction time for short-term-starved cells did not increase progressively with increasing quinoline concentration. Experiments with starved cells are important for the development of accurate predictive models of contaminant transport in the subsurface because starvation, which induces a cell physiology that simulates the in situ physiology of many bacteria, may affect the induction process.
Detection of a Single Genetically Modified Bacterial Cell in Soil by Using Charge Coupled Device-Enhanced Microscopy
Authors: Silcock Deborah J. , Waterhouse Rosemary N. , Glover L. Anne , Prosser James I. , Killham Kenneth
Published: January 2020

Genes encoding bioluminescence from
Vibrio harveyi
were cloned into
Pseudomonas syringae
pv. phaseoli-cola, resulting in high levels of bioluminescence. After inoculation of sterile and nonsterile soil slurries with bioluminescent
P. syringae
, cells could not be identified by conventional light microscopy. However, when we used charge coupled device-enhanced microscopy, bioluminescent single cells were detected easily in dark fields despite masking by soil particulate matter, and in addition, the extent of competition from indigenous soil bacteria could be monitored. The technique which we describe offers great potential for tracking and determining the spatial distribution of genetically marked microorganisms in the environment.
Characterization of Lactococcus lactis phage antigens
Authors: Schouler C , Bouet C , Ritzenthaler P , Drouet X , Mata M
Published: January 2020

Phage phi 197 is representative of a widespread lactococcal phage group characterized by a particular morphology (prolate head with a noncontractile tail). In order to develop an immunoenzymatic phage detection test, fusion proteins containing beta-galactosidase fused to epitopes of phage phi 197 structural proteins were constructed by cloning random DNA fragments from the phage genome upstream of a lacZ gene on a plasmid vector. Recombinant plasmids containing certain fragments encoded the synthesis of fusion proteins which react with polyclonal antibodies against the phage and confer a Lac+ phenotype on Escherichia coli. Three different epitopes were represented; phage-specific DNA fragments encoding these epitopes were mapped at three locations on the phage genome, and their nucleotide sequences were determined. Two fused phage antigens were conformational epitopes, whereas the phage epitope of protein encoded by the recombinant plasmid designated pOA17 was a denaturation-resistant epitope. This epitope was very immunogenic. Protein encoded by plasmid pOA17 was synthesized in large amounts from a strong promoter. Antibodies raised against this hybrid protein were used to identify the 46-kDa minor phage protein which provides the epitope. Antibody cross-reactivity of phages related to phi 197 showed that this epitope is well conserved in this genetic group.
Isolation and characterization of a variety of microcystins from seven strains of the cyanobacterial genus Anabaena
Authors: Sivonen K , Namikoshi M , Evans W R , Carmichael W W , Sun F , Rouhiainen L , Luukkainen R , Rinehart K L
Published: January 2020

Hepatotoxins (microcystins) from seven freshwater Anabaena strains originating from three different Finnish lakes and one lake in Norway were isolated by high-performance liquid chromatography and characterized by amino acid analysis and fast atom bombardment mass spectrometry. All strains produced three to seven different microcystins. A total of 17 different compounds were isolated, of which 8 were known microcystins. The known compounds identified from six strains were MCYST (microcystin)-LR, [D-Asp3]MCYST-LR, [Dha7]MCYST-LR, [D-Asp3,Dha7]MCYST-LR, MCYST-RR, [D-Asp3]MCYST-RR, [Dha7]MCYST-RR, and [D-Asp3,Dha7]MCYST-RR. With the exception of MCYST-LR and [D-Asp3]MCYST-LR, this is the first time that isolation of these toxins from Anabaena strains has been reported. Three of the strains produced one to three toxins as minor components which could not be identified. Anabaena sp. strain 66 produced four unidentified toxins. The other Anabaena strains always contained both MCYST-LR and MCYST-RR and/or their demethyl variants. Quantitative differences between toxins within and between strains were detected; at times MCYST-LR and at other times MCYST-RR or demethyl derivatives thereof were the most abundant toxins found in a strain.
Growth and Survival of Genetically Manipulated
Lactobacillus plantarum
in Silage
Authors: Sharp R. , O'Donnell A. G. , Gilbert H. G. , Hazlewood G. P.
Published: January 2020

