bluuebunny/crossref_metadata_2025 huggingface dataset:
Assay for the enzymatic conversion of indoleacetic acid to 3-methylindole in a ruminal Lactobacillus species
Authors: Honeyfield D C , Carlson J R
Published: January 2020

An assay to measure the rate of enzymatic formation of 3-methylindole (3MI) from indoleacetic acid (IAA) in Lactobacillus sp. strain 11201 was developed. The reaction mixture contained 50 micrograms of microbial protein per ml (range, 25 to 100 mg/ml), essential low-molecular-weight reaction ingredients, and radiolabeled IAA as substrate (range, 0 to 2 mM IAA). The reaction was anaerobic for 25 min at 39 degrees C. The apparent Michaelis-Menten constants were: Km, 0.14 mM IAA; and Vmax, 64 nmol 3MI.mg-1.min-1. The inhibitors avidin, aminopterin, and EDTA had no effect on the 3MI-forming enzyme. Dithionite stimulated the 3MI-forming enzyme. The product of the reaction, 3MI, acted as a noncompetitive inhibitor of the enzyme. Enzyme activity was associated with the cell wall fraction after sonication; treatment with the French press; or treatment with detergents, proteolytic enzymes, and EDTA.
Die-Away Kinetic Analysis of the Capacity of Epilithic and Planktonic Bacteria from Clean and Polluted River Water To Biodegrade Sodium Dodecyl Sulfate
Authors: Anderson David J. , Day Martin J. , Russell Nicholas J. , White Graham F.
Published: January 2020

The capacities of epilithic and planktonic river bacterial populations to degrade sodium dodecyl sulfate (SDS) in samples taken at two times during 1987 from one clean and four polluted sites in a South Wales river were estimated in die-away tests under simulated environmental conditions. There was a relatively slow disappearance of SDS in die-away tests for both planktonic and epilithic populations taken from the clean source site, as compared with those taken from the downstream polluted sites, for which the rate of biodegradation was accelerated, sometimes after an apparent initial lag period. The kinetic components contributing to the die-away curves were quantified by nonlinear regression analysis in which the experimental data were fitted to a variety of possible kinetic models. All samples except for one from the polluted sites best fitted a model which describes the biodegradation of SDS at concentrations well below its

K
m

by bacteria whose growth is exponential and unaffected by the addition of a test substrate. Each sample from the clean source site fitted a different model, but there was generally little or no growth on endogenous carbon. A consideration of the numerical values of constants derived from the modeling of epilithic and planktonic populations from polluted sites indicated clearly that the biodegradative capacity of epilithic bacterial populations towards SDS is more stable than that of planktonic bacterial populations.
Comparison of growth, acetate production, and acetate inhibition of Escherichia coli strains in batch and fed-batch fermentations
Authors: Luli G W , Strohl W R
Published: January 2020

The growth characteristics and acetate production of several Escherichia coli strains were compared by using shake flasks, batch fermentations, and glucose-feedback-controlled fed-batch fermentations to assess the potential of each strain to grow at high cell densities. Of the E. coli strains tested, including JM105, B, W3110, W3100, HB101, DH1, CSH50, MC1060, JRG1046, and JRG1061, strains JM105 and B were found to have the greatest relative biomass accumulation, strain MC1060 accumulated the highest concentrations of acetic acid, and strain B had the highest growth rates under the conditions tested. In glucose-feedback-controlled fed-batch fermentations, strains B and JM105 produced only 2 g of acetate.liter-1 while accumulating up to 30 g of biomass.liter-1. Under identical conditions, strains HB101 and MC1060 accumulated less than 10 g of biomass.liter-1 and strain MC1060 produced 8 g of acetate.liter-1. The addition of various concentrations of sodium acetate to the growth medium resulted in a logarithmic decrease, with respect to acetate concentration, in the growth rates of E. coli JM105, JM105(pOS4201), and JRG1061. These data indicated that the growth of the E. coli strains was likely to be inhibited by the acetate they produced when grown on media containing glucose. A model for the inhibition of growth of E. coli by acetate was derived from these experiments to explain the inhibition of acetate on E. coli strains at neutral pH.
Large-scale purification of staphylococcal enterotoxins A, B, and C2 by dye ligand affinity chromatography
Authors: Brehm R D , Tranter H S , Hambleton P , Melling J
Published: January 2020

A simple method for the purification of staphylococcal enterotoxins A (SEA), B (SEB), and C2 (SEC2) from fermentor-grown cultures was developed. The toxins were purified by pseudo-affinity chromatography by using the triazine textile dye "Red A" and gave overall yields of 49% (SEA), 44% (SEB), and 53% (SEC2). The purified toxins were homogeneous when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but isoelectric focusing of the preparations revealed the microheterogeneity associated with these toxins. The SEA and SEB preparations each consisted of two isoelectric forms with pI values of 7.3 and 6.8 (SEA) and 8.9 and 8.55 (SEB); in contrast, SEC2 contained five different isoelectric forms, with pI values ranging between 7.6 and 6.85. The pattern of elution of the isoelectric forms from the column indicated a cationic-exchange process involved in the binding of toxin to Red A. Such a method forms the basis of a high-yielding, rapid means of purifying the staphylococcal enterotoxins that can easily be adapted to large-scale production.
Biodegradation of azo and heterocyclic dyes by Phanerochaete chrysosporium
Authors: Cripps C , Bumpus J A , Aust S D
Published: January 2020

