bluuebunny/crossref_metadata_2025 huggingface dataset:
Toxin production by Bacillus cereus dairy isolates in milk at low temperatures
Authors: Christiansson A , Naidu A S , Nilsson I , Wadström T , Pettersson H E
Published: January 2020

A total of 136 strains of Bacillus cereus isolated from milk and cream were evaluated for toxin production based on HeLa S3, Vero, and human embryonic lung (HEL) cell cytotoxicity in vitro. HEL cell monolayers were more susceptible than the other two cell lines. The percentage of isolates exhibiting HEL cytotoxicity was similar (43.0 and 48.4%) when the strains were grown in brain heart infusion broth containing 0.1% glucose (BHIG) at 7 and 24 h, respectively, at 30 degrees C. In milk, only 21.8% of isolates showed HEL cytotoxicity at 7 h, and the number increased significantly to 73.2% at 24 h at 30 degrees C. Further, 102 toxin-positive isolates were acclimatized to grow at 8 degrees C in milk. Ninety-four (92.2%) of the strains produced HEL cytotoxicity of various degrees with no strict correlation to bacterial cell numbers and also elicited vascular permeability reaction in rabbit skin. Under aerated growth conditions (agitation, 200 rpm) B. cereus elicited cytotoxicity in BHIG and in milk at temperatures of 30, 15, and 8 degrees C. However, in nonaerated (stagnant) cultures toxin production was diminished (BHIG) or completely lost (milk) at all temperatures. Toxin production at 8 degrees C was evaluated in two different types of commercial cardboard milk packages by inoculation with a potent toxigenic dairy isolate. No detectable HEL cytotoxicity was observed in milk in any of the packages either at stagnant conditions or during mechanical shaking. However, the same strain produced cytotoxin in whipped cream at 8 degrees C.
Survival of Pseudomonas putida UWC1 containing cloned catabolic genes in a model activated-sludge unit
Authors: McClure N C , Weightman A J , Fry J C
Published: January 2020

The possibility of the accidental or deliberate release of genetically engineered microorganisms into the environment has accentuated the need to study their survival in, and effect on, natural habitats. In this study, Pseudomonas putida UWC1 harboring a non-self-transmissible plasmid, pD10, encoding the breakdown of 3-chlorobenzoate was shown to survive in a fully functioning laboratory-scale activated-sludge unit (ASU) for more than 8 weeks. The ASU maintained a healthy, diverse protozoal population throughout the experiment, and the introduced strain did not adversely affect the functioning of the unit. Although plasmid pD10 was stably maintained in the host bacterium, the introduced strain did not enhance the degradation of 3-chlorobenzoate in the ASU. When reisolated from the ASU, derivatives of strain UWC1 (pD10) were identified which were able to transfer plasmid pD10 to a recipient strain, P. putida PaW340, indicating the in situ transfer of mobilizing plasmids from the indigenous population to the introduced strain. Results from plate filter matings showed that bacteria present in the activated-sludge population could act as recipients for plasmid pD10 and actively expressed genes carried on the plasmid. Some of these activated-sludge transconjugants gave higher rates of 3-chlorobenzoate breakdown than did strain UWC1(pD10) in batch culture.
Model to predict aerial dispersal of bacteria during environmental release
Authors: Knudsen G R
Published: January 2020

Risk assessment for genetically engineered bacteria sprayed onto crops includes determination of off-site dispersal and deposition. The ability to predict microbial dispersal patterns is essential to characterize the uncertainty (risk) associated with environmental release of recombinant organisms. Toward this end, a particle dispersal model was developed to predict recovery of bacteria on fallout plates at various distances and directions about a test site. The microcomputer simulation incorporates particle size distribution, wind speed and direction, turbulence, evaporation, sedimentation, and mortality, with a time step of 0.5 s. The model was tested against data reported from three field applications of nonrecombinant bacteria and two applications of recombinant bacteria. Simulated dispersal of 10(5) particles was compared with reported deposition measurements. The model may be useful in defining appropriate populations of organisms for release, methods of release or application, characteristics of a release site that influence containment or dispersal, and in developing an appropriate sampling methodology for monitoring the dispersal of organisms such as genetically engineered bacteria.
Monohydroxylation of phenol and 2,5-dichlorophenol by toluene dioxygenase in Pseudomonas putida F1
Authors: Spain J C , Zylstra G J , Blake C K , Gibson D T
Published: January 2020

Pseudomonas putida F1 contains a multicomponent enzyme system, toluene dioxygenase, that converts toluene and a variety of substituted benzenes to cis-dihydrodiols by the addition of one molecule of molecular oxygen. Toluene-grown cells of P. putida F1 also catalyze the monohydroxylation of phenols to the corresponding catechols by an unknown mechanism. Respirometric studies with washed cells revealed similar enzyme induction patterns in cells grown on toluene or phenol. Induction of toluene dioxygenase and subsequent enzymes for catechol oxidation allowed growth on phenol. Tests with specific mutants of P. putida F1 indicated that the ability to hydroxylate phenols was only expressed in cells that contained an active toluene dioxygenase enzyme system. 18O2 experiments indicated that the overall reaction involved the incorporation of only one atom of oxygen in the catechol, which suggests either a monooxygenase mechanism or a dioxygenase reaction with subsequent specific elimination of water.
Microbial growth and accumulation in industrial metal-working fluids
Authors: Mattsby-Baltzer I , Sandin M , Ahlström B , Allenmark S , Edebo M , Falsen E , Pedersen K , Rodin N , Thompson R A , Edebo L
Published: January 2020

