bluuebunny/crossref_metadata_2025 huggingface dataset:
Restriction enzyme analysis of Listeria monocytogenes strains associated with food-borne epidemics
Authors: Wesley I V , Ashton F
Published: January 2020

Listeria monocytogenes (serotype 4b) has caused four major food-borne epidemics in North America. In this study, L. monocytogenes isolates from the Nova Scotia (Canada), Boston (Mass.), and Los Angeles (Calif.) outbreaks were examined by restriction enzyme analysis with the endonuclease HhaI. Human isolates (n = 32) from the 1981 Canadian outbreak were compared with a strain recovered from coleslaw, which was epidemiologically incriminated as the vehicle of infection. After HhaI digestion, 29 of 32 isolates exhibited the restriction enzyme pattern of the reference coleslaw isolate. The restriction enzyme patterns of the nine clinical isolates from the 1983 Massachusetts outbreak were identical to each other but differed from those of raw milk isolates recovered from sources supplying the pasteurizer. Isolates (n = 48) from the 1985 California outbreak were evaluated. The restriction enzyme patterns of the L. monocytogenes isolates from humans and from the suspect cheese samples were identical to those of four of five cheese factory environmental isolates. Isolates from each of these outbreaks exhibited a restriction enzyme pattern that was characteristic of that outbreak. The case with which restriction enzyme analysis can be applied to all serotypes of L. monocytogenes argues for its use in the epidemiology of L. monocytogenes.
Identification of
para
-Cresol as a Growth Factor for
Methanoplanus endosymbiosus
Authors: Poirot Chris C. M. , Van Alebeek Gert-Jan W. M. , Keltjens Jan T. , Vogels Godfried D.
Published: January 2020

Growth of the methanogenic bacterium
Methanoplanus endosymbiosus
is dependent on the presence of ruminal fluid. Ruminal fluid could be replaced by the eluate of a rumen-derived anaerobic digester. From the eluate of the digester, a growth-stimulatory component was purified and identified as
p
-cresol. Authentic
p
-cresol supported a half-maximal growth rate of the organism at 50 nM concentration.
Evaluation of an Escherichia coli host strain for enumeration of F male-specific bacteriophages
Authors: Debartolomeis J , Cabelli V J
Published: January 2020

A method was developed for the selective enumeration of F male-specific bacteriophages in samples of environmental waters. The host strain for the phages, Escherichia coli HS(pFamp)R, has three antibiotic resistance markers, ampicillin on the Famp plasmid, which codes for pilus production, and streptomycin and nalidixic acid on the chromosome. The strain is resistant to coliphages T2 to T7 and phi X174. More than 95% of the phages from environmental samples which plaqued on the host strain were F male specific. The host bacterium had a higher plaquing efficiency than E. coli K-12 Hfr for F-specific phages in stock suspensions and sewage effluents. The F male-specific phage levels in prechlorinated, secondary-treated sewage effluents generally were about 10(3) to 10(4) PFU/100 ml. The levels in the influents to the sewage treatment plants and in septic tank contents were about 10(5) PFU/100 ml. RNA-containing phages composed about 90% of the total F-specific phage population in sewage effluents.
Expression, localization, and functional analysis of polychlorinated biphenyl degradation genes cbpABCD of Pseudomonas putida
Authors: Khan A A , Walia S K
Published: January 2020

Genes of Pseudomonas putida strains that are capable of degrading polychlorinated biphenyls were cloned in the plasmid vector pUC19. The resultant hybrid plasmid, pAW6194, contained cbpABCD genes on a 9.0-kb DNA fragment that was necessary for the catabolism of polychlorinated biphenyls. These genes were further subcloned on an 8.0-kb HindIII fragment of pAW540. Degradation of 3-chlorobiphenyl, 2,4-dichlorobiphenyl, and 2,4,5-trichlorobiphenyl into a chloro derivative of benzoic acid was found in Escherichia coli harboring chimeric plasmid pAW540. Expression of cbpA (biphenyl dioxygenase, 6.2 U/mg of protein) and cbpC (3-phenylcatechol dioxygenase, 611.00 U/mg of protein) genes was also found in E. coli containing the hybrid plasmid pAW540. These enzyme activities were up to 10-fold higher than those found in P. putida OU83. These results led us to conclude that cbpABCD genes of P. putida OU83 were encoded on cloned DNA and expressed in E. coli. Whether the expression of cbpABCD genes of P. putida OU83 was driven by its own promoters located on the cloned DNA or by the lacZ promoter of pUC19 was examined by subcloning a 8.0-kb DNA fragment encoding the cbpABCD genes, in both orientations, in the HindIII site of the promoter probe vector pKK232-8. The resulting recombinant plasmids, pAW560 and pAW561, expressed cbpABCD genes and conferred chloramphenicol resistance only in E. coli harboring pAW560, indicating that the expression of chloramphenicol acetyltransferase is independent of cbpABCD gene expression.(ABSTRACT TRUNCATED AT 250 WORDS)
Selection of cyanobacteria isolated from mosquito breeding sites as a potential food source for mosquito larvae
Authors: Thiery I , Nicolas L , Rippka R , Tandeau de Marsac N
Published: January 2020

