bluuebunny/crossref_metadata_2025 huggingface dataset:
Degradation of 2-hydroxybiphenyl and 2,2'-dihydroxybiphenyl by Pseudomonas sp. strain HBP1
Authors: Kohler H P , Kohler-Staub D , Focht D D
Published: January 2020

Pseudomonas sp. strain HBP1 was found to grow on 2-hydroxy- and 2,2'-dihydroxy-biphenyl as the sole carbon and energy sources. The first step in the degradation of these compounds was catalyzed by an NADH-dependent monooxygenase. The enzyme inserted a hydroxyl group adjacent to the already existing hydroxyl group to form 2,3-dihydroxybiphenyl when acting on 2-hydroxybiphenyl and to form 2,2',3-trihydroxybiphenyl when acting on 2,2'-dihydroxybiphenyl. To be substrates of the monooxygenase, compounds required a 2-hydroxyphenyl-R structure, with R being a hydrophobic group (e.g., methyl, ethyl, propyl, sec-butyl, phenyl, or 2-hydroxyphenyl). Several chlorinated hydroxybiphenyls served as pseudosubstrates by effecting consumption of NADH and oxygen without being hydroxylated. Further degradation of 2,3-dihydroxy- and 2,2',3-trihydroxybiphenyl involved meta cleavage, with subsequent formation of benzoate and salicylate, respectively.
Identification of a δ-Endotoxin Gene Product Specifically Active against
Spodoptera littoralis
Bdv. among Proteolysed Fractions of the Insecticidal Crystals of
Bacillus thuringiensis
subsp.
aizawai
7.29
Authors: Lecadet M.-M. , Sanchis V. , Menou G. , Rabot P. , Lereclus D. , Chaufaux J. , Martouret D.
Published: January 2020

At least three different insecticidal crystal protein genes were shown to be expressed in
Bacillus thuringiensis
subsp.
aizawai
7.29, a strain that is potentially active against the cotton leafworm
Spodoptera littoralis
Bdv. Among crude K-60 fractions (60- to 70-kilodalton [kDa] molecules) that were products of proteolysed crystals containing the active domains of the protoxin molecules, we were able to distinguish several distinct components on the basis of their antigenic relationship and their larvicidal properties. A purified fraction designated SF2 was a 61-kDa component specifically active against
Pieris brassicae
L. and homologous to the
B. thuringiensis
subsp.
berliner
1715 plasmid-encoded crystal protein. A second fraction designated SF1 was composed of 63- and 65-kDa polypeptides and was specifically active against
S. littoralis.
The SF1 fraction and particularly the 65-kDa component were not antigenically related to the 61-kDa component. The purified fractions were compared with the products of three different crystal protein genes we previously cloned from total DNA of
B. thuringiensis
subsp.
aizawai
, among them a new type of crystal protein gene encoding a protein that is specifically active against
S. littoralis
and other insects of the Noctuidae family. This approach led us to consider the 65-kDa component as a minimum active part of a δ-endotoxin that is encoded by this new gene. Products of the two other cloned genes can be correlated with the 61- and 63-kDa components, respectively. Thus, in
B. thuringiensis
subsp.
aizawai
7.29, multiple δ-endotoxin genes of different structural types direct the synthesis of several δ-endotoxins with different host specificities which were identified as components of the insecticidal crystals.
N-Acetyl-
d
-Glucosamine-Mediated Regulation of Extracellular Protease in the Entomopathogenic Fungus Beauveria bassiana
Authors: Bidochka Michael J. , Khachatourians George G.
Published: January 2020

The entomopathogenic fungus
Beauveria bassiana
GK2016 grown in a liquid medium incorporating gelatin as the sole carbon and nitrogen source produced an extracellular serine protease (molecular weight, 35,000; pI ca. 10). Without gelatin,
B. bassiana
could utilize
N
-acetyl-
d
-glucosamine (GlcNAc; 2-acetamido-2-deoxy-
d
-glucose) as the sole source of carbon and nitrogen, and GlcNAc availability increased the storage carbohydrate content in mycelia. Synthesis of protease was repressed in gelatin medium containing GlcNAc at levels of >1.07 μmol mg of fungal dry weight
−1
. At levels below this, protease synthesis was initiated; subsequently, free amino nitrogen appeared in the medium and diauxic growth was observed. Slow feeding with GlcNAc (35.34 μg ml
−1
h
−1
) did not repress protease synthesis nor did GlcNAc accumulate in the medium above 0.5 mg ml
−1
. Increasing the rate of release of GlcNAc (83.51 μg ml
−1
h
−1
) resulted in the accumulation of GlcNAc in the medium to 2.0 mg ml
−1
, a 45% increase in growth and a decrease in protease synthesis by about 81%. Free amino acids generated from the hydrolysis of gelatin did not repress protease synthesis. These data are interpreted in terms of known interaction of
B. bassiana
with insect cuticular components. We suggest that the action of extracellular chitinases synthesized by
B. bassiana
on insect cuticle, and pursuant release of GlcNAc, may have important consequences on the regulation of other extracellular catabolic enzymes such as the protease.
Dynamics of Soil Denitrifier Populations: Relationships between Enzyme Activity, Most-Probable-Number Counts, and Actual N Gas Loss
Authors: Martin Kendall , Parsons Laura L. , Murray Robert E. , Smith M. Scott
Published: January 2020