The dynamics of microbial growth in metal-working fluids (MWF) and the effect of the addition of biocides were studied in large fluid systems, in this case, one central tank which holds 150 m3. In this system, populations of Pseudomonas pseudoalcaligenes (greater than 10(8) CFU/ml) were sustained for a year, although large quantities of biocides were added. Quantitation of 3-OH lauric acid, a marker for many Pseudomonas spp., by gas chromatography indicated that the bacterial biomass exceeded the viable counts by approximately 15 times. Fungi were grown on several occasions, the dominating genera being Fusarium and Candida. Soon after the old MWF was removed and the tank was provided with fresh MWF, which consisted of an emulsion of mineral oil in water, there was a massive growth of P. pseudoalcaligenes that reached levels of greater than 10(8) bacteria per ml. Initially, only low concentrations of other species were found for some weeks. After this period, different enterobacteria and other gram-negative rods often appeared at high concentrations (10(7) and 10(8) bacteria per ml, respectively). Bacteria identified as P. pseudoalcaligenes showed great variation with respect to colony morphology and a certain heterogeneity with respect to biochemical characteristics. Certain bacterial species grew as microcolonies on metal strips immersed in the circulating MWF, but P. pseudoalcaligenes was not recovered from this habitat. The total bacterial count in the air surrounding the machines in the metal-working shop showed an inverse relation to increasing distance from the machine. The concentration of bacteria in the air varied because of the number of machines in use, temperature, and humidity.(ABSTRACT TRUNCATED AT 250 WORDS)
Human origin of Bacteroides fragilis bacteriophages present in the environment
Authors: Tartera C , Lucena F , Jofre J
Published: January 2020

Bacteroides fragilis HSP40 phages have been detected in waters with various levels of fecal contamination of human origin. The average numbers of B. fragilis phages present in sewage water reached 5.3 x 10(3) per 100 ml of water. We found a number 1,000 times lower in a river contaminated with domestic sewage only, in which the levels of fecal coliforms and fecal streptococci were 10,000 times lower than those found in raw sewage. In addition, B. fragilis phages were not found in significant numbers in slaughterhouse wastewaters. They were not present in fecal-polluted waters containing fecal contamination from wildlife only. Although the number of B. fragilis phages present in contaminated waters was lower than the number of coliphages, their presence indicated human fecal contamination. It is also shown that Bacteroides phages are only able to multiply under anaerobic conditions in the presence of nutrients, and they cannot multiply in natural waters and sediments.
Biological degradation of 2,4-dichlorophenoxyacetic acid: chloride mass balance in stirred tank reactors
Authors: Kelly M P , Hallberg K B , Tuovinen O H
Published: January 2020

A mass balance was developed for the degradation of 2,4-dichlorophenoxyacetic acid by a mixed culture. Batch culture experiments showed the degradation to be an acid-producing step. Inorganic chloride concentration consistently correlated with the expected value and with base consumption to maintain a constant pH.
Iron-Binding Compounds Produced by Wood-Decaying Basidiomycetes
Authors: Fekete Frank A. , Chandhoke Vikas , Jellison Jody
Published: January 2020

The chrome azurol-S universal siderophore assay and the rapid paper electrophoresis siderophore assay were used to screen 10 wood-decaying basidiomycete isolates for the formation of iron-chelating compounds. All 10 isolates were positive for chrome azurol-S reactivity on solid plating medium and in liquid cultures, and 9 of the 10 isolates produced fluorescent iron-binding compounds in the paper electrophoresis assay.
Most-probable-number procedures for enumerating ruminal bacteria, including the simultaneous estimation of total and cellulolytic numbers in one medium
Authors: Dehority B A , Tirabasso P A , Grifo A P
Published: January 2020

Based on results from eight experiments, no overall difference was found between roll tube and three- and five-tube most-probable-number (MPN) methods for estimating total numbers of ruminal bacteria. However, standard errors for the replicate means within an experiment were higher with the MPN procedures. Visual growth and pH were the criteria used for scoring the MPN tubes. Total numbers were significantly higher in MPN medium containing 40% ruminal fluid, as compared with a complete medium without ruminal fluid. By using a broth medium containing ball-milled cellulose and soluble carbohydrates as energy sources, it was possible to estimate both total and cellulolytic ruminal bacterial numbers in the same MPN series. Disappearance of cellulose and decrease in pH were used to determine growth. Values did not differ from those obtained in separate MPN assays. By using this method, diurnal changes in total and cellulolytic bacterial numbers were estimated in sheep fed forage or a concentrate-type diet.
Degradation of chlorinated aliphatic hydrocarbons by Methylosinus trichosporium OB3b expressing soluble methane monooxygenase
Authors: Oldenhuis R , Vink R L , Janssen D B , Witholt B
Published: January 2020

Degradation of trichloroethylene (TCE) by the methanotrophic bacterium Methylosinus trichosporium OB3b was studied by using cells grown in continuous culture. TCE degradation was a strictly cometabolic process, requiring the presence of a cosubstrate, preferably formate, and oxygen. M. trichosporium OB3b cells degraded TCE only when grown under copper limitation and when the soluble methane monooxygenase was derepressed. During TCE degradation, nearly total dechlorination occurred, as indicated by the production of inorganic chloride, and only traces of 2,2,2-trichloroethanol and trichloroacetaldehyde were produced. TCE degradation proceeded according to first-order kinetics from 0.1 to 0.0002 mM TCE with a rate constant of 2.14 ml min-1 mg of cells-1. TCE concentrations above 0.2 mM inhibited degradation in cell suspensions of 0.42 mg of cells ml-1. Other chlorinated aliphatics were also degraded by M. trichosporium OB3b. Dichloromethane, chloroform, 1,1-dichloroethane, and 1,2-dichloroethane were completely degraded, with the release of stoichiometric amounts of chloride. trans-1,2-Dichloroethylene, cis-1,2-dichloroethylene, and 1,2-dichloropropane were completely converted, but not all the chloride was released because of the formation of chlorinated intermediates, e.g., trans-2,3-dichlorooxirane, cis-2,3-dichlorooxirane, and 2,3-dichloropropanol, respectively. 1,1,1-Trichloroethane, 1,1-dichloroethylene, and 1,3-dichloropropylene were incompletely converted, and the first compound yielded 2,2,2-trichloroethanol as a chlorinated intermediate. The two perchlorinated compounds tested, carbon tetrachloride and tetrachloroethylene, were not converted.
Microbial Utilization of Naturally Occurring Hydrocarbons at the Guaymas Basin Hydrothermal Vent Site
Authors: Bazylinski Dennis A. , Wirsen Carl O. , Jannasch Holger W.
Published: January 2020

