bluuebunny/crossref_metadata_2025 huggingface dataset:
Purification and properties of an acetoacetyl coenzyme A-reacting phosphotransbutyrylase from Clostridium beijerinckii ("Clostridium butylicum") NRRL B593
Authors: Thompson D K , Chen J S
Published: January 2020

During the study of acetoacetyl coenzyme A (CoA)-reacting enzymes of Clostridium beijerinckii NRRL B593, a phosphate-dependent acetoacetyl-CoA-utilizing activity was detected in protein fractions devoid of thiolase and phosphotransacetylase. Further purification of this acetoacetyl-CoA-utilizing activity yielded an enzyme which may be designated as phosphotransbutyrylase (PTB; phosphate butyryltransferase [EC 2.3.1.19]). PTB from C. beijerinckii NRRL B593 was purified 160-fold with a yield of 14% and, with the best fractions, purified 190-fold to near homogeneity. It showed a native Mr of 205,000 and a subunit Mr of 33,000. PTB activity was sensitive to pH changes within the physiological range of 6 to 8. PTB exhibited a broad substrate specificity. The Km values at pH 7.5 for butyryl-CoA, acetoacetyl-CoA, and acetyl-CoA were 0.04, 1.10, and 3.33 mM, respectively. The Vmax values with butyryl-CoA and acetoacetyl-CoA were comparable, but the Vmax/Km was higher for butyryl-CoA than for acetoacetyl-CoA. An apparent Km of 6.5 mM for phosphate was obtained with butyryl-CoA as the cosubstrate, whereas it was 12.9 mM with acetoacetyl-CoA as the cosubstrate. It remains to be established whether the putative compound acetoacetyl phosphate is produced in the PTB-catalyzed reaction with acetoacetyl-CoA.
Effects of Jet Fuel Spills on the Microbial Community of Soil
Authors: Song Hong-Gyu , Bartha Richard
Published: January 2020

Hydrocarbon residues, microbial numbers, and microbial activity were measured and correlated in loam soil contaminated by jet fuel spills resulting in 50 and 135 mg of hydrocarbon g of soil
−1
. Contaminated soil was incubated at 27°C either as well-aerated surface soil or as poorly aerated subsurface soil. In the former case, the effects of bioremediation treatment on residues, microbial numbers, and microbial activity were also assessed. Hydrocarbon residues were measured by quantitative gas chromatography. Enumerations included direct counts of metabolically active bacteria, measurement of mycelial length, plate counts of aerobic heterotrophs, and most probable numbers of hydrocarbon degraders. Activity was assessed by fluorescein diacetate (FDA) hydrolysis. Jet fuel disappeared much more rapidly from surface soil than it did from subsurface soil. In surface soil, microbial numbers and mycelial length were increased by 2 to 2.5 orders of magnitude as a result of jet fuel contamination alone and by 3 to 4 orders of magnitude as a result of the combination of jet fuel contamination and bioremediation. FDA hydrolysis was stimulated by jet fuel and bioremediation, but was inhibited by jet fuel alone. The latter was traced to an inhibition of the FDA assay by jet fuel biodegradation products. In subsurface soil, oxygen limitation strongly attenuated microbial responses to jet fuel. An increase in the most probable numbers of hydrocarbon degraders was accompanied by a decline in other aerobic heterotrophs, so that total plate counts changed little. The correlations between hydrocarbon residues, microbial numbers, and microbial activity help in elucidating microbial contributions to jet fuel elimination from soil.
Bioremediation Potential of Terrestrial Fuel Spills
Authors: Song Hong-Gyu , Wang Xiaoping , Bartha Richard
Published: January 2020

A bioremediation treatment that consisted of liming, fertilization, and tilling was evaluated on the laboratory scale for its effectiveness in cleaning up a sand, a loam, and a clay loam contaminated at 50 to 135 mg g of soil
−1
by gasoline, jet fuel, heating oil, diesel oil, or bunker C. Experimental variables included incubation temperatures of 17, 27, and 37°C; no treatment; bioremediation treatment; and poisoned evaporation controls. Hydrocarbon residues were determined by quantitative gas chromatography or, in the case of bunker C, by residual weight determination. Four-point depletion curves were obtained for the described experimental variables. In all cases, the disappearance of hydrocarbons was maximal at 27°C and in response to bioremediation treatment. Poisoned evaporation controls underestimated the true biodegradation contribution, but nevertheless, they showed that biodegradation makes only a modest contribution to gasoline disappearance from soil. Bunker C was found to be structurally recalcitrant, with close to 80% persisting after 1 year of incubation. The three medium distillates, jet fuel, heating oil, and diesel oil, increased in persistence in the listed order but responded well to bioremediation treatment under all test conditions. With bioremediation treatment, it should be possible to reduce hydrocarbons to insignificant levels in contaminated soils within one growing season.
Evaluation of the Mast ID and API 50CH systems for identification of Listeria spp
Authors: Kerr K G , Rotowa N A , Hawkey P M , Lacey R W
Published: January 2020

