bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of Amino Acids and Vitamins in Nutrition of Mesophilic
Methanococcus
spp
Authors: Whitman William B. , Sohn SeeHyang , Kuk SeungUk , Xing RuYe
Published: January 2020

In this study we found that autotrophic methanococci similar to
Methanococcus maripaludis
obtained up to 57% of their cellular carbon from exogenous amino acids. About 85% of the incorporation was into protein. Primarily nonpolar and basic amino acids and glycine were incorporated; only small amounts of acidic and some polar amino acids were taken up. An additional 10% of the incorporation was into the nucleic acid fraction. Because little
14
CO
2
was formed from the
14
C-amino acids, little metabolism of the amino acids occurred. Therefore the growth stimulation by amino acids was probably due to the sparing of anabolic energy requirements. Of the amino acids incorporated, only alanine was also a sole nitrogen source for these methanococci. In contrast,
Methanococcus vannielii
and “
Methanococcus aeolicus
” are autotrophic methanococci which did not incorporate amino acids and did not utilize alanine as a sole nitrogen source. Although glutamine served as a sole nitrogen source for the autotrophic methanococci and
Methanococcus voltae
, a heterotrophic methanococcus, growth was due to chemical deamination in the medium.
M. voltae
requires leucine and isoleucine for growth. However, these amino acids were not significant nitrogen sources, and alanine was not a sole nitrogen source for the growth of
M. voltae.
The branched-chain amino acids were not extensively metabolized by
M. voltae.
Pantoyl lactone and pantoic acid were readily incorporated by
M. voltae.
The intact vitamin pantothenate was neither stimulatory to growth nor incorporated. In conclusion, although amino acids and vitamins are nutritionally important to both autotrophic and heterotrophic methanococci, generally they are not subject to extensive catabolism.
Conjugative mobilization as an alternative vector delivery system for lactic streptococci
Authors: Romero D A , Slos P , Robert C , Castellino I , Mercenier A
Published: January 2020

Due to the current variability in applying polyethylene glycol-mediated protoplast transformation to lactic streptococci, a study was undertaken to assess the feasibility of conjugative mobilization as an alternative method for vector delivery. By using the broad-host-range conjugative plasmid pVA797, the partially homologous cloning vector pVA838 was successfully introduced into various strains of Streptococcus lactis, Streptococcus cremoris, Streptococcus lactis subsp. diacetylactis, Streptococcus thermophilus, and Streptococcus faecalis. Frequencies ranged from 10(-2) to 10(-6) transconjugants per recipient. Both pVA797 and pVA838 were acquired intact, without alteration in functionality. Also, the shuttle vector pSA3, which shares partial homology with pVA797, was mobilized via conjugation. The use of S. lactis LM2301 as the intermediate donor allowed the use of physiologic and metabolic characteristics for recipient differentiation. The construction of a vector containing a "DNA cassette" conferring mobilization and the resolution, segregation, and stability of the cointegrates, pVA797, pVA838, and pSA3, are also reported.
Nuclear magnetic resonance spectroscopy reveals the metabolic origins of proline excreted by an Escherichia coli derivative during growth on [13C]acetate
Authors: Crawford A , Hunter B K , Wood J M
Published: January 2020

We have used 13C nuclear magnetic resonance to monitor acetate metabolism in a proline-overproducing strain of Escherichia coli growing on 13C-labeled acetate. The conversion of 13C-labeled acetate to proline by actively dividing cells was followed in vivo, and the site specificity of the incorporation of the acetate carbons in the proline was determined from spectra of butanol extracts of the growth media. The degree of incorporation of deuterium from partially deuterated water into various sites on the proline was monitored from the beta-deuterium-shifted signals in the 13C spectra. The spectra provide information on the origin of the carbons and the protons during proline biosynthesis.
Transformations of chloroguaiacols, chloroveratroles, and chlorocatechols by stable consortia of anaerobic bacteria
Authors: Neilson A H , Allard A S , Lindgren C , Remberger M
Published: January 2020

Metabolically stable consortia of anaerobic bacteria obtained by enrichment of sediment samples with 3,4,5-trimethoxybenzoate (TMBA), 3,4,5-trihydroxybenzoate (gallate [GA]), or 5-chlorovanillin (CV) were used to study the anaerobic transformation of a series of chloroveratroles, chloroguaiacols, and chlorocatechols used as cosubstrates. Experiments were carried out with growing cultures, and the following pathways were demonstrated for metabolism of the growth substrates: (i) TMBA produced GA, which was further degraded without the formation of aromatic intermediates; (ii) GA formed pyrogallol, which was stable to further transformation; and (iii) CV was degraded by a series of steps involving de-O-methylation, oxidation of the aldehyde group, and decarboxylation to 3-chlorocatechol before ring cleavage. Mono-de-O-methylation of the cosubstrates occurred rapidly in the order 4,5,6-trichloroguaiacol greater than 3,4,5-trichloroguaiacol approximately 3,4,5-trichloroveratrole approximately tetrachloroveratrole greater than tetrachloroguaiacol and was concomitant with degradation of the growth substrates. For the polymethoxy compounds--chloroveratroles, 1,2,3-trichloro-4,5,6-trimethoxybenzene, and 4,5,6-trichlorosyringol--de-O-methylation took place sequentially. The resulting chlorocatechols were stable to further transformation until the cultures had exhausted the growth substrates; selective dechlorination then occurred with the formation of 3,5-dichlorocatechol from 3,4,5-trichlorocatechol and of 3,4,6-trichlorocatechol from tetrachlorocatechol. 2,4,5-, 2,4,6-, and 3,4,5-trichoroanisole and 2,3,4,5-tetrachloroanisole were de-O-methylated, but the resulting chlorophenols were resistant to dechlorination. These results extend those of a previous study with spiked sediment samples and their endogenous microflora and illustrate some of the transformations of chloroguaiacols and chlorocatechols which may be expected to occur in anaerobic sediments.
Competitive Exclusion of Epiphytic Bacteria by Ice
−
Pseudomonas syringae
Mutants
Authors: Lindow Steven E.
Published: January 2020