The growth and persistence of two genetically manipulated forms of
Lactobacillus plantarum
NCDO (National Collection of Dairy Organisms) 1193 have been monitored in grass silage. Both recombinants contained pSA3, a shuttle vector for gram-positive organisms that encodes erythromycin resistance. In one of the recombinants, pSA3 was integrated onto the chromosome, whereas in the other, a pSA3 derivative designated pM25, which contains a
Clostridium thermocellum
cellulase gene cloned into pSA3, was maintained as an extrachromosomal element. This extrachromosomal element is a plasmid. Rifampin-resistant mutants were selected for the recombinants and the parent strain. When applied to minisilos at a rate of 10
6
CFU/g of grass, both the recombinants and the parent strain proliferated to dominate the epiphytic microflora and induced an increase in the decline in pH compared with that of the noninoculated silos. The presence of extra genetic material did not appear to disadvantage the bacterium in comparison with the parent strain. The selective recovery of both strains by using rifampin and erythromycin was confirmed by Southern hybridization. Interestingly, the free plasmid (pM25) appeared more stable in silage than was expected from studies in MRS broth. The plasmid was retained by 85% of the rifampin-resistant
L. plantarum
colonies isolated from a day 30 silo. These data answer an important question by showing that genetically manipulated recombinants of
L. plantarum
can proliferate and compete with epiphytic lactic acid bacteria in silage.
Genus- and species-specific identification of mycoplasmas by 16S rRNA amplification
Authors: van Kuppeveld F J , van der Logt J T , Angulo A F , van Zoest M J , Quint W G , Niesters H G , Galama J M , Melchers W J
Published: January 2020

Systematic computer alignment of mycoplasmal 16S rRNA sequences allowed the identification of variable regions with both genus- and species-specific sequences. Species-specific sequences of Mycoplasma collis were elucidated by asymmetric amplification and dideoxynucleotide sequencing of variable regions, using primers complementary to conserved regions of 16S rRNA. Primers selected for Mycoplasma pneumoniae, M. hominis, M. fermentans, Ureaplasma urealyticum, M. pulmonis, M. arthritidis, M. neurolyticum, M. muris, and M. collis proved to be species specific in the polymerase chain reaction. The genus-specific primers reacted with all mycoplasmal species investigated as well as with members of the genera Ureaplasma, Spiroplasma, and Acholeplasma. No cross-reaction was observed with members of the closely related genera Streptococcus, Lactobacillus, Bacillus, and Clostridium or with any other microorganism tested. On the basis of the high copy number of rRNA, a highly sensitive polymerase chain reaction assay was developed in which the nucleic acid content equivalent to a single organism could be detected.
Metabolism of 3-methylindole by a methanogenic consortium
Authors: Gu J D , Berry D F
Published: January 2020

A methanogenic 3-methylindole (3-MI)-degrading consortium, enriched from wetland soil, completely mineralized 3-MI. Degradation proceeded through an initial hydroxylation reaction forming 3-methyloxindole. The consortium was unable to degrade oxindole or isatin, suggesting a new pathway for 3-MI fermentation.
Culture and decontamination methods affecting enumeration of phages infecting Bacteroides fragilis in sewage
Authors: Tartera C , Araujo R , Michel T , Jofre J
Published: January 2020

A new medium has been adapted for the growth of Bacteroides fragilis so that its phages can be recovered from environmental samples, and its efficiency has been assessed. Polyvinylidene difluoride membranes allow significantly higher recoveries among different membrane filters used to decontaminate the samples. In all cases, a number of phages remain in the filters and a percentage of them can be recovered by treatment with an eluant.
Isolation of Endophytic
Streptomyces
Strains from Surface-Sterilized Roots
Authors: Sardi P. , Saracchi M. , Quaroni S. , Petrolini B. , Borgonovi G. E. , Merli S.
Published: January 2020

When the roots of 28 plant species were surface sterilized and incubated on agar medium, endophytic actinomycetes in the root cortex were observed by direct microscopic observation and pure culture techniques.
Use of inducible disaccharidases to assess the importance of different carbohydrate sources for Bacteroides ovatus growing in the intestinal tracts of germfree mice
Authors: Valentine P J , Salyers A A
Published: January 2020

Patterns of disaccharidase expression were used to determine which polysaccharides were the major sources of carbohydrate for Bacteroides ovatus growing in the intestinal tracts of monocolonized germfree mice. Results indicate that B. ovatus grows on a variety of different carbohydrates, which are present in low concentrations, rather than relying on one type of carbohydrate as the major carbohydrate source.
Temperature-dependent induction of an acid-inducible stimulon of Escherichia coli in broth
Authors: Hassani M , Pincus D H , Bennett G N , Hirshfield I N
Published: January 2020