A multipoint inoculation technique (Mast ID) for the identification and species determination of Listeria monocytogenes (sensu strictu) and six other species of the genus Listeria was evaluated. This was compared with the commercially available API 50CH system. Both methods successfully identified all 123 strains tested. The Mast ID system is inexpensive and utilizing a multipoint inoculation technique permits the screening of up to 21 isolates per 9-cm petri dish. The API 50CH system was more expensive and time consuming and is therefore suitable only for the examination of smaller numbers of strains.
Stereoselective fungal metabolism of methylated anthracenes
Authors: Cerniglia C E , Campbell W L , Fu P P , Freeman J P , Evans F E
Published: January 2020

The metabolism of 9-methylanthracene (9-MA), 9-hydroxymethylanthracene (9-OHMA), and 9,10-dimethylanthracene (9,10-DMA) by the fungus Cunninghamella elegans ATCC 36112 is described. The metabolites were isolated by high-performance liquid chromatography and characterized by UV-visible, mass, and 1H nuclear magnetic resonance spectral techniques. The compounds 9-MA and 9,10-DMA were metabolized by two pathways, one involving initial hydroxylation of the methyl group(s) and the other involving epoxidation of the 1,2- and 3,4- aromatic double bond positions, followed by enzymatic hydration to form hydroxymethyl trans-dihydrodiols. For 9-MA metabolism, the major metabolites identified were trans-1,2-dihydro-1,2-dihydroxy and trans-3,4-dihydro-3,4-dihydroxy derivatives of 9-MA and 9-OHMA. 9-OHMA was also metabolized to trans-1,2- and 3,4-dihydrodiol derivatives. The absolute configuration and optical purity were determined for each of the trans-dihydrodiols formed by fungal metabolism and compared with previously published circular dichroism spectral data obtained from rat liver microsomal metabolism of 9-MA, 9-OHMA, and 9,10-DMA. Circular dichroism spectral analysis revealed that the major enantiomer for each dihydrodiol was predominantly in the S,S configuration, in contrast to the predominantly R,R configuration of the trans-dihydrodiol formed by mammalian enzyme systems. These results indicate that C. elegans metabolizes methylated anthracenes in a highly stereoselective manner that is different from that reported for rat liver microsomes.
Effects of Metals on
Streptomyces coelicolor
Growth and Actinorhodin Production
Authors: Abbas Ala S. , Edwards Clive
Published: January 2020

Actinorhodin production by
Streptomyces coelicolor
was used as a model system to study the effects of metals on growth and polyketide synthesis in a streptomycete. Numerous metals were tested in cultures grown in liquid media. Mercury and cadmium were highly toxic, and copper, nickel, and lead were less so, but all tended to inhibit both growth and antibiotic synthesis to a similar extent. Unexpectedly, manganese, cobalt, zinc, and, to a lesser extent, chromium caused complex effects that in general resulted in some enhancement of growth yield but a reduction in antibiotic titers. These complex effects meant that cobalt, manganese, and zinc had lower 50% inhibitory concentrations for antibiotic yields compared with those for biomass. The physiologically active divalent cations calcium and magnesium were also tested. Calcium at high concentrations was particularly effective in reducing antibiotic titers and enhancing growth yields. By adding calcium at different phases of growth, it could be demonstrated that it was most effective in reducing the antibiotic yield when added during the early growth phase. Addition during the antibiotic-producing phase resulted in little reduction of final actinorhodin titers.
Pyrite oxidation by thermophilic archaebacteria
Authors: Larsson L , Olsson G , Holst O , Karlsson H T
Published: January 2020

Three species of thermophilic archaebacteria of the genera Sulfolobus (Sulfolobus acidocaldarius and S. solfataricus) and Acidianus (Acidianus brierleyi) were tested for their ability to oxidize pyrite and to grow autotrophically on pyrite, to explore their potential for use in coal desulfurization. Only A. brierleyi was able to oxidize and grow autotrophically on pyrite. Jarosite was formed during the pyrite oxidation, resulting in the precipitation of sulfate and iron. The medium composition affected the extent of jarosite formation.
Enhanced Growth of Wheat and Soybean Plants Inoculated with
Azospirillum brasilense
Is Not Necessarily Due to General Enhancement of Mineral Uptake
Authors: Bashan Yoav , Harrison S. Kent , Whitmoyer Robert E.
Published: January 2020