The growth of ice nucleation-active and near-isogenic ice nucleation-deficient (Ice
−
)
Pseudomonas syringae
strains coexisting on leaf surfaces was examined to determine whether competition was sufficient to account for antagonism of phylloplane bacteria. The ice nucleation frequency spectra of 46 Ice
−
P. syringae
mutants, obtained after mutagenesis with ethyl methanesulfonate, differed both quantitatively and qualitatively, but the mutants could be grouped into four distinct phenotypic classes. The numbers of ice nucleation-active bacteria and ice nuclei active at −5�C were reduced on plants colonized with Ice
−
P. syringae
mutant strains before challenge inoculations with an Ice
+
P. syringae
wild-type strain. Frost injury to plants pretreated with Ice
−
P. syringae
strains was also reduced significantly compared with that to control plants and was correlated with the population size of the Ice
+
P. syringae
strain and with the numbers of ice nuclei active at −5�C. An Ice
−
P. syringae
strain colonized leaves, flowers, and young fruit of pears in field experiments and significantly reduced the colonization of these tissues by Ice
+
P. syringae
strains and
Erwinia amylovora
as compared with untreated trees.
Localizations of aluminum in soybean bacteroids and seeds
Authors: Roth L E , Dunlap J R , Stacey G
Published: January 2020

Aluminum, long known to be detrimental to soybean productivity, was localized in the polyphosphate granules (PPG) of bacteroids in root nodules of soybean plants. By using energy-dispersive X-ray analysis, bacteroids in early infections were shown to have typical PPG constituents. However, in PPG in older infections and after the bacteroids were digested intracellularly, aluminum was also detected. These results indicate that aluminum accumulates in PPG after a period when organisms have been resident in host cells and that high levels of aluminum were present in the bacteroids at the time of their demise. At least some of the aluminum in these laboratory-grown plants could have come from the seeds used.
Effect of Proteolytic Enzymes on Transfection and Transformation of
Streptococcus lactis
Protoplasts
Authors: Woskow Steven A. , Kondo Jeffery K.
Published: January 2020

With both chymotrypsin and mutanolysin used to form protoplasts, consistent transformation frequencies of 10
4
to 10
5
transformants and transfectants per μg of DNA were achieved. The procedure was used to transform protoplasts of
Streptococcus cremoris
CS224 at low frequency (5 transformants per μg of DNA).

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Freeze-Thawing of
Aquaspirillum magnetotacticum
Cells Selectively Releases Periplasmic Proteins
Authors: Paoletti Lawrence C. , Short Kevin A. , Blakemore Nancy , Blakemore Richard P.
Published: January 2020

Cells of the gram-negative bacterium
Aquaspirillum magnetotacticum
, when suspended in buffer and freeze-thawed, produced pinkish orange supernatant fluid. The fluid contained ≤2.0% of total extractable outer membrane component 2-keto-3-deoxyoctonate or of the cytoplasmic membrane marker succinic dehydrogenase. Electrophoretic banding patterns and difference spectra of proteins and hemoproteins released by freeze-thawing cells were distinct from those of membrane-associated substances and similar to those of periplasmic substances obtained by applying conventional fractionation methods to this organism.
Metabolism of Lignin Model Compounds of the Arylglycerol-β-Aryl Ether Type by
Pseudomonas acidovorans
D
3
Authors: Vicu�a R. , Gonz�lez B. , Mozuch M. D. , Kirk T. Kent
Published: January 2020

A natural bacterial isolate that we have classified as
Pseudomonas acidovorans
grows on the lignin model compounds 1-(3,4-dimethoxyphenyl)-2-(2-methoxyphenoxy)propane-1,3-diol (compound 1) and 1-(4-hydroxy-3-methoxyphenyl)-2-(2-methoxyphenoxy)propane-1,3-diol (compound 1′), as well as on the corresponding 1-oxo compounds (2 and 2′) as sole sources of carbon and energy. Metabolic intermediates present in cultures growing on compound 1 included compound 2, 2-methoxyphenol (guaiacol [compound 3]), β-hydroxypro-pioveratrone (compound 4), acetoveratrone (compound 5), and veratric acid (compound 6). Also identified were compounds 1′, 2′, β-hydroxypropiovanillone (compound 4′), and acetovanillone (compound 5′), indicating that 4-O demethylation also occurs. The phenolic intermediates were the same as those found in cultures growing on compound 1′. Compounds 2 and 2′ were in part also reduced to compounds 1 and 1′, respectively. Compound 3 was shown to be derived from the 2-methoxyphenoxy moiety. A suggested degradation scheme is as follows: compound 1→2→(3 + 4)→5→6 (and similarly for 1′). In this scheme, the key reaction is cleavage of the ether linkage between C-2 (C
β
) of the phenylpropane moiety and the 2-methoxyphenoxy moiety in compounds 2 and 2′ (i.e., β-aryl ether cleavage). On the basis of compounds identified, viz., 3 and 4 (4′), cleavage appears formally to be reductive. Because this is unlikely, the initial cleavage products probably were not detected. The implications of these results for the enzyme(s) responsible are discussed.
Kinetics of p-nitrophenol mineralization by a Pseudomonas sp.: effects of second substrates
Authors: Schmidt S K , Scow K M , Alexander M
Published: January 2020