To better understand temporal variability in soil denitrification, denitrifying enzyme activity (DEA) and denitrifier populations (as determined by most-probable-number [MPN] counts) were measured in field and laboratory experiments. Measurements of DEA and MPN provided highly contradictory indications of denitrifier dynamics. In laboratory incubations, under conditions favoring active denitrification, the synthesis of new denitrifying enzymes and the actual amount of denitrification were closely related. In other experiments, however, both DEA and MPN counts were poor indicators of actual denitrification. In some cases, we found significant increases in DEA but no significant production of N gas. Except with unnaturally high substrate amendments, changes in DEA were small relative both to the persistently high DEA background and to changes in MPN. As estimated by MPN counts, denitrifier populations increased significantly during denitrification events. It was apparent that only a small fraction of the denitrifiers were included in the MPN counts, but it appeared that this isolatable fraction increased during periods of active denitrifier growth. Use of DEA as an index of biomass of cells which have synthesized denitrifying enzymes suggested that denitrifier populations were persistent, stable, and much larger than indicated by MPN procedures.
Carbon allocation in wild-type and Glc+ Rhodobacter sphaeroides under photoheterotrophic conditions
Authors: Macler B A , Bassham J A
Published: January 2020

The photosynthetic bacterium Rhodobacter sphaeroides is capable of producing H2 via nitrogenase when grown photoheterotrophically in the absence of N2. By using 14C-labeled malate, it was found that greater than 95% of this substrate was catabolized completely to CO2 during H2 production. About 60% of this catabolism was associated with H2 biosynthesis, while almost 40% provided reductant for other cellular purposes. Thus, only a small fraction of malate provided carbon skeletons. The addition of ammonium, which inhibited nitrogenase activity, increased substrate conversion into carbon skeletons threefold. Catabolism of malate occurred primarily via the tricarboxylic acid cycle, but gluconeogenesis was also observed. The wild-type organism grew poorly on glucose, accumulated gluconate and 2-keto-3-deoxygluconate, and did not produce H2. More than 50% of metabolized glucose appeared in carbon skeletons or in storage compounds. A glucose-utilizing mutant was five times more effective in utilizing this substrate. This mutant produced H2 from glucose, using 74% of metabolized substrate for this purpose. Glucose converted to storage products or to other carbon skeletons was reduced to 8%. Fixation of CO2 competed directly with H2 production for reducing equivalents and ATP. Refixation of CO2 released from these substrates under H2-producing conditions was, at most, 10 to 12%. Addition of ammonium increased refixation of respired CO2 to 83%. Patterns of carbon flow of fixation products were associated with the particular strains and culture conditions.
Production of
d
-Aminoacylase from
Alcaligenes denitrificans
subsp.
xylosoxydans
MI-4
Authors: Moriguchi Mitsuaki , Ideta Kohtaro
Published: January 2020

A bacterial strain that produces
d
-aminoacylase was isolated from soil and identified as
Alcaligenes denitrificans
subsp.
xylosoxydans
MI-4.
l
-Aminoacylase activity in this strain was only 1 to 2% of
d
-aminoacylase activity.
d
-Aminoacylase was inducibly produced.
N
-Acetyl-
dl
-leucine was the best inducer, and the
d
-isomer had the ability to induce the enzyme. Enzymatic resolution of
N
-acetyl-
dl
-methionine with the crude enzyme was carried out, and the
d
/
l
ratio in the resolved methionine was approximately 100/7, suggesting that resolution with crude enzymes may become possible by removing small amounts of the contaminated
l
-form with
l
-amino acid oxidase.
Cross-species transmission of Giardia spp.: inoculation of beavers and muskrats with cysts of human, beaver, mouse, and muskrat origin
Authors: Erlandsen S L , Sherlock L A , Januschka M , Schupp D G , Schaefer F W , Jakubowski W , Bemrick W J
Published: January 2020

Giardia cysts isolated from humans, beavers, mice, and muskrats were tested in cross-species transmission experiments for their ability to infect either beavers or muskrats. Giardia cysts, derived from multiple symptomatic human donors and used for inoculation of beavers or muskrats, were shown to be viable by incorporation of fluorogenic dyes, excystation, and their ability to produce infections in the Mongolian gerbil model. Inoculation of beavers with 5 x 10(5) Giardia lamblia cysts resulted in the infection of 75% of the animals (n = 8), as judged by the presence of fecal cysts or intestinal trophozoites at necropsy. The mean prepatent period was 13.1 days. An infective dose experiment, using 5 x 10(1) to 5 x 10(5) viable G. lamblia cysts collected by fluorescence-activated cell sorting, demonstrated that doses of between, less than 50, and less than 500 viable cysts were required to produce infection in beavers. Scanning electron microscopy of beaver small intestine revealed that attachment of G. lamblia trophozoites produced lesions in the microvillous border. Inoculation of muskrats with G. lamblia cysts produced infections when the dose of cysts was equal to or greater than 1.25 x 10(5). The inoculation of beavers with Giardia ondatrae or Giardia muris cysts did not produce any infection; however, the administration to muskrats of Giardia cysts of beaver origin resulted in the infection of 62% of the animals (n = 8), with a prepatent period of 5 days. Our results demonstrated that beavers and muskrats could be infected with Giardia cysts derived from humans, but only by using large numbers of cysts.(ABSTRACT TRUNCATED AT 250 WORDS)
Transformation of tetrachloromethane to dichloromethane and carbon dioxide by Acetobacterium woodii
Authors: Egli C , Tschan T , Scholtz R , Cook A M , Leisinger T
Published: January 2020