The capacity of
Azospirillum brasilense
to enhance the accumulation of K
+
, P, Ca
2+
, Mg
2+
, S, Na
+
, Mn
2+
, Fe
2+
, B, Cu
2+
, and Zn
2+
in inoculated wheat and soybean plants was evaluated by using two different analytical methods with five
A. brasilense
strains originating from four distinct geographical regions. A
Pseudomonas
isolate from the rhizosphere of
Zea mays
seedlings was included as a control. All
A. brasilense
strains significantly improved wheat and soybean growth by increasing root and shoot dry weight and root surface area. The degree of plant response to inoculation varied among the different strains of
A. brasilense.
All strains were capable of colonizing roots, but the best root colonizer,
Pseudomonas
sp., had no effect on plant growth. The numbers of organisms of Brazilian strains Sp-245 and Sp-246 colonizing roots were similar regardless of the host plant. Numbers of organisms for the other strains were directly dependent on the host plant. The main feature characterizing mineral accumulation in inoculated plants was that all inoculation treatments changed the mineral balance of the plants, but in an inconsistent manner. Enhancement of mineral uptake by plants also varied among strains to a great extent and was directly dependent on the strain-plant combination; i.e., a strain capable of increasing accumulation of a particular ion in one plant species or cultivar often lacked the ability to do so in another. Minerals in inoculated plants were not evenly distributed in different plant tissues, and the changes varied among groups of plants within each bacterial strain inoculation treatment. We suggest that, although
A. brasilense
strains are capable of changing the mineral balance and content of plants, it is unlikely that this ability is a general mechanism responsible for plant improvement by
A. brasilense.
Comparison of phenotypic diversity and DNA heterogeneity in a population of soil bacteria
Authors: Torsvik V , Salte K , Sørheim R , Goksøyr J
Published: January 2020

The phenotypic diversity of about 200 bacterial strains isolated from soil was compared with the genotypic diversity of the same population. The strains were phenotypically characterized by the API 20B test system. The results of these tests were subjected to cluster analysis, which revealed 41 biotypes at 80% similarity. The five dominating biotypes contained 43% of the strains. The phenotypic diversity as determined by the Shannon index, equitability, rarefaction, and cumulative differences was high, but indicated some dominant biotypes. The genetic diversity was measured by reassociation of mixtures of denatured DNA isolated from the bacterial strains (C0t plots). The observed genetic diversity was high. Reassociation of DNA from all bacterial strains together revealed that the population contained heterologous DNA equivalent to 20 totally different bacterial genomes (i.e., genomes that have no homology). This study showed that reassociation of DNA isolated from a collection of bacteria gave a good estimate of the diversity of the collection and that there was good agreement with different phenotypic diversity measures. The Shannon index in particular has features in common with the genetic diversity measure presented here.
Kinetics of Pseudomonas aeruginosa adhesion to 304 and 316-L stainless steel: role of cell surface hydrophobicity
Authors: Vanhaecke E , Remon J P , Moors M , Raes F , De Rudder D , Van Peteghem A
Published: January 2020

Fifteen different isolates of Pseudomonas aeruginosa were used to study the kinetics of adhesion to 304 and 316-L stainless steel. Stainless steel plates were incubated with approximately 1.5 X 10(7) CFU/ml in 0.01 M phosphate-buffered saline (pH 7.4). After the plates were rinsed with the buffer, the number of adhering bacteria was determined by a bioluminescence assay. Measurable adhesion, even to the electropolished surfaces, occurred within 30 s. Bacterial cell surface hydrophobicity, as determined by the bacterial adherence to hydrocarbons test and the contact angle measurement test, was the major parameter influencing the adhesion rate constant for the first 30 min of adhesion. A parabolic relationship between the CAM values and the logarithm of the adhesion rate constants (In k) was established. No correlation between either the salt aggregation or the improved salt aggregation values and the bacterial adhesion rate constants could be found. Since there was no significant correlation between the bacterial electrophoretic mobilities and the In k values, the bacterial cell surface charge seemed of minor importance in the process of adhesion of P. aeruginosa to 304 and 316-L stainless steel.
Enzymatic profiles of 11 barophilic bacteria under in situ conditions: evidence for pressure modulation of phenotype
Authors: Straube W L , O'Brien M , Davis K , Colwell R R
Published: January 2020

Barophilic bacteria are microorganisms that grow preferentially (facultative barophiles) or exclusively (obligate barophiles) under elevated hydrostatic pressure. Barophilic bacteria have been isolated from a variety of deep-sea environments. Attempts to characterize these organisms have been hampered by a lack of appropriate methodologies. A colorimetric method for the detection of 19 constitutively expressed enzymes under in situ conditions of pressure and temperature has been devised, using a simple modification of the commercially available API ZYME enzyme assay kit. By using this method, enzyme profiles of 11 barophilic isolates, including an obligate barophile, were determined. Nine of the 10 facultatively barophilic isolates examined exhibited a change of phenotype in at least one enzyme reaction when tested at 1 atm (1 atm = 101.29 kPa), compared with results obtained under in situ pressure. The assay is simple and rapid and allows for direct determination of enzyme activity under conditions of high pressure and low temperature.
Formation of a clear zone on tannin-treated brain heart infusion agar by a Streptococcus sp. isolated from feces of koalas
Authors: Osawa R
Published: January 2020

