bluuebunny/crossref_metadata_2025 huggingface dataset:
Cloning of a Gene Cluster from
Cellvibrio mixtus
which Codes for Cellulase, Chitinase, Amylase, and Pectinase
Authors: Wynne E. C. , Pemberton J. M.
Published: January 2020

The soil isolate
Cellvibrio mixtus
UQM2294 degraded a variety of polysaccharides including microcrystalline cellulose. Among 6,000 cosmid clones carrying
C. mixtus
DNA, constructed in
Escherichia coli
with pHC79, 50 expressed the ability to degrade one or more of the following substrates: carboxymethyl cellulose, chitin, pectin (polygalacturonic acid), cellobiose, and starch. These degradative genes are encoded in a single 94.1-kilobase segment of the
C. mixtus
genome; a preliminary order of the genes is starch hydrolysis, esculin hydrolysis, cellobiose utilization, chitin hydrolysis, carboxymethyl cellulose hydrolysis, and polygalacturonic acid hydrolysis. A restriction endonuclease cleavage map was constructed, and the genes for starch, carboxymethyl cellulose, cellobiose, chitin, and pectin hydrolysis were subcloned.
Mutagenesis of Protoplasts and Regeneration of Mycelium in the Mushroom
Volvariella volvacea
Authors: Mukherjee M. , Sengupta S.
Published: January 2020

Regenerating protoplasts were obtained from mycelial culture of the mushroom
Volvariella volvacea
by the action of the lytic enzyme Novozym 234 in the presence of 0.01 M phosphate buffer (pH 6.0) containing 0.6 M NaCl. Regeneration was found to be poor in liquid medium, but more than 50% regeneration was achieved on solid 2% agar medium overlaid with 0.5% agar. Protoplasts of
V. volvacea
were found to be highly sensitive to the killing action of both UV irradiation and
N
-methyl-
N
′-nitro-
N
-nitrosoguanidine. However, no morphological or auxotrophic mutants could be obtained from protoplasts by chemical mutagenesis. Four types of morphological mutants and one auxotrophic (adenine-negative) mutant were obtained from UV-irradiated protoplasts. The adenine-negative mutant of
V. volvacea
was found to be stable, not losing auxotrophy on repeated subculture.
Chlorpromazine as permeabilizer and reagent for detection of microbial peroxidase and peroxidaselike activities
Authors: Galeazzi L , Turchetti G , Grilli G , Groppa G , Giunta S
Published: January 2020

Chlorpromazine was used to perform a test for the detection of microbial peroxidase activities. The compound acts as both a cell permeabilizer and a reagent in the procedure developed which allows the detection of peroxidase and peroxidase like reactions both semiquantitatively in whole cell determinations and quantitatively in cell-free supernatants.
Chlorinated fatty acid distribution in Mycobacterium convolutum phospholipids after growth on 1-chlorohexadecane
Authors: Murphy G L , Perry J J
Published: January 2020

The composition of phospholipids from Mycobacterium convolutum R22 was determined after growth at two temperatures (20 and 30 degrees C) with 1-chlorohexadecane as the substrate. Comparisons were made with the phospholipids of cells grown on n-hexadecane. Phosphatidylinositolmannosides and phosphatidylethanolamine (PE) were the major phospholipids in n-hexadecane-grown cells. In 1-chlorohexadecane-grown cells, phosphatidylinositolmannosides were approximately half of the total phospholipids, with lesser amounts of PE and cardiolipin (CL). The relative level of PE was greater at 20 degrees C (versus that at 30 degrees C) after growth on either substrate. A determination was made of structure and positional distribution of constituent fatty acid in both CL and PE. The relative amount of unsaturated fatty acid was higher at 20 degrees C. There were two C16:1 fatty acids (C16:1 delta 9 and C16:1 delta 11), and these had positional preferences in both CL and PE. The positional sites of chlorinated fatty acids differed in both CL and PE at the two temperatures. The results confirm that microorganisms can specifically distribute chlorinated fatty acids into cellular phospholipids.
Naphthalene biodegradation in environmental microcosms: estimates of degradation rates and characterization of metabolites
Authors: Heitkamp M A , Freeman J P , Cerniglia C E
Published: January 2020

Naphthalene biodegradation was investigated in microcosms containing sediment and water collected from three ecosystems which varied in past exposure to anthropogenic and petrogenic chemicals. Mineralization half-lives for naphthalene in microcosms ranged from 2.4 weeks in sediment chronically exposed to petroleum hydrocarbons to 4.4 weeks in sediment from a pristine environment. Microbiological analysis of sediments indicated that hydrocarbon-utilizing microbial populations also varied among ecosystems and were 5 to 12 times greater in sediment after chronic petrogenic chemical exposure than in sediment from an uncontaminated ecosystem. Sediment from an ecosystem exposed to agricultural chemicals had a mineralization half-life of 3.2 weeks for naphthalene and showed about a 30-fold increase in heterotrophic bacterial populations in comparison to uncontaminated sediments, but only a 2- to 3-fold increase in hydrocarbon-degrading bacteria. Analysis of organic solvent-extractable residues from the microcosms by high-pressure liquid chromatography detected polar metabolites which accounted for 1 to 3% of the total radioactivity. Purification of these residues by thin-layer chromatography and further analysis by gas chromatography-mass spectrometry indicated that cis-1,2-dihydroxy-1,2-dihydronaphthalene, 1-naphthol, salicylic acid, and catechol were metabolites of naphthalene. These results provide useful estimates for the rates of naphthalene mineralization in different natural ecosystems and on the degradative pathway for microbial metabolism of naphthalene in freshwater and estuarine environments.
Filter Paper Dot-Immunobinding Assay for Detection of
Spiroplasma citri
Authors: Fletcher Jacqueline
Published: January 2020