Five anaerobic bacteria were tested for their abilities to transform tetrachloromethane so that information about enzymes involved in reductive dehalogenations of polychloromethanes could be obtained. Cultures of the sulfate reducer Desulfobacterium autotrophicum transformed some 80 microM tetrachloromethane to trichloromethane and a small amount of dichloromethane in 18 days under conditions of heterotrophic growth. The acetogens Acetobacterium woodii and Clostridium thermoaceticum in fructose-salts and glucose-salts media, respectively, degraded some 80 microM tetrachloromethane completely within 3 days. Trichloromethane accumulated as a transient intermediate, but the only chlorinated methanes recovered at the end of the incubation were 8 microM dichloromethane and traces of chloromethane. Desulfobacter hydrogenophilus and an autotrophic, nitrate-reducing bacterium were unable to transform tetrachloromethane. Reduction of chlorinated methanes was thus observed only in the organisms with the acetyl-coenzyme A pathway. Experiments with [14C]tetrachloromethane were done to determine the fate of this compound in the acetogen A. woodii. Radioactivity in an 11-day heterotrophic culture was largely (67%) recovered in CO2, acetate, pyruvate, and cell material. In experiments with cell suspensions to which [14C]tetrachloromethane was added, 14CO2 appeared within 20 s as the major transformation product. A. woodii thus catalyzes reductive dechlorinations and transforms tetrachloromethane to CO2 by a series of unknown reactions.
Isolation of
Rhizobium loti
Strain-Specific DNA Sequences by Subtraction Hybridization
Authors: Bjourson A. J. , Cooper J. E.
Published: January 2020

Mixed-phase (heterogeneous) and single-phase (homogeneous) DNA subtraction-hybridization methods were used to isolate specific DNA probes for closely related
Rhizobium loti
strains. In the heterogeneous method, DNA from the prospective probe strain was repeatedly hybridized to a mixture of DNA from cross-hybridizing strains (subtracter DNA) which was immobilized on an epoxy-activated cellulose matrix. Probe strain sequences which shared homology with the matrix-bound subtracter DNA hybridized to it, leaving unique probe strain sequences in the mobile phase. In the homogeneous method, probe strain sequences were hybridized in solution to biotinylated, mercurated subtracter DNA. Biotinylated, mercurated subtracer DNA and probe strain sequences hybridized to it were removed by two-step affinity chromatography on streptavidin-agarose and thiol-Sepharose. The specificity of the sequences remaining after subtraction hybridization by both methods was assessed and compared by colony hybridization with
R. loti
strains. Both methods allowed the rapid isolation of strain-specific DNA fragments which were suitable for use as probes.
Role of sublethal injury in decline of bacterial populations in lake water
Authors: Gurijala K R , Alexander M
Published: January 2020

Following their addition to lake water, the populations of Escherichia coli and of antibiotic-resistant strains of Pseudomonas fluorescens, Agrobacterium tumefaciens, Micrococcus flavus, Rhizobium meliloti, and Klebsiella pneumoniae declined rapidly, as determined by counting on media containing antibacterial compounds. The estimates of population sizes were occasionally higher if procedures were used that permitted possible resuscitation of injured cells. No resuscitation procedure yielded consistently higher estimates of populations of surviving cells than the use of selective media alone. The patterns of survival of the test bacteria in lake water amended with eucaryotic inhibitors were essentially the same whether a resuscitation procedure was used or not, and the patterns of survival in sterile lake water or buffer were the same whether counts were made on selective media or on media without antibacterial agents. On the basis of the methods used to show sublethal injury caused by stress, we suggest that such injury to the test bacteria is not a significant factor involved in their decline in lake water.
Delivery system for creation of one-step in vivo lac gene fusions in Pseudomonas spp. involved in biological control
Authors: O'Sullivan D J , O'Gara F
Published: January 2020

The suicide plasmid pVA838 carrying the operon fusion transposon Tn5-lac was used as a delivery system to introduce Tn5-lac into Pseudomonas sp. strain M114. Random, in vivo lac gene fusions were successfully isolated in a one-step conjugation approach with this vector system. Tn5-lac-containing exconjugants were recovered at a frequency of approximately 10(-7) per recipient. However, when the mating temperature was increased from the normal growth temperature (28 degrees C) to 34 degrees C, the frequency was increased to approximately 10(-4) per recipient. A number of in vivo lac gene fusions were isolated and characterized in strain M114, a potentially important bacterium for biological control purposes.
Survival of Escherichia coli and Salmonella spp. in estuarine environments
Authors: Rhodes M W , Kator H
Published: January 2020

Survival of Escherichia coli and Salmonella spp. in estuarine waters was compared over a variety of seasonal temperatures during in situ exposure in diffusion chambers. Sublethal stress was measured by both selective-versus-resuscitative enumeration procedures and an electrochemical detection method. E. coli and Salmonella spp. test suspensions, prepared to minimize sublethal injury, were exposed in a shallow tidal creek and at a site 7.1 km further downriver. Bacterial die-off and sublethal stress in filtered estuarine water were inversely related to water temperature. Salmonella spp. populations exhibited significantly less die-off and stress than did E. coli at water temperatures of less than 10 degrees C. Although the most pronounced reductions (ca. 3 log units) in test bacteria occurred during seasonally warm temperatures in the presence of the autochthonous microbiota, 10(2) to 10(4) test cells per ml remained after 2 weeks of exposure to temperatures of greater than 15 degrees C. Reductions in test bacteria were associated with increases in the densities of microflagellates and plaque-forming microorganisms. These studies demonstrated the survival potential of enteric bacteria in estuarine waters and showed that survival was a function of interacting biological and physical factors.
Synthetic oligodeoxyribonucleotide probes for detection of Listeria monocytogenes
Authors: Datta A R , Wentz B A , Shook D , Trucksess M W
Published: January 2020