Gram-positive cocci, isolated from the feces of koalas and identified as Streptococcus bovis biotype I, formed a distinct clear zone on tannin-treated brain heart infusion agar, suggesting that this isolate has the unique characteristic of degrading the tannin-protein complex.
Growth, Respiration, and Polypeptide Patterns of
Bradyrhizobium
sp. (
Arachis
) Strain 3G4b20 from Succinate- or Oxygen-Limited Continuous Cultures
Authors: Allen George C. , Elkan Gerald H.
Published: January 2020

Succinate- or oxygen-limited continuous cultures were used to study the influences of different concentrations of dissolved oxygen and ammonia on the growth, respiration, and polypeptide patterns of
Bradyrhizobium
sp. (
Arachis
) strain 3G4b20. During succinate-limited growth, molar growth yields on succinate (
Y
succ
) ranged from 38.9 to 44.4 g (dry weight) of cells mol of succinate
−1
and were not greatly influenced by changes in dilution rates or changes in the oxygen concentrations that we tested. Succinate, malate, and fumarate induced the highest rates of oxygen uptake in all of the steady states in which the supply rates of (NH
4
)
2
SO
4
ranged between 322 and 976 μmol h
−1
. However, the amino acids aspartate, asparagine, and glutamate could also be used as respiratory substrates, especially when the (NH
4
)
2
SO
4
supply rate was decreased to 29 μmol h
−1
. Glutamine-dependent respiration was seen only when the (NH
4
)
2
SO
4
supply rate was 29 μmol h
−1
and thus appears to be under tight ammonia control. Nitrogenase activity was detected only when the culture was switched from a succinate-limited steady state to an oxygen-limited steady state. Comparison of major silver-stained proteins from three steady states by two-dimensional gel electrophoresis revealed that nearly 60% were affected by oxygen and 24% were affected by ammonia. These data are consistent with reports that oxygen has a major regulatory role over developmental processes in
Rhizobium
sp. and
Bradyrhizobium
sp.
Isolation and Characterization of a
Pseudomonas
Strain That Restricts Growth of Various Phytopathogenic Fungi
Authors: Jayaswal Radheshyam K. , Fernandez Marcel A. , Schroeder Ralph G.
Published: January 2020

The characterization of a novel
Pseudomonas
strain exhibiting antagonism towards many important corn fungal pathogens is presented. This strain was isolated from the caryopses of the grass
Tripsacum dactyloides
and was identified as
Pseudomonas cepacia.
The antagonistic activity is due to the production of an antifungal compound. The chromatographic properties of this partially purified compound isolated from growth medium differ from those reported previously for other pseudomonads. The suppression of the growth of economically important phytopathogens by this strain and by the partially purified compound indicates a potential biocontrol agent.
Identification and Plant Interaction of a
Phyllobacterium
sp., a Predominant Rhizobacterium of Young Sugar Beet Plants
Authors: Lambert Bart , Joos Henk , Dierickx Sabine , Vantomme Robert , Swings Jean , Kersters Karel , Van Montagu Marc
Published: January 2020

The second most abundant bacterium on the root surface of young sugar beet plants was identified as a
Phyllobacterium
sp. (
Rhizobiaceae
) based on a comparison of the results of 39 conventional identification tests, 167 API tests, 30 antibiotic susceptibility tests, and sodium dodecyl sulfate-polyacrylamide gel electrophoretic fingerprints of total cellular proteins with type strains of
Phyllobacterium myrsinacearum
and
Phyllobacterium rubiacearum.
It was found on 198 of 1,100 investigated plants between the 2nd and 10th leaf stage on three different fields in Belgium and one field in Spain. Densities ranged from 2 × 10
4
to 2 × 10
8
CFU/g of root. Five isolates exerted a broad-spectrum in vitro antifungal activity. DNA-DNA hybridizations showed that
Phyllobacterium
sp. does not contain DNA sequences that are homologous with the attachment genes
chvA, chvB
, the transferred-DNA (T-DNA) hormone genes
iaaH
and
ipt
from
Agrobacterium tumefaciens, iaaM
from
A. tumefaciens
and
Pseudomonas savastanoi
, or the nitrogenase genes
nifHDK
from
Klebsiella pneumoniae. Phyllobacterium
sp. produces indolylacetic acid in in vitro cultures and induces auxinlike effects when cocultivated with callus tissue of tobacco. When
Phyllobacterium
sp. was transformed with a Ti plasmid derivative, it gained the capacity to induce tumors on
Kalanchoe daigremontiana.
The potential role of
Phyllobacterium
sp. in this newly recognized niche is discussed.
Transport of malic acid and other dicarboxylic acids in the yeast Hansenula anomala
Authors: Côrte-Real M , Leão C
Published: January 2020