The Guaymas Basin (Gulf of California; depth, 2,000 m) is a site of hydrothermal activity in which petroliferous material is formed by thermal alteration of deposited planktonic and terrestrial organic matter. We investigated certain components of these naturally occurring hydrocarbons as potential carbon sources for a specific microflora at these deep-sea vent sites. Respiratory conversion of [1-
14
C]hexadecane and [1(4,5,8)-
14
C]naphthalene to
14
CO
2
was observed at 4°C and 25°C, and some was observed at 55°C, but none was observed at 80°C. Bacterial isolates were capable of growing on both substrates as the sole carbon source. All isolates were aerobic and mesophilic with respect to growth on hydrocarbons but also grew at low temperatures (4 to 5°C). These results correlate well with previous geochemical analyses, indicating microbial hydrocarbon degradation, and show that at least some of the thermally produced hydrocarbons at Guaymas Basin are significant carbon sources to vent microbiota.
Efficacy of Phospholipid Analysis in Determining Microbial Biomass in Sediments
Authors: Findlay Robert H. , King Gary M. , Watling Les
Published: January 2020

Improvements in the analysis of lipid-bound phosphates resulted in a simplified and sensitive method for determining microbial biomass in sediments. Sensitivity was enhanced over previous methods by use of a dye, malachite green, which when complexed with phosphomolybdate at low pH has a high extinction coefficient (at 610 nm). The use of a persulfate oxidation technique to liberate phosphate from lipids increased the simplicity and safety of the method relative to the traditional perchloric acid digestions. The modified method was both accurate (yielding quantitative recoveries of cells added to sediments) and precise (coefficient of variation of less than 5% for cells and sediments). A comparison with an epifluorescence technique indicated that the analysis of lipid-bound phosphate was more rapid and less tedious and could be successfully applied to a wider variety of sediment types. An estimate of the lipid-bound phosphate-to-carbon conversion factor based on a diverse enrichment culture from sediments suggested that previous factors for pure cultures may have been too low.
Characterization of Large, Autotrophic
Beggiatoa
spp. Abundant at Hydrothermal Vents of the Guaymas Basin
Authors: Nelson Douglas C. , Wirsen Carl O. , Jannasch Holger W.
Published: January 2020

Filamentous bacteria, identified as members of the genus
Beggiatoa
by gliding motility and internal globules of elemental sulfur, occur in massive aggregations at the deep-sea hydrothermal vents of the Guaymas Basin, Gulf of California. Cell aggregates covering the surface of sulfide-emanating sediments and rock chimneys were collected by DS R/V
Alvin
and subjected to shipboard and laboratory experiments. Each sample collected contained one to three discrete width classes of this organism usually accompanied by a small number of “flexibacteria” (width, 1.5 to 4 μm). The average widths of the
Beggiatoa
classes were 24 to 32, 40 to 42, and 116 to 122 μm. As indicated by electron microscopy and cell volume/protein ratios, the dominant bacteria are hollow cells, i.e., a thin layer of cytoplasm surrounding a large central liquid vacuole. Activities of Calvin-cycle enzymes indicated that at least two of the classes collected possess autotrophic potential. Judging from temperature dependence of enzyme activities and whole-cell CO
2
incorporation, the widest cells were mesophiles. The narrowest
Beggiatoa
sp. was either moderately thermophilic or mesophilic with unusually thermotolerant enzymes. This was consistent with its occurrence on the flanks of hot smoker chimneys with highly variable exit temperatures. In situ CO
2
fixation rates, sulfide stimulation of incorporation, and autoradiographic studies suggest that these
Beggiatoa
spp. contribute significantly as lithoautrophic primary producers to the Guaymas Basin vent ecosystems.
Formation of delta 2- and delta 3-cholenoic acids from bile acid 3-sulfates by a human intestinal Fusobacterium strain
Authors: Robben J , Janssen G , Merckx R , Eyssen H
Published: January 2020

We isolated two strains of an unnamed Fusobacterium species from human intestinal microflora, which stereospecifically transformed bile acid 3-sulfates into C-3-unsubstituted, ring A-unsaturated bile acids. Both 3 alpha- and 3 beta-sulfates of 5 beta-bile acids were metabolized to delta 3-5 beta-cholenoic acids; 3 beta-sulfates of 5 alpha-bile acids were converted into a mixture of delta 2-5 alpha-bile acids and 3 alpha-hydroxy-5 alpha-bile acids, whereas 3 alpha-sulfates of 5 alpha-bile acids were left intact. Unsulfated bile acids were not transformed into unsaturated derivatives. These strains differ from previously isolated intestinal bacteria, which desulfated bile acid sulfates without further transformation.
Correlation of nitrogen metabolism with biosurfactant production by Pseudomonas aeruginosa
Authors: Mulligan C N , Gibbs B F
Published: January 2020