A rapid filter paper dot-immunobinding assay was adapted to detect the wall-less mollicute
Spiroplasma citri
in medium, plants, or insects. Filter paper spotted with sample was incubated first in dilute antiserum, then in protein A-peroxidase, and finally in a substrate of 4-chloro-1-naphthol plus hydrogen peroxide. The detection limit averaged 2.3 � 10
10
CFU/ml in cultures, and
S. citri
was detected in single infected leafhoppers. This assay was less sensitive but more rapid and economical than an enzyme-linked immunosorbent assay.
Effects of irradiation on growth and toxigenicity of Clostridium botulinum types A and B inoculated onto chicken skins
Authors: Dezfulian M , Bartlett J G
Published: January 2020

This study was conducted to examine the effects of 0.3-Mrad irradiation on growth and toxigenicity of Clostridium botulinum types A and B on chicken skins. Irradiation followed by aerobic or anaerobic incubation at 30 degrees C extended the shelf life of skin samples and delayed growth and toxin production by C. botulinum. During 2 weeks of incubation at 10 degrees C, the irradiated and nonirradiated C. botulinum spores failed to grow or produce toxin.
Autoconditioning factor relieves ethanol-induced growth inhibition of Saccharomyces cerevisiae
Authors: Walker-Caprioglio H M , Parks L W
Published: January 2020

Viable Saccharomyces cerevisiae suspended in medium containing growth-inhibiting concentrations of ethanol produce a metabolite that relieves growth inhibition. This autoconditioning of the medium by yeasts is due to the formation of small amounts (0.01%, vol/vol) of acetaldehyde. The effect is duplicated precisely in fresh medium by the addition of acetaldehyde. Acetaldehyde does not increase the yield of or accelerate ethanol production by the organism. Ethanol-induced modifications of membrane order in the plasma membranes, as measured by steady-state fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene, were not resolved by exogenously added acetaldehyde.
Formation and Regeneration of Protoplasts of the Actinorhizal Nitrogen-Fixing Actinomycete
Frankia
Authors: Tisa Louis S. , Ensign J. C.
Published: January 2020

Procedures for forming and regenerating protoplasts of four
Frankia
strains are described. Cells obtained from growth medium containing 0.1% glycine were digested with lysozyme (250 μg/ml) in a medium containing 0.5 M sucrose, 5.0 mM CaCl
2
, and 5.0 mM MgCl
2
. Protoplasts were formed during 15 to 120 min of digestion at 25�C. Optimum conditions for protoplast regeneration involved placing protoplasts on a layer of complex growth medium containing 0.3 M sucrose, 5.0 mM CaCl
2
, and 5.0 mM MgCl
2
which was overlaid with a layer of 0.8% low-melting-point agarose containing 0.5 M sucrose, 5.0 mM MgCl
2
, and 5.0 mM CaCl
2
. The maximum regeneration efficiency was 36.9% for strain CpI1, 1.3% for strain ACN1
AG
, 27% for strain EAN1pec, and 20% for strain EuI1c.
Production of staphylococcal enterotoxin in mixed cultures
Authors: Noleto A L , Malburg J�nior L M , Bergdoll M S
Published: January 2020

Two Staphylococcus aureus strains were grown in brain-heart infusion (BHI) broth and a meat medium with Bacillus cereus, Streptococcus faecalis, Escherichia coli, and Pseudomonas aeruginosa. Both S. aureus strains grew well and produced enterotoxin in the presence of S. faecalis in BHI broth; however, enterotoxin production was observable in the meat medium only when the S. aureus inoculum was greater than the S. faecalis inoculum. S. aureus FRI-100 grown with B. cereus produced enterotoxin in both media only when the S. aureus inoculum was much higher than the B. cereus inoculum (10 versus 10(4) CFU), whereas S. aureus FRI-196E produced enterotoxin in both media at all inoculum combinations except in the meat medium, when the inocula of the two organisms were the same. S. aureus grown with E. coli in BHI broth produced enterotoxin at all inoculum combinations except when the E. coli inoculum was greater than the S. aureus inoculum; however, in the meat medium, enterotoxin was produced only when the S. aureus inoculum was much greater than the E. coli inoculum (10 versus 10(4) CFU), S. aureus FRI-100 grown with P. aeruginosa in either medium produced enterotoxin only when the S. aureus inoculum was much greater than the P. aeruginosa inoculum (10 versus 10(3) or 10(4) CFU). It can be concluded from these results that enterotoxin production is unlikely in mixed cultures unless the staphylococci outnumber the other contaminating organisms.
Seasonal Variation in Population Density and Heterotrophic Activity of Attached and Free-Living Bacteria in Coastal Waters
Authors: Iriberri Juan , Unanue Marian , Barcina Isabel , Egea Luis
Published: January 2020

The abundance and heterotrophic activity of attached and free-living bacteria were examined seasonally in coastal water. Heterotrophic activity was determined by the uptake of [
14
C]glucose. The density of attached bacteria was always minor, not showing a seasonal variation, whereas the free-living bacteria were more numerous and showed a marked seasonal variation, their density being higher under warmer conditions. The contribution of the attached bacteria to the total assimilation of [
14
C]glucose (from 10 to 38%) was lower than that of the free-living bacteria, neither of them showing a seasonal variation. On a cellular basis, attached bacteria were more active, since they assimilated more [
14
C]glucose and showed, under warmer conditions, a higher cellular volume (0.102 versus 0.047 μm
3
). We consider that the factors responsible for these observations were the amount and quality of the particulate material, the different availability of organic matter for the two types of bacteria, and in a fundamental way, the variation in water temperature.
Correlation of Direct Viable Counts with Heterotrophic Activity for Marine Bacteria
Authors: Kogure Kazuhiro , Simidu Usio , Taga Nobuo , Colwell Rita R.
Published: January 2020