DL-Malic acid-grown cells of the yeast Hansenula anomala formed a saturable transport system that mediated accumulative transport of L-malic acid with the following kinetic parameters at pH 5.0: Vmax, 0.20 nmol.s-1.mg (dry weight)-1; Km, 0.076 mM L-malate. Uptake of malic acid was accompanied by proton disappearance from the external medium with rates that followed Michaelis-Menten kinetics as a function of malic acid concentration. Fumaric acid, alpha-ketoglutaric acid, oxaloacetic acid, D-malic acid, and L-malic acid were competitive inhibitors of succinic acid transport, and all induced proton movements that followed Michaelis-Menten kinetics, suggesting that all of these dicarboxylates used the same transport system. Maleic acid, malonic acid, oxalic acid, and L-(+)-tartaric acid, as well as other Krebs cycle acids such as citric and isocitric acids, were not accepted by the malate transport system. Km measurements as a function of pH suggested that the anionic forms of the acids were transported by an accumulative dicarboxylate proton symporter. The accumulation ratio at pH 5.0 was about 40. The malate system was inducible and was subject to glucose repression. Undissociated succinic acid entered the cells slowly by simple diffusion. The permeability of the cells by undissociated acid increased with pH, with the diffusion constant increasing 100-fold between pH 3.0 and 6.0.
Expression of Bacillus thuringiensis delta-endotoxin genes during vegetative growth
Authors: Mettus A M , Macaluso A
Published: January 2020

Bacillus thuringiensis delta-endotoxin (crystal protein) genes are normally expressed only during sporulation. It is possible to produce crystal protein during vegetative growth by placing B. thuringiensis crystal protein genes downstream of a strong vegetative promoter. By removing a possible transcriptional terminator of the tetracycline resistance gene of pBC16 and inserting a multiple cloning site, delta-endotoxin genes can be cloned downstream from the tetracycline resistance gene promoter. This construct allows for readthrough transcription from the strong vegetative promoter. Crystal protein is then produced during vegetative growth as well as during sporulation in both B. thuringiensis and Bacillus megaterium. This construct also allows for production of delta-endotoxin in B. thuringiensis strains that do not normally produce delta-endotoxin because of a defect in sporulation.
Surface and virulence properties of environmental Vibrio cholerae non-O1 from Albufera Lake (Valencia, Spain)
Authors: Amaro C , Toranzo A E , González E A , Blanco J , Pujalte M J , Aznar R , Garay E
Published: January 2020

A total of 140 environmental Vibrio cholerae non-O1 isolates, together with several culture collection strains from both environmental and clinical sources, were studied in relation to hemagglutination, surface hydrophobicity, and the enzymatic, hemolytic, cytotoxic, and enterotoxic activities of their extracellular products. A total of 78 and 62% of the strains produced hemagglutinins and exohemagglutinins, respectively. Four different hemagglutinating and two exohemagglutinating activities were found by using eight sugars in the inhibition assays. Cell-bound mannose-sensitive hemagglutination was detected mainly in chicken blood, whereas fucose-sensitive hemagglutination was recorded only in human blood. Cell-bound hemagglutinin resistant to all sugars tested was the only one related to surface hydrophobicity. The surface properties varied along the growth curves. The non-O1 strains displayed strong enzymatic and hemolytic activities, except for esculin hydrolysis. Of 26 non-O1 isolates selected for cytotoxin and enterotoxin production, 23 showed a wide spectrum of cytotoxic effects on cell lines of poikilothermic and homoiothermic species, but they were weakly enterotoxigenic in the infant mouse test. All extracellular products of cytotoxic strains were proteolytic, lipolytic, and hemolytic, and a high percentage produced hemagglutination of chicken blood. The cytotoxic factors in the non-O1 strains analyzed were not R plasmid mediated.
Nitrogen Fixation in Continuous Culture with NH
4
Cl-Containing Media
Authors: Fritzsche C. , Niemann E.-G.
Published: January 2020

N balance and
15
N dilution were determined from growth of
Azospirillum brasilense
Sp7 and two unidentified gram-negative nitrogen-fixing microorganisms in continuous culture supplied with NH
4
Cl. At the 1.1 and 2.2 mM NH
4
Cl steady states (N-to-C ratios of 1:68 and 1:34, respectively), the organisms grew with NH
4
Cl and N
2
as N sources simultaneously under carbon limitation. No ammonium could be detected in the supernatant of these cultures.
Simplified Henry technique for initial recognition of Listeria colonies
Authors: Lachica R V
Published: January 2020