A 500-base-pair DNA fragment of a presumptive beta-hemolysin gene of Listeria monocytogenes has been used to identify this organism by a modified colony hybridization technique. We have cloned this DNA fragment into M13 bacteriophage vectors and sequenced it by a dideoxynucleotide sequencing technique. From this sequencing information, several oligodeoxyribonucleotides were synthesized and used as synthetic probes to identify L. monocytogenes. The probes were specific for L. monocytogenes and did not react with any other Listeria strains in a colony hybridization assay. In particular, one of these probes (AD07) was used to detect L. monocytogenes in artificially contaminated raw-milk and soft-cheese samples.
Physiological characterization of strain DCB-1, a unique dehalogenating sulfidogenic bacterium
Authors: Stevens T O , Linkfield T G , Tiedje J M
Published: January 2020

Strain DCB-1 is an obligately anaerobic bacterium which carries out the reductive dehalogenation of halobenzoates and was previously known to grow only on pyruvate plus 20% ruminal fluid. When various electron acceptors were supplied, thiosulfate and sulfite were found to stimulate growth. Sulfide was produced from thiosulfate. Cytochrome c and desulfoviridin were detected. The mol% G+C was 49 (at the thermal denaturation temperature). Of 55 carbon sources tested, only pyruvate supported growth as the sole carbon source in mineral medium. Lactate, acetate, L- and D-malate, glycerol, and L- and D-arabinose stimulated growth when supplemented with 10% ruminal fluid and 20 mM thiosulfate. In mineral medium, pyruvate was converted to acetate and lactate, with small amounts of succinate and fumarate accumulating transiently. During growth with thiosulfate, all of these products accumulated transiently. Addition of excess hydrogen to pyruvate-grown cultures resulted in diversion of carbon to formate, lactate, and butyrate, which caused a decrease in cell yield. We conclude that strain DCB-1 is a new type of sulfidogenic bacterium.
Reconstitution of the gastrointestinal microflora of lactobacillus-free mice
Authors: Tannock G W , Crichton C , Welling G W , Koopman J P , Midtvedt T
Published: January 2020

A colony of mice that do not harbor lactobacilli in their digestive tracts but whose intestinal microflora is otherwise functionally similar to that of conventional animals was derived. Methods used to reconstitute the intestinal microflora of the mice included inoculation of the animals with cultures of specific microbes, noncultivable microbes attached to epithelial cells, and cecal contents from conventional mice treated with chloramphenicol. Twenty-six microflora-associated characteristics were monitored by using relatively simple tests to determine the microflora status of the mice.
Dependence of tetrachloroethylene dechlorination on methanogenic substrate consumption by Methanosarcina sp. strain DCM
Authors: Fathepure B Z , Boyd S A
Published: January 2020

Tetrachloroethylene (perchloroethylene, PCE) is a suspected carcinogen and a common groundwater contaminant. Although PCE is highly resistant to aerobic biodegradation, it is subject to reductive dechlorination reactions in a variety of anaerobic habitats. The data presented here clearly establish that axenic cultures of Methanosarcina sp. strain DCM dechlorinate PCE to trichloroethylene and that this is a biological reaction. Growth on methanol, acetate, methylamine, and trimethylamine resulted in PCE dechlorination. The reductive dechlorination of PCE occurred only during methanogenesis, and no dechlorination was noted when CH4 production ceased. There was a clear dependence of the extent of PCE dechlorination on the amount of methanogenic substrate (methanol) consumed. The amount of trichloroethylene formed per millimole of CH4 formed remained essentially constant for a 20-fold range of methanol concentrations and for growth on acetate, methylamine, and trimethylamine. These results suggest that the reducing equivalents for PCE dechlorination are derived from CH4 biosynthesis and that the extent of chloroethylene dechlorination can be enhanced by stimulating methanogenesis. It is proposed that electrons transferred during methanogenesis are diverted to PCE by a reduced electron carrier involved in methane formation.
Survival of coliforms and bacterial pathogens within protozoa during chlorination
Authors: King C H , Shotts E B , Wooley R E , Porter K G
Published: January 2020

The susceptibility of coliform bacteria and bacterial pathogens to free chlorine residuals was determined before and after incubation with amoebae and ciliate protozoa. Viability of bacteria was quantified to determine their resistance to free chlorine residuals when ingested by laboratory strains of Acanthamoeba castellanii and Tetrahymena pyriformis. Cocultures of bacteria and protozoa were incubated to facilitate ingestion of the bacteria and then were chlorinated, neutralized, and sonicated to release intracellular bacteria. Qualitative susceptibility of protozoan strains to free chlorine was also assessed. Protozoa were shown to survive and grow after exposure to levels of free chlorine residuals that killed free-living bacteria. Ingested coliforms Escherichia coli, Citrobacter freundii, Enterobacter agglomerans, Enterobacter cloacae, Klebsiella pneumoniae, and Klebsiella oxytoca and bacterial pathogens Salmonella typhimurium, Yersinia enterocolitica, Shigella sonnei, Legionella gormanii, and Campylobacter jejuni had increased resistance to free chlorine residuals. Bacteria could be cultured from within treated protozoans well after the time required for 99% inactivation of free-living cells. All bacterial pathogens were greater than 50-fold more resistant to free chlorine when ingested by T. pyriformis. Escherichia coli ingested by a Cyclidium sp., a ciliate isolated from a drinking water reservoir, were also shown to be more resistant to free chlorine. The mechanism that increased resistance appeared to be survival within protozoan cells. This study indicates that bacteria can survive ingestion by protozoa. This bacterium-protozoan association provides bacteria with increased resistance to free chlorine residuals which can lead to persistence of bacteria in chlorine-treated water. We propose that resistance to digestion by predatory protozoa was an evolutionary precursor of pathogenicity in bacteria and that today it is a mechanism for survival of fastidious bacteria in dilute and inhospitable aquatic environments.
Examination of Genetic Relatedness of Marine
Synechococcus
spp. by Using Restriction Fragment Length Polymorphisms
Authors: Douglas Susan E. , Carr Noel
Published: January 2020