One way to increase the persistence of larvicidal toxins in mosquito breeding sites is to clone the corresponding genes in microorganisms, such as cyanobacteria, which could serve as a source of food for the larvae. We isolated and cultured 10 strains of cyanobacteria from three mosquito breeding sites along the French Mediterranean coast. Most of the strains were tolerant to a relatively wide range of salt concentrations, and all of them were totally or partially resistant to at least four of the five biological or chemical larvicides used in the local mosquito control program. Six unicellular strains from these habitats and Synechococcus strain PCC 7942, a strain maintained for more than 10 years under laboratory conditions, were assessed for ingestion and digestion by larvae Culex pipiens and Anopheles gambiae mosquitoes. The numbers of cells ingested and digested were dependent on the cyanobacterial strain and varied with the mosquito species. Three of the new isolates, Synechococcus strain PCC 8905 and Synechocystis strains PCC 8906 and PCC 8912, were ingested and digested rapidly by larvae of both mosquito species. Since these strains are also tolerant to larvicides and relatively resistant to elevated salt concentrations, they meet the basic requirements for potential recipients of bacterial genes that encode endotoxins.
Expression of an L-alanine dehydrogenase gene in Zymomonas mobilis and excretion of L-alanine
Authors: Uhlenbusch I , Sahm H , Sprenger G A
Published: January 2020

An approach to broaden the product range of the ethanologenic, gram-negative bacterium Zymomonas mobilis by means of genetic engineering is presented. Gene alaD for L-alanine dehydrogenase (EC 1.4.1.1.) from Bacillus sphaericus was cloned and introduced into Z. mobilis. Under the control of the strong promoter of the pyruvate decarboxylase (pdc) gene, the enzyme was expressed up to a specific activity of nearly 1 mu mol . min -1 . mg of protein -1 in recombinant cells. As a results of this high L-alanine dehydrogenase activity, growing cells excreted up to 10 mmol of alanine per 280 mmol of glucose utilized into a mineral salts medium. By the addition of 85 mM NH4+ to the medium, growth of the recombinant cells stopped, and up to 41 mmol alanine was secreted. As alanine dehydrogenase competed with pyruvate decarboxylase (PDC) (EC 4.1.1.1.) for the same substrate (pyruvate), PDC activity was reduced by starvation for the essential PDC cofactor thiamine PPi. A thiamine auxotrophy mutant of Z. mobilis which carried the alaD gene was starved for 40 h in glucose-supplemented mineral salts medium and then shifted to mineral salts medium with 85 mM NH4+ and 280 mmol of glucose. The recombinants excreted up to 84 mmol of alanine (7.5 g/liter) over 25 h. Alanine excretion proceeded at an initial velocity of 238 nmol . min-1 . mg [dry weight]-1. Despite this high activity, the excretion rate seemed to be a limiting factor, as the intracellular concentration of alanine was as high as 260 mM at the beginning of the excretion phase and decreased to 80 to 90 mM over 24 h.
Cloning, sequencing, and expression of the N-acyl-D-mannosamine dehydrogenase gene from Flavobacterium sp. strain 141-8 in Escherichia coli
Authors: Yamamoto-Otake H , Koyama Y , Horiuchi T , Nakano E
Published: January 2020

The gene coding for N-acyl-D-mannosamine dehydrogenase (NAM-DH) from Flavobacterium sp. strain 141-8 was cloned and expressed under the control of a lac promoter in Escherichia coli JM109. The DNA sequence of the gene was determined, and an open reading frame encoding a polypeptide composed of 272 amino acid residues (Mr, 27,473) was identified. The E. coli transformants which showed over 200-fold higher NAM-DH activity than did the Flavobacterium strain produced the enzyme as a protein fused with beta-galactosidase. Despite being a fusion, NAM-DH produced by E. coli transformants appeared unchanged in pH optimum, Km, and substrate specificity from Flavobacterium sp. strain 141-8. This newly recombinant enzyme may be applicable to the quantitative determination of sialic acid in serum.
Polyethylene glycol-induced internalization of bacteria into fungal protoplasts: electron microscopic study and optimization of experimental conditions
Authors: Guerra-Tschuschke I , Martín I , González M T
Published: January 2020

We studied the mechanism of internalization of Escherichia coli into Saccharomyces cerevisiae induced by polyethylene glycol (PEG) and optimized the experimental conditions. Transmission electron microscope studies revealed that the principal factor involved in the internalization was the degree of cell aggregation attained. Internalization occurred mainly by an endocytosis-like mechanism and took place during the elimination of PEG. The optimum conditions were to treat a mixed pellet of both microorganisms with 15% PEG and then gradually dilute the polymer. The same conditions were applied to E. coli and Aspergillus nidulans, with similar results.
Biotransformation of the Pentahydroxy Flavone Quercetin by
Rhizobium loti
and
Bradyrhizobium
Strains (
Lotus
)
Authors: Rao J. R. , Sharma N. D. , Hamilton J. T. G. , Boyd D. R. , Cooper J. E.
Published: January 2020

Lotus
rhizobia catabolized quercetin in an arabinose-based medium via a novel form of C-ring cleavage, yielding phloroglucinol and protocatechuic acid. Conservation of the A and B rings of the flavone suggests that a chalcone could be formed as a transient intermediate.
Performance characterization of a model bioreactor for the biodegradation of trichloroethylene by Pseudomonas cepacia G4
Authors: Folsom B R , Chapman P J
Published: January 2020

Pseudomonas cepacia G4 grown in chemostats with phenol demonstrated constant specific degradation rates for both phenol and trichloroethylene (TCE) over a range of dilution rates. Washout of cells from chemostats was evident at a dilution rate of 0.2 h-1 at 28 degrees C. Increased phenol concentrations in the nutrient feed led to increased biomass production with constant specific degradation rates for both phenol and TCE. The addition of lactate to the phenol feed led to increased biomass production but lowered specific phenol and TCE degradation rates. The maximum potential for TCE degradation was about 1.1 g per day per g of cell protein. Cell growth and degradation kinetic parameters were used in the design of a recirculating bioreactor for TCE degradation. In this reactor, the total amount of TCE degraded increased as either reaction time or biomass was increased. TCE degradation was observed up to 300 microM TCE with no significant decreases in rates. On the average, this reactor was able to degrade 0.7 g of TCE per day per g of cell protein. These results demonstrate the feasibility of TCE bioremediation through the use of bioreactors.
Typing of human, animal, food, and environmental isolates of Listeria monocytogenes by multilocus enzyme electrophoresis
Authors: Boerlin P , Piffaretti J C
Published: January 2020