Viable-bacteria counts, heterotrophic activity, and substrate responsiveness of viable bacteria have been used to measure microbial activity. However, the relationship between these parameters is not clear. Thus, the direct viable count (DVC) method was used to analyze seawater samples collected from several different geographical locations. Samples collected from offshore waters of the South China Sea and western Pacific Ocean yielded DVC that indicated the presence of surface and subsurface peaks of viable, substrate-responsive bacteria which could be correlated with turnover rates of amino acids obtained by using uniformly
14
C-labeled amino acids. DVC were always less than total viable counts (acridine orange direct counts), and the DVC subsurface peak occurred close to and within the chlorophyll
a
zone, suggesting algal-bacterial interactions within the layer. For comparison with the open-ocean samples, selected substrates were used to determine the response of viable bacteria present in seawater samples collected near an ocean outfall of the Barceloneta Regional Waste Treatment Plant, Barceloneta, Puerto Rico. The number of specific substrate-responsive bacteria at the outfall stations varied depending on the substrate used and the sampling location. Changes in the population size or physiological condition of the bacteria were detected and found to be associated with the presence of pharmaceutical waste.
Diel Interactions of Oxygenic Photosynthesis and N
2
Fixation (Acetylene Reduction) in a Marine Microbial Mat Community
Authors: Bebout Brad M. , Paerl Hans W. , Crocker Kenneth M. , Prufert Leslie E.
Published: January 2020

Diel variations in N
2
fixation (acetylene reduction), CO
2
fixation, and oxygen concentrations were measured, on three separate occasions, in a marine microbial mat located on Shackleford Banks, North Carolina. Nitrogenase activity (NA) was found to be inversely correlated with CO
2
fixation and, in two of the three diel periods studied, was higher at night than during the day. Oxygen concentrations within the top 3 mm of the mat ranged from 0 to 400 μM on a diel cycle; anaerobic conditions generally persisted below 4 mm. NA in the mat was profoundly affected by naturally occurring oxygen concentrations. Experimentally elevated oxygen concentrations resulted in a significant depression of NA, whereas the addition of the Photosystem II inhibitor 3(3,4-dichlorophenyl)-1,1-dimethylurea decreased oxygen concentrations within the mat and resulted in a significant short-term enhancement of NA. Mat N
2
-fixing microorganisms include cyanobacteria and heterotrophic, photoautotrophic, and chemolithotrophic eubacteria. Measured (whole-mat) NA is probably due to a combination of the NA of each of these groups of organisms. The relative contributions of each group to whole-mat NA probably varied during diel and seasonal (successional) cycles. Reduced compounds derived from photosynthetic CO
2
fixation appeared to be an important source of energy for NA during the day, whereas heterotrophic or chemolithotrophic utilization of reduced compounds appeared to be an important source of energy for NA at night, under reduced ambient oxygen concentrations. Previous estimates of N
2
fixation calculated on the basis of daytime measurements may have seriously underestimated diel and seasonal nitrogen inputs in mat systems.
Wood Degradation by White Rot Fungi: Cytochemical Studies Using Lignin Peroxidase-Immunoglobulin-Gold Complexes
Authors: Garcia Susana , Latge Jean Paul , Prevost Marie Christine , Leisola Matti
Published: January 2020

Using an anti-lignin peroxidase antiserum-protein A-gold complex, we found lignin peroxidase mainly intracellularly in several white rot fungi colonizing sawdust under laboratory conditions. This enzyme was also present in fungi found in naturally decayed wood. However, in all cases, lignin peroxidase was located mainly inside the fungal cells. Labeled lignin peroxidase did not bind to the lignocellulosic samples tested, with the exception of poplar milled-wood lignin. These results are discussed in relation to the role of lignin peroxidase during wood degradation.
Measurement of the effects of cadmium stress on protozoan grazing of bacteria (bacterivory) in activated sludge by fluorescence microscopy
Authors: Hoffman R L , Atlas R M
Published: January 2020

The effect of cadmium stress on protozoan bacterivory in sewage sludge was measured by experimentally exposing sludge communities to 0 to 150 mg of Cd per liter for up to 6 h and then determining the rates of protozoan grazing on bacteria, using a double-staining technique and epifluorescence microscopy. Bacterivory was measured by incubating the sludge with fluorescently labeled bacterium-sized latex beads and directly observing ingestion of the beads and bacterial cells in the sludge by epifluorescence microscopy of preserved samples. Staining with 4',6-diamidino-2-phenylindole and acridine orange permitted the simultaneous determination of protozoan numbers and bacterivory activity as estimated by the number of bacterial cells and bacterium-sized latex beads ingested by the representative ciliate Aspidisca costata. Enumeration with latex beads proved to be an effective way of estimating bacterivory in sludges subjected to heavy-metal stress. This technique should prove useful for determining the effects of other chemical stresses on protozoan numbers and bacterivory in organic-rich environments. Although the number of protozoa declined significantly only after exposure to 100 mg of Cd per liter for 4 h, grazing, as indicated by bead ingestion, was significantly inhibited by Cd concentrations of greater than 25 mg/liter in less than 1 h, and exposure to 100 mg of Cd per liter effectively stopped protozoan grazing within 1 h of exposure. Protozoan ingestion of latex beads and bacteria was inversely correlated to Cd concentration and exposure time. The reduction of protozoan bacterivory by Cd provides a possible explanation for the increase in suspended bacteria in the effluents of metal-stressed treatment facilities.
Isolation and characterization of Streptococcus cremoris Wg2-specific promoters
Authors: van der Vossen J M , van der Lelie D , Venema G
Published: January 2020