The Henry oblique transmitted-light viewing technique was modified to provide a more precise, convenient, and familiar manner with which to read (score) and recognize colonies of listeriae by their distinct bluish cast. The simplified technique involved illuminating each colony directly with a high-intensity lamp while viewing it with a hand lens at a precise angle in place of a scanning light microscope.
Composition and Thermal Properties of Membrane Lipids in Cryptoendolithic Lichen Microbiota from Antarctica
Authors: Finegold Leonard , Singer Michael A. , Federle Thomas W. , Vestal J. Robie
Published: January 2020

In Antarctica there exists a cryptoendolithic microbiota which survives extremely low temperatures. Fatty acid analysis of the membrane phospholipids of this microbial community showed a predominance of polyunsaturated fatty acids. Artificial membranes made from the purified community phospholipids remained fluid to below −20°C and had unusual hydration properties.
Microbiological quality and safety of zoo food
Authors: Richter E R , al-Sheddy I
Published: January 2020

Two types of commercial products for feeding zoo animals (a frozen meat product, referred to as zoo food, and a dry product, referred to as dry food) were microbiologically examined for spoilage organisms (aerobic, psychrotrophic, coliform, Escherichia coli, mold, and yeasts) and pathogens (Salmonella spp., Listeria monocytogenes, and Campylobacter jejuni). Levels of microorganisms in frozen ground zoo food were compared with those in frozen ground beef and frozen ground turkey meat. The level of microbial contaminants in frozen ground zoo meat was found to be similar to that in frozen ground beef and higher than that in frozen ground turkey meat. Sixty percent of the frozen zoo meat samples were Salmonella positive, and all of the samples were L. monocytogenes positive. Dry zoo food was documented to have microbial levels lower than those in frozen zoo meat; the pathogen levels were less than 1/25 g of food. Defrosting zoo meat at 10, 25, and 37 degrees C for 24 h showed that 10 degrees C is the best temperature for defrosting frozen ground zoo meat loaves (length, 9 in. [22.8 cm]; radius, 2 in. [5.1 cm]) without affecting the microbiological quality or safety of the product.
Utilization of Glycerol as a Hydrogen Acceptor by
Lactobacillus reuteri
: Purification of 1,3-Propanediol:NAD
+
Oxidoreductase
Authors: Talarico Todd L. , Axelsson Lars T. , Novotny James , Fiuzat Mitra , Dobrogosz Walter J.
Published: January 2020

Lactobacillus reuteri
utilizes exogenously added glycerol as a hydrogen acceptor during carbohydrate fermentations, resulting in higher growth rates and cell yields than those obtained during growth on carbohydrates alone. Glycerol is first converted to 3-hydroxypropionaldehyde by a coenzyme B
12
-dependent glycerol dehydratase and then reduced to 1,3-propanediol by an NAD
+
-dependent oxidoreductase. The latter enzyme was purified and determined to have a molecular weight of 180,000; it is predicted to exist as a tetramer of identical 42,000-molecular-weight subunits.
Metabolism of One-Carbon Compounds by the Ruminal Acetogen
Syntrophococcus sucromutans
Authors: Doré J. , Bryant M. P.
Published: January 2020

Syntrophococcus sucromutans
is the predominant species capable of O demethylation of methoxylated lignin monoaromatic derivatives in the rumen. The enzymatic characterization of this acetogen indicated that it uses the acetyl coenzyme A (Wood) pathway. Cell extracts possess all the enzymes of the tetrahydrofolate pathway, as well as carbon monoxide dehydrogenase, at levels similar to those of other acetogens using this pathway. However, formate dehydrogenase could not be detected in cell extracts, whether formate or a methoxyaromatic was used as electron acceptor for growth of the cells on cellobiose. Labeled bicarbonate, formate, [1-
14
C] pyruvate, and chemically synthesized
O
-[
methyl
-
14
C]vanillate were used to further investigate the catabolism of one-carbon (C
1
) compounds by using washed-cell preparations. The results were consistent with little or no contribution of formate dehydrogenase and pointed out some unique features. Conversion of formate to CO
2
was detected, but labeled formate predominantly labeled the methyl group of acetate. Labeled CO
2
readily exchanged with the carboxyl group of pyruvate but not with formate, and both labeled CO
2
and pyruvate predominantly labeled the carboxyl group of acetate. No CO
2
was formed from O demethylation of vanillate, and the acetate produced was position labeled in the methyl group. The fermentation pattern and specific activities of products indicated a complete synthesis of acetate from pyruvate and the methoxyl group of vanillate.
Efficacy of beta-glucuronidase assay for identification of Escherichia coli by the defined-substrate technology
Authors: Rice E W , Allen M J , Edberg S C
Published: January 2020