The relatedness of several marine
Synechococcus
spp. was estimated by DNA hybridization. Strains isolated from various geographical locations and representing a diversity of DNA base compositions and phycobiliprotein profiles were compared by restriction fragment length polymorphisms for a number of genes. DNAs from two marine red algae and a cryptomonad alga (which exhibit a phycobiliprotein composition similar to that of the marine
Synechococcus
spp.) and
Synechococcus
strain PCC6301 (
Anacystis nidulans
) were also included in the comparison. Strains WH8008, WH8018, and WH7805 were shown to be very similar to one another, as were strains WH7802 and WH7803. Strains WH8110 and WH5701 were clearly unrelated to any of the other strains, and no marine
Synechococcus
isolate showed any similarity to the freshwater
Synechococcus
strain PCC6301 or the eucaryotic algae. The method is relatively straightforward and sensitive and uses a variety of basic molecular biology techniques. Its utility in ascertaining the genetic relatedness and diversity of marine
Synechococcus
spp. and possible extension to field studies are discussed.
Dehalogenation in marine sediments containing natural sources of halophenols
Authors: King G M
Published: January 2020

Halophenols such as 2,4-dibromophenol (DBP) occur naturally in some marine sediments, as a consequence of various animal and algal activities. In an earlier study, DBP was observed in the burrow microenvironment of the hemichordate Saccoglossus kowalewskii. At the concentrations found in the burrow lining, aerobic respiration appeared to be inhibited significantly relative to anaerobic catabolism. This effect, as well as factors contributing to the degradation of DBP, has been documented further here. Results from the addition of radiolabeled DBP to oxic and anoxic sediment slurries and growth experiments with aerobic and anaerobic enrichments suggested that aerobes did not significantly metabolize DBP and that concentrations likely to be encountered on the inner surfaces of the burrow wall were inhibitory. In contrast, only minimal inhibition of growth occurred for anaerobes exposed to 1 mM DBP; in addition, DBP was substantially degraded in both enrichments and sediments under anaerobic conditions. Dehalogenation with the consequent production of phenol appeared to initiate anaerobic degradation. Sulfate-reducing bacteria did not dehalogenate DBP but appeared to degrade phenol. Decreased bacterial numbers and marked differences in the concentration and chemical speciation of iron in sediments from S. kowalewskii burrows may be attributed to toxic effects of DBP on aerobic bacteria.
Colonization of Tomato Plants by Two Agrocin-Producing Strains of
Agrobacterium tumefaciens
Authors: Macrae Sharmane , Thomson Jennifer A. , Van Staden Johannes
Published: January 2020

For a bacterium to be a successful biocontrol agent against crown gall disease, it must produce an effective agrocin specific for
Agrobacterium tumefaciens
and be able to colonize host plants efficiently. The colonization abilities of K84 and J73, successful and potential biocontrolling strains, respectively, were compared both in vivo and in vitro. Both strains produced fibrils attaching them to tomato root surfaces and had similar colonization efficiencies up to 14 days after inoculation. However, the ability of J73 to colonize plants for longer periods was significantly less than that of K84. Thus, the presence of fibrils is not sufficient to ensure colonization. No correlation was found between hydrophobicity and colonization.
Induction of Yellow Pigmentation in
Serratia marcescens
Authors: Trias Joaquim , Viñas Miquel , Guinea Jesús , Lorén José G.
Published: January 2020

The appearance of yellow pigmentation in nonpigmented strains of
Serratia
sp. has been demonstrated to be due to the production of a muconic acid, 2-hydroxy-5-carboxymethylmuconic acid semialdehyde. The 3,4-dihydroxyphenylacetate 2,3-dioxygenase responsible for the synthesis of this muconic acid was induced in all strains tested. Another muconic acid, the β-
cis-cis
-carboxymuconic acid, could also be synthesized from 3,4-dihydroxybenzoate, but this product was not colored. Mutants that were unable to grow on tyrosine and produced yellow pigment were isolated from nonpigmented strains. These mutants had properties similar to those of the yellow-pigmented strains. The ability to produce pigment may be more widespread among
Serratia marcescens
strains than is currently known.
Restoration of colony-forming activity in osmotically stressed Escherichia coli by betaine
Authors: Roth W G , Leckie M P , Dietzler D N
Published: January 2020

Exposure of Escherichia coli to 0.8 M NaCl caused a rapid and large decrease in colony-forming activity. When such osmotically upshocked cells were exposed to betaine, colony-forming activity was restored. Betaine was able to restore colony-forming activity even when chloramphenicol inhibited protein synthesis. Thus, restoration was not the result of cell turnover. The cells were not killed by exposure to 0.8 M NaCl, because during exposure they accumulated ATP intracellularly. Betaine treatment caused this cellular ATP to decrease to a lower level. This work may provide the foundation for a simple plating procedure to quantitatively detect nonculturable E. coli in ocean beach recreational waters.
Tn
5
-Mediated Cloning of a Genetic Region from
Pseudomonas putida
Involved in the Stimulation of Plant Root Elongation
Authors: Lifshitz Ran , Guilmette Helene , Kozlowski Maya
Published: January 2020