Biodegradation of Orange II, Tropaeolin O, Congo Red, and Azure B in cultures of the white rot fungus, Phanerochaete chrysosporium, was demonstrated by decolarization of the culture medium, the extent of which was determined by monitoring the decrease in absorbance at or near the wavelength maximum for each dye. Metabolite formation was also monitored. Decolorization of these dyes was most extensive in ligninolytic cultures, but substantial decolorization also occurred in nonligninolytic cultures. Incubation with crude lignin peroxidase resulted in decolorization of Azure B, Orange II, and Tropaeolin O but not Congo Red, indicating that lignin peroxidase is not required in the initial step of Congo Red degradation.
Identification of Deep-Sea-Sediment Bacteria Which Produce Tetrodotoxin
Authors: Do H. K. , Kogure K. , Simidu U.
Published: January 2020

Forty-nine bacterial strains were isolated from deep-sea sediments. Among them, 22 strains were shown by the tissue culture assay method to produce sodium channel blockers. For some strains, high-performance liquid chromatography analysis confirmed that the blocker was tetrodotoxin. Tetrodotoxin-producing bacteria seem to be widespread in marine sediment.
Same-day identification scheme for colonies of Listeria monocytogenes
Authors: Lachica R V
Published: January 2020

A diagnostic scheme is described for the same-day identification of food-borne cells of Listeria monocytogenes that emerge in 40 h at 30 degrees C as large colonies, representatives of which are used to advantage as heavy inocula on agar plates for the rapid determination of hemolytic activity and acidification of rhamnose and xylose. Additional tests consisting of phase-contrast microscopy for cell morphology and motility, the catalase production test, and the KOH viscosity test in place of Gram staining complete the rapid identification scheme.
Trichloroethylene degradation by two independent aromatic-degrading pathways in Alcaligenes eutrophus JMP134
Authors: Harker A R , Kim Y
Published: January 2020

The bacterium Alcaligenes eutrophus JMP134(pJP4) degrades trichloroethylene (TCE) by a chromosomal phenol-dependent pathway and by the plasmid-encoded 2,4-dichlorophenoxyacetic acid pathway. The two pathways were independent and exhibited different rates of removal and capacities for quantity of TCE removed. The phenol-dependent pathway was more rapid (0.2 versus 0.06 nmol of TCE removed per min per mg of protein) and consumed all detectable TCE. The 2,4-dichlorophenoxyacetic acid-dependent pathway removed 40 to 60% of detectable TCE.
Chemical germination of native and cation-exchanged bacterial spores with trifluoperazine
Authors: Sacks L E
Published: January 2020

The calmodulin antagonist trifluoperazine and its analog chlorpromazine, both amphipaths, induced chemical germination of spores of various species, as do many surfactants. Cation load can greatly influence this response. Calmodulin antagonism does not seem to be involved. A new fluorometric assay for dipicolinic acid based on the fluorescence of the dipicolinic acid chelate of Tb3+ was found to be simple and sensitive.
Inhibition Enzyme-Linked Immunosorbent Assay for Detection of
Pseudomonas fluorescens
Proteases in Ultrahigh-Temperature-Treated Milk
Authors: Clements Richard S. , Wyatt David M. , Symons Michael H. , Ewings Kenneth N.
Published: January 2020

An inhibition enzyme-linked immunosorbent assay was developed to detect low levels of the proteases extracted from four strains of
Pseudomonas fluorescens.
The assay detected between 0.24 and 7.8 ng of protease per ml of ultrahigh-temperature-treated milk and could be completed within 6 h. It could be used as a framework for a test system for quantifying spoilage proteases in dairy products.
Purification and Characterization of Glycerol Dehydratase from
Lactobacillus reuteri
Authors: Talarico Todd L. , Dobrogosz Walter J.
Published: January 2020

A coenzyme B
12
-dependent glycerol dehydratase from
Lactobacillus reuteri
has been purified and characterized. The dehydratase has a molecular weight of approximately 200,000, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis yielded a single major band with a molecular weight of 52,000.
K

m

values for substrates and coenzyme B
12
were in the millimolar and the submicromolar range, respectively.
Efficient DNA transformation of Bradyrhizobium japonicum by electroporation
Authors: Hattermann D R , Stacey G
Published: January 2020

Intact cells of Bradyrhizobium japonicum USDA 110 were transformed with a 30-kilobase plasmid to efficiencies of 10(6) to 10(7) transformants per microgram by high-voltage electroporation. The technique was reliable and simple, with single colonies arising from transformed cells within 5 days of antibiotic selection. Plasmid DNA from B. japonicum transformed the Bradyrhizobium (Arachis) sp. with high efficiency, while the same plasmid extracted from Escherichia coli transformed B. japonicum at very low efficiency. The electrical conditions that resulted in the highest efficiencies were high voltage (10.5 to 12.5 kV/cm) and short pulse length (6 to 7 ms). A linear increase in the number of transformants was observed as DNA concentration was increased over 4 orders of magnitude; saturation appeared to begin between 120 ng/ml and 1.2 micrograms/ml. This novel method of transformation should enhance B. japonicum genetic research by providing a valuable alternative to conjugal mating, which is currently the only efficient, widely used means of introducing DNA into this organism.
Purification and Characterization of an Extracellular Acid Proteinase from the Ectomycorrhizal Fungus
Hebeloma crustuliniforme
Authors: Zhu Hong , Guo Da-Cheng , Dancik Bruce P.
Published: January 2020