The kinetics of simultaneous mineralization of p-nitrophenol (PNP) and glucose by Pseudomonas sp. were evaluated by nonlinear regression analysis. Pseudomonas sp. did not mineralize PNP at a concentration of 10 ng/ml but metabolized it at concentrations of 50 ng/ml or higher. The Ks value for PNP mineralization by Pseudomonas sp. was 1.1 micrograms/ml, whereas the Ks values for phenol and glucose mineralization were 0.10 and 0.25 micrograms/ml, respectively. The addition of glucose to the media did not enable Pseudomonas sp. to mineralize 10 ng of PNP per ml but did enhance the degradation of higher concentrations of PNP. This enhanced degradation resulted from the simultaneous use of glucose and PNP and the increased rate of growth of Pseudomonas sp. on glucose. The Monod equation and a dual-substrate model fit these data equally well. The dual-substrate model was used to analyze the data because the theoretical assumptions of the Monod equation were not met. Phenol inhibited PNP mineralization and changed the kinetics of PNP mineralization so that the pattern appeared to reflect growth, when in fact growth was not occurring. Thus, the fitting of models to substrate depletion curves may lead to erroneous interpretations of data if the effects of second substrates on population dynamics are not considered.
Anaerobic bacteria that dechlorinate perchloroethene
Authors: Fathepure B Z , Nengu J P , Boyd S A
Published: January 2020

In this study, we identified specific cultures of anaerobic bacteria that dechlorinate perchlorethene (PCE). The bacteria that significantly dechlorinated PCE were strain DCB-1, an obligate anaerobe previously shown to dechlorinate chlorobenzoate, and two strains of Methanosarcina. The rate of PCE dechlorination by DCB-1 compared favorably with reported rates of trichloroethene bio-oxidation by methanotrophs. Even higher PCE dechlorination rates were achieved when DCB-1 was grown in a methanogenic consortium.
Evaluation of the Minitek system for characterization of Bacillus species
Authors: Sullivan N M , Mills D C , Riemann H P , Arnon S S
Published: January 2020

Minitek (BBL Microbiology Systems, Cockeysville, Md.) substrate disks were evaluated as alternatives to conventional tests for the characterization of Bacillus species. Results were compared for 10 reference isolates and 87 isolates from food sources. The overall agreement of results between the Minitek and conventional tests was 92% for reference strains and 86% for food isolates.
Arginine catabolism in Lactobacillus sake isolated from meat
Authors: Montel M C , Champomier M C
Published: January 2020

Lactobacillus sake isolated from meat can hydrolyze arginine via the arginine deiminase pathway. Two enzymes, arginine deiminase and ornithine transcarbamylase, have been revealed by detection of their reaction products, citrulline and ornithine, respectively. The production of citrulline depends on the concentration of glucose in a synthetic medium; it does not occur when the concentration of glucose is 27.5 mM or higher.
Effects of pentachlorophenol and some of its known and possible metabolites on different species of bacteria
Authors: Ruckdeschel G , Renner G , Schwarz K
Published: January 2020

The antibacterial activity of pentachlorophenol and 35 of its known or possible metabolites against 30 different species of bacteria was tested. In comparison with pentachlorophenol, no increase of inhibitory activity was found for any of the chlorinated anisoles tested (except for pentachloroanisole against Streptomyces spp.), 2-chlorophenol, 2,6-dichlorophenol, 2,3,6- and 2,4,6-trichlorophenol, 2,3,5,6-tetrachlorophenol, tetrachloro-1,4- and -1,3-benzenediol (except for the 1,3-isomer against Streptomyces spp.), tetrachloro-1,3-dimethoxybenzene, and tetrachloro-1,3-benzenediol diacetate. Two chlorophenols, five dichlorophenols, four trichlorophenols, two tetrachlorophenols, and tetrachloro-1,2-benzenediol were more active than pentachlorophenol against some, but not all, of the strains tested.
Hemolytic activity of and lethal toxin production by environmental strains of Vibrio parahaemolyticus
Authors: Sarkar B L , Kumar R , De S P , Pal S C
Published: January 2020

Repeated subculturing of Kanagawa-negative strains of Vibrio parahaemolyticus on Wagatsuma agar induced the production of a hemolysin which was not the thermostable direct hemolysin. Crude hemolysin exhibited a 30 to 40% lethal toxicity in mice after intraperitoneal injection. A 21-kilodalton protein band was observed with all the environmental isolates in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Results suggested that a certain percentage of environmental strains of V. parahaemolyticus is responsible for pathogenesis.
Increased Phosphorus Uptake by Wheat and Field Beans Inoculated with a Phosphorus-Solubilizing
Penicillium bilaji
Strain and with Vesicular-Arbuscular Mycorrhizal Fungi
Authors: Kucey R. M. N.
Published: January 2020

Greenhouse and field experiments were conducted to test the effect of a P-solubilizing isolate of
Penicillium bilaji
on the availability of Idaho rock phosphate (RP) in a calcareous soil. Under controlled greenhouse conditions, inoculation of soils with
P. bilaji
along with RP at 45 μg of P per g of soil resulted in plant dry matter production and P uptake by wheat (
Triticum aestivum
) and beans (
Phaseolus vulgaris
) that were not significantly different from the increases in dry matter production and P uptake caused by the addition of 15 μg of P per g of soil as triple superphosphate. Addition of RP alone had no effect on plant growth. Addition of vesicular-arbuscular mycorrhizal fungi was necessary for maximum effect in the sterilized soil in the greenhouse experiment. Under field conditions, a treatment consisting of RP (20 kg of P per ha of soil) plus
P. bilaji
plus straw resulted in wheat yields and P uptake equivalent to increases due to the addition of monoammonium phosphate added at an equivalent rate of P. RP added alone had no effect on wheat growth or P uptake. The results indicate that a biological system of RP solubilization can be used to increase the availability of RP added to calcareous soils.
Plant Cell Wall Carbohydrates as Substrates for
Azospirillum brasiliense
Authors: Myers Mary L. , Hubbell David H.
Published: January 2020