Transposon (Tn
5
) mutagenesis was applied to
Pseudomonas putida
GR12-2R3, which promotes root elongation (a phenotype designated Pre) of
Brassica campestris
under gnotobiotic conditions. Of 3,000 Tn
5
transconjugants, only one mutant that lost Pre activity but remained prototrophic and capable of plant root colonization was detected. This mutant was complemented by plasmid pRE53, which contained a 15.0-kilobase DNA insert isolated from a parental strain. The complemented mutant regained full Pre activity comparable to that of the wild type.
Effect of inorganic nutrients on the acclimation period preceding mineralization of organic chemicals in lake water
Authors: Jones S H , Alexander M
Published: January 2020

The addition of phosphate, nitrate, or sulfate (each at 10 mM) decreased the acclimation period for the mineralization of low concentrations of p-nitrophenol (PNP) in lake water. Added phosphate shortened the acclimation period for biodegradation of 2 ng to 2 micrograms of PNP per ml in various lake water samples and of 2,4-dichlorophenoxyacetate at 100 ng/ml. Added P enhanced the rate of growth of PNP-mineralizing microorganisms in waters containing 200 ng or 2 micrograms of PNP per ml. We suggest that the effect of P on the acclimation period results from an increase in the growth rate of the initially small population of microorganisms able to mineralize the synthetic chemicals.
Toxin profiles of Vibrio cholerae non-O1 from environmental sources in Calcutta, India
Authors: Nair G B , Oku Y , Takeda Y , Ghosh A , Ghosh R K , Chattopadhyay S , Pal S C , Kaper J B , Takeda T
Published: January 2020

A collection of Vibrio cholerae non-O1 isolated from the aquatic environs of Calcutta, a cholera-hyperendemic area, were examined for the production of cholera toxin (CT), Shiga-like toxins (Vero toxins), heat-stable enterotoxin, and hemolysins. Two (0.5%) V. cholerae non-O1 isolates produced CT. The DNA from both these isolates also hybridized with a DNA probe containing sequences encoding the A subunit of CT. None of the strains produced Shiga-like toxins or heat-stable enterotoxin. Hemolytic activity was observed in 89.7% of the strains, of which 36.1% exhibited biological activity in the suckling mouse. However, none of them produced a hemolysin that cross-reacted with the thermostable direct hemolysin of Vibrio parahaemolyticus. It appears from this study that a small percentage of environmental V. cholerae non-O1 strains do possess the potential for causing cholera-like diarrhea.
Control of Lignin Peroxidase Production by
Phanerochaete chrysosporium
INA-12 by Temperature Shifting
Authors: Asther Marcel , Capdevila Cécile , Corrieu Georges
Published: January 2020

There are two temperature optima connected with lignin peroxidase synthesis by
Phanerochaete chrysosporium
INA-12. One, at 37°C, is for the mycelium-growing phase; the other, at 30°C, is for the lignin peroxidase-producing phase. One of six extracellular proteins with ligninase activity increased when cultures were grown at 30°C for the entire fermentation period or when cultures were grown at 37°C for the first 2 days of incubation and then shifted to 30°C, compared with the activity of control cultures grown at 37°C for the entire fermentation period. The unsaturation of fatty acid (Δ/mole) of
P. chrysosporium
INA-12 mycelium decreased from 1.25 to 1.03 when the growth temperature was shifted from 20 to 40°C.
Characterization of an Endoglucanase from
Pseudomonas fluorescens
subsp.
cellulosa
Produced in
Escherichia coli
and Regulation of the Expression of Its Cloned Gene
Authors: Lejeune André , Courtois Stéphane , Colson Charles
Published: January 2020

Several enzymatic properties of an endoglucanase produced in
Escherichia coli
by a gene from
Pseudomonas fluorescens
subsp.
cellulosa
were investigated. Gel filtration revealed a single peak of
M
r
36,000 with endoglucanase activity. The pH optimum of the enzyme was 7.0. Carboxymethyl cellulose and barley β-glucan (mixed β-1,3 and 1,4 linkages) were good substrates, but not laminarin (β-1,3 linkages), amylose, filter paper, microcrystalline cellulose (Avicel), or cellotriose. The mode of action was typical of an “endo”-acting enzyme. Taken together, these properties do not correspond to those of any of the endoglucanases described in
P. fluorescens
subsp.
cellulosa
. Consequently, the gene was designated
egIX
. The enzyme was sensitive to end-product inhibition by cellobiose but was only moderately inhibited by glucose. The enzyme was formed constitutively in
E. coli
throughout the growth phase. Urea had no effect on endoglucanase synthesis, but glucose acted as a catabolite repressor. The formation of the enzyme in
E. coli
was partially dependent on cyclic AMP.
Assessment of recovery efficiency of beef extract reagents for concentrating viruses from municipal wastewater sludge solids by the organic flocculation procedure
Authors: Safferman R S , Rohr M E , Goyke T
Published: January 2020