Hebeloma crustuliniforme
produced an extracellular acid proteinase in a liquid medium containing bovine serum albumin as the sole nitrogen source. The proteinase was purified 26-fold with 20% activity recovery and was shown to have a molecular weight of 37,800 (as indicated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis) and an isoelectric point of 4.8 ± 0.2. The enzyme was most active at 50°C and pH 2.5 against bovine serum albumin and was stable in the absence of substrates at temperatures up to 45°C and pHs between 2.0 and 5.0. Pepstatin A, diazoacetyl-
dl
-norleucine methylester, metallic ions Fe
2+
and Fe
3+
, and phenolic acids severely inhibited the enzyme activity, while antipain, leupeptin,
N
-α-
p
-tosyl-
l
-lysine chloromethyl ketone, and trypsin inhibitor inhibited the activity moderately. The proteinase hydrolyzed bovine serum albumin and cytochrome
c
rapidly compared with casein and azocasein but failed to hydrolyze any of the low-molecular-weight peptide derivatives tested.
Characterization of Propionibacterium plasmids
Authors: Rehberger T G , Glatz B A
Published: January 2020

Plasmid DNAs from 15 Propionibacterium strains were characterized by using restriction endonuclease analyses, DNA-DNA hybridizations, and curing experiments. Restriction endonuclease analysis identified seven distinct plasmids (pRGO1 through pRGO7). Detailed restriction maps were constructed for four of these plasmids. DNA-DNA hybridization analysis revealed that plasmids pRGO1 and pRGO2 had extensive sequence homology and that both were homologous to pRGO7 and to similar sequences of pRGO5. Plasmids pRGO4 and pRGO6 did not have any significant sequence homology with any of the other plasmids. Plasmid pRGO3 had partial sequence homology only with pRGO7. Curing of plasmids pRGO1, pRGO2, and pRGO5 was achieved by treatment with acriflavin, but we failed to identify any plasmid-encoded bacteriocin production, carbohydrate fermentation, or antibiotic resistance. However, physical evidence was obtained that tentatively linked the clumping phenotype of Propionibacterium jensenii P38 with plasmid pRGO5.
Phosphonate utilization by bacterial cultures and enrichments from environmental samples
Authors: Schowanek D , Verstraete W
Published: January 2020

A selection of axenic microbial strains and a variety of environmental samples were investigated with respect to the utilization of a series of natural and xenobiotic phosphonates as the sole phosphorus source for growth. Phosphonate degradation was observed only with bacteria and not with eucaryotic microorganisms. All representatives of the phosphonates examined supported bacterial growth, with the exception of methylphosphonate diethylester. Yet, distinctly different phosphonate utilization patterns were noted between phosphonate-positive strains. C-P bond cleavage by a photosynthetic bacterium is reported for the first time; growing photoheterotrophically, Rhodobacter capsulatus ATCC 23782 was able to utilize 2-aminoethylphosphonate and alkylphosphonates. Bacteria with the potential to utilize at least one of the phosphonate moieties from the xenobiotic phosphonates Dequest 2010, Dequest 2041, and Dequest 2060 were detected in all environments, with only two exceptions for Dequest 2010. Phosphonate P utilization to an extent of 94 and 97%, for Dequest 2010 and Dequest 2041, respectively, provided evidence that a complete breakdown of these compounds with respect to the C-P bond cleavage can be achieved by some bacteria. The results suggest that phosphonate-utilizing bacteria are ubiquitous, and that selected strains can degrade phosphonates that are more complex than those described previously.
Incidence of urea-hydrolyzing Vibrio parahaemolyticus in Willapa Bay, Washington
Authors: Kaysner C A , Abeyta C , Stott R F , Lilja J L , Wekell M M
Published: January 2020

A high incidence (71.5%) of Vibrio parahaemolyticus was found in samples of water, oysters, and sediment from a Washington State estuary which produces a significant amount of commercial product. Strains of V. parahaemolyticus capable of hydrolyzing urea comprised 58.4% of all V. parahaemolyticus isolates tested. Values for fecal coliforms were within certification criteria for commercial harvest and were not correlated with levels of V. parahaemolyticus.
Production of the Antibiotic Phenazine-1-Carboxylic Acid by Fluorescent
Pseudomonas
Species in the Rhizosphere of Wheat
Authors: Thomashow Linda S. , Weller David M. , Bonsall Robert F. , Pierson Leland S.
Published: January 2020

Pseudomonas fluorescens
2-79 and
P. aureofaciens
30-84 produce the antibiotic phenazine-1-carboxylic acid and suppress take-all, an important root disease of wheat caused by
Gaeumannomyces graminis
var.
tritici.
To determine whether the antibiotic is produced in situ, wheat seeds were treated with strain 2-79 or 30-84 or with phenazine-nonproducing mutants or were left untreated and then were sown in natural or steamed soil in the field or growth chamber. The antibiotic was isolated only from roots of wheat colonized by strain 2-79 or 30-84 in both growth chamber and field studies. No antibiotic was recovered from the roots of seedlings grown from seeds treated with phenazine-nonproducing mutants or left untreated. In natural soils, comparable amounts of antibiotic (27 to 43 ng/g of root with adhering soil) were recovered from roots colonized by strain 2-79 whether or not the pathogen was present. Roots of plants grown in steamed soil yielded larger bacterial populations and more antibiotic than roots from natural soils. In steamed and natural soils, roots from which the antibiotic was recovered had significantly less disease than roots with no antibiotic, indicating that suppression of take-all is related directly to the presence of the antibiotic in the rhizosphere.
Construction of lactose-utilizing Xanthomonas campestris and production of xanthan gum from whey
Authors: Fu J F , Tseng Y H
Published: January 2020

Xanthomonas campestris pv. campestris possesses a low level of beta-galactosidase and therefore is not able to grow and produce significant amounts of xanthan gum in a medium containing lactose as the sole carbon source. In this study, a beta-galactosidase expression plasmid was constructed by ligating an X. campestris phage phi LO promoter with pKM005, a ColE1 replicon containing Escherichia coli lacZY genes and the lpp ribosome-binding site. It was then inserted into an IncP1 broad-host-range plasmid, pLT, and subsequently transferred by conjugation to X. campestris 17, where it was stably maintained. The lacZ gene under the control of the phage promoter was expressed at a high level, enabling the cells to grow in a medium containing lactose. Production of xanthan gum in lactose or diluted whey by the engineered strain was evaluated, and it was found to produce as much xanthan gum in these substrates as the cells did in a medium containing glucose.
Salmonella-TEK, a rapid screening method for Salmonella species in food
Authors: Van Poucke L S
Published: January 2020