Carbohydrate components (simple sugars and polysaccharides) of cell walls of pearl millet (
Pennisetum americanum
L., cv. Gahi) were studied as potential substrates for the root-associated diazotroph
Azospirillum brasiliense
Sp. 7. Simple sugars were utilized, but no evidence was obtained to support the suggestion that the polysaccharide components tested might serve as substrates for growth following hydrolysis by the associated azospirilla.
Extracellular Enzyme Activities during Lignocellulose Degradation by
Streptomyces
spp.: A Comparative Study of Wild-Type and Genetically Manipulated Strains
Authors: Ramachandra Muralidhara , Crawford Don L. , Pometto Anthony L.
Published: January 2020

The wild-type ligninolytic actinomycete
Streptomyces viridosporus
T7A and two genetically manipulated strains with enhanced abilities to produce a water-soluble lignin degradation intermediate, an acid-precipitable polymeric lignin (APPL), were grown on lignocellulose in solid-state fermentation cultures. Culture filtrates were periodically collected, analyzed for APPL, and assayed for extracellular lignocellulose-catabolizing enzyme activities. Isoenzymes were analyzed by polyacrylamide gel electrophoresis and activity staining on the gels. Two APPL-overproducing strains, UV irradiation mutant T7A-81 and protoplast fusion recombinant SR-10, had higher and longer persisting peroxidase, esterase, and endoglucanase activities than did the wild-type strain T7A. Results implicated one or more of these enzymes in lignin solubilization. Only mutant T7A-81 had higher xylanase activity than the wild type. The peroxidase was induced by both lignocellulose and APPL. This extracellular enzyme has some similarities to previously described ligninases in fungi. This is the first report of such an enzyme in
Streptomyces
spp. Four peroxidase isozymes were present, and all catalyzed the oxidation of 3,4-dihydroxyphenylalanine, while one also catalyzed hydrogen peroxide-dependent oxidation of homoprotocatechuic acid and caffeic acid. Three constitutive esterase isozymes were produced which differed in substrate specificity toward α-naphthyl acetate and α-naphthyl butyrate. Three endoglucanase bands, which also exhibited a low level of xylanase activity, were identified on polyacrylamide gels as was one xylanase-specific band. There were no major differences in the isoenzymes produced by the different strains. The probable role of each enzyme in lignocellulose degradation is discussed.
Clostridium acetobutylicum
Mutants That Produce Butyraldehyde and Altered Quantities of Solvents
Authors: Rogers Palmer , Palosaari Neil
Published: January 2020

Spontaneous mutants of
Clostridium acetobutylicum
NRRL B643 that were resistant to allyl alcohol (AA) were selected and characterized. These mutants contained 10- to 100-fold reduced activities of butanol and ethanol alcohol dehydrogenase. The AA mutants formed two groups and produced no ethanol. Type 1 AA mutants produced significant amounts of a new solvent, butyraldehyde, and contained normal levels of the coenzyme A-dependent butyraldehyde dehydrogenase (BAD). Type 2 AA mutants produced no significant butyraldehyde and lower levels of all solvents, and they contained 45- to 100-fold lower activity levels of BAD. Following ethyl methanesulfonate mutagenesis, low-acid-producing (Acid
−
) mutants were selected and characterized as superinduced solvent producers, yielding more than 99% of theoretical glucose carbon as solvents and only small amounts of acetate and butyrate. Following ethyl methanesulfonate mutagenesis, 13 sporulation-negative (Spo
−
) mutants were characterized; and 3 were found to produce only butyrate and acetate, a minor amount of acetone, and no alcohols. These Spo
−
mutants contained reduced butanol dehydrogenase activity and no BAD enzyme activity. The data support the view that the type 2 AA, the Acid
−
, and the Spo
−
mutants somehow alter normal regulated expression of the solvent pathway in
C. acetobutylicum
.
Quantitative assessment of vaginal microflora during use of tampons of various compositions
Authors: Onderdonk A B , Zamarchi G R , Rodriguez M L , Hirsch M L , Muñoz A , Kass E H
Published: January 2020

Although the effect of vaginal tampons on the microbial flora during menstruation has recently been studied by several investigators, quantitative effects attributable to particular tampon fibers have received less attention. The purposes of the present study were (i) to determine and then to compare the effects of polyacrylate rayon tampons and viscose rayon tampons on the normal vaginal flora, (ii) to compare quantitative bacterial counts obtained from these tampons with those obtained from concomitant vaginal swabs, and (iii) to determine whether either of these tampon types alters the vaginal microflora when compared with the microflora in the same women using all-cotton tampons or external catamenial pads. Tampon and swab samples were obtained at predetermined times from 18 women for an average of seven menstrual cycles. Samples consisting of swabs from women wearing menstrual pads were compared with swab and tampon samples taken at predetermined times during the menstrual cycle from women using cotton, polyacrylate rayon, or viscose rayon tampons. Samples were analyzed for total aerobic, facultative, and anaerobic bacterial counts. Statistical evaluation of the results indicated that, on the whole, total bacterial counts decreased during menstruation and that the numbers of bacteria in tampons tended to be lower than those in swab samples taken at the same time. The tampon type had little effect on the vaginal microflora.
Identification and Characterization of Rhizosphere-Competent Bacteria of Wheat
Authors: Juhnke M. Elliott , Mathre D. E. , Sands D. C.
Published: January 2020