By cloning MboI fragments in the promoter selection vector pGKV210, which replicates in Streptococcus lactis, Bacillus subtilis, and Escherichia coli and carries a promoterless chloramphenicol acetyltransferase gene, we obtained a number of fragments endowed with promoter activity, partly by direct selection for chloramphenicol resistance in S. lactis IL1403 and partly by selection in B. subtilis. Five fragments were sequenced, and the promoters were mapped with S1 nuclease. The promoters agreed with the E. coli promoter consensus and the B. subtilis vegetative sigma 43 promoter consensus. The promoters were preceded by an A + T-rich region (ranging from 64 to 78% A + T). S1 nuclease mapping data showed that the transcriptional start point in three of the fragments was at a TAG sequence 5 to 9 nucleotides downstream from the promoter. Three fragments carried an open reading frame preceded by a ribosome-binding site which can be recognized by E. coli, B. subtilis, and S. lactis ribosomes.
Bacillus subtilis generates a major specific deletion in pAM beta 1
Authors: van der Lelie D , Venema G
Published: January 2020

pAM beta 1, a 26.5-kilobase plasmid originally isolated from Streptococcus faecalis, was conjugally transferred from Streptococcus lactis to Bacillus subtilis. No conjugal transfer of pAM beta 1 from B. subtilis to S. lactis was observed. In addition, pAM beta 1 which had been reintroduced in S. lactis after cycling through B. subtilis had lost its conjugal transferability to Streptococcus cremoris, although under the same conditions noncycled pAM beta 1 was transferred at high efficiency. Restriction and Southern blot analyses showed that pAM beta 1 had suffered one major, specific 10.6-kilobase deletion and several minor but also specific deletions in B. subtilis. Comparing the major deletion derivative, delta pAM beta 1, with B. subtilis strains which have been reported to contain pAM beta 1 showed that these strains also contained delta pAM beta 1. Hybridization experiments showed that the deleted fragment was not transposed to the B. subtilis chromosome. Based on the size of the minor deletion derivatives from pAM beta 1, it is suggested that these use a different origin of replication in B. subtilis.
Detection of hepatitis A virus by hybridization with single-stranded RNA probes
Authors: Jiang X , Estes M K , Metcalf T G
Published: January 2020

An improved method of dot-blot hybridization to detect hepatitis A virus (HAV) was developed with single-stranded RNA (ssRNA) probes. Radioactive and nonradioactive ssRNA probes were generated by in vitro transcription of HAV templates inserted into the plasmid pGEM-1. 32P-labeled ssRNA probes were at least eightfold more sensitive than the 32P-labeled double-stranded cDNA counterparts, whereas biotin-labeled ssRNA probes showed a sensitivity comparable with that of the 32P-labeled double-stranded cDNA counterparts. Hybridization of HAV with the ssRNA probes at high stringency revealed specific reactions with a high signal-to-noise ratio. The differential hybridization reactions seen with probes of positive and negative sense (compared with HAV genomic RNA) were used to detect HAV in clinical and field samples. A positive/negative ratio was introduced as an indicator that permitted a semiquantitative expression of a positive HAV reaction. Good agreement of this indicator was observed with normal stool samples and with HAV-seeded samples. By using this system, HAV was detected in estuarine and freshwater samples collected from a sewage-polluted bayou in Houston and a saltwater tributary of Galveston Bay.
Regulation of beta-glucosidase in Bacteroides ruminicola by a different mechanism: growth rate-dependent derepression
Authors: Strobel H J , Russell J B
Published: January 2020

Bacteroides ruminicola B(1)4, a predominant ruminal and cecal bacterium, was grown in batch and continuous cultures, and beta-glucosidase activity was measured by following the hydrolysis of p-nitrophenyl-beta-glucopyranoside. Specific activity was high when the bacterium was grown in batch cultures containing cellobiose, mannose, or lactose (greater than 286 U/g of protein). Activity was reduced approximately 90% when the organism was grown on glucose, sucrose, fructose, maltose, or arabinose. The specific activity of cells fermenting glucose was initially low but increased as glucose was depleted. When glucose was added to cultures growing on cellobiose, beta-glucosidase synthesis ceased immediately. Catabolite repression by glucose was not accompanied by diauxic growth and was not relieved by cyclic AMP. Since glucose-grown cultures eventually exhibited high beta-glucosidase activity, cellobiose was not needed as an inducer. Catabolite repression explained beta-glucosidase activity of batch cultures and high-dilution-rate chemostats where glucose accumulated, but it could not account for activity at slow dilution rates. Maximal beta-glucosidase activity was observed at a dilution rate of approximately 0.35 h-1, and cellobiose-limited chemostats showed a 15-fold decrease in activity as the dilution rate declined. An eightfold decline was observed in glucose-limited chemostats. Since inducer availability was not a confounding factor in glucose-limited chemostats, the growth rate-dependent derepression could not be explained by other mechanisms.
Flagellar Motility Confers Epiphytic Fitness Advantages upon
Pseudomonas syringae
Authors: Haefele Douglas M. , Lindow Steven E.
Published: January 2020