A direct relationship between increased glutamine synthetase activity and enhanced biosurfactant production was found in Pseudomonas aeruginosa grown in nitrate and Proteose Peptone media. A chloramphenicol-tolerant strain showed a twofold increase in biosurfactant production and glutamine synthetase activity. Increased ammonium and glutamine concentrations repressed both phenomena.
Semisolid media for isolation of Salmonella spp. from coastal waters
Authors: Perales I , Audicana A
Published: January 2020

The use of two semisolid media (semisolid Rappaport and semisolid Rappaport-Vassiliadis) for the isolation of Salmonella spp. from coastal waters was compared with the use of conventional media. Two hundred and fifty six samples were studied; Salmonella spp. were detected in 83. The semisolid media were the most sensitive, detecting 73 samples instead of the 53 detected by the conventional media (P less than 0.001). The rate of isolation of Salmonella spp. showed an increase of 56.6% when the semisolid media were added to the conventional media, it being possible to detect 54.2% of the total organisms 1 day earlier.
Identification of a Locus Upstream from the Hydrogenase Structural Genes That Is Involved in Hydrogenase Expression in
Bradyrhizobium japonicum
Authors: Novak Patricia D. , Maier Robert J.
Published: January 2020

A locus involved in the expression of the uptake hydrogenase system of
Bradyrhizobium japonicum
was identified adjacent to genes encoding the hydrogenase subunits. A cloned fragment of DNA was used to complement to autotrophy a Hup
−
putative regulatory mutant of
B. japonicum
. The mutant strain lacked hydrogenase activity and synthesized low levels of the large subunit of hydrogenase as determined by Western gels. Tn
5
-induced mutagenesis located the region within the fragment which was necessary for complementation of the mutant phenotype. The locus identified is adjacent to that encoding the small subunit of hydrogenase; its right border is <0.5 kilobase upstream from the hydrogenase transcriptional start site, and its left border is between 1 and 2.5 kilobases from that start site. However, the locus is outside the region previously shown to contain
hup
-related genes of
B. japonicum
. Thus, the localization of this gene describes a previously unidentified
hup
-related gene on a region of DNA not previously shown to contain
hup
-specific DNA.
Action of a Fluoranthene-Utilizing Bacterial Community on Polycyclic Aromatic Hydrocarbon Components of Creosote
Authors: Mueller J. G. , Chapman P. J. , Pritchard P. H.
Published: January 2020

Cultures enriched by serial transfer through a mineral salts medium containing fluoranthene were used to establish a stable, seven-member bacterial community from a sandy soil highly contaminated with coal tar creosote. This community exhibited an ability to utilize fluoranthene as the sole carbon source for growth, as demonstrated by increases in protein concentration and changes in absorption spectra when grown on fluoranthene in liquid culture. Biotransformation of other polycyclic aromatic hydrocarbons (PAHs) was verified by demonstrating their disappearance from an artificial PAH mixture by capillary gas chromatography. When grown on fluoranthene as the sole carbon source and subsequently exposed to fluoranthene plus 16 additional PAHs typical of those found in creosote, this community transformed all PAHs present in this defined mixture. After 3 days of incubation, 13 of the original 17 PAH components were degraded to levels below the limit of detection (10 ng/liter). Continued incubation resulted in extensive degradation of the remaining four compounds. The ability of this community to utilize a high-molecular-weight PAH as the sole carbon source, in conjunction with its ability to transform a diverse array of PAHs, suggests that it may be of value in the bioremediation of environments contaminated with PAHs, such as those impacted by creosote.
Physiological and Environmental Studies of Sclerotium Formation and Maturation in Isolates of
Morchella crassipes
Authors: Volk Thomas J. , Leonard Thomas J.
Published: January 2020

This study provides a set of nutritional and environmental parameters suitable for the growth of morel (
Morchella crassipes
) sclerotia in the laboratory, using a modification of the jar method of Ower et al. (U.S. patent 4,594,809, June 1986). The optimum nutritional and environmental conditions for morel sclerotium formation and maturation as determined in this study consist of a layer of rye grain supplemented with peptone, yeast extract, trace elements, and Casamino Acids overlaid with perforated aluminum foil and covered with a layer of nutrient-poor soil medium in an 8-oz. (ca. 237-ml) glass jar in the dark. We noted that addition of asparagine or aspartic acid as a nitrogen source to the rye also had a beneficial effect on sclerotium formation, while addition of carbon sources had no significant effect.
Mechanism of Proteinase Release from
Lactococcus lactis
subsp.
cremoris
Wg2
Authors: Laan Harry , Konings Wil N.
Published: January 2020

The procedure generally used for the isolation of extracellular, cell-associated proteinases of
Lactococcus lactis
species is based on the release of the proteinases by repeated incubation and washing of the cells in a Ca
2+
-free buffer. For
L. lactis
subsp.
cremoris
Wg2, as many as five incubations for 30 min at 29°C are needed in order to liberate 95% of the proteinase. Proteinase release was not affected by chloramphenicol, which indicates that release is not the result of protein synthesis during the incubations. Ca
2+
inhibited, while ethylene glycol-bis(β-aminoethyl ether)-
N,N,N′,N′
-tetraacetic acid (EGTA) stimulated, proteinase release from the cells. The pH optimum for proteinase release ranged between 6.5 and 7.5, which was higher than the optimum pH of the proteinase measured for casein hydrolysis (i.e., 6.4). Treatment of cells with the serine proteinase inhibitor phenylmethylsulfonyl fluoride prior to the incubations in Ca
2+
-free buffer reduced the release of the proteinase by 70 to 80%. The residual proteinase remained cell associated but could be removed by the addition of active
L. lactis
subsp.
cremoris
Wg2 proteinase. This suggests that proteinase release from cells of
L. lactis
subsp.
cremoris
Wg2 is the result of autoproteolytic activity. From a comparison of the N-terminal amino acid sequence of the released proteinase with the complete amino acid sequence determined from the nucleotide sequence of the proteinase gene, a protein of 180 kilodaltons would be expected. However, a proteinase with a molecular weight of 165,000 was found, which indicated that further hydrolysis had occurred at the C terminus.
In vivo protocol for testing efficacy of hand-washing agents against viruses and bacteria: experiments with rotavirus and Escherichia coli
Authors: Ansari S A , Sattar S A , Springthorpe V S , Wells G A , Tostowaryk W
Published: January 2020