To obtain rhizosphere-competent bacteria which could subsequently be modified for the development of biological control agents, bacteria were isolated from the rhizosphere and rhizoplane of wheat and barley plants by standard techniques. Of these isolates, 60 were selected for field testing as spring wheat seed inoculants in 1985. Isolates were marked genetically for resistance to antibiotics via selection of spontaneous mutants to detect and monitor isolates in the field. Forty-three days after planting, the average log
10
CFU/mg (dry weight) of roots and rhizosphere soil for the mutant isolates sampled ranged from 0 to 3.4. Twenty mutant isolates were retested in 1986. A total of 4 isolates were not detected, but the other 16 had an average root colonization value of log
10
2.1 CFU and a range of log
10
0.9 CFU to log
10
3.2 CFU when sampled 32 days after planting. The average colonization value dropped to log
10
1.1 CFU 51 days later. Some isolates detected previously were not detected in the second sampling; others had root colonization values similar to those obtained in the first sampling. Mutant isolates of rhizosphere bacteria included
Bacillus pumilus, Bacillus subtilis, Pseudomonas fluorescens, Streptomyces
spp.,
Xanthomonas maltophilia
, and a saprophytic coryneform. Mixtures of isolates from different genera and species were compatible on seeds and roots.
Use of oligonucleotide probes to study the relatedness of delta-endotoxin genes among Bacillus thuringiensis subspecies and strains
Authors: Prefontaine G , Fast P , Lau P C , Hefford M A , Hanna Z , Brousseau R
Published: January 2020

Fifteen Bacillus thuringiensis strains representing 13 serotypes were screened with five oligodeoxyribonucleotide probes specific for certain regions of two published sequences and one unpublished sequence of B. thuringiensis delta-endotoxin genes. Of the 15 cultures, 14 hybridized with at least one probe; the B. thuringiensis subsp. thompsoni strain alone did not hybridize. Two B. thuringiensis subsp. kurstaki strains of commercial interest, HD-1 and NRD-12, were found to be so closely related as to be indistinguishable with this technique; the same situation was found with strains from B. thuringiensis subspp. dendrolimus and sotto. Five strains were identified as probably containing only one endotoxin gene. A probe specific for the gene from the B. thuringiensis subsp. kurstaki HD-73 strain hybridized to only 3 of the 15 cultures tested. The hybridization data suggest that the DNA sequences coding for the C-terminal region of the endotoxin protein are as well conserved as those coding for the N-terminal toxic portion.
Preparation and characterization of the deepoxy trichothecenes: deepoxy HT-2, deepoxy T-2 triol, deepoxy T-2 tetraol, deepoxy 15-monoacetoxyscirpenol, and deepoxy scirpentriol
Authors: Swanson S P , Rood H D , Behrens J C , Sanders P E
Published: January 2020

The production of deepoxy metabolites of the trichothecene mycotoxins T-2 toxin and diacetoxyscirpenol, including deepoxy HT-2 (DE HT-2), deepoxy T-2 triol, deepoxy T-2 tetraol, deepoxy 15-monoacetoxyscirpenol, and deepoxy scirpentriol is described. The metabolites were prepared by in vitro fermentation with bovine rumen microorganisms under anaerobic conditions and purified by normal and reverse-phase high-pressure liquid chromatography. Capillary gas chromatographic retention times and mass spectra of the derivatized metabolites were obtained. The deepoxy metabolites were significantly less toxic to brine shrimp than were the corresponding epoxy analogs. Polyclonal and monoclonal T-2 antibodies were examined for cross-reactivity to several T-2 metabolites. Both HT-2 and DE HT-2 cross-reacted with mouse immunoglobulin monoclonal antibody 15H6 to a greater extent than did T-2 toxin. Rabbit polyclonal T-2 antibodies displayed greater specificity to T-2 toxin compared with the monoclonal antibody, with relative cross-reactivities of only 17.4, 14.6, and 9.2% for HT-2, DE HT-2, and deepoxy T-2 triol, respectively. Cross-reactivity of both antibodies was weak for T-2 triol, T-2 tetraol, 3'OH T-2, and 3'OH HT-2.
Solubilization of Leonardite by an Extracellular Fraction from
Coriolus versicolor
Authors: Pyne John W. , Stewart Dorothy L. , Fredrickson James , Wilson Bary W.
Published: January 2020

Coriolus versicolor
has previously been shown to degrade leonardite, an oxidized form of lignite. An extracellular fraction containing protein purified from a
C. versicolor
culture solubilized leonardite in vitro. Expression of the activity did not require the presence of leonardite and appeared during idiophase. During ion-exchange and gel filtration column chromatography, leonardite-biosolubilizing activity eluted with syringaldazine oxidase activity and with protein, as measured by
A
280
and the biuret protein assay. Syringaldazine is a substrate of the polyphenol oxidase formed by
C. versicolor
. Comparison of leonardite-biosolubilizing activity with the effects of chelators and surface-active agents on leonardite showed that biosolubilization was not due to either surfactant or chelating ability. Heat treatment of the preparation at 60°C for 30 min significantly reduced both syringaldazine oxidase and leonardite-biosolubilizing activities. Cyanide, azide, and thioglycolate, which are known inhibitors of syringaldazine oxidase activity of
C. versicolor
, also inhibited leonardite biosolubilization. From these data, we conclude that the purified protein fraction from
C. versicolor
contains a syringaldazine oxidase activity that participates in leonardite biosolubilization by enzymatic action.
Enumeration of Vibrio cholerae O1 in Bangladesh waters by fluorescent-antibody direct viable count
Authors: Brayton P R , Tamplin M L , Huq A , Colwell R R
Published: January 2020