In order to elucidate some aspects of the epidemiology of listeriosis in Switzerland, 181 strains of Listeria monocytogenes isolated from humans, animals, food, and the environment have been analyzed by multilocus enzyme electrophoresis at 21 enzyme loci. The clone responsible for several recent food-borne outbreaks in Switzerland and in North America (marked by electrophoretic type 1 and serovar 4b) has been found frequently among strains isolated from animals. Thus, animals may represent a major source of diffusion of this clone in the environment and in food, in which it has been found only sporadically, however. Two other unrelated clones (including strains belonging to serovars 1/2b and 1/2c) have often been isolated from meat but not from animals. These findings indicate that contamination of meat with L. monocytogenes might originate mainly from the environment in which it is processed rather than from animals themselves. This could explain the differences in the distribution of L. monocytogenes serovars isolated from meat and from animals.
Chemical Characterization of Polysaccharide from the Slime Layer of the Cyanobacterium
Microcystis flos-aquae
C3-40
Authors: Plude John L. , Parker Dorothy L. , Schommer Olivia J. , Timmerman Robert J. , Hagstrom Stephanie A. , Joers James M. , Hnasko Robert
Published: January 2020

Macromolecular material from the slime layer of the cyanobacterium
Microcystis flos-aquae
C3-40 was defined as material that adhered to cells during centrifugation in growth medium but was dislodged by washing with deionized water and retained within dialysis tubing with a molecular-weight cutoff of 3,500. At each step of this isolation procedure, the slime was observed microscopically. Cells in the centrifugal pellet were surrounded by large amounts of slime that excluded negative stain, whereas cells that had been washed with water lacked visible slime. Two independently isolated lots of slime contained no detectable protein (<1%, wt/wt) and consisted predominantly of anthrone-reacting polysaccharide. Sugars in a hydrolysate of slime polysaccharide were derivatized with trimethylsilylimidazole and examined by gas chromatography-mass spectrometry. The composition of the slime polysaccharide was 1.5% (wt/wt) galactose, 2.0% glucose, 3.0% xylose, 5.0% mannose, 5.5% rhamnose, and 83% galacturonic acid. This composition resembles that of the plant polysaccharide pectin, which was treated in parallel as a control. Consistent with earlier indications that
M. flos-aquae
slime preferentially binds certain cations, the ratio of Fe to Na in the dialyzed slime was 10
4
times that in the growth medium. The composition of the slime is discussed with respect to possible mechanisms of cation binding in comparison with other cyanobacterial exopolysaccharides and pectin.
Effects of increased transaldolase activity on D-xylulose and D-glucose metabolism in Saccharomyces cerevisiae cell extracts
Authors: Senac T , Hahn-Hägerdal B
Published: January 2020

In vitro metabolism of D-xylulose and D-glucose in extracts obtained from D-glucose- and D-xylulose-fermenting Saccharomyces cerevisiae cells was investigated with 10- and 100-fold-increased activity of the enzyme transaldolase (EC 2.2.1.2). The rate of sugar consumption was the same in most cases, whereas the rate of ethanol formation decreased with increased levels of transaldolase. The formation of glycerol, pentitols, and acetic acid was not dependent on added transaldolase but was dependent on the sugar used as the growth substrate and on the sugar used in the in vitro metabolism experiments. The carbon balance showed that the dissimilated carbon could not be accounted for in products when transaldolase was added. The concentration of D-fructose-1,6.-diphosphate in the extracts was not influenced by added transaldolase but was higher with D-xylulose than with D-glucose. Levels of pyruvate, comparable with the two substrates, decreased with increasing levels of transaldolase. Exogenously added transaldolase decreased D-sedoheptulose-7-phosphate levels when D-xylulose was the substrate. The results are discussed in relation to the dissimilation of carbon through the upper part of glycolysis and the pentose phosphate pathway.
Diversity and Population Dynamics of Methanogenic Bacteria in a Granular Consortium
Authors: Visser Frans A. , van Lier Jules B. , Macario Alberto J. L. , de Macario Everly Conway
Published: January 2020

Upflow anaerobic sludge blanket bioreactor granules were used as an experimental model microbial consortium to study the dynamics and distribution of methanogens. Immunologic methods revealed a considerable diversity of methanogens that was greater in mesophilic granules than in the same granules 4 months after a temperature shift from 38 to 55°C. During this period, the sizes of the methanogenic subpopulations changed with distinctive profiles after the initial reduction caused by the shift. Methanogens antigenically related to
Methanobrevibacter smithii
PS and ALI,
Methanobacterium hungatei
JF1, and
Methanosarcina thermophila
TM1 increased rapidly, reached a short plateau, and then fell to lower concentrations that persisted for the duration of the experiment. A methanogen related to
Methanogenium cariaci
JR1 followed a similar profile at the beginning, but it soon diminished below detection levels.
Methanothrix
rods weakly related to the strain Opfikon increased rapidly, reaching a high-level, long-lasting plateau. Two methanogens related to
Methanobrevibacter arboriphilus
AZ and
Methanobacterium thermoautotrophicum
ΔH emerged from very low levels before the temperature shift and multiplied to attain their highest numbers 4 months after the shift. Histochemistry and immunohistochemistry revealed thick layers, globular clusters, and lawns of variable density which were distinctive of the methanogens related to
M. thermoautotrophicum
ΔH,
M. thermophila
TM1, and
M. arboriphilus
AZ and
M. soehngenii
Opfikon, respectively, in thin sections of granules grown at 55°C for 4 months. Mesophilic granules showed a different pattern of methanogenic subpopulations.
Control of the Lysine Biosynthesis Sequence in
Corynebacterium glutamicum
as Analyzed by Overexpression of the Individual Corresponding Genes
Authors: Cremer Josef , Eggeling Lothar , Sahm Hermann
Published: January 2020