Ten antiseptic formulations, an unmedicated liquid soap, and tap water alone were compared for their capacities to eliminate human rotavirus from the finger pads of adult volunteers; three of the antiseptics, the soap, and the tap water alone were also tested against Escherichia coli. A fecal suspension of virus or bacterium was placed on each finger pad and air dried. The contaminated site was exposed to the test product for 10 s, rinsed in tap water, and dried on a paper towel. The residual virus or bacterium was then eluted. Selected agents were also tested by an analogous whole-hand method by which the entire palm surfaces of both hands were contaminated. Alcohols (70%) alone or with Savlon reduced the virus titer by greater than 99%, whereas the reductions by Proviodine, Dettol, and Hibisol ranged from 95 to 97%. Aqueous solutions of chlorhexidine gluconate were significantly less effective for virus removal or inactivation than 70% alcohol solutions. Furthermore, Savlon in water (1:200) was found to be much less effective in eliminating the virus (80.6%) than the bacterium (98.9%). The tap water alone and the soap reduced the virus titers by 83.6 and 72.5% and the bacterial titers by 90 and 68.7%, respectively. The results of the whole-hand method agreed well with those of the finger pad protocol. We conclude that the finger pad method is a suitable model for testing the in vivo efficacy of hand-washing agents and emphasize the need for using appropriate test viruses and bacteria.
Aromatic hydroxylation and sulfation of 5-hydroxyflavone by Streptomyces fulvissimus
Authors: Ibrahim A R , Abul-Hajj Y J
Published: January 2020

The conversion of 5-hydroxyflavone by various microorganisms was studied. Among them, Streptomyces fulvissimus was the sole microbe which produced a new polar metabolite from 5-hydroxyflavone in addition to 5,4-dihydoxy- and 5,3,4-trihydroxyflavone. The structure of this polar metabolite was determined to be 5,4-dihydroxyflavone-4-sulfate on the basis of mass, infrared, and nuclear magnetic resonance spectroscopies. These results demonstrate that S. fulvissimus catalyzes sulfation at the 4 position of 4,5-dihydroxyflavone.
Cloning and Expression of a
Schwanniomyces occidentalis
α-Amylase Gene in
Saccharomyces cerevisiae
Authors: Wang Tsung Tsan , Lin Long Liu , Hsu Wen Hwei
Published: January 2020

An α-amylase gene (
AMY
) was cloned from
Schwanniomyces occidentalis
CCRC 21164 into
Saccharomyces cerevisiae
AH22 by inserting
Sau
3AI-generated DNA fragments into the
Bam
HI site of YEp16. The 5-kilobase insert was shown to direct the synthesis of α-amylase. After subclones containing various lengths of restricted fragments were screened, a 3.4-kilobase fragment of the donor strain DNA was found to be sufficient for α-amylase synthesis. The concentration of α-amylase in culture broth produced by the
S. cerevisiae
transformants was about 1.5 times higher than that of the gene donor strain. The secreted α-amylase was shown to be indistinguishable from that of
Schwanniomyces occidentalis
on the basis of molecular weight and enzyme properties.
Two Bacillus beta-mannanases having different COOH termini are produced in Escherichia coli carrying pMAH5
Authors: Akino T , Kato C , Horikoshi K
Published: January 2020

The nucleotide sequence was determined for the alkalophilic Bacillus sp. strain AM-001 beta-mannanase gene which produced two beta-mannanases (A and B) in Escherichia coli transformants. The putative beta-mannanase gene was 1,539 base pairs long and encoded a mature beta-mannanase protein of 487 amino acids and a signal peptide of 26 amino acids. The COOH-terminal amino acid of beta-mannanase A is an arginine residue located at amino acid 513 of the deduced amino acid sequence, and that of beta-mannanase B is a valine residue located at amino acid 365. Deletion derivatives having 1,098 base pairs from the ATG start codon maintained the beta-mannanase activity of the encoded polypeptide. However, clones harboring DNA fragments (1,051 base pairs) shorter than the gene which encoded beta-mannanase B (1,095 base pairs) did not exhibit the beta-mannanase activity. The simultaneous production of both beta-mannanases A and B in an E. coli transformant was demonstrated by the maxicell procedure.
Substrate interactions during aerobic biodegradation of benzene
Authors: Arvin E , Jensen B K , Gundersen A T
Published: January 2020