The role of flagellar motility in determining the epiphytic fitness of an ice-nucleation-active strain of
Pseudomonas syringae
was examined. The loss of flagellar motility reduced the epiphytic fitness of a normally motile
P. syringae
strain as measured by its growth, survival, and competitive ability on bean leaf surfaces. Equal population sizes of motile parental or nonmotile mutant
P. syringae
strains were maintained on bean plants for at least 5 days following the inoculation of fully expanded primary leaves. However, when bean seedlings were inoculated before the primary leaves had expanded and bacterial populations on these leaves were quantified at full expansion, the population size of the nonmotile derivative strain reached only 0.9% that of either the motile parental or revertant strain. When fully expanded bean primary leaves were coinoculated with equal numbers of motile and nonmotile cells, the population size of a nonmotile derivative strain was one-third of that of the motile parental or revertant strain after 8 days. Motile and nonmotile cells were exposed in vitro and on plants to UV radiation and desiccating conditions. The motile and nonmotile strains exhibited equal resistance to both stresses in vitro. However, the population size of a nonmotile strain on leaves was less than 20% that of a motile revertant strain when sampled immediately after UV irradiation. Epiphytic populations of both motile and nonmotile
P. syringae
declined under desiccating conditions on plants, and after 8 days, the population size of a nonmotile strain was less than one-third that of the motile parental or revertant strain.
Plasmid-Associated Bacteriocin Production and Sucrose Fermentation in
Pediococcus acidilactici
Authors: Gonzalez Carlos F. , Kunka Blair S.
Published: January 2020

Production of bacteriocin activity designated pediocin PA-1 was associated with the presence of a 6.2-megadalton plasmid in
Pediococcus acidilactici
PAC1.0. The bacteriocin exhibited activity against
P. acidilactici, P. pentosaceus, Lactobacillus plantarum, L. casei, L. bifermentans
, and
Leuconostoc mesenteroides
subsp.
dextranicum.
Partial characterization of pediocin PA-1 is described. The molecular weight of pediocin PA-1 was ca. 16,500. Additionally, strain PAC1.0 was found to contain a 23-megadalton plasmid associated with sucrose-fermenting ability.
Stereoselective fungal metabolism of 7,12-dimethylbenz[a]anthracene: identification and enantiomeric resolution of a K-region dihydrodiol
Authors: McMillan D C , Fu P P , Cerniglia C E
Published: January 2020

Syncephalastrum racemosum UT-70 and Cunninghamella elegans ATCC 36112 metabolized 7,12-dimethylbenz[a]anthracene (7,12-DMBA) to hydroxymethyl metabolites as well as 7-hydroxymethyl-12-methylbenz[a]anthracene trans-3,4-, -5,6-, -8,9-, and -10,11-dihydrodiols. The 7,12-DMBA metabolites were isolated by reversed-phase high-performance liquid chromatography and identified by their UV-visible absorption, mass, and nuclear magnetic resonance spectral characteristics. A comparison of the circular dichroism spectra of the K-region (5,6-position) dihydrodiol of both fungal strains with those of the 7,12-DMBA 5S,6S-dihydrodiol formed from 7,12-DMBA by rat liver microsomes indicated that the major enantiomer of the 7-hydroxymethyl-12-methylbenz[a]anthracene trans-5,6-dihydrodiol formed by both fungal strains had a 5R,6R absolute stereochemistry. Direct resolution of the fungal trans-5,6-dihydrodiols by chiral stationary-phase high-performance liquid chromatography indicated that the ratios of the R,R and S,S enantiomers were 88:12 and 77:23 for S. racemosum and C. elegans, respectively. These results indicate that the fungal metabolism of 7,12-DMBA at the K region (5,6-position) is highly stereoselective and different from that reported for mammalian enzyme systems.
Isolation of intact high-molecular-weight DNA by using guanidine isothiocyanate
Authors: Lippke J A , Strzempko M N , Raia F F , Simon S L , French C K
Published: January 2020

A method for obtaining high-molecular-weight chromosomal DNA from Bacteroides intermedius and Bacteroides gingivalis is described. This technique is a modification of the guanidine isothiocyanate isolation procedure for RNA and should be useful for isolating intact DNA from organisms with high endogenous nuclease activity.
Two-level factorial screening for influence of temperature, pH, and aeration on production of Serratia marcescens nuclease
Authors: Jepsen P K , Riise E , Biedermann K , Kristensen P C , Emborg C
Published: January 2020

Two high-nuclease-yielding mutants of Serratia marcescens, derived by chemical mutagenesis (W280, W355), and two strains with the pBR322 plasmid 403-SD2, carrying a nuclease gene and a chloramphenicol resistance gene [Escherichia coli CSH50(403-SD2) and S. marcescens CH30(403-SD2)] were investigated for nuclease production in a factorial shake flask experiment, with temperature (30 and 37 degrees C), pH (with or without CaCO3 tablets), and aeration (with or without baffles) as variable conditions. Yields varied 10-fold depending on the conditions investigated.
Stable Carbon Isotope Fractionation by
Methanosarcina barkeri
during Methanogenesis from Acetate, Methanol, or Carbon Dioxide-Hydrogen
Authors: Krzycki J. A. , Kenealy W. R. , DeNiro M. J. , Zeikus J. G.
Published: January 2020

Methanosarcina barkeri
was cultured on methanol, H
2
-CO
2
, and acetate, and the
13
C/
12
C ratios of the substrates and the methane produced from them were determined. The discrimination against
13
C in methane relative to substrate decreased in the order methanol > CO
2
> acetate. The isotopic fractionation for methane derived from acetate was only one-third of that observed with methanol as the substrate. The data presented indicate that the last enzyme of methanogenesis, methylreductase, is not the primary site of isotopic discrimination during methanogenesis from methanol or CO
2
. These results also support biogeochemical interpretations that gas produced in environments in which acetate is the primary methane precursor will have higher
13
C/
12
C ratios than those from environments where other substrates predominate.
Nodulation and Competition for Nodulation of Selected Soybean Genotypes among
Bradyrhizobium japonicum
Serogroup 123 Isolates
Authors: Keyser H. H. , Cregan P. B.
Published: January 2020