A micro-enzyme-linked immunosorbent assay (micro-ELISA) using the Salmonella-TEK screen kit was tested for the detection of Salmonella spp. in pure cultures as well as in 30 artificially contaminated food samples and in 45 naturally contaminated food samples. Different raw, fleshy foods and processed foods were used as test products. The artificially contaminated minced meat samples were preenriched in buffered peptone water, and after incubation, different selective enrichment broths were tested. The micro-ELISA optical density values after enrichment and isolation of the different broths were very analogous. The quickest method to detect Salmonella spp. in different foods is to enrich them with Salmosyst broth, which reduces the total analysis time to 31 h. The Salmonella-TEK kit for Salmonella spp. provides a promising test for the detection of Salmonella antigens in food even when they are present at a low concentration (1 to 5 CFU/25 g). The cross-reaction of the anti-Salmonella antibodies, especially to other gram-negative bacteria, is nil.
Identification and localization of 3-phenylcatechol dioxygenase and 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate hydrolase genes of Pseudomonas putida and expression in Escherichia coli
Authors: Khan A A , Walia S K
Published: January 2020

The bphC and bphD genes of Pseudomonas putida involved in the catabolism of polychlorinated biphenyls or biphenyl were identified, localized, and studied for expression in Escherichia coli. This was achieved by cloning a 2.4-kilobase (kb) DNA fragment of recombinant cosmid pOH101 into HindIII site of pUC plasmids downstream of a lacZ promoter and measuring the enzyme activities of 3-phenylcatechol dioxygenase (3-PDase; a product of bphC) and the meta-cleavage product 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate hydrolase (a product of bphD). The amount of 3-PDase produced in E. coli was about 20 times higher than that of the enzyme produced by the parent, P. putida. Determination of expression of the bphC and bphD genes through their own promoter sequences or by using the lacZ promoter of pUC plasmids was done by cloning the DNA that encodes bphC and bphD genes in a HindIII site of a promoter selection vector (pKK232-8) upstream of the gene for chloramphenicol acetyltransferase (CAT). The recombinant plasmid (pAW787) constructed by inserting the 2.4-kb DNA in pKK232-8 expressed both 3-PDase and CAT activities. Another hybrid construct (pAW786) in which the DNA insert was cloned in the opposite orientation lacked CAT activity but produced normal amounts of 3-PDase activity. On the basis of these results, we suggest that the bphC and bphD genes were expressed by using promoter sequences that are independent of the promoter that expresses CAT activity in E. coli. The locations of the bphC and bphD genes were determined by insertional inactivation of the open reading frames of structural genes bphC and bphD by Tn5 mutagenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
Evaluation of DNA probes for detection of Shiga-like-toxin-producing Escherichia coli in food and calf fecal samples
Authors: Samadpour M , Liston J , Ongerth J E , Tarr P I
Published: January 2020

The use of DNA probes for Shiga-like toxin I (SLT-I) and SLT-II for detection of SLT-producing Escherichia coli (SLTEC) in foods and calf fecal samples was evaluated. Enrichment cultures were prepared from food or fecal samples. Colonies formed by plating of enrichment cultures were probed for SLTEC by colony hybridization. Alternatively, enrichment cultures were analyzed for SLTEC presence by dot blot. The lowest detected concentration of SLTEC in sample homogenates inoculated with E. coli O157:H7 corresponded to 1.3 CFU/g of sample. Of the 44 food samples and 28 fecal samples from dairy calves tested by the colony hybridization method, 4 food samples, including ground beef, raw goat milk, blueberries, and surimi-based delicatessen salad, and 9 calf fecal samples were positive with the SLT probes. The dot blot technique yielded results within 48 h and can be used as a fast and sensitive method of detection for SLTEC in foods and calf fecal samples. The colony hybridization technique took 3 to 4 days but permits recovery of the positive colonies when desired.
Cellulose digestion and cellulase regulation and distribution in Fibrobacter succinogenes subsp. succinogenes S85
Authors: Huang L , Forsberg C W
Published: January 2020

Fibrobacter succinogenes subsp. succinogenes S85 initiated growth on microcrystalline cellulose without a lag whether inoculated from a glucose, cellobiose, or cellulose culture. During growth on cellulose, there was no accumulation of soluble carbohydrate. When the growth medium contained either glucose or cellobiose in combination with microcrystalline cellulose, there was a lag in cellulose digestion until all of the soluble sugar had been utilized, suggesting an end product feedback mechanism that affects cellulose digestion. Cl-stimulated cellobiosidase and periplasmic cellodextrinase were produced under all growth conditions tested, indicating constitutive synthesis. Both cellobiosidases were cell associated until the stationary phase of growth, whereas proteins antigenically related to the Cl-stimulated cellobiosidase and a proportion of the endoglucanase were released into the extracellular culture fluid during growth, irrespective of the substrate. Immunoelectron microscopy of cells with a polyclonal antibody to Cl-stimulated cellobiosidase as the primary antibody and 10-nm-diameter gold particles conjugated to goat anti-rabbit antibodies as the second antibody revealed protrusions of the outer surface which were selectively labeled with gold, suggesting that Cl-stimulated cellobiosidase was located on the protrusions. These data support the contention that the protrusions have a role in cellulose hydrolysis; however, this interpretation is complicated by reactivity of the antibodies with a large number of other proteins that possess related antigenic epitopes.
DNA Probes Show Genetic Variation in Cyanobacterial Symbionts of the
Azolla
Fern and a Closer Relationship to Free-Living
Nostoc
Strains than to Free-Living
Anabaena
Strains
Authors: Plazinski Jacek , Zheng Qi , Taylor Rona , Croft Lynn , Rolfe Barry G. , Gunning Brian E. S.
Published: January 2020