This study dealt with the interactions with benzene degradation of the following aromatic compounds in a mixed substrate: toluene, o-xylene, naphthalene, 1,4-dimethylnaphthalene, phenanthrene, and pyrrole. The experiment was performed as a factorial experiment with simple batch cultures. The effect of two different types of inocula was tested. One type of inoculum was grown on a mixture of aromatic hydrocarbons; the other was grown on a mixture of aromatic hydrocarbons and nitrogen-, sulfur-, and oxygen-containing aromatic compounds (NSO compounds), similar to some of the compounds identified in creosote waste. The culture grown on the aromatic hydrocarbons and NSO compounds was much less efficient in degrading benzene than the culture grown on only aromatic hydrocarbons. The experiments indicated that toluene- and o-xylene-degrading bacteria are also able to degrade benzene, whereas naphthalene-, 1,,4-dimethylnaphthalene-, and phenanthrene-degrading bacteria have no or very little benzene-degrading ability. Surprisingly, the stimulating effect of toluene and o-xylene was true only if the two compounds were present alone. In combination an antagonistic effect was observed, i.e., the combined effect was smaller than the sum from each of the compounds. The reason for this behavior has not been identified. Pyrrole strongly inhibited benzene degradation even at concentrations of about 100 to 200 micrograms/liter. Future studies will investigate the generality of these findings.
Simple Method To Detect β-Galactosidase
Authors: Bhowmik Tarun , Marth Elmer H.
Published: January 2020

A simple and rapid method was developed to detect β-galactosidase by using α- or β-naphthyl-β-
d
-galactopyranoside as substrate and fast garnet GBC as a dye coupler following polyacrylamide gel electrophoresis. This method was specific for β-galactosidase but not for phospho-β-galactosidase.
Plasmid homologies in Edwardsiella ictaluri
Authors: Reid W S , Boyle J A
Published: January 2020

Plasmids from all available non-channel catfish isolates of Edwardsiella ictaluri were classified by gel electrophoresis and hybridization methods. All isolates, regardless of source, contained classes of homologous plasmids with similar but not identical sizes.
Evidence from Liposome Encapsulation for Transport-Limited Microbial Metabolism of Solid Alkanes
Authors: Miller R. M. , Bartha R.
Published: January 2020

The recalcitrance of xenobiotics may be caused by an absence of transforming enzymes or by their inability to enter microbial cells. A nondestructive method for differentiating between these two possibilities is described. The solid
n
-alkanes octadecane (C
18
) and hexatriacontane (C
36
) were encapsulated into phosphatidylcholine bilayers (liposomes). The uptake and metabolism rates of encapsulated and unencapsulated substrates were then compared. During 1 h at 25°C, a
Pseudomonas
isolate took up 1.3% of radiolabeled and unencapsulated C
18
(solid state) versus 23.5% of labeled and encapsulated C
18
. Growth at 25°C occurred with an apparent

k
s

of 2453 ± 148 mg/liter. Liposome encapsulation decreased this

K
s

to 60 ± 12 mg/liter. At 34°C, growth on C
18
(liquid state) occurred with an apparent

K
s

of 819 ± 83 mg/liter and on the readily available carbon source succinate,

K
s

values were 80 ± 10 and 13 ± 7 mg/liter at 25 and 34°C, respectively. At 25°C, the isolate grew on C
36
with an apparent

K
s

of 2,698 ± 831 mg/liter. Liposome encapsulation decreased the

K
s

more than 60-fold to 41 ± 7 mg/liter, resulting in the complete utilization of 400 mg of C
36
per liter in 16 h. Since controls excluded the metabolic utilization of phosphatidylcholine, the results clearly identify transport limitation as the cause for C
36
recalcitrance.
Coenzyme A transferase from Clostridium acetobutylicum ATCC 824 and its role in the uptake of acids
Authors: Wiesenborn D P , Rudolph F B , Papoutsakis E T
Published: January 2020

Coenzyme A (CoA) transferase from Clostridium acetobutylicum ATCC 824 was purified 81-fold to homogeneity. This enzyme was stable in the presence of 0.5 M ammonium sulfate and 20% (vol/vol) glycerol, whereas activity was rapidly lost in the absence of these stabilizers. The kinetic binding mechanism was Ping Pong Bi Bi, and the Km values at pH 7.5 and 30 degrees C for acetate, propionate, and butyrate were, respectively, 1,200, 1,000, and 660 mM, while the Km value for acetoacetyl-CoA ranged from about 7 to 56 microM, depending on the acid substrate. The Km values for butyrate and acetate were high relative to the intracellular concentrations of these species; consequently, in vivo enzyme activity is expected to be sensitive to changes in those concentrations. In addition to the carboxylic acids listed above, this CoA transferase was able to convert valerate, isobutyrate, and crotonate; however, the conversion of formate, n-caproate, and isovalerate was not detected. The acetate and butyrate conversion reactions in vitro were inhibited by physiological levels of acetone and butanol, and this may be another factor in the in vivo regulation of enzyme activity. The optimum pH of acetate conversion was broad, with at least 80% of maximal activity from pH 5.9 to greater than 7.8. The purified enzyme was a heterotetramer with subunit molecular weights of about 23,000 and 25,000.
Bioconversion of Gelatin to Methane by a Coculture of
Clostridium collagenovorans
and
Methanosarcina barkeri
Authors: Jain M. K. , Zeikus J. G.
Published: January 2020

A simple, stable, and transferable coculture of
Clostridium collagenovorans
and
Methanosarcina barkeri
that readily degraded gelatin into methane and carbon dioxide was developed. In monoculture,
C. collagenovorans
fermented all of the amino acids in gelatin except proline into acetate and carbon dioxide as the main products, with hydrogen, isovalerate, and isobutyrate detected in trace amounts (<1 mM). In coculture with
M. barkeri
, gelatin was transformed into methane and carbon dioxide, with varying levels of intermediary acetate formed as a function of incubation time. Various complex proteinaceous polymers could be readily transformed into methane and carbon dioxide at 30 to 40°C by a stable coculture which did not require exogenous growth factor additions. In addition, the coculture was readily transferable and preserved in the viable state for long periods, and methanogenesis could be initiated rapidly without the need for exogenous pH control.
Degradation of p-chlorotoluene by a mutant of Pseudomonas sp. strain JS6
Authors: Haigler B E , Spain J C
Published: January 2020