Twenty recently obtained field isolates of
Bradyrhizobium japonicum
serogroup 123 were tested for their nodule mass production on the standard commercial soybean (
Glycine max
(L.) Merr. cv. Williams) and on two soybean plant introduction (PI) genotypes previously determined to restrict nodulation by strain USDA 123. Four of the field isolates showed similar restricted nodulation on the two genotypes, while all 20 isolates produced a normal amount of nodules on
G. max
cv. Williams. Serological analyses with adsorbed fluorescent antibodies showed that members of the 123 serotype ranked low in nodulation of the two PIs, in contrast to members of serotypes 127 and 129. Competition studies on the PIs indicated that isolates which were restricted were not competitive for nodule occupancy against strain USDA 110. However, unrestricted isolates of serogroup 123 were very competitive against USDA 110. On
G. max
cv. Williams, all serogroup 123 isolates tested were very competitive against USDA 110.
Detection of low-enterotoxin-producing Staphylococcus aureus strains
Authors: Kokan N P , Bergdoll M S
Published: January 2020

Culture supernatant fluids from 26 (23.6%) monkey feeding test-positive Staphylococcus aureus strains, negative for enterotoxins by gel diffusion, were positive by enzyme-linked immunosorbent assay for one or more of the identified enterotoxins. Staphylococcal enterotoxin D (SED) was produced by 23 (88.5%) strains, SED and SEA were produced in two strains, and SED and SEC were produced in one strain. One strain produced only SEA, and two strains produced only SEC.
Preparation and regeneration of bacteriophage PL-1 enzyme-induced Lactobacillus casei protoplasts
Authors: Watanabe K , Hayashida M , Nakashima Y , Hayashi S
Published: January 2020

Lactobacillus casei ATCC 27092 protoplasts were obtained by treatment with a bacteriolytic enzyme, which was produced in the PL-1 phage lysates of this bacterium, at 37 degrees C for 1.5 h in 50 mM Tris hydrochloride buffer (pH 7.2) containing 20% sucrose and 10 mM MgSO4. The protoplasts lacked the cell wall layer but retained the ability to transport L-[3H]glutamine. The frequency of regeneration was about 1%, whereas about 99% of the cells were osmotically sensitive.
Intestinal colonization of laboratory rats with Oxalobacter formigenes
Authors: Daniel S L , Hartman P A , Allison M J
Published: January 2020

Six strains of Oxalobacter formigenes (anaerobic oxalate-degrading bacteria) were examined for their ability to colonize the gastrointestinal tracts of adult laboratory rats. These rats did not harbor O. formigenes. Strain OxCR6, isolated from the cecal contents of a laboratory rat that was naturally colonized by oxalate-degrading bacteria, colonized the ceca and colons of adult rats fed a diet that contained 4.5% sodium oxalate. Five days after rats were inoculated intragastrically with 10(9) viable cells of strain OxCR6, oxalate degradation rates in cecal and colonic contents increased by 19 and 40 times, respectively. Viable counts of strain OxCR6 from these rats averaged 10(8)/g (dry weight) of cecal contents. Strain OxCR6 was not detected in the cecal contents of inoculated rats fed diets that contained less than 3.0% sodium oxalate. Strains of O. formigenes isolated from the cecal contents of swine, guinea pigs, and wild rats and from human feces also colonized the ceca of laboratory rats; a ruminal strain failed to colonize the rat cecum.
Induction of Cellulolytic and Xylanolytic Enzyme Systems in
Streptomyces
spp
Authors: MacKenzie C. Roger , Bilous Doris , Schneider Henry , Johnson Kenneth G.
Published: January 2020

Extracellular enzyme preparations from
Streptomyces flavogriseus
and
Streptomyces olivochromogenes
cultures grown on cellulose contained primarily cellulase activities, but similar preparations from cultures grown on xylan-containing materials possessed high levels of both cellulase and xylanase activities. Growth conditions that gave high endoxylanase levels also resulted in the production of enzymes involved in the hydrolysis of the nonxylose components of xylan. Specific acetyl xylan esterase activities were identified in enzyme preparations from both organisms. Both organisms also produced α-
l
-arabinofuranosidase activity that was not associated with endoxylanase activity. Other activities produced were α-
l
-
O
-methylglucuronidase and ferulic acid esterase. The latter enzyme was produced only by
S. olivochromogenes
and is an activity which has not previously been identified as a component of hemicellulase preparations.
Fermentation of xylans by Butyrivibrio fibrisolvens and other ruminal bacteria
Authors: Hespell R B , Wolf R , Bothast R J
Published: January 2020