Twenty-two isolates of
Anabaena azollae
derived from seven
Azolla
species from various geographic and ecological sources were characterized by DNA-DNA hybridization. Cloned DNA fragments derived from the genomic sequences of three different
A. azollae
isolates were used to detect restriction fragment length polymorphism among all symbiotic anabaenas. DNA clones were radiolabeled and hybridized against southern blot transfers of genomic DNAs of different isolates of
A. azollae
digested with restriction endonucleases. Eight DNA probes were selected to identify the
Anabaena
strains tested. Two were strain specific and hybridized only to
A. azollae
strains isolated from
Azolla microphylla
or
Azolla caroliniana.
One DNA probe was section specific (hybridized only to anabaenas isolated from
Azolla
ferns representing the section
Euazolla
), and five other probes gave finer discrimination among anabaenas representing various ecotypes of
Azolla
species. These cloned genomic DNA probes identified 11 different genotypes of
A. azollae
isolates. These included three endosymbiotic genotypes within
Azolla filiculoides
species and two genotypes within both
A. caroliniana
and
Azolla pinnata
endosymbionts. Although we were not able to discriminate among anabaenas extracted from different ecotypes of
Azolla nilotica, Azolla mexicina, Azolla rubra
and
Azolla microphylla
species, each of the endosymbionts was easily identified as a unique genotype. When total DNA isolated from free-living
Anabaena
sp. strain PCC7120 was screened, none of the genomic DNA probes gave detectable positive hybridization. Total DNA of
Nostoc cycas
PCC7422 hybridized with six of eight genomic DNA fragments. These data imply that the dominant symbiotic organism in association with
Azolla
spp. is more closely related to
Nostoc
spp. than to free-living
Anabaena
spp.
Evaluation and Reliability of a Simplified Method for Identification of Food-Borne Yeasts
Authors: Rohm Harald , Lechner Frieda
Published: January 2020

A simplified identification key described by Deak and Beuchat (T. Deak and L. R. Beuchat, J. Food Prot. 50:243-264, 1987) and the computer method of Barnett et al. (J. A. Barnett, R. W. Payne, and D. Yarrow,
Yeast Identification Program
, 1985) were used to identify 12 reference strains and 382 yeasts isolated from cultured milk products. Because the simplified key failed to account for species variability with regard to physiological, morphological, and sexual reproduction characteristics, poor agreement of the identification results was obtained. A reevaluation of the basic theoretical assumptions of the simplified key only confirmed the practical results and indicates that this identification method is unsatisfactory
Enterotoxin production by staphylococci isolated from healthy goats
Authors: Valle J , Gomez-Lucia E , Piriz S , Goyache J , Orden J A , Vadillo S
Published: January 2020

The ability of 342 staphylococcal isolates from different anatomical sites in healthy goats to produce staphylococcal enterotoxins (SE) was investigated. SE were produced by 74.3% of the 70 coagulase-positive strains and by 22% of the coagulase-negative strains studied. Most enterotoxigenic strains were isolated from the skin of udders and teats and from milk. SEC was the SE type most frequently produced, either alone (67.9%) or in combination with others. Five coagulase-negative species not previously reported as SE producers were identified (Staphylococcus chromogenes, S. warneri, S. sciuri, S. saprophyticus, and S. lentus). SEA, SEB, and SEC were detected in the milk of 17 of the 133 healthy goats studied. These results suggest that the goat is an important reservoir of enterotoxigenic staphylococci, most of which produce SEC.
Thymidine uptake, thymidine incorporation, and thymidine kinase activity in marine bacterium isolates
Authors: Jeffrey W H , Paul J H
Published: January 2020

One assumption made in bacterial production estimates from [3H]thymidine incorporation is that all heterotrophic bacteria can incorporate exogenous thymidine into DNA. Heterotrophic marine bacterium isolates from Tampa Bay, Fla., Chesapeake Bay, Md., and a coral surface microlayer were examined for thymidine uptake (transport), thymidine incorporation, the presence of thymidine kinase genes, and thymidine kinase enzyme activity. Of the 41 isolates tested, 37 were capable of thymidine incorporation into DNA. The four organisms that could not incorporate thymidine also transported thymidine poorly and lacked thymidine kinase activity. Attempts to detect thymidine kinase genes in the marine isolates by molecular probing with gene probes made from Escherichia coli and herpes simplex virus thymidine kinase genes proved unsuccessful. To determine if the inability to incorporate thymidine was due to the lack of thymidine kinase, one organism, Vibrio sp. strain D19, was transformed with a plasmid (pGQ3) that contained an E. coli thymidine kinase gene. Although enzyme assays indicated high levels of thymidine kinase activity in transformants, these cells still failed to incorporate exogenous thymidine into DNA or to transport thymidine into the cells. These results indicate that the inability of certain marine bacteria to incorporate thymidine may not be solely due to the lack of thymidine kinase activity but may also be due to the absence of thymidine transport systems.
Purification and characterization of purine nucleoside phosphorylase from Proteus vulgaris
Authors: Surette M , Gill T , MacLean S
Published: January 2020