The gene cluster that codes for feedback-resistant aspartate kinase (
lysC
α and
lysC
β) and aspartate semialdehyde dehydrogenase (
asd
) was cloned from a mutant strain of
Corynebacterium glutamicum.
Its functional analysis by subcloning, enzyme assays, and type of aspartate kinase regulation enabled the isolation of a fragment for separate expression of the feedback-resistant kinase without aspartate semialdehyde dehydrogenase expression. This was used together with other clones constructed (J. Cremer, L. Eggeling, and H. Sahm, Mol. Gen. Genet. 220:478-480, 1990) to overexpress individually each of the six genes that convert aspartate to lysine. Analysis of lysine formation revealed that overexpression of the feedback-resistant kinase alone suffices to achieve lysine formation (38 mM). Also, sole overexpression of wild-type dihydrodipicolinate synthase resulted in lysine formation but in a lower amount (11 mM). The other four enzymes had no effect on lysine secretion. With a plasmid overexpressing both relevant enzymes together, a further increase in lysine yield was obtained. This shows that of the six enzymes that convert aspartate to lysine the kinase and the synthase are responsible for flow control in the wild-type background and can be useful for construction of lysine-producing strains.
Assay for detection and enumeration of genetically engineered microorganisms which is based on the activity of a deregulated 2,4-dichlorophenoxyacetate monooxygenase
Authors: King R J , Short K A , Seidler R J
Published: January 2020

An assay system was developed for the enumeration of genetically engineered microorganisms expressing a deregulated 2,4-dichlorophenoxyacetate (TFD) monooxygenase, which converts phenoxyacetate (PAA) to phenol. In PAA-amended cultures of Pseudomonas aeruginosa PAO1C(pRO103) and Pseudomonas putida PPO301(pRO103), strains which express a deregulated TFD monooxygenase, phenol production was proportional to cell number. Phenol was reacted, under specific conditions, with a 4-aminoantipyrine dye to form an intensely colored dye-phenol complex (AAPPC), which when measured spectrophotometrically could detect as few as 10(3) cells per ml. This assay was corroborated by monitoring the disappearance of PAA and the accumulation of phenol by high-performance liquid chromatography and gas chromatography. The AAPPC assay was modified for use with plate cultures and clearly distinguished colonies of PPO301(pRO103) and PAO1C(pRO103) from a strain expressing a regulated TFD monooxygenase. Colonies of P. putida PPO301(pRO101) remained cream colored, while colonies of PPO301(pRO103) and PAO1C(pRO103) turned a distinct red.
Incompatibility of
Lactobacillus
Vectors with Replicons Derived from Small Cryptic
Lactobacillus
Plasmids and Segregational Instability of the Introduced Vectors
Authors: Posno M. , Leer R. J. , van Luijk N. , van Giezen M. J. F. , Heuvelmans P. T. H. M. , Lokman B. C. , Pouwels P. H.
Published: January 2020

Three new
Lactobacillus
vectors based on cryptic
Lactobacillus
plasmids were constructed. The shuttle vector pLP3537 consists of a 2.3-kb plasmid from
Lactobacillus pentosus
MD353, an erythromycin resistance gene from
Staphylococcus aureus
plasmid pE194, and pUC19 as a replicon for
Escherichia coli
. The vectors pLPE317 and pLPE323, which do not contain
E. coli
sequences, were generated by introducing the erythromycin resistance gene of pE194 into a 1.7- and a 2.3-kb plasmid from
L. pentosus
MD353, respectively. These vectors and the shuttle vector pLP825 (M. Posno, R. J. Leer, J. M. M. van Rijn, B. C. Lokman, and P. H. Pouwels, p. 397-401,
in
A. T. Ganesan and J. A. Hoch, ed., Genetics and biotechnology of bacilli, vol. 2, 1988) could be introduced by electroporation into
Lactobacillus casei, L. pentosus, L. plantarum, L. acidophilus, L. fermentum
, and
L. brevis
strains with similar efficiencies. Transformation efficiencies were strain dependent and varied from 10
2
to 10
7
transformants per μg of DNA. Plasmid DNA analysis of
L. pentosus
MD353 transformants revealed that the introduction of pLP3537 or pLPE323 was invariably accompanied by loss of the endogenous 2.3-kb plasmid. Remarkably, pLPE317 could only be introduced into an
L. pentosus
MD353 strain that had been previously cured of its endogenous 1.7-kb plasmid. The curing phenomena are most likely to be explained by the incompatibility of the vectors and resident plasmids.
Lactobacillus
vectors are generally rapidly lost when cells are cultivated in the absence of selective pressure. However, pLPE323 is stable in three of four
Lactobacillus
strains tested so far.
Hydrogen Oxidation by the Host-Controlled Uptake Hydrogenase Phenotype of
Bradyrhizobium japonicum
in Symbiosis with Soybean Host Plants
Authors: van Berkum Peter , Sloger Charles
Published: January 2020

Symbioses between uptake hydrogenase host-regulated (Hup-hr) phenotypes of
Bradyrhizobium japonicum
and exotic, agronomically unadapted soybean germ plasm were examined for expression of uptake hydrogenase activity. Determinations for hydrogen evolution and uptake hydrogenase activity identified five plant introduction (PI) lines which formed hydrogen-oxidizing symbioses with strains USDA 61 and PA3 6c. Hup-hr strains belonging to serogroup 94 expressed uptake hydrogenase activity in symbioses with PI 181696 and PI 219655 at rates sufficient to prevent hydrogen from escaping the nodules. The identification of soybean germ plasm forming hydrogen-oxidizing symbioses with Hup-hr bradyrhizobia potentially has implications for enhancing nitrogen fixation efficiency in soybean production.
Chemical control of interstrain competition for soybean nodulation by Bradyrhizobium japonicum
Authors: Cunningham S , Kollmeyer W D , Stacey G
Published: January 2020