The ability of Butyrivibrio fibrisolvens and other ruminal bacteria (6 species, 18 strains) to ferment a crude xylan from wheat straw or to ferment xylans from larchwood or oat spelts was studied. Liquid cultures were monitored for carbohydrate utilization, cell growth (protein), and fermentation acid production. B. fibrisolvens 49, H17c, AcTF2, and D1 grew almost as well on one or more of the xylans as they did on cellobiose-maltose. B. fibrisolvens 12, R28, A38, X10C34, ARD22a, and X6C61 exhibited moderate growth on xylans. Partial fermentation of xylans was observed with Bacteroides ruminicola B14, Bacteroides succinogenes S85, Ruminococcus albus 7, Ruminococcus flavefaciens C94 and FD1, and Succinivibrio dextrinosolvens 22B. All xylans tested appeared to have a small fraction of carbohydrate that supported low levels of growth of nonxylanolytic strains such as Selenomonas ruminantium HD4. Compared to growth on hexoses, the same array of fermentation acids was produced upon growth on xylans for most strains; however, reduced lactate levels were observed for B. fibrisolvens 49 and Selenomonas ruminantium HD4. Measurements of enzyme activities of B. fibrisolvens AcTF2, 49, H17c, and D1 indicated that the xylobiase activities were cell associated and that the xylanase activities were predominantly associated with the culture fluid. The pattern of expression of these enzymes varied both between strains and between the carbon sources on which the strains were grown.
Comparison of four membrane filter methods for fecal coliform enumeration in tropical waters
Authors: Santiago-Mercado J , Hazen T C
Published: January 2020

Four membrane filter methods for the enumeration of fecal coliforms were compared for accuracy, specificity, and recovery. Water samples were taken several times from 13 marine, 1 estuarine, and 4 freshwater sites around Puerto Rico, from pristine waters and waters receiving treated and untreated sewage and effluent from a tuna cannery and a rum distillery. Differences of 1 to 3 orders of magnitude in the levels of fecal coliforms were observed in some samples by different recovery techniques. Marine water samples gave poorer results, in terms of specificity, selectivity, and comparability, than freshwater samples for all four fecal coliform methods used. The method using Difco m-FC agar with a resuscitation step gave the best overall results; however, even this method gave higher false-positive error, higher undetected-target error, lower selectivity, and higher recovery of nontarget organisms than the method using MacConkey membrane broth, the worst method for temperate waters. All methods tested were unacceptable for the enumeration of fecal coliforms in tropical fresh and marine waters. Thus, considering the high densities of fecal coliforms observed at most sites in Puerto Rico by all these methods, it would seem that these density estimates are, in many cases, grossly overestimating the degree of recent fecal contamination. Since Escherichia coli appears to be a normal inhabitant of tropical waters, fecal contamination may be indicated when none is present. Using fecal coliforms as an indicator is grossly inadequate for the detection of recent human fecal contamination and associated pathogens in both marine and fresh tropical waters.
Testing for bacterial resistance to arsenic in monitoring well water by the direct viable counting method
Authors: Zelibor J L , Doughten M W , Grimes D J , Colwell R R
Published: January 2020

Direct viable counting of metal-resistant bacteria (DVCMR) has been found to be useful in both enumerating and differentiating metal-resistant and metal-sensitive strains of bacteria. The DVCMR bioassay was used to detect effects of low and high concentrations of arsenic and arsenicals on bacterial populations in groundwater. The level of resistance of the bacterial populations to arsenate was determined by the DVCMR bioassay, and the results showed a linear correlation with the total arsenic concentrations in the monitoring well water samples; no correlation was observed by culture methods with the methods employed. Bacteria resistant to 2,000 micrograms of arsenate per ml were isolated from all monitoring well water samples studied. Strains showed similar antibiotic and heavy-metal profiles, suggesting that the arsenic was not a highly selective pressure for arsenic alone. The monitoring well water samples were amended with arsenate and nutrients to determine the biotransformation mechanisms involved. Preliminary results suggest that bacteria indigenous to the monitoring well water samples did not directly transform, i.e., precipitate or volatilize, dissolved arsenic. It was concluded that arsenic contamination of the groundwater can be monitored by the DVCMR bioassay.
Adsorption of DNA to sand and variable degradation rates of adsorbed DNA
Authors: Lorenz M G , Wackernagel W
Published: January 2020

Adsorption and desorption of DNA and degradation of adsorbed DNA by DNase I were studied by using a flowthrough system of sand-filled glass columns. Maximum adsorption at 23 degrees C occurred within 2 h. The amounts of DNA which adsorbed to sand increased with the salt concentration (0.1 to 4 M NaCl and 1 mM to 0.2 M MgCl2), salt valency (Na+ less than Mg2+ and Ca2+), and pH (5 to 9). Maximum desorption of DNA from sand (43 to 59%) was achieved when columns were eluted with NaPO4 and NaCl for 6 h or with EDTA for 1 h. DNA did not desorb in the presence of detergents. It is concluded that adsorption proceeded by physical and chemical (Mg2+ bridging) interaction between the DNA and sand surfaces. Degradability by DNase I decreased upon adsorption of transforming DNA. When DNA adsorbed in the presence of 50 mM MgCl2, the degradation rate was higher than when it adsorbed in the presence of 20 mM MgCl2. The sensitivity to degradation of DNA adsorbed to sand at 50 mM MgCl2 decreased when the columns were eluted with 0.1 mM MgCl2 or 100 mM EDTA before application of DNase I. This indicates that at least two types of DNA-sand complexes with different accessibilities of adsorbed DNA to DNase I existed. The degradability of DNA adsorbed to minor mineral fractions (feldspar and heavy minerals) of the sand differed from that of quartz-adsorbed DNA.
Cloning of the
Pachysolen tannophilus
Xylulokinase Gene by Complementation in
Escherichia coli
Authors: Stevis Panayiotis E. , Huang James J. , Ho Nancy W. Y.
Published: January 2020

The gene coding for xylulokinase has been isolated from the yeast
Pachysolen tannophilus
by complementation of
Escherichia coli
xylulokinase (
xylB
) mutants. Through subcloning, the gene has been localized at one end of a 3.2-kilobase
Eco
RI-
Pst
I fragment. Expression of the cloned gene was insensitive to glucose inhibition. Furthermore, the cloned gene did not cross-hybridize with
E. coli
and
Saccharomyces cerevisiae
xylulokinase genes.
Growth determinations for unattached bacteria in a contaminated aquifer
Authors: Harvey R W , George L H
Published: January 2020