Purine nucleoside phosphorylase was isolated and purified from cell extracts of Proteus vulgaris recovered from spoiling cod fish (Gadus morhua). The molecular weight and isoelectric point of the enzyme were 120,000 +/- 2,000 and pH 6.8. The Michaelis constant for inosine as substrate was 3.9 x 10(-5). Guanosine also served as a substrate (Km = 2.9 x 10(-5). However, the enzyme was incapable of phosphorylizing adenosine. Adenosine proved to be useful as a competitive inhibitor and was used as a ligand for affinity chromatography of purine nucleoside phosphorylase following initial purification steps of gel filtration and ion-exchange chromatography.
Nonradioactive Screening Method for Isolation of Disease-Specific Probes To Diagnose Plant Diseases Caused by Mycoplasmalike Organisms
Authors: Lee Ing-Ming , Davis Robert E. , DeWitt Natalie D.
Published: January 2020

DNA fragments of tomato big bud (BB) mycoplasmalike organism (MLO) in diseased periwinkle plants (
Catharanthus roseus
L.) were cloned to pSP6 plasmid vectors and amplified in
Escherichia coli
JM83. A nonradioactive method was developed and used to screen for MLO-specific recombinants. Cloned DNA probes were prepared by nick translation of the MLO recombinant plasmids by using biotinylated nucleotides. The probes all hybridized with nucleic acid from BB MLO-infected, but not healthy, plants. Results from dot hybridization analyses indicated that several MLOs, e.g., those of Italian tomato big bud, periwinkle little leaf, and clover phyllody, are closely related to BB MLO. The Maryland strain of aster yellows and maize bushy stunt MLOs are also related to BB MLO. Among the remaining MLOs used in this study,
Vinca
virescence and elm yellows MLOs may be very distantly related, if at all, to BB MLO. Potato witches' broom, clover proliferation, ash yellows, western X, and Canada X MLOs are distantly related to BB MLO. Southern hybridization analyses revealed that BB MLO contains extrachromosomal DNA that shares sequence homologies with extrachromosomal DNAs from aster yellows and periwinkle little leaf MLOs.
Effects of propionate and acetate additions on solvent production in batch cultures of Clostridium acetobutylicum
Authors: Hüsemann M H , Papoutsakis E T
Published: January 2020

Addition of acetate or propionate to uncontrolled-pH batch cultures does not affect the initiation of solventogenesis but does enhance final solvent concentrations compared with those of unchallenged cultures. This observation can be explained in terms of the increased buffering capacity of the medium brought about by the added acids, resulting in protection against premature growth inhibition due to low culture pH values at the end of the fermentation. The uptake of propionic acid from the medium does not proceed solely via the coenzyme A-transferase pathway, since less acetone than propanol is formed. Therefore, at least 50% of the propionic acid is taken up through the reversed kinase-phosphotransbutyrylase reaction pathway.
Occurrence of beta-glutamate, a novel osmolyte, in marine methanogenic bacteria
Authors: Robertson D E , Roberts M F , Belay N , Stetter K O , Boone D R
Published: January 2020

The unusual compound beta-aminoglutaric acid (beta-glutamate) has been identified by 13C nuclear magnetic resonance spectroscopy in soluble extracts of marine methanogenic bacteria. We examined several methanogen species representing nine genera and found that beta-glutamate occurred in methanococci and two methanogenium strains (Methanogenium cariaci JR1 and "Methanogenium anulus" AN9). The presence of this compound in the methanococci examined was further restricted to thermophilic members of the genus Methanococcus, including Methanococcus thermolithotrophicus strains, Methanococcus jannaschii, and "Methanococcus igneus." The two Methanogenium strains examined were mesophiles. Studies using Methanococcus thermolithotrophicus showed that levels of beta-glutamate in cells of that species were not affected by variation in growth temperature (40 to 65 degrees C), NH4+ (2 to 80 mM), Mg2+ (10 to 50 mM), or K+ (2 to 10 mM) in the medium. In contrast, soluble pools of beta-glutamate and L-alpha-glutamate (the other major free amino acid in all the methanococci) were proportional to NaCl levels in the growth medium. This dependence of beta-glutamate and L-alpha-glutamate concentrations on salt levels in the medium suggests that they function as osmolytes in these cells.
Supplemental Substrate Enhancement of 2,4-Dinitrophenol Mineralization by a Bacterial Consortium
Authors: Hess T. F. , Schmidt S. K. , Silverstein J. , Howe B.
Published: January 2020

A
Janthinobacterium
sp. and an actinomycete, both capable of mineralizing 2,4-dinitrophenol (DNP), were used to construct a consortium to mineralize DNP in nonaxenic bench-scale sequencing batch reactors (SBRs). Average

K
m

values for DNP mineralization by pure cultures of the
Janthinobacterium
sp. and the actinomycete were 0.01 and 0.13 μg/ml, respectively, and the average maximum specific growth rate (μ
max
) values for them were 0.06 and 0.23/h, respectively. In the presence of NH
4
Cl, nitrite accumulation in pure culture experiments and in the SBRs was stoichiometric to initial DNP concentration and the addition of nitrogen enhanced DNP mineralization in the SBRs. Mineralization of 10 μg of DNP per ml was further enhanced in SBRs by the addition of glucose at concentrations of 100 and 500 μg/ml but not at 10 μg/ml. Possible mechanisms for this enhanced DNP mineralization in SBRs were suggested by kinetic analyses and biomass measurements. Average μ
max
values for DNP mineralization in the presence of 0, 10, 100, and 500 μg of glucose per ml were 0.33, 0.13, 0.42, and 0.59/h, respectively. In addition, there was greater standing biomass in reactors amended with glucose. At steady-state operation, all SBRs contained heterogeneous microbial communities but only one organism, an actinomycete, that was capable of mineralizing DNP. This research demonstrates the usefulness of supplemental substrates for enhancing the degradation of toxic chemicals in bioreactors that contain heterogeneous microbial communities.
Cloning and expression of Clostridium acetobutylicum ATCC 824 acetoacetyl-coenzyme A:acetate/butyrate:coenzyme A-transferase in Escherichia coli
Authors: Cary J W , Petersen D J , Papoutsakis E T , Bennett G N
Published: January 2020