A field trial to enumerate Vibrio cholerae O1 in aquatic environments in Bangladesh was conducted, comparing fluorescent-antibody direct viable count with culture detection by the most-probable-number index. Specificity of a monoclonal antibody prepared against the O1 antigen was assessed and incorporated into the fluorescence staining method. All pond and water samples yielded higher counts of viable V. cholerae O1 by fluorescent-antibody direct viable count than by the most-probable-number index. Fluorescence microscopy is a more sensitive detection system than culture methods because it allows the enumeration of both culturable and nonculturable cells and therefore provides more precise monitoring of microbiological water quality.
Production Rate of Planktonic Bacteria in the North Basin of Lake Biwa, Japan
Authors: Nagata Toshi
Published: January 2020

Vertical and seasonal variations in the cell number and production rate of planktonic bacteria were investigated at a pelagic site (water depth, ca. 72 m) of the north basin of Lake Biwa during April to October 1986. The [
methyl
-
3
H]thymidine uptake rate into a cold trichloroacetic acid-insoluble fraction and the frequency of dividing cells (FDCs) were measured for each sample as indices of the bacterial production rate. The seasonal data of bacterial number, thymidine uptake rate, and bacterial growth rate based on the FDCs were correlated with one another (rank correlation analysis,
P
< 0.05). These bacterial variables were not correlated positively with the chlorophyll
a
concentration. Vertically, the maxima of both bacterial number and the thymidine uptake rate were found in the euphotic zone. The direct counting of bacteria and the measurements of thymidine uptake rate combined with the size-fractionation method revealed that more than 90% of the bacterial biomass and production rate were attributed to unattached bacteria throughout the investigation period. The carbon flux estimates of bacterial production were less certain due to the variability of the conversion factor for the thymidine uptake method and that of the calibration for the FDC method, but even when the conservative range of bacterial net production rate was used (5 to 60 μg of carbon per liter per day), it can be suggested that bacterial net production in the investigated area was a significant fraction (ca. 30%) of the level of the primary production rate in the same water basin.
Gas chromatography analysis of cellular fatty acids and neutral monosaccharides in the identification of lactobacilli
Authors: Rizzo A F , Korkeala H , Mononen I
Published: January 2020

Cellular fatty acids and monosaccharides in a group of 14 lactobacilli were analyzed by gas chromatography and the identity of the components was confirmed by gas chromatography-mass spectrometry. From the same bacterial sample, both monosaccharides and fatty acids were liberated by methanolysis, and in certain experiments, fatty acids alone were released by basic hydrolysis. The results indicate that basic hydrolysis gave more comprehensive information about the fatty acids, but the analysis of monosaccharides was found to be much more useful in distinguishing between different species of lactobacilli. The method described allowed differentiation of 11 of 14 Lactobacillus species, and even single colonies isolated from agar plates could be used for analysis without subculturing.
Biosynthesis of Indole-3-Acetic Acid by the Pine Ectomycorrhizal Fungus
Pisolithus tinctorius
Authors: Frankenberger W. T. , Poth M.
Published: January 2020

Previous work has indicated that anatomical and morphological changes (stunting and dichotomy) in roots of various conifers may be influenced by plant-growth-regulating substances secreted by mycorrhizae. Indole-3-acetic acid (IAA) has been tentatively identified as a major auxin produced by some selected ectomycorrhizae. We report the isolation and detection of IAA as a secondary metabolite from
Pisolithus tinctorius
by thin-layer chromatography, high-performance liquid chromatography (HPLC), enzyme-linked immunosorbent (monoclonal antibody) assay (ELISA), and unequivocal identification by gas chromatography-mass spectrometry (GC-MS). The thin-layer chromatography methods for auxin isolation described here are novel, with the use of heptane-acetone-glacial acetic acid as the migrating solvent and formaldehyde, H
2
SO
4
, and vanadate in detection. The acidic extract of the culture supernatant was methylated with ethereal diazomethane to detect IAA as methyl-3-IAA by HPLC, ELISA, and GC-MS. The quantitative amount of IAA detected ranged from 4 to 5 μmol liter
−1
by HPLC and ELISA. Another unidentified metabolite was detected by GC-MS with a typical indole nucleus (
m/z
= 130), indicating that it could be an intermediate in auxin metabolism. Plant response (
Pseudotsuga menziesii,
Douglas fir) was monitored upon inoculation of
P. tinctorius
and
l
-tryptophan. There was a consistent increase in plant height and stem diameter as a result of the two treatments, with statistical differences in dry weights of the shoots and roots.
Evidence for Genetic Exchange and Recombination of
Rhizobium
Symbiotic Plasmids in a Soil Population
Authors: Schofield Peter R. , Gibson Alan H. , Dudman William F. , Watson John M.
Published: January 2020

A soil population of 16
Rhizobium leguminosarum
bv.
trifolii
isolates was characterized by using three Sym (for symbiotic) plasmid-specific DNA hybridization probes: (i) an
R. leguminosarum
bv.
trifolii
-specific, repeated-sequence probe; (ii) a
nifHDK
gene probe, and (iii) a
nod
gene probe. A predominant Sym plasmid family was identified among the isolates. Three other unrelated Sym plasmid families were also identified. The isolates were also classified either by using a chromosomal DNA hybridization probe or by serological relatedness to 25 different
R. leguminosarum
bv.
trifolii
antisera. With either method, it was possible to group the 16 soil isolates into identical or related families. However, the correlation between the two techniques was not high. Irrespective of the means used to classify the bacterial host strain, it was possible to identify the same Sym plasmids in unrelated strains, as well as unrelated Sym plasmids in identical host strains. These data indicate that, within this soil population, there has been genetic exchange of Sym plasmids, and in one instance the hybridization pattern indicates that in vivo recombination of two different Sym plasmids may have occurred. Symbiotic effectiveness tests on red, strawberry, and subterranean clovers clearly differentiated the isolates. In general, the pattern of response was similar within groupings on the basis of Sym plasmid and chromosomal profiles but different between such groups.
Variability in Effectiveness of Rhizobia during Culture and in Nodules
Authors: Weaver R. W. , Wright S. F.
Published: January 2020