Previous research has shown that a significant limitation to the agricultural use of improved rhizobial inoculant strains is competition from the indigenous soil population. In this work, we sought to test whether chemical inhibitors of flavonoid-induced nod gene expression in Bradyrhizobium japonicum could be identified and utilized to affect interstrain competition for nodulation of soybeans. Approximately 1,000 structural and functional analogs of the known, natural inducers of nod gene expression were tested on six strains of B. japonicum containing a nodY-lacZ fusion. We successfully identified effective inhibitors of nodY expression. The addition of the inhibitor 7-hydroxy-5-methylflavone significantly inhibited nodulation by a sensitive strain and could be used to effectively manipulate the competition between strains for soybean nodulation. However, this work also uncovered significant limitations for the practical use of this methodology. For example, despite the almost universal induction response to the identified natural inducers, there was a wide variability among strains in their response to any specific inhibitor. Given this unexpected variability, the cost of registration of an agronomic chemical, and the potential for the development of resistant field populations, it is unlikely that chemical inhibitors can be successfully applied to a field situation.
Mutants of Pseudomonas cepacia G4 defective in catabolism of aromatic compounds and trichloroethylene
Authors: Shields M S , Montgomery S O , Cuskey S M , Chapman P J , Pritchard P H
Published: January 2020

Pseudomonas cepacia G4 possesses a novel pathway of toluene catabolism that is shown to be responsible for the degradation of trichloroethylene (TCE). This pathway involves conversion of toluene via o-cresol to 3-methylcatechol. In order to determine the enzyme of toluene degradation that is responsible for TCE degradation, chemically induced mutants, blocked in the toluene ortho-monooxygenase (TOM) pathway of G4, were examined. Mutants of the phenotypic class designated TOM A- were all defective in their ability to oxidize toluene, o-cresol, m-cresol, and phenol, suggesting that a single enzyme is responsible for conversion of these compounds to their hydroxylated products (3-methylcatechol from toluene, o-cresol, and m-cresol and catechol from phenol) in the wild type. Mutants of this class did not degrade TCE. Two other mutant classes which were blocked in toluene catabolism, TOM B-, which lacked catechol-2,3-dioxygenase, and TOM C-, which lacked 2-hydroxy-6-oxoheptadienoic acid hydrolase activity, were fully capable of TCE degradation. Therefore, TCE degradation is directly associated with the monooxygenation capability responsible for toluene, cresol, and phenol hydroxylation.
Production and Degradation of Oxalic Acid by Brown Rot Fungi
Authors: Espejo Eduardo , Agosin Eduardo
Published: January 2020

Our results show that all of the brown rot fungi tested produce oxalic acid in liquid as well as in semisolid cultures.
Gloeophyllum trabeum
, which accumulates the lowest amount of oxalic acid during decay of pine holocellulose, showed the highest polysaccharide-depolymerizing activity. Semisolid cultures inoculated with this fungus rapidly converted
14
C-labeled oxalic acid to CO
2
during cellulose depolymerization. The other brown rot fungi also oxidized
14
C-labeled oxalic acid, although less rapidly. In contrast, semisolid cultures inoculated with the white rot fungus
Coriolus versicolor
did not significantly catabolize the acid and did not depolymerize the holocellulose during decay. Semisolid cultures of
G. trabeum
amended with desferrioxamine, a specific iron-chelating agent, were unable to lower the degree of polymerization of cellulose or to oxidize
14
C-labeled oxalic acid to the extent or at the rate that control cultures did. These results suggest that both iron and oxalic acid are involved in cellulose depolymerization by brown rot fungi.
Construction of an Acid-Tolerant
Rhizobium leguminosarum
Biovar Trifolii Strain with Enhanced Capacity for Nitrogen Fixation
Authors: Chen Hancai , Richardson Alan E. , Gartner Elena , Djordjevic Michael A. , Roughley Rodney J. , Rolfe Barry G.
Published: January 2020

Strain ANU1173 is an acid-tolerant
Rhizobium leguminosarum
biovar trifolii strain that is able to nodulate subterranean clover plants growing in agar culture at pH 4.4 At pH 6.5, its symbiotic effectiveness in association with
Trifolium subterraneum
cv. Mt. Barker was 80% relative to that of strain ANU794, a Sm
r
derivative of the commercial inoculant
R. leguminosarum
bv. trifolii TA1. Strain ANU1173 contained four indigenous megaplasmids, the smallest of these being the symbiotic (Sym) plasmid. The critical pH requirement for growth of strain ANU1173 in laboratory media was shown not to be associated with this plasmid. When the Sym plasmid of strain ANU1173(pSym-1173) was mobilized into a Nod
-
strain of
R. leguminosarum
bv. viciae, the plasmid conferred to the transconjugant a level of symbiotic effectiveness in association with
T. subterraneum
that was similar to that observed with ANU1173. The symbiotic effectiveness of strain ANU1173 was improved by first curing pSym-1173 (generating strain ANU1184) and replacing it with another
R. leguminosarum
bv. trifolii Sym plasmid, pBR1AN. Subterranean clover plants inoculated with strain ANU1184 (pBR1AN) exhibited a 35 or 53% increase in acetylene reduction activity and a 20 or 17% increase in dry weight when grown at pH 6.5 and pH 4.4, respectively, compared with plants inoculated with strain ANU1173 and grown under the same pH conditions. It was further shown that pBR1AN was stably maintained in strain ANU1184 under free-living and symbiotic conditions. These results indicate that it is possible to construct an acid-tolerant strain of
R. leguminosarum
bv. trifolii with an enhanced capacity for nitrogen fixation.
Nutritional complementation of oxidative glucose metabolism in Escherichia coli via pyrroloquinoline quinone-dependent glucose dehydrogenase and the Entner-Doudoroff pathway
Authors: Adamowicz M , Conway T , Nickerson K W
Published: January 2020