The induction of the inducible lysyl-tRNA synthetase, LysU, and the inducible lysine and arginine decarboxylases of Escherichia coli K-12 grown in AC broth to a pH of 5.5 or less is temperature dependent, being distinctly lower at 24 than at 37 degrees C. This induction does not appear to be under HtpR control.
Virulence characteristics of clinical and environmental isolates of Vibrio vulnificus
Authors: Stelma G N , Reyes A L , Peeler J T , Johnson C H , Spaulding P L
Published: January 2020

Twenty-four randomly selected clinical and environmental Vibrio vulnificus isolates were tested for virulence in iron-overloaded mice (250 mg of iron dextran per kg of body weight). The log10 50% lethal doses of 17 isolates were lower by greater than or equal to 3.5 log10 units in iron-overloaded mice than in control mice. These isolates were classified as virulent. The 50% lethal doses of these virulent isolates were also lower in mice that were immunosuppressed by treatment with cyclophosphamide (150 mg/kg). Four of the seven isolates initially classified as avirulent were virulent in mice that were simultaneously iron overloaded and immunosuppressed. These isolates were classified as moderately virulent. The remaining three isolates were avirulent under all conditions. The incidence of virulent strains among clinical and environmental isolates did not differ. The virulent isolates produced high titers of hemolysin, were resistant to inactivation by serum complement, produced phenolate siderophore, and utilized transferrin-bound iron. The moderately virulent isolates differed from the virulent isolates only in their increased sensitivity to inactivation by serum complement. The avirulent isolates differed from those of the other two classes in their inability to either produce significant amounts of phenolate siderophore or utilize transferrin-bound iron. A modified agar plate diffusion method for transferrin-bound iron utilization was developed to differentiate the two classes of virulent isolates from the avirulent isolates in vitro.
A model study of factors involved in adhesion of Pseudomonas fluorescens to meat
Authors: Piette J P , Idziak E S
Published: January 2020

A study was undertaken to investigate the factors involved in the adhesion of Pseudomonas fluorescens to model meat surfaces (tendon slices). Adhesion was fast (less than 2.5 min) and was not suppressed by killing the cells with UV, gamma rays, or heat, indicating that physiological activity was not required. In various salt solutions (NaCl, KCl, CaCl2, MgCl2), adhesion increased with increasing ionic strength up to 10 to 100 mM, suggesting that, at low ionic strengths, electrostatic interactions were involved in the adhesion process. At higher ionic strengths (greater than 10 to 100 mM) or in the presence of Al3+ ions, adhesion was sharply reduced. Selectively blocking of carboxyl or amino groups at the cell surface by chemical means did not affect adhesion. These groups are therefore not directly involved in an adhesive bond with tendon. Given a sufficient cell concentration (10(10) CFU.ml-1) in the adhesion medium, the surface of tendon was almost entirely covered with adherent bacteria. This suggests that if the adhesion is specific, the attachment sites on the tendon surface must be located within collagen or proteoglycan molecules.
Loss of allosteric control but retention of the bifunctional catalytic competence of a fusion protein formed by excision of 260 base pairs from the 3' terminus of pheA from Erwinia herbicola
Authors: Xia T , Zhao G , Jensen R A
Published: January 2020

A bifunctional protein denoted as the P protein and encoded by pheA is widely present in purple gram-negative bacteria. This P protein carries catalytic domains that specify chorismate mutase (CM-P) and prephenate dehydratase. The instability of a recombinant plasmid carrying a pheA insert cloned from Erwinia herbicola resulted in a loss of 260 bp plus the TAA stop codon from the 3' terminus of pheA. The plasmid carrying the truncated pheA gene (denoted pheA*) was able to complement an Escherichia coli pheA auxotroph. pheA* was shown to be a chimera composed of the residual 5' part of pheA (901 bp) and a 5-bp fragment from the pUC18 vector. The new fusion protein (PheA*) retained both chorismate mutase and prephenate dehydratase activities. PheA* had a calculated subunit molecular weight of 33,574, in comparison to the 43,182-molecular-weight subunit size of PheA. The deletion did not affect the ability of PheA* to assume the native dimeric configuration of PheA. Both the CM-P and prephenate dehydratase components of PheA* were insensitive to L-phenylalanine inhibition, in contrast to the corresponding components of PheA. L-Phenylalanine protected both catalytic activities of PheA from thermal inactivation, and this protective effect of L-phenylalanine upon the PheA* activities was lost. PheA* was more stable than PheA to thermal inactivation; this was more pronounced for prephenate dehydratase than for CM-P. In the presence of dithiothreitol, the differential resistance of PheA* prephenate dehydratase to thermal inactivation was particularly striking.(ABSTRACT TRUNCATED AT 250 WORDS)
Heat and cold shock protein synthesis in arctic and temperate strains of rhizobia
Authors: Cloutier J , Prévost D , Nadeau P , Antoun H
Published: January 2020