Pseudomonas sp. strain JS6 grows on chlorobenzene, p-dichlorobenzene, or toluene as a sole source of carbon and energy. It does not grow on p-chlorotoluene (p-CT). Growth on glucose in the presence of p-CT resulted in the accumulation of 4-chloro-2,3-dihydroxy-1-methylbenzene (3-chloro-6-methylcatechol), 4-chloro-2,3-dihydroxy-1-methylcyclohexa-4,6-diene (p-CT dihydrodiol), and 2-methyl-4-carboxymethylenebut-2-en-4-olide (2-methyl dienelactone). Strain JS21, a spontaneous mutant capable of growth on p-CT, was isolated from cultures of strain JS6 after extended exposure to p-CT. In addition to growing on p-CT, JS21 grew on all of the substrates that supported growth of the parent strain, including p-dichlorobenzene, chlorobenzene, benzene, toluene, benzoate, p-hydroxybenzoate, phenol, and ethylbenzene. The pathway for degradation of p-CT by JS21 was investigated by respirometry, isolation of intermediates, and assay of enzymes in cell extracts. p-CT was converted to 3-chloro-6-methylcatechol by dioxygenase and dihydrodiol dehydrogenase enzymes. 3-Chloro-6-methylcatechol underwent ortho ring cleavage catalyzed by a catechol 1,2-dioxygenase to form 2-chloro-5-methyl-cis,cis-muconate, which was converted to 2-methyl dienelactone. A dienelactone hydrolase converted 2-methyl dienelactone to 2-methylmaleylacetic acid. Preliminary results indicate that a change in wild-type induction patterns allows JS21 to grow on p-CT.
Production of 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine from 2',3'-dideoxyuridine and the corresponding purine bases by resting cells of Escherichia coli AJ 2595
Authors: Shirae H , Kobayashi K , Shiragami H , Irie Y , Yasuda N , Yokozeki K
Published: January 2020

A novel microbial method for the production of 2',3'-dideoxynucleosides by transdideoxyribosylation has been developed. By screening microorganisms producing 2',3'-dideoxyadenosine (DDA) from 2',3'-dideoxyuridine (DDU) and adenine, Escherichia coli AJ 2595 was selected as the best producer. Optimal pH and temperature for the DDA-producing reaction were ca. 6.5 and 50 degrees C, respectively. Pi seemed to be an essential factor for the reaction, and its optimal concentration was ca. 25 mM. Moreover, polyethylene glycol had a notable effect on DDA production. Under the best conditions established, 52 mM DDA was obtained from 100 mM DDU and 100 mM adenine after 48 h of incubation from resting cells of E. coli AJ 2595. This strain could also produce 2',3'-dideoxynucleosides, such as 2',3'-dideoxyinosine (DDI), 2',3'-dideoxyguanosine, and 2',3'-dideoxythymidine, from DDU and the corresponding bases. In particular, this strain could produce DDI in high yield (ca. 32 mM from 100 mM DDU and 100 mM hypoxanthine) after 24 h of incubation. However, 2',3'-dideoxycytidine was not produced from DDU and cytosine by resting cells of E. coli AJ 2595.
Effect of Light on the Cell Cycle of a Marine
Synechococcus
Strain
Authors: Armbrust E. V. , Bowen J. D. , Olson R. J. , Chisholm S. W.
Published: January 2020

Light-dependent regulation of cell cycle progression in the marine cyanobacterium
Synechococcus
strain WH-8101 was demonstrated through the use of flow cytometry. Our results show that, similar to eucaryotic cells, marine
Synechococcus
spp. display two gaps in DNA synthesis, at the beginning and at the end of the cell cycle. Progression through each of these gaps requires light, and their durations lengthen under light limitation.
Expression of a cloned Bacillus thuringiensis delta-endotoxin gene in Bacillus subtilis
Authors: Calogero S , Albertini A M , Fogher C , Marzari R , Galizzi A
Published: January 2020

The entire coding region of the Bacillus thurigiensis HD73 crystal protein gene was subcloned from plasmid pJWK20 into the integration vector pUG2-15. This plasmid expresses chloramphenicol resistance when integrated into the Bacillus subtilis chromosome in the outH locus near the recE region. The correct molecular organization of the integrated plasmid was verified by hybridization to Southern blots of chromosomal DNA digests. Production of the toxic crystal protein was monitored at different time points during the life cycle of B. subtilis. Toxicity assays against Anagasta (Ephestia) larvae, direct electron microscopy crystal detection, and immunoblotting assays proved that the expression of the gene in B. subtilis is time regulated and restricted mainly to the sporulation stage. RNase protection experiments defined the transcription initiation start point and the transcription timing. All tests were made in a strain containing one to three copies of the integrated plasmid and in a strain subjected to an amplification regimen.
Survival of
Azorhizobium caulinodans
in the Soil and Rhizosphere of Wetland Rice under
Sesbania rostrata
-Rice Rotation
Authors: Ladha J. K. , Garcia Minviluz , Miyan S. , Padre Agnes T. , Watanabe I.
Published: January 2020