Coenzyme A (CoA)-transferase (acetoacetyl-CoA:acetate/butyrate:CoA-transferase [butyrate-acetoacetate CoA-transferase] [EC 2.8.3.9]) of Clostridium acetobutylicum ATCC 824 is an important enzyme in the metabolic shift between the acid-producing and solvent-forming states of this organism. The purification and properties of the enzyme have recently been described (D. P. Weisenborn, F. B. Rudolph, and E. T. Papoutsakis, Appl. Environ. Microbiol. 55:323-329, 1989). The genes encoding the two subunits of this enzyme have been cloned by using synthetic oligodeoxynucleotide probes designed from amino-terminal sequencing data from each subunit of the CoA-transferase. A bacteriophage lambda EMBL3 library of C. acetobutylicum DNA was prepared and screened by using these probes. Subsequent subcloning experiments established the position of the structural genes for CoA-transferase. Complementation of Escherichia coli ato mutants with the recombinant plasmid pCoAT4 (pUC19 carrying a 1.8-kilobase insert of C. acetobutylicum DNA encoding CoA-transferase activity) enabled the transformants to grow on butyrate as a sole carbon source. Despite the ability of CoA-transferase to complement the ato defect in E. coli mutants, Southern blot and Western blot (immunoblot) analyses showed that neither the C. acetobutylicum genes encoding CoA-transferase nor the enzyme itself shared any apparent homology with its E. coli counterpart. Polypeptides of Mr of the purified CoA-transferase subunits were observed by Western blot and maxicell analysis of whole-cell extracts of E. coli harboring pCoAT4. The proximity and orientation of the genes suggest that the genes encoding the two subunits of CoA-transferase may form an operon similar to that found in E. coli.(ABSTRACT TRUNCATED AT 250 WORDS)
Degradation of 2-bromobenzoic acid by a strain of Pseudomonas aeruginosa
Authors: Higson F K , Focht D D
Published: January 2020

A strain of Pseudomonas aeruginosa producing 2-bromobenzoic acid, designated 2-BBZA, was isolated by enrichment culture from municipal sewage. It degraded all four 2-halobenzoates as well as certain 3-halo- and dihalobenzoates, though none of the 4-halobenzoates supported growth of this organism. 3-Hydroxybenzoate and 3-chlorocatechol were respective inhibitors of salicylate and catechol oxidation: when each was added separately to resting cells incubated with 2-bromobenzoate, salicylate and catechol were found. Oxygen uptake data suggest that the same dehalogenase may be involved in the oxidation of 2-bromo-, 2-chloro-, and 2-iodobenzoates.
Effect of growth rate and hydrophobicity on bacteria surviving protozoan grazing
Authors: Gurijala K R , Alexander M
Published: January 2020

Measurements were made of the predation by Tetrahymena thermophila on several bacterial species in media containing heat-killed Escherichia coli cells to serve as an alternative prey. If grazing pressure was initially not intense on a mixture of bacterial species, the species that survived protozoan feeding at greater densities were those that grew quickly before the onset of active predation. If members of several species were incubated individually at similar initial densities with actively grazing T. thermophila, some species survived at ca. 10(4)/ml, some survived at ca. 10(2)/ml, and others were eliminated. Members of the first two groups but not the third group were able to multiply in the medium in the absence of the protozoan, but the growth rates in the protozoan-free medium did not correlate with the number of survivors. However, the species that persisted at the higher densities possessed highly hydrophobic cell surfaces. The size of the surviving population of four bacterial species whose growth was prevented by chloramphenicol correlated with the initial cell density that was incubated with T. thermophila. It is concluded that the individual species surviving predation on a mixture of species is related to the capacity of the bacterium to grow, the hydrophobicity of its cell surface, and the population density of the species before the onset of intense grazing.
Xylose uptake by the ruminal bacterium Selenomonas ruminantium
Authors: Williams D K , Martin S A
Published: January 2020

Selenomonas ruminantium HD4 does not use the phosphoenolpyruvate phosphotransferase system to transport xylose (S. A. Martin and J. B. Russell, J. Gen. Microbiol. 134:819-827, 1988). Xylose uptake by whole cells of S. ruminantium HD4 was inducible. Uptake was unaffected by monensin or lasalocid, while oxygen, o-phenanthroline, and HgCl2 were potent inhibitors. Menadione, antimycin A, and KCN had little effect on uptake, and acriflavine inhibited uptake by 23%. Sodium fluoride decreased xylose uptake by 10%, while N,N'-dicyclohexylcarbodiimide decreased uptake by 31%. Sodium arsenate was a strong inhibitor (83%), and these results suggest the involvement of a high-energy phosphate compound and possibly a binding protein in xylose uptake. The protonophores carbonyl cyanide m-chlorophenylhydrazone, 2,4-dinitrophenol, and SF6847 inhibited xylose uptake by 88, 82, and 43%, respectively. The cations Na+ and K+ did not stimulate xylose uptake. The kinetics of xylose uptake were nonlinear, and it appeared that more than one uptake mechanism may be involved or that two proteins (i.e., a binding protein and permease protein) with different affinities for xylose were present. Excess (10 mM) glucose, sucrose, or maltose decreased xylose uptake less than 40%. Uptake was unaffected at extracellular pH values between 6.0 and 8.0, while pH values of 5.0 and 4.0 decreased uptake 28 and 24%, respectively. The phenolic monomers p-coumaric acid and vanillin inhibited growth on xylose and xylose uptake more than ferulic acid did. The predominant end products resulting from the fermentation of xylose were lactate (7.5 mM), acetate (4.4 mM), and propionate (5.1 nM), and the Yxylose was 24.1 g/mol.
Molecular cloning of the 130-kilodalton mosquitocidal delta-endotoxin gene of Bacillus thuringiensis subsp. israelensis in Bacillus sphaericus
Authors: Trisrisook M , Pantuwatana S , Bhumiratana A , Panbangred W
Published: January 2020