In 1976, Kilian and Bulow described the association of beta-glucuronidase with the genus Escherichia (97% positive) and suggested that a beta-glucuronidase assay would be a useful identification test. Since that report, papers about the sensitivity and specificity of this enzyme for the identification of Escherichia coli from clinical sources, food, seawater, potable-water supplies, and various environmental sources have appeared. A study was undertaken to determine the efficacy and specificity of the defined-substrate technology beta-glucuronidase (Colilert) assay for the identification of this species from fecal samples. A total of 460 human, 105 cow, and 55 horse E. coli isolates were tested. Results showed 95.5% beta-glucuronidase-positive isolates in 24 h and 99.5% positive after 28 h of incubation. Only one E. coli isolate was negative. There were no significant differences in the percentage of beta-glucuronidase-positive isolates among the human or animal isolates. There were no non-E. coli isolates that were positive. All subjects carried beta-glucuronidase-positive E. coli.
Pathogenicity of Listeria monocytogenes grown on crabmeat
Authors: Brackett R E , Beuchat L R
Published: January 2020

The pathogenicity of Listeria monocytogenes as influenced by growth on crabmeat at 5 and 10 degrees C was studied. Crabmeat was inoculated with L. monocytogenes V7 (ca. 10(4) CFU/g) and incubated for up to 14 days at 5 and 10 degrees C. At selected incubation times, L. monocytogenes was removed from crabmeat by washing with 0.1 M potassium phosphate buffer (pH 7.0), and populations were determined by surface plating on LiCl-phenylethanol-moxalactam agar. Buffered suspensions were then centrifuged, and the resulting pellets were suspended in phosphate buffer containing 10% glycerol and stored at -18 degrees C. Thawed, diluted suspensions of cells were tested for pathogenicity by intraperitoneal injection into immunocompromised and nonimmunocompromised mice. L. monocytogenes cells recovered from crabmeat and then recultured in tryptose phosphate broth (TPB), as well as cells which had not been passed through crabmeat but had been cultured in TPB, were likewise harvested, suspended in buffered 10% glycerol, frozen, thawed, diluted, and tested for pathogenicity by intraperitoneal injection. Growth on crabmeat at 5 and 10 degrees C did not have a significant effect on pathogenicity. The population of L. monocytogenes necessary to kill about 50% of the immunocompromised mice in each test set within 7 days was about 10(4) CFU, and this result was not significantly affected by storage temperature of the crabmeat or type of substrate, i.e., crabmeat or TPB, on which it had grown.
Regulation and distribution of Fibrobacter succinogenes subsp. succinogenes S85 endoglucanases
Authors: McGavin M , Lam J , Forsberg C W
Published: January 2020

The distribution of endoglucanase activities in cultures of Fibrobacter succinogenes subsp. succinogenes S85 grown on different carbon sources was examined by a variety of biochemical and immunological techniques. Total culture endoglucanase activity was primarily cell associated and was expressed constitutively, although synthesis of endoglucanase 1 (EG1) was repressed by cellobiose. Western immunoblotting showed that EG1 and EG3 were released into the culture fluid during growth, while EG2 remained largely associated with the cell. Subcellular localization showed low endoglucanase activity in the periplasmic fraction and similar, high levels in the cytoplasmic and membrane fractions. Western immunoblotting showed that EG2 was absent from the periplasmic fraction. Data from immunoelectron microscopy with either polyclonal or monoclonal antibody to EG2 revealed a high density of gold labeling at sites where there was a disruption in the regular features of the cell surface, such as in blebbing or physical tearing of the membrane. When cells were grown on cellulose, there was a high density of labeling on the cellulose but not on the cells, indicating that EG2 has limited exposure at the cell surface. On the basis of these data, export of enzymes from their intracellular locations appears to occur via three different mechanisms: a specific secretory pathway independent of cellulose, a secretory mechanism which is mediated by contact with cellulose, and a generalized blebbing process that occurs irrespective of the carbon source.
Adhesive Properties of a Symbiotic Bacterium from a Wood-Boring Marine Shipworm
Authors: Imam Syed H. , Greene Richard V. , Griffin Harold L.
Published: January 2020