This study was designed to assess the capacity of beef extract reagents to form flocs suitable for virus adsorption. Reagent comparisons resulted in the establishment of a modified organic flocculation procedure to concentrate viruses desorbed from sewage sludge solids with currently available modified powdered beef extracts. The method, based on supplementation with paste beef extract floc, achieved virus recoveries comparable to those obtained with powdered beef extract produced before a 1979 change in the manufacturing process. When primary settled sludge solids originating from mostly domestic waste were eluted with an unsupplemented modified powdered beef extract, high virus recovery efficiency was observed upon concentration by organic flocculation. This appreciable increase might have been due to floc-forming substances that were present in the primary settled sludge. These substances did not appear to be present in settled sludge collected from biologically treated wastes. Apparently, the floc-forming substances had been either removed or substantially altered during biological treatment.
Molecular Studies on the Role of a Root Surface Agglutinin in Adherence and Colonization by
Pseudomonas putida
Authors: Anderson Anne J. , Habibzadegah-Tari Pouran , Tepper Craig S.
Published: January 2020

Pseudomonas putida
aggressively colonizes root surfaces and is agglutinated by a root surface glycoprotein. Mutants of
P. putida
derived chemically or by Tn
5
insertion demonstrated enhanced or decreased agglutinability. Two nonagglutinable Tn
5
mutants (Agg
−
) and two mutants with enhanced agglutinability (Agg
s
) possessed Tn
5
in unique restriction sites. Agg
−
mutants colonized root surfaces of seedlings grown from inoculated seeds, but at levels lower than those observed with the Agg
+
parent. In short-term binding studies, Agg
−
cells adhered at levels that were 20- to 30-fold less than those for Agg
+
parental cells. These data suggest that the agglutination interaction plays a role in the attachment of
P. putida
to root surfaces.
Oxidative Pathway from Squalene to Geranylacetone in
Arthrobacter
sp. Strain Y-11
Authors: Ikeguchi Naoki , Nihira Takuya , Kishimoto Atsuko , Yamada Yasuhiro
Published: January 2020

The reaction pathway from squalene to
trans
-geranylacetone in
Arthrobacter
sp. strain Y-11 was studied. The enzyme or enzymes catalyzing squalene degradation were found to be membrane bound. Stoichiometric analysis of a cell-free system revealed that the ratio of squalene to
trans
-geranylacetone changed from 1:2 to 1:1 as the reaction proceeded, indicating two steps in geranylacetone formation. The initial step was found to be oxygenase catalyzed, from the absolute requirement for molecular oxygen in geranylacetone formation and the incorporation of
18
O into geranylacetone under
18
O
2
atmosphere. By using [
3
H]squalene as the substrate, we detected an intermediate in the pathway and identified it as 5,9,13-trimethyltetradeca-4,8,12-trienoic acid by mass spectrometry, infrared spectrometry, nuclear magnetic resonance spectrometry, and chemical synthesis. We deduced that squalene was first oxidatively cleaved to geranylacetone and the intermediate, and that the intermediate was further metabolized to geranylacetone. We also synthesized some of the presumptive metabolites, such as 4,8,12-trimethyltrideca-4,8,12-trien-2-one, and confirmed that they served as active precursors for geranylacetone formation. Based on these lines of evidence, we present here the pathway from squalene to
trans
-geranylacetone in
Arthrobacter
sp. strain Y-11.
Expression of Different Levels of Ethanologenic Enzymes from
Zymomonas mobilis
in Recombinant Strains of
Escherichia coli
Authors: Ingram L. O. , Conway T.
Published: January 2020

The expression of
Zymomonas mobilis
genes encoding pyruvate decarboxylase and alcohol dehydrogenase II in
Escherichia coli
converted this organism from the production of organic acids to the production of ethanol. Ethanol was produced during both anaerobic and aerobic growth. The extent to which these ethanologenic enzymes were expressed correlated with the extent of ethanol production. The replacement of organic acids with ethanol as a metabolic product during aerobic and anaerobic growth resulted in dramatic increases in final cell density, indicating that these acids (and the associated decline in pH) are more damaging than the production of ethanol. Of the plasmids examined, the best plasmid for growth and ethanol production expressed pyruvate decarboxylase and alcohol dehydrogenase II at levels of 6.5 and 2.5 IU/mg of total cell protein, respectively.
Prediction of Substrate Removal Rates of Attached Microorganisms and of Relative Contributions of Attached and Suspended Communities at Field Sites
Authors: Lewis David L. , Gattie David K.
Published: January 2020

A mathematical model composed of a direct proportionality relationship between bulk water velocities and field-determined second-order microbial transformation rate coefficients, and the relative rate coefficient of a benchmark chemical, was developed for estimating the substrate removal rates of rapidly degraded chemicals by attached organisms in shallow (<1 m deep) aquatic ecosystems. Data from 31 field experiments involving the addition of 2,4-dichlorophenoxyacetic acid methyl ester (2,4-DME) in nine field areas were used to determine a field-derived second-order rate coefficient for microbial transformation of the ester. By using 2,4-DME as a benchmark chemical, the model was used to predict microbial transformation rates of the butoxyethyl ester of 2,4-dichlorophenoxyacetic acid (2,4-DBE) at five other field sites. The predicted half-lives of 2,4-DBE varied 1,500-fold and were within about a threefold range or less of the measured half-lives. Under conditions of mass transport limitation, the contributions of attached microorganisms relative to total microbial activities at various field sites were related to the ratio of water velocity,
U
, and depth,
D
, showing that historical definitions of ecosystems according to flow and depth characteristics are also valid for describing the process-related structure of ecosystems. An equation was developed for predicting the relative contributions of attached and suspended communities with values of
U
and
D
for lotic and lentic ecosystems. On the basis of this equation, attached microorganisms were expected to be insignificant in deep lentic ecosystems and suspended microorganisms were expected to be insignificant in shallow lotic systems for the same process carried out by both populations. Neglecting epiphytic microorganisms, both suspended and attached organisms were expected to be significant in wetlands.
Enumeration of Tn5 mutant bacteria in soil by using a most- probable-number-DNA hybridization procedure and antibiotic resistance
Authors: Fredrickson J K , Bezdicek D F , Brockman F J , Li S W
Published: January 2020