A 3.7-kilobase (kb) XbaI fragment harboring the cryIVB gene (L. Thorne, F. Garduno, T. Thompson, D. Decker, M. A. Zounes, M. Wild, A. M. Walfield, and T. J. Pollock, J. Bacteriol. 166:801-811, 1986) which encoded a 130-kilodalton (kDa) mosquitocidal toxin from a 110-kb plasmid of Bacillus thuringiensis subsp. israelensis 4Q2-72 was cloned into pUC12 and transformed into Escherichia coli. The clone with a recombinant plasmid (designated pBT8) was toxic to Aedes aegypti larvae. The fragment (3.7 kb) was ligated into pBC16 (tetracycline resistant [Tcr]) and transformed by the method of protoplast transformation into Bacillus sphaericus 1593 and 2362, which were highly toxic to Anopheles and Culex mosquito larvae but less toxic to Aedes larvae. After cell regeneration on regeneration medium, the Tcr plasmids from transformants (pBTC1) of both strains of B. sphaericus were prepared and analyzed. The 3.7-kb XbaI fragment from the B. thuringiensis subsp. israelensis plasmid was shown to be present by agarose gel electrophoresis and Southern blot hybridization. In addition, B. sphaericus transformants produced a 130-kDa mosquitocidal toxin which was detected by Western (immuno-) blot analysis with antibody prepared against B. thuringiensis subsp. israelensis 130-kDa mosquitocidal toxin. The 50% lethal concentrations of the transformants of strains 1593 and 2362 against A. aegypti larvae were 2.7 X 10(2) and 5.7 X 10(2) cells per ml, respectively. This level of toxicity was comparable to the 50% lethal concentration of B. thuringiensis subsp. israelensis but much higher than that of B. sphaericus 1593 and 2362 (4.7 X 10(4) cells per ml) against A. aegypti larvae.(ABSTRACT TRUNCATED AT 250 WORDS)
Effects of temperature and incubation period on production of fumonisin B1 by Fusarium moniliforme
Authors: Alberts J F , Gelderblom W C , Thiel P G , Marasas W F , Van Schalkwyk D J , Behrend Y
Published: January 2020

The kinetics of the production of fumonisin B1 (FB1) by Fusarium moniliforme MRC 826 in corn cultures was investigated as a function of fungal growth at various incubation temperatures. The growth rate of F. moniliforme, as measured by ergosterol concentration, was higher at 25 degrees C than at 20 degrees C, reaching a stationary phase after 4 to 6 weeks in both cases. FB1 production commenced after 2 weeks during the active growth phase, continued to increase during the stationary phase, and decreased after 13 weeks. The overall maximal yield of FB1 (17.9 g/kg, dry weight) was obtained in corn cultures incubated at 20 degrees C for 13 weeks, but it was not significantly (P greater than 0.05) higher than the maximum yield (16.5 g/kg, dry weight) obtained at 25 degrees C after 11 weeks. However, a significantly (P less than 0.05) higher mean yield was detected at 25 degrees C (9.5 g/kg, dry weight) than at 20 degrees C (8.7 g/kg, dry weight). Production reached a plateau after 7 weeks of incubation at 25 degrees C or 9 weeks of incubation at 20 degrees C. The maximal production of FB1 at 30 degrees C was very low (0.6 g/kg, dry weight). FB1 was also found to be heat stable, as there was no reduction in the FB1 concentration after boiling culture material of F. moniliforme MRC 826.
Construction and Use of a Nonradioactive DNA Hybridization Probe for Detection of
Pseudomonas syringae
pv. Tomato on Tomato Plants
Authors: Cuppels D. A. , Moore R. A. , Morris V. L.
Published: January 2020

Pseudomonas syringae
pv. tomato, the causal agent for bacterial speck of tomato, produces the phytotoxin coronatine. A 5.3-kilobase
XhoI
fragment from the chromosomal region controlling toxin production was cloned into the plasmid pGB2, and the resulting recombinant plasmid, pTPR1, was tested for its ability to serve as a diagnostic probe for
P. syringae
pv. tomato. In a survey of 75 plant-associated bacteria, pTPR1 hybridized exclusively to those strains that produced coronatine. The detection limit for this probe, which was labeled with the Chemiprobe nonradioactive reporter system, was approximately 4 × 10
3
CFU of lesion bacteria. During the 1989 growing season, a total of 258 leaf and fruit lesions from nine tomato fields were screened for
P. syringae
pv. tomato by using pTPR1 and the culture method of detection. The best agreement between the two methods, 90%, occurred early in the season with samples taken from relatively young (5-week-old) plants. Young plants also had a higher percentage of
P. syringae
pv. tomato-positive lesions.
P. syringae
pv. tomato was the only coronatine producer recovered from the nine tomato fields. All 244
P. syringae
pv. tomato strains isolated during this study reacted strongly with the probe. The
P. syringae
pv. tomato population of healthy field tomato leaves was determined by a pTPR1 colony hybridization procedure. Every probe-positive colony that was isolated and characterized was identified as
P. syringae
pv. tomato. The pTPR1 probe should expedite disease diagnosis and facilitate epidemiological studies of this pathogen. It also should aid in screening transplant seedlings for bacterial speck infestation.