Growth rates of unattached bacteria in groundwater contaminated with treated sewage and collected at various distances from the source of contamination were estimated by using frequency of dividing cells and tritiated-thymidine uptake and compared with growth rates obtained with unsupplemented, closed-bottle incubations. Estimates of bacterial generation times [(In 2)/mu] along a 3-km-long transect in oxygen-depleted (0.1 to 0.7 mg of dissolved oxygen liter-1) groundwater ranged from 16 h at 0.26 km downgradient from an on-land, treated-sewage outfall to 139 h at 1.6 km and correlated with bacterial abundance (r2 = 0.88 at P less than 0.001). Partitioning of assimilated thymidine into nucleic acid generally decreased with distance from the contaminant source, and one population in heavily contaminated groundwater assimilated little thymidine during a 20-h incubation. Several assumptions commonly made when frequency of dividing cells and tritiated-thymidine uptake are used were not applicable to the groundwater samples.
Influence of Physical Disruption on Growth of Attached Bacteria
Authors: Murray Robert E. , Cooksey Keith E. , Priscu John C.
Published: January 2020

Attached bacterial populations cultured without an exogenous carbon source or grown in conjunction with attached diatoms incorporated [
3
H]thymidine at a rate between four and five times lower than that of replicate bacterial populations which were dispersed before being assayed.
Estimation of Bacterial Nitrate Reduction Rates at In Situ Concentrations in Freshwater Sediments
Authors: Hordijk Cornelis A. , Snieder Marchel , van Engelen Johannes J. M. , Cappenberg Thomas E.
Published: January 2020

A method was developed to follow bacterial nitrate reduction in freshwater sediments by using common high-performance liquid chromatographic equipment. The low detection limit (14 pmol) of the method enabled us to study concentration profiles and reaction kinetics under natural conditions. Significant nitrate concentrations (1 to 27 μM) were observed in the sediment of Lake Vechten during the nonstratified period; the concentration profiles showed a successive depletion of oxygen, nitrate, and sulfate with depth. The profiles were restricted to the upper 3 cm of the sediment which is rich in organics and loosely structured. Nitrate reduction in the sediment-water interface followed first-order reaction kinetics at in situ concentrations. Remarkably high potential nitrate-reducing activity was observed in the part of the sediment in which nitrate did not diffuse. This activity was also observed throughout the whole year. Estimates of
K

m

varied between 17 and 100 μM and
V
max
varied between 7.2 and 36 μmol cm
−3
day
−1
for samples taken at different depths. The diffusion coefficient of nitrate ([10 ± 0.4] × 10
−6
cm
2
s
−1
) across the sediment-water interface was estimated by a constant-source technique and applied to a mathematical model to estimate the net nitrate reduction during the nonstratified period. In this period, observed nitrate reduction rates by the model, 0.2 to 0.4 mmol m
−2
day
−1
, were lower than those found for oxygen (27 mmol m
−2
day
−1
) and sulfate (0.4 mmol m
−2
day
−1
). During the summer stratification, nitrate was absent in the sediment and reduction could not be estimated by the model.
Transformation of toluene and benzene by mixed methanogenic cultures
Authors: Grbić-Galić D , Vogel T M
Published: January 2020

The aromatic hydrocarbons toluene and benzene were anaerobically transformed by mixed methanogenic cultures derived from ferulic acid-degrading sewage sludge enrichments. In most experiments, toluene or benzene was the only semicontinuously supplied carbon and energy source in the defined mineral medium. No exogenous electron acceptors other than CO2 were present. The cultures were fed 1.5 to 30 mM unlabeled or 14C-labeled aromatic substrates (ring-labeled toluene and benzene or methyl-labeled toluene). Gas production from unlabeled substrates and 14C activity distribution in products from the labeled substrates were monitored over a period of 60 days. At least 50% of the substrates were converted to CO2 and methane (greater than 60%). A high percentage of 14CO2 was recovered from the methyl group-labeled toluene, suggesting nearly complete conversion of the methyl group to CO2 and not to methane. However, a low percentage of 14CO2 was produced from ring-labeled toluene or from benzene, indicating incomplete conversion of the ring carbon to CO2. Anaerobic transformation pathways for unlabeled toluene and benzene were studied with the help of gas chromatography-mass spectrometry. The intermediates detected are consistent with both toluene and benzene degradation via initial oxidation by ring hydroxylation or methyl oxidation (toluene), which would result in the production of phenol, cresols, or aromatic alcohol. Additional reactions, such as demethylation and ring reduction, are also possible. Tentative transformation sequences based upon the intermediates detected are discussed.
Particle Counter Determination of Bacterial Biomass in Seawater
Authors: Kogure Kazuhiro , Koike Isao
Published: January 2020

The applicability of the Elzone particle counter to the determination of marine bacterial biomass was investigated. The biomass of bacterial pure cultures and a mixed natural population were followed by using the particle counter, a CHN analyzer, and an ATP analyzer. The particle counter showed the precise size distribution of number and volume of submicron-size particles in seawater. For the pure cultured bacterial strains, the conversion factor from volume to carbon is 0.209 mg of C per mm
3
, and for natural bacterial cells of >0.6 μm in diameter, it is 0.184 mg of C per mm
3
. It is recommended that 0.2 be used as the conversion factor for both pure cultured marine bacterial cells and natural bacteria from coastal and near-shore marine environments.