Investigations were made into the utility of DNA hybridization in conjunction with a microdilution most-probable-number procedure for the enumeration of Rhizobium spp. and Pseudomonas putida in soil. Isolates of Rhizobium spp. and P. putida carrying the transposon Tn5 were added to sterile and nonsterile Burbank sandy loam soil and enumerated over time. Soil populations of rhizobia were enumerated by colony hybridization, most-probable-number-DNA hybridization procedure, plate counts, plant infectivity most probable number, and fluorescent antibody counts. Population values compared well for all methods at 5 and 30 days after the addition of cells, although the fluorescent antibody method tended to overestimate the viable population. In nonsterile soil, most-probable-number-DNA hybridization procedure enumerated as few as 10 P. putida Tn5 cells g of soil-1 and 100 R. leguminosarum bv. phaseoli Tn5 cells g of soil-1 and should have utility for following the fate of genetically engineered microorganisms released to the environment. Among the Kmr isolates containing Tn5, approximately 5% gave a dark, more intense autoradiograph when probed with 32P-labeled pGS9 DNA, which facilitated their detection in soil. Hybridization with a pCU101 probe (pGS9 without Tn5) indicated that donor plasmid sequences were being maintained in the bacterial chromosome. Transposon-associated antibiotic resistance was also utilized as a phenotypic marker. Tn5 vector-integrate mutants were successfully enumerated at low populations (10 to 100 cells g of soil-1) in soil by both phenotypic (Kmr) and genotypic (DNA probe) analysis. However, determination of the stability of Tn5 or Tn5 and vector sequences in the bacteria is necessary.
Relationship of Intracellular Coenzyme F
420
Content to Growth and Metabolic Activity of
Methanobacterium bryantii
and
Methanosarcina barkeri
Authors: Heine-Dobbernack E. , Schoberth S. M. , Sahm H.
Published: January 2020

The use of F
420
as a parameter for growth or metabolic activity of methanogenic bacteria was investigated. Two representative species of methanogens were grown in batch culture:
Methanobacterium bryantii
(strain M.o.H.G.) on H
2
and CO
2
, and
Methanosarcina barkeri
(strain Fusaro) on methanol or acetate. The total intracellular content of coenzyme F
420
was followed by high-resolution fluorescence spectroscopy. F
420
concentration in
M. bryantii
ranged from 1.84 to 3.65 μmol · g of protein
−1
; and in
M. barkeri
grown with methanol it ranged from 0.84 to 1.54 μmol · g
−1
depending on growth conditions. The content of F
420
in
M. barkeri
was influenced by a factor of 2 depending on the composition of the medium (minimal or complex) and by a factor of 3 to 4 depending on whether methanol or acetate was used as the carbon source. A comparison of F
420
content with protein, cell dry weight, optical density, and specific methane production rate showed that the intracellular content of F
420
approximately followed the increase in biomass in both strains. In contrast, no correlation was found between specific methane production rate and intracellular F
420
content. However, qCH
4
(F
420
), calculated by dividing the methane production rate by the coenzyme F
420
concentration, almost paralleled qCH
4
(protein). These results suggest that F
420
may be used as a specific parameter for estimating the biomass, but not the metabolic activity, of methanogens; hence qCH
4
(F
420
) determined in mixed populations with complex carbon substrates must be considered as measure of the actual methanogenic activity and not as a measure of potential activity.
Purification and Properties of Primary and Secondary Alcohol Dehydrogenases from
Thermoanaerobacter ethanolicus
Authors: Bryant Frank O. , Wiegel Juergen , Ljungdahl Lars G.
Published: January 2020

Thermoanaerobacter ethanolicus
(ATCC 31550) has primary and secondary alcohol dehydrogenases. The two enzymes were purified to homogeneity as judged from sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration. The apparent
M
r
s of the primary and secondary alcohol dehydrogenases are 184,000 and 172,000, respectively. Both enzymes have high thermostability. They are tetrameric with apparently identical subunits and contain from 3.2 to 5.5 atoms of Zn per subunit. The two dehydrogenases are NADP dependent and reversibly convert ethanol and 1-propanol to the respective aldehydes. The
V

m

values with ethanol as a substrate are 45.6 μmol/min per mg for the primary alcohol dehydrogenase and 13 μmol/min per mg for the secondary alcohol dehydrogenase at pH 8.9 and 60°C. The primary enzyme oxidizes primary alcohols, including up to heptanol, at rates similar to that of ethanol. It is inactive with secondary alcohols. The secondary enzyme is inactive with 1-pentanol or longer chain alcohols. Its best substrate is 2-propanol, which is oxidized 15 times faster than ethanol. The secondary alcohol dehydrogenase is formed early during the growth cycle. It is stimulated by pyruvate and has a low
K

m

for acetaldehyde (44.8 mM) in comparison to that of the primary alcohol dehydrogenase (210 mM). The latter enzyme is formed late in the growth cycle. It is postulated that the secondary alcohol dehydrogenase is largely responsible for the formation of ethanol in fermentations of carbohydrates by
T. ethanolicus.