Two glucose-negative Escherichia coli mutants (ZSC113 and DF214) were unable to grow on glucose as the sole carbon source unless supplemented with pyrroloquinoline quinone (PQQ). PQQ is the cofactor for the periplasmic enzyme glucose dehydrogenase, which converts glucose to gluconate. Aerobically, E. coli ZSC113 grew on glucose plus PQQ with a generation time of 65 min, a generation time about the same as that for wild-type E. coli in a defined glucose-salts medium. Thus, for E. coli ZSC113 the Enter-Doudoroff pathway was fully able to replace the Embden-Meyerhof-Parnas pathway. In the presence of 5% sodium dodecyl sulfate, PQQ no longer acted as a growth factor. Sodium dodecyl sulfate inhibited the formation of gluconate from glucose but not gluconate metabolism. Adaptation to PQQ-dependent growth exhibited long lag periods, except under low-phosphate conditions, in which the PhoE porin would be expressed. We suggest that E. coli has maintained the apoenzyme for glucose dehydrogenase and the Entner-Doudoroff pathway as adaptations to an aerobic, low-phosphate, and low-detergent aquatic environment.
Use of an A-T-rich DNA clone for identification and detection of Peronosclerospora sorghi
Authors: Yao C L , Magill C W , Frederiksen R A
Published: January 2020

A recombinant plasmid, pMLY12-1, screened from a Peronosclerospora sorghi library hybridizes only to DNA of P. sorghi, or to DNA from leaves infected with P. sorghi, not to DNA of P. sorghi Thailand isolate, P. philippinensis, P. sacchari, or P. maydis. The terminal sequences of the 1.3-kb insert, which appears to contain mitochondrial DNA, are 85% A and T. No polymorphisms were detected when the probe was hybridized to Southern blots containing DNA from P. sorghi pathotype 1, pathotype 3, or a Botswana isolate digested with any of the eight restriction endonucleases tested. The banding patterns were the same whether DNA was extracted directly from the fungus or from infected leaves.
Reductive dechlorination of Tri- and tetrachloroethylenes depends on transition from aerobic to anaerobic conditions
Authors: Kästner M
Published: January 2020

Aerobic enrichment cultures from contaminated groundwaters dechlorinated trichloroethylene (TCE) (14.6 mg/liter; 111 mumol/liter) and tetrachloroethylene (PCE) (16.2 mg/liter; 98 mumol/liter) reductively within 4 days after the transition from aerobic to anaerobic conditions. The transformation products were equimolar amounts of cis-1,2-dichloroethylene and traces of 1,1-dichloroethylene. No other chlorinated product and no methane were detected. The change was accompanied by the release of sulfide, which caused a decrease in the redox potential from 0 to -150 mV. In sterile control experiments, sulfide led to the abiotic formation of traces of 1,1-dichloroethylene without cis-1,2-dichloroethylene production. The reductive dechlorination of PCE via TCE depended on these specific transition conditions after consumption of the electron acceptor oxygen or nitrate. Repeated feeding of TCE or PCE to cultures after the change to anaerobic conditions yielded no further dechlorination. Only aerobic subcultures with an air/liquid ratio of 1:4 maintained dechlorination activities; anaerobic subcultures showed no transformation. Bacteria from noncontaminated sites showed no reduction under the same conditions.
Potential for thermophilic (50 degrees C) anaerobic dechlorination of pentachlorophenol in different ecosystems
Authors: Larsen S , Hendriksen H V , Ahring B K
Published: January 2020

Thermophilic (50 degrees C) anaerobic biodegradation of pentachlorophenol (PCP) was investigated by using different inocula from natural ecosystems and anaerobic digesters. The inocula tested were three freshwater sediments, four anaerobic sewage sludge samples from digesters treating sludge from wastewater plants with various industrial inputs, and digested manure from an anaerobic reactor. Only one digested-sludge sample and the manure sample were from thermophilic environments. The initial PCP concentration was 7.5 or 37.5 microM. After 8 months, PCP had disappeared from the sediment samples and various, less chlorinated intermediates were present. Additions of extra PCP were degraded within 4 weeks, and a maximal observed dechlorination rate of 1.61 mumol/liter/day in the vials with addition of 7.5 microM PCP and 7.50 mumol/liter/day in the vials with addition of 37.5 microM PCP were measured for a freshwater sediment. In contrast, only 2.8 to 17.5% of the initial PCP added had disappeared from the sludge samples after 8 months of incubation. The complex pattern of intermediates formed indicated that the dechlorination of PCP proceeded via different pathways, involving at least two different populations in the dechlorination processes.
Examination of retail chickens and sausages in Britain for vero cytotoxin-producing Escherichia coli
Authors: Smith H R , Cheasty T , Roberts D , Thomas A , Rowe B
Published: January 2020

Samples from chickens and pork sausages were examined for the presence of Vero cytotoxin-producing Escherichia coli by using DNA probes for the Vero cytotoxin genes. Hybridization was detected in 25% of the 184 sausage samples, but none of the chickens was positive. No E. coli O157:H7 strains were isolated, and serotyping showed that the Vero cytotoxin-producing E. coli strains belonged to eight different O serogroups and that six strains had an unidentifiable O antigen.
Hyphomonas
spp.,
Shewanella
spp., and Other Marine Bacteria Lack Heterogeneous (Ladderlike) Lipopolysaccharides
Authors: Sledjeski Darren D. , Weiner Ronald M.
Published: January 2020