The survival of indigenous and introduced strains of
Azorhizobium caulinodans
in flooded soil and in the rice rhizosphere, where in situ
Sesbania rostrata
was incorporated before the rice crop, is reported. The azorhizobia studied were both root and stem nodulating. In a pot experiment, two crop cycles each of inoculated and noninoculated
Sesbania
-rice were compared with two crop cycles of flooded fallow-rice. In a field experiment, the effect of repeated incorporation of in situ
S. rostrata
in the
Sesbania
-rice sequence was studied. Soils in which inoculated
S. rostrata
was incorporated contained about 3,000 times more azorhizobia than did soils in the flooded fallow treatment and about 50 times more azorhizobia than did soils in the noninoculated
Sesbania
treatment. Azorhizobial numbers in the inoculated
Sesbania
treatment declined toward rice harvest but remained much higher than in the flooded fallow-rice treatment. Repeated incorporation of
S. rostrata
increased the population density of indigenous soil azorhizobia, whereas the population of inoculated strain ORS571 (Str
r
Spc
r
) declined to an undetectable level; this finding suggested low competitiveness by the introduced strain. In the incorporated
Sesbania
treatment, the rice rhizosphere harbored significantly more
A. caulinodans
and supported higher nitrogenase activity per plant than did the rhizosphere of the flooded fallow-rice treatment. Sterile rice seedlings inoculated with
A. caulinodans
showed nitrogenase activity comparable to that of seedlings inoculated with
Azospirillum lipoferum
34H, a rice root isolate. Rhizobia from
Sesbania aculeata, Sesbania sesban
, a
Trifolium
sp., and
Vigna unguiculata
did not support appreciable nitrogenase activity.
Biotin-labeled plasmid DNA probes for detection of epithelium-associated strains of lactobacilli
Authors: Tannock G W
Published: January 2020

Biotin-labeled DNA probes prepared from whole plasmids (5.5 and 4.8 kilobases) of two lactobacillus strains (Lactobacillus delbrueckii 21 and Lactobacillus reuteri 100-23) were used to detect the homologous bacteria in microtome-cryostat-prepared sections of murine forestomach. The forestomach sections were incubated on nylon filter membranes placed on agar plates and, after lysis of the lactobacillus cells and denaturation of their DNA, were used in hybridization experiments with the strain-specific DNA probes. Hybridization of the probes to membranes containing sections from lactobacillus-free mice did not occur. The probes detected the presence of homologous strains of lactobacilli in sections cut from the forestomach of mice harboring one or both of the strains.
Rhizobium
Strain Identification and Quantification in Commercial Inoculants by Immunoblot Analysis
Authors: Olsen Perry E. , Rice Wendell A.
Published: January 2020

An immunoblot procedure for the strain-specific quantitative analysis of commercial
Rhizobium
inoculants was developed. The technique greatly reduced the time required for inoculant analysis. Correlation between immunoblot analysis and traditional plant nodule grow-out most-probable-number techniques was
r
= 0.90 for 16 commercial alfalfa inoculants tested.
Cell-free mercury volatilization activity from three marine caulobacter strains
Authors: Ji G Y , Salzberg S P , Silver S
Published: January 2020

Three mercury-resistant marine Caulobacter strains showed an inducible mercury volatilization activity. Cell-free mercury volatilization (mercuric reductase) from these three marine Caulobacter strains was characterized and compared with enzyme activities determined by plasmids of Escherichia coli and Staphylococcus aureus. The temperature sensitivity of the Caulobacter mercuric reductase was greater than that of mercuric reductase from other gram-negative sources. Cell-free enzyme activity required NADH or NADPH, with NADPH functioning much better at lower concentrations than NADH. The Km for the Caulobacter enzyme was 4 microM Hg2+. Ag+ was a competitive inhibitor of Caulobacter mercuric reductase (Ki = 0.2 microM Ag+), as with previously studied enzymes. Arsenite was a noncompetitive inhibitor of the Caulobacter enzyme with a Ki of 75 microM AsO2-.
In Vitro Evolution and Preliminary Characterization of a Cadmium-Resistant Population of
Chlamydomonas reinhardtii
Authors: Nagel Klaus , Voigt Jürgen
Published: January 2020

A cadmium-tolerant population of
Chlamydomonas reinhardtii
was derived from a Cd-sensitive cell wall-deficient strain by long-term selection in liquid culture. A comparison of Cd-sensitive and Cd-tolerant cells revealed that Cd tolerance was due to genetically determined alterations of metabolism rather than to increased efficiency of a detoxification system.
Effect of temperature on aflatoxin production in Mucuna pruriens seeds
Authors: Roy A K , Chourasia H K
Published: January 2020

This paper describes the effect of temperature on the level of aflatoxin production in Mucuna pruriens seeds. The highest level of aflatoxin B1 (1.75 micrograms/g) was detected in the samples incubated at 25 degrees C for three weeks. At 20, 30, and 35 degrees C, aflatoxin levels were 0.30 to 0.56, 0.37 to 1.20, and 0.26 to 0.65 micrograms/g, respectively. The lowest concentration of aflatoxin B1 (0.10 to 0.29 microgram/g) was produced at 15 degrees C.
The suicide phenomenon in motile aeromonads
Authors: Namdari H , Cabelli V J
Published: January 2020

Certain strains of motile Aeromonas species, including all those of Aeromonas caviae examined, were shown to be suicidal. When they were grown in the presence of glucose at both 30 and 37 degrees C, there was rapid die-off of the organisms after 12 h of incubation, and viable cells generally could not be recovered after 24 h. It was shown that this phenomenon was due to the production of relatively high levels of acetic acid by these strains, even during growth under highly aerobic conditions, and to the greater susceptibility of these strains to acetic acid-mediated death. Suicide did not occur when the pH was maintained above 6.5 or in the presence of high concentration of Pi. These observations were consistent with our inability to isolate suicidal Aeromonas spp. from acidic lakes in New England and with their recovery from alkaline waters in Israel and from sewage. Suicidal aeromonads appear to be better adapted than the nonsuicidal biotypes to anaerobic growth in low-nutrient environments.