Adhesive properties of a cellulolytic, nitrogen-fixing bacterium isolated from a marine shipworm by Waterbury et al. (J. B. Waterbury, C. B. Calloway, and R. D. Turner, Science 221:1401-1403, 1983) are described.
35
S-labeled cells of the shipworm bacterium bound preferentially to Whatman no. 1 cellulose filter paper, compared with its binding to other cellulose substrata or substrata lacking cellulose. The ability of the bacteria to bind to Whatman no. 1 filter paper was significantly reduced by glutaraldehyde or heat treatment of cells. Pretreatment of cells with azide, valinomycin, gramicidin-D, bis-hexafluoroacetylacetone (1799), or carbonyl cyanide-
p
-trifluoromethoxyphenylhydrazone inhibited adhesion activity. Cells pretreated with pronase or trypsin also exhibited reduced binding activity, but chymotrypsin and peptidase had no effect on adhesion activity. Cellodextrins and methyl cellulose 15 inhibited the adhesion of shipworm bacteria to filter paper, whereas glucose, cellobiose, and soluble carboxymethyl cellulose had no significant effect. The divalent cation chelators EDTA and EGTA [ethylene glycol-bis(β-aminoethyl ether)-
N,N,N′N′
-tetraacetic acid] had little or no effect on adhesive properties of shipworm bacteria. Also, preabsorbing the substratum with extracellular endoglucanase isolated from the shipworm bacterium or 1% bovine serum albumin had no apparent effect on bacterial binding. Low concentrations (0.01%) of sodium dodecyl sulfate solubilized a fraction from whole cells, which appeared to be involved in cellular binding activity. After removal of sodium dodecyl sulfate, several proteins in this fraction associated with intact cells. These cells exhibited up to 50% enhanced binding to filter paper in comparison to cells which had not been exposed to the sodium dodecyl sulfate-solubilized fraction.
Copper-induced production of copper-binding supernatant proteins by the marine bacterium Vibrio alginolyticus
Authors: Harwood-Sears V , Gordon A S
Published: January 2020

Growth of the marine bacterium Vibrio alginolyticus is temporarily inhibited by micromolar levels of copper. During the copper-induced lag phase, supernatant compounds which complex and detoxify copper are produced. In this study two copper-inducible supernatant proteins having molecular masses of ca. 21 and 19 kilodaltons (CuBP1 and CuBP2) were identified; these proteins were, respectively, 25 and 46 times amplified in supernatants of copper-challenged cultures compared with controls. Experiments in which chloramphenicol was added to cultures indicated that there was de novo synthesis of these proteins in response to copper. When supernatants were separated by gel permeation chromatography, CuBP1 and CuBP2 coeluted with a copper-induced peak in copper-binding activity. CuBP1 and CuBP2 from whole supernatants were concentrated and partially purified by using a copper-charged immobilized metal ion affinity chromatography column, confirming the affinity of these proteins for copper. A comparison of cell pellets and supernatants demonstrated that CuBP1 was more concentrated in supernatants than in cells. Our data are consistent with a model for a novel mechanism of copper detoxification in which excretion of copper-binding protein is induced by copper.
Oligonucleotide Probes That Hybridize with rRNA as a Tool To Study
Frankia
Strains in Root Nodules
Authors: Hahn Dittmar , Starrenburg Marjo J. C. , Akkermans Antoon D. L.
Published: January 2020

Oligonucleotide probes that hybridize with specific sequences in variable regions of the 16S rRNA of the nitrogen-fixing actinomycete
Frankia
were used for the identification of
Frankia
strains in nodules.
Frankia
cells were released from plant tissue by grinding glutaraldehyde-fixed root nodules in guanidine hydrochloride solution. rRNA was obtained after sonication, precipitation with ethanol, and purification by phenolchloroform extraction. Degradation of rRNA, evident in Northern blots, did not affect hybridization with the oligonucleotides. Nodules of about 1 mg (fresh weight) provided sufficient rRNA for reliable detection of the
Frankia
strain. The utility of this rRNA extraction method was tested in a competition experiment between two effective
Frankia
strains on cloned
Alnus glutinosa
plants.
Coupled Metabolic and Photolytic Pathway for Degradation of Pyridinedicarboxylic Acids, Especially Dipicolinic Acid
Authors: Amador José A. , Taylor Barrie F.
Published: January 2020

Three isomers of pyridinedicarboxylic acid (PDCA) (2,3-, 2,5-, and 2,6-PDCA) were partially oxidized by marine bacteria when grown aerobically on the corresponding phthalate analogs. The metabolites, unlike the parent PDCAs, absorbed light in the solar actinic range (wavelengths greater than 300 nm) and were readily degraded in sunlight. The principal product from 2,6-PDCA (dipicolinic acid) metabolism was extracted from a culture fluid, purified by column chromatography, and analyzed by UV-visible, infrared, and
13
C nuclear magnetic resonance spectroscopy. The compound was identified as 2,3-dihydroxypicolinic acid (2,3-DHPA). 2,3-DHPA was photolyzed in aqueous solution (pH 8.0) with a half-life of 100 min. Eight photoproducts, three of which were photolabile, were detected by high-performance liquid chromatography. Ammonia was also photoproduced from 2,3-DHPA. Analysis of the photoproducts by UV-visible spectroscopy and by high-performance liquid chromatography of 