The lipopolysaccharides (LPS) of 19 marine bacteria were examined for size heterogeneities by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis in conjunction with an LPS-specific silver staining method. Fifteen marine bacteria had an R-type LPS instead of the ladderlike LPS array characteristic of most bacteria. In addition, three marine bacteria also had a single large LPS molecule. Without constraints (e.g., surface masking), R-type LPS, a more hydrophobic molecule, predominates in
Shewanella
and
Hyphomonas
species and in other marine bacteria.
Conjugal transfer of a shuttle vector from the human colonic anaerobe Bacteroides uniformis to the ruminal anaerobe Prevotella (Bacteroides) ruminicola B(1)4
Authors: Shoemaker N B , Anderson K L , Smithson S L , Wang G R , Salyers A A
Published: January 2020

Prevotella ruminicola (formerly Bacteroides ruminicola) is an anaerobic, gram-negative, polysaccharide-degrading bacterium which is found in the rumina of cattle. Since P. ruminicola is thought to make an important contribution to digestion of plant material in rumina, the ability to alter this strain genetically might help improve the efficiency of rumen fermentation. However, previously there has been no way to introduce foreign DNA into P. ruminicola strains. In this study we transferred a shuttle vector, pRDB5, from the colonic species Bacteroides uniformis to P. ruminicola B(1)4. The transfer frequency was 10(-6) to 10(-7) per recipient. pRDB5 contains sequences from pBR328, a cryptic colonic Bacteroides plasmid pB8-51, and a colonic Bacteroides tetracycline resistance (Tcr) gene. pRDB5 was mobilized out of B. uniformis by a self-transmissible Bacteroides chromosomal element designated Tcr Emr 12256. pRDB5 replicated in Escherichia coli as well as in Bacteroides spp. and was also mobilized from E. coli to B. uniformis by using IncP plasmid R751. However, direct transfer from E. coli to P. ruminicola B(1)4 was not detected. Thus, to introduce cloned DNA into P. ruminicola B(1)4, it was necessary first to mobilize the plasmid from E. coli to B. uniformis and then to mobilize the plasmid from B. uniformis to P. ruminicola B(1)4.
Carboxylation of o-cresol by an anaerobic consortium under methanogenic conditions
Authors: Bisaillon J G , Lépine F , Beaudet R , Sylvestre M
Published: January 2020

The metabolism of o-cresol under methanogenic conditions by an anaerobic consortium known to carboxylate phenol to benzoate was investigated. After incubation with the consortium at 29 degrees C for 59 days, o-cresol was transformed to 3-methylbenzoic acid, which was not further metabolized by the consortium. Proteose peptone in the culture medium was essential for the transformation of o-cresol. In addition, a transient compound detected in the culture was identified as 4-hydroxy-3-methylbenzoic acid. o-Cresol-6d was transformed by the consortium to deuterated hydroxy-methylbenzoic acid and deuterated methylbenzoic acid. These results demonstrate that o-cresol is carboxylated in the para position relative to the phenolic hydroxyl group and dehydroxylated by the anaerobic consortium.
Effects of Fungal Root Pathogens on the Population Dynamics of Biocontrol Strains of Fluorescent Pseudomonads in the Wheat Rhizosphere
Authors: Mazzola Mark , Cook R. James
Published: January 2020

The influences of
Gaeumannomyces graminis
var.
tritici
(which causes take-all of wheat),
Rhizoctonia solani
AG-8 (which causes rhizoctonia root rot of wheat),
Pythium irregulare, P. aristosporum
, and
P. ultimum
var.
sporangiiferum
(which cause pythium root rot of wheat) on the population dynamics of
Pseudomonas fluorescens
2-79 and Q72a-80 (bicontrol strains active against take-all and pythium root rot of wheat, respectively) in the wheat rhizosphere were examined. Root infection by either
G. graminis
var.
tritici
or
R. solani
resulted in populations of both bacterial strains that were equal to or significantly larger than their respective populations maintained on roots in the absence of these pathogens. In contrast, the population of strain 2-79 was significantly smaller on roots in the presence of any of the three
Pythium
species than on noninfected roots and was often below the limits of detection (50 CFU/cm of root) on
Pythium
-infected roots after 40 days of plant growth. In the presence of either
P. aristosporum
or
P. ultimum
var.
sporangiiferum
, the decline in the population of Q72a-80 was similar to that observed on noninfected roots; however, the population of this strain declined more rapidly on roots infected by
P. irregulare
than on noninfected roots. Application of metalaxyl (which is selectively inhibitory to
Pythium
spp.) to soil naturally infestated with
Pythium
spp. resulted in significantly larger rhizosphere populations of the introduced bacteria over time than on plants grown in the same soil without metalaxyl. It is apparent that root infections by fungal pathogens may either enhance or depress the population of fluorescent pseudomonads introduced for their control, with different strains of pseudomonads reacting differentially to different genera and species of the root pathogens.
Enhancement of the Potential To Utilize Octopine in the Nonfluorescent
Pseudomonas
sp. Strain 92
Authors: Gill Santokh Singh , Boivin Rodolphe , Dion Patrice
Published: January 2020

The nonfluorescent
Pseudomonas
sp. strain 92 requires the presence of a supplementary carbon source for growth on octopine, whereas the spontaneous mutant RB100 has acquired the capacity to utilize this opine as the sole carbon and nitrogen source. Insertional mutagenesis of RB100 with transposon Tn
5
generated mutants which were unable to grow on octopine and others which grew slowly on this substrate. Both types of mutants yielded revertants that had regained the ability to utilize octopine. Some of the revertants had lost the transposon, whereas in others the transposon was retained but with rearrangements of the insertion site. Genes of octopine catabolism from strain 92 were cloned on a cosmid vector to generate pK3. The clone pK3 conferred the ability to utilize octopine as the sole carbon and nitrogen source on the host
Pseudomonas putida
KT2440. Although they conferred an equivalent growth phenotype, the mutant genes carried by RB100 and the cloned genes on pK3 differed in their regulation. Utilization of [
14
C]octopine was inducible by octopine in RB100 and was constitutive in KT2440(pK3).
Toxicity of Bacillus thuringiensis to laboratory populations of the olive fruit fly (Dacus oleae)
Authors: Karamanlidou G , Lambropoulos A F , Koliais S I , Manousis T , Ellar D , Kastritsis C
Published: January 2020