We compared heat shock proteins (HSPs) and cold shock proteins (CSPs) produced by different species of Rhizobium having different growth temperature ranges. Several HSPs and CSPs were induced when cells of three arctic (psychrotrophic) and three temperate (mesophilic) strains of rhizobia were shifted from their optimal growth temperatures (arctic, 25 degrees C; temperate, 30 degrees C) to shock temperatures outside their growth temperature ranges. At heat shock temperatures, three major HSPs of high molecular weight (106,900, 83,100, and 59,500) were present in all strains for all shock treatments (29, 32, 36.4, 38.4, 40.7, 41.4, and 46.4 degrees C), with the exception of temperate strains exposed to 46.4 degrees C, in which no protein synthesis was detected. Cell survival of arctic and temperate strains decreased markedly with the increase of shock temperature and was only 1% at 46.4 degrees C. Under cold shock conditions, five proteins (52.0, 38.0, 23.4, 22.7, and 11.1 kDa) were always present for all treatments (-2, -5, and -10 degrees C) in arctic strains. Among temperate strains, five CSPs (56.1, 37.1, 34.4, 17.3, and 11.1 kDa) were present at temperatures down to 0 degrees C. The 34.4- and the 11.1-kDa components were present in all temperate strains at -5 degrees C and in one strain at -10 degrees C. Survival of all strains decreased with cold shock temperatures but was always higher than 50%. These results show that rhizobia can synthesize proteins at temperatures not permissive for growth. In all shock treatments, no correspondence between the number of HSPs or CSPs produced and rhizobial survival was found.(ABSTRACT TRUNCATED AT 250 WORDS)
Characterization of a novel Pseudomonas sp. that mineralizes high concentrations of pentachlorophenol
Authors: Radehaus P M , Schmidt S K
Published: January 2020

A pentachlorophenol (PCP)-mineralizing bacterium was isolated from polluted soil and identified as Pseudomonas sp. strain RA2. In batch cultures, Pseudomonas sp. strain RA2 used PCP as its sole source of carbon and energy and was capable of completely degrading this compound as indicated by radiotracer studies, stoichiometric release of chloride, and biomass formation. Pseudomonas sp. strain RA2 was able to mineralize a higher concentration of PCP (160 mg liter-1) than any previously reported PCP-degrading pseudomonad. At a PCP concentration of 200 mg liter-1, cell growth was completely inhibited and PCP was not degraded, although an active population of Pseudomonas sp. RA2 was still present in these cultures after 2 weeks. The inhibitory effect of PCP was partially attributable to its effect on the growth rate of Pseudomonas sp. strain RA2. The highest specific growth rate (mu = 0.09 h-1) was reached at a PCP concentration of 40 mg liter-1 but decreased at higher or lower PCP concentrations, with the lowest mu (0.05 h-1) occurring at 150 mg liter-1. Despite this reduction in growth rate, total biomass production was proportional to PCP concentration at all PCP concentrations degraded by Pseudomonas sp. RA2. In contrast, final cell density was reduced to below expected values at PCP concentrations greater than 100 mg liter-1. These results indicate that, in addition to its effect as an uncoupler of oxidative phosphorylation, PCP may also inhibit cell division in Pseudomonas sp. strain RA2.(ABSTRACT TRUNCATED AT 250 WORDS)
Degradation of 2,4-dinitrophenol by two Rhodococcus erythropolis strains, HL 24-1 and HL 24-2
Authors: Lenke H , Pieper D H , Bruhn C , Knackmuss H J
Published: January 2020

Two Rhodococcus erythropolis strains, HL 24-1 and HL 24-2, were isolated from soil and river water by their abilities to utilize 2,4-dinitrophenol (0.5 mM) as the sole source of nitrogen. Although succinate was supplied as a carbon and energy source during selection, both isolates could utilize 2,4-dinitrophenol also as the sole source of carbon. Both strains metabolized 2,4-dinitrophenol under concomitant liberation of stoichiometric amounts of nitrite and 4,6-dinitrohexanoate as a minor dead-end metabolite.