The ability of three strains of
Bradyrhizobium
sp. (
Vigna
) to fix dinitrogen in symbiotic association with siratro (
Macropitilium atropurpureum
) was measured after culture in broth and after isolation from nodules. Seven transfers were made between the initial broth culture and the final broth culture. A total of 40 single-colony isolates were obtained from cultures 1 and 7 to test effectiveness. Variation in dinitrogen-fixing effectiveness of the population of one strain did not change on culturing, whereas there was considerable variation in effectiveness of populations of the other two strains. Generally, single-colony isolates from individual nodules had similar levels of effectiveness, but some exceptions occurred. Isolates from different nodules formed by the same
Bradyrhizobium
strain often differed in their effectiveness.
Simple method for screening aflatoxin-producing molds by UV photography
Authors: Yabe K , Ando Y , Ito M , Terakado N
Published: January 2020

UV absorption by aflatoxins was monitored in GY agar medium by UV photography. In the UV photographs, aflatoxin-producing molds were identified as gray or black colonies, whereas aflatoxin-nonproducing molds appeared as white colonies. By cellophane transplantation experiments and silica gel thin-layer chromatography, the products absorbing UV light substantially were found to be mainly aflatoxins B1 and G1 excreted from the mold mycelium into the agar medium. UV absorption did not occur when the agar medium contained aflatoxin-noninducible carbon sources instead of glucose. Various inhibitors of aflatoxin production, such as dichlorovos and dimethyl sulfoxide, also decreased the intensity of UV absorption. These results indicate that this technique can be used as a simple, safe, and rapid method of screening aflatoxin-producing molds.
Increased susceptibility and reduced phytoalexin accumulation in drought-stressed peanut kernels challenged with Aspergillus flavus
Authors: Wotton H R , Strange R N
Published: January 2020

Three genotypes of peanut (Arachis hypogaea L.), with ICG numbers 221, 1104, and 1326, were grown in three replicate plots and drought stressed during the last 58 days before harvest by withholding irrigation water. Within each plot there were eight levels of stress ranging from 1.1 to 25.9 cm of water. Kernels harvested from the plots were hydrated to 20% moisture and challenged with Aspergillus flavus. Fungal colonization, aflatoxin content, and phytoalexin accumulation were measured. Fungal colonization of non-drought-stressed kernels virtually ceased by 3 days after inoculation, when the phytoalexin concentration exceeded 50 micrograms/g (fresh weight) of kernels, but the aflatoxin concentration continued to rise exponentially for an additional day. When fungal colonization, aflatoxin production, and phytoalexin accumulation were measured 3 days after drought-stressed material was challenged, the following relationships were apparent. Fungal colonization was inversely related to water supply (r varied from -0.848 to -0.904, according to genotype), as was aflatoxin production (r varied from -0.876 to -0.912, according to genotype); the phytoalexin concentration was correlated with water supply when this exceeded 11 cm (r varied from 0.696 to 0.917, according to genotype). The results are discussed in terms of the critical role played by drought stress in predisposing peanuts to infection by A. flavus and the role of the impaired phytoalexin response in mediating this increased susceptibility.
Evaluation of Slime-Producing Bacteria in Oil Field Core Flood Experiments
Authors: Geesey G. G. , Mittelman M. W. , Lieu V. T.
Published: January 2020

Epifluorescence microscopy and carbohydrate determinations indicated that the decrease in permeability of oil reservoir sand to reclaimed sewage water was partially the result of biological plugging. Filtration and biocide addition studies demonstrated that the increase in bacterial densities and slime concentrations in flooded oil field cores appeared to be due to both deposition from the reclaimed water and in situ microbial growth and slime production. Although these biological components increased throughout the cores during flooding, the region where the water entered the core exhibited the highest cell densities and slime concentrations. The approach described in this report should be useful in predicting the potential of a water source to induce biological plugging of oil reservoir sand.
Selection of Protease-Positive and Protease-Negative Variants of
Streptococcus cremoris
Authors: Hugenholtz Jeroen , Splint Rob , Konings Wil N. , Veldkamp Hans
Published: January 2020

Protease-negative variants were shown to outcompete the wild-type strains of
Streptococcus cremoris
E
8
, HP, and Wg
2
at pH values higher than 6.0 in milk. For
S. cremoris
E
8
this process was studied in more detail. At lower pH values the wild type had a selective advantage. This pH-dependent selection was not found in all media tested. The poor growth of the protease-negative variant at low pH was not due to lower internal pH values. By growing
S. cremoris
E
8
and Wg
2
in acidified milk (pH 5.9) the proteolytic activity of the cultures could be stabilized. In continuous cultures under amino acid limitation the wild type
S. cremoris
E
8
and HP strains had a selective advantage over the protease-negative variants at low dilution rates (
D
< 0.2) at all pH values of the medium. This was apparently due to a lower affinity-constant (

K
s

) of the protease-positive variants for amino acids. Finally, a high fraction of protease-positive variants could be maintained in continuous cultures by using a growth medium with low concentrations of casein as a nitrogen source. At high dilution rates nearly all cells were protease positive.
Microorganisms capable of metabolizing the herbicide metolachlor
Authors: Saxena A , Zhang R W , Bollag J M
Published: January 2020

We screened several strains of microorganisms and microbial populations for their ability to mineralize or transform the herbicide metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)-acetami de] because such cultures would potentially be useful in the cleanup of contaminated sites. Although we used various inocula and enrichment culture techniques, we were not able to isolate microorganisms that could mineralize metolachlor. However, strains of Bacillus circulans, Bacillus megaterium, Fusarium sp., Mucor racemosus, and an actinomycete were found to transform metolachlor. Several metabolites could be determined with high-performance liquid chromatography. The tolerance of the strains to high concentrations of metolachlor was also evaluated for the usefulness of the strains for decontamination. Tolerance of the actinomycete to metolachlor concentrations over 200 ppm (200 micrograms/ml) was low and could not be increased by doubling the sucrose concentration in the growth medium or by using a large biomass as inoculum. However, a Fusarium sp. could grow and transform metolachlor up to a concentration of 300 ppm.
Symbiotic Expression of Cosmid-Borne
Bradyrhizobium japonicum
Hydrogenase Genes
Authors: Lambert Grant R. , Harker Alan R. , Cantrell Michael A. , Hanus F. Joe , Russell Sterling A. , Haugland Richard A. , Evans Harold J.
Published: January 2020

The expression of cosmid-borne
Bradyrhizobium japonicum
hydrogenase genes in alfalfa, clover, and soybean nodules harboring
Rhizobium
transconjugants was studied. Cosmid pHU52 conferred hydrogen uptake (Hup) activity in both free-living bacteria and in nodules on the different plant hosts, although in nodules the instability of the cosmid resulted in low levels of Hup activity. In contrast, cosmid pHU1, which does not confer Hup activity on free-living bacteria, gave a Hup
+
phenotype in nodules on alfalfa and soybean. Nodules formed by
B. japonicum
USDA 123Spc(pHU1) recycled about 90% of nitrogenase-mediated hydrogen evolution. Both subunits of hydrogenase (30- and 60-kilodalton polypeptides) were detected in enzyme-linked immunosorbent assays of bacteroid preparations from nodules harboring
B. japonicum
strains with pHU1 or pHU52. Neither pHU53 nor pLAFR1 conferred detectable Hup activity in either nodules or free-living bacteria. Based on the physical maps of pHU1 and pHU52, it is suggested that a 5.5-kilobase
Eco
RI fragment unique to pHU52 contains a gene or part of a gene required for Hup activity in free-living bacteria but not in nodules. This conclusion is supported by the observation that two Tn
5
insertions in the chromosome of
B. japonicum
USDA 122DES obtained by marker exchange with Tn
5
-mutagenized pHU1 abolished Hup activity in free-living bacteria but not in nodules.
Additional characteristics of one-carbon-compound utilization by Eubacterium limosum and Acetobacterium woodii
Authors: Sharak Genthner B R , Bryant M P
Published: January 2020

Growth characteristics of Eubacterium limosum and Acetobacterium woodii during one-carbon-compound utilization were investigated. E. limosum RF grew with formate as the sole energy source. Formate also replaced a requirement for CO2 during growth with methanol. Growth with methanol required either rumen fluid, yeast extract, or acetate, but their effects were not additive. Cultures were adapted to grow in concentrations of methanol of up to 494 mM. Growth occurred with methanol in the presence of elevated levels of Na+ (576 mM). The pH optima for growth with methanol, H2-CO2, and carbon monoxide were similar (7.0 to 7.2). Growth occurred with glucose at a pH of 4.7, but not at 4.0. The apparent Km values for methanol and hydrogen were 2.7 and 0.34 mM, respectively. The apparent Vmax values for methanol and hydrogen were 1.7 and 0.11 mumol/mg of protein X min-1, respectively. The Ks value for CO was estimated to be less than 75 microM. Cellular growth yields were 70.5, 7.1, 3.38, and 0.84 g (dry weight) per mol utilized for glucose, methanol, CO, and hydrogen (in H2-CO2), respectively. E. limosum was also able to grow with methoxylated aromatic compounds as energy sources. Glucose apparently repressed the ability of E. limosum to use methanol, hydrogen, or isoleucine but not CO. Growth with mixtures of methanol, H2, CO, or isoleucine was not diauxic. The results, especially the relatively high apparent Km values for H2 and methanol, may indicate why E. limosum does not usually compete with rumen methanogens for these energy sources.(ABSTRACT TRUNCATED AT 250 WORDS)
Bacterial Growth in Mixed Cultures on Dissolved Organic Carbon from Humic and Clear Waters
Authors: Tranvik Lars J. , H�fle Manfred G.
Published: January 2020

Interactions between bacterial assemblages and dissolved organic carbon (DOC) from different sources were investigated. Mixed batch cultures were set up with water from a humic and a clear-water lake by a 1:20 dilution of the bacterial assemblage (1.0 μm of prefiltered lake water) with natural medium (sterile filtered lake water) in all four possible combinations of the two waters and their bacterial assemblages. Bacterial numbers and biomass, DOC, thymidine incorporation, ATP, and uptake of glucose and phenol were followed in these cultures. Growth curves and exponential growth rates were similar in all cultures, regardless of inoculum or medium. However, bacterial biomass produced was double in cultures based on water from the humic lake. The fraction of DOC consumed by heterotrophic bacteria during growth was in the same range, 15 to 22% of the total DOC pool, in all cultures. Bacterial growth efficiency, calculated from bacterial biomass produced and DOC consumed, was in the order of 20%. Glucose uptake reached a peak during exponential growth in all cultures. Phenol uptake was insignificant in the cultures based on the clear-water medium, but occurred in humic medium cultures after exponential growth. The similarity in the carbon budgets of all cultures indicated that the source of the bacterial assemblage did not have a significant effect on the overall carbon flux. However, fluxes of specific organic compounds differed, as reflected by glucose and phenol uptake, depending on the nature of the DOC and the bacterial assemblage.