A survey of Bacillus thuringiensis recovered from the environments of olive groves in Greece was carried out. Of 80 soil samples, 24 were found to contain B. thuringiensis with parasporal crystal inclusions; these were tested for toxicity against the olive fruit fly (Dacus oleae). Mortality levels of larvae caused by the different isolates varied from 7 to 87%. Higher levels of mortality were observed if a mixture of relatively pure crystals and spores was used compared with the mortality resulting from either fraction alone. We were able to show that the toxicity of the most active isolate is likely to be specific for D. oleae.
Photosynthetic Carbon Incorporation and Turnover in Antarctic Cryptoendolithic Microbial Communities: Are They the Slowest-Growing Communities on Earth?
Authors: Johnston Carl G. , Vestal J. Robie
Published: January 2020

The main forms of terrestrial life in the cold, desolate Ross Desert of Antarctica are lichen-dominated or cyanobacterium-dominated cryptoendolithic (hidden in rock) microbial communities. Though microbial community biomass (as measured by extractable lipid phosphate) was well within the range of values determined for other microbial communities, community lipid carbon turnover times (calculated from community lipid biomass, rates of community photosynthetic carbon incorporation into lipids versus temperature, and the in situ temperature record) were among the longest on Earth (ca. 20,000 years). When the temperature is above freezing and moisture is present, moderate rates of photosynthesis can be measured. Lichen communities had a psychrophilic temperature response (maximal rate of 4.5 ng of C h
-1
m
-2
at 10°C) while cyanobacteria communities had maximal rates at 20 to 30°C (3 ng of C h
-1
m
-2
). These extraordinarily slowly growing communities were not nutrient limited. No significant changes in photosynthetic metabolism were observed upon additions of 100 nM to 1 mM nitrate, ammonium, phosphate, and manganese. These simple, tenacious microbial communities demonstrate strategies of survival under conditions normally considered too extreme for life.
Purification and Properties of a Thermoactive Glucoamylase from
Clostridium thermosaccharolyticum
Authors: Specka Ulrike , Mayer Frank , Antranikian Garabed
Published: January 2020

A bacterial glucoamylase was purified from the anaerobic thermophilic bacterium
Clostridium thermosaccharolyticum
and characterized. The enzyme, which was purified 63-fold, with a yield of 36%, consisted of a single subunit with an apparent molecular mass of 75 kDa. The purified enzyme was able to attack α-1,4- and α-1,6-glycosidic linkages in various α-glucans, liberating glucose with a β-anomeric configuration. The purified glucoamylase, which was optimally active at 70°C and pH 5.0, attacked preferentially polysaccharides such as starch, glycogen, amylopectin, and maltodextrin. The velocity of oligosaccharide hydrolysis decreased with a decrease in the size of the substrate. The

K
m

values for starch and maltose were 18 mg/ml and 20 mM, respectively. Enzyme activity was not significantly influenced by Ca
2+
, EDTA, or α- or β-cyclodextrins.
Isolation and characterization of p-coumaroyl esterase from the anaerobic fungus Neocallimastix strain MC-2
Authors: Borneman W S , Ljungdahl L G , Hartley R D , Akin D E
Published: January 2020

An extracellular p-coumaroyl esterase produced by the anaerobic fungus Neocallimastix strain MC-2 released p-coumaroyl groups from 0-[5-0-((E)-p-coumaroyl)-alpha-L-arabinofuranosyl]-(1----3)-0-beta -D-xylopyranosyl-(1----4)-D-xylopyranose (PAXX). The esterase was purified 121-fold from culture medium in successive steps involving ultrafiltration column chromatography on S-sepharose and hydroxylapatite, isoelectric focusing, and gel filtration. The native enzyme had an apparent mass of 11 kDa under nondenaturing conditions and a mass of 5.8 kDa under denaturing conditions, suggesting that the enzyme may exist as a dimer. The isoelectric point was 4.7, and the pH optimum was 7.2. The purified esterase had 100 times more activity towards PAXX than towards the analogous feruloyl ester (FAXX). The apparent Km and Vmax of the purified p-coumaroyl esterase for PAXX at pH 7.2 and 40 degrees C were 19.4 microM and 5.1 microM min(-1), respectively. p-Coumaroyl tetrasaccharides isolated from plant cell walls were hydrolyzed at rates similar to that for PAXX, whereas a dimer of PAXX was hydrolyzed at a rate 20-fold lower, yielding 4,4'-dihydroxy-alpha-truxillic acid as an end product. Ethyl and methyl p-coumarates were hydrolyzed at very slow rates, if at all. The purified esterase released p-coumaroyl groups from finely, but not coarsely, ground plant cell walls, and this activity was enhanced by the addition of xylanase and other cell wall-degrading enzymes.
Evidence for Bacterial Chemotaxis to Cyanobacteria from a Radioassay Technique
Authors: Kangatharalingam N. , Wang Lizhu , Priscu John C.
Published: January 2020

Lyngbya birgei
and
Aphanizomenon flos-aquae
elicited a significant chemotactic attraction of
Aeromonas hydrophila
compared with controls lacking cyanobacteria. There was a positive exponential relationship between biomass (chlorophyll
a
) of
L. birgei
and
A. flos-aquae
and chemotactic attraction of
A. hydrophila.
The assay equipment was simple and reliable and could be used to study bacterial chemotaxis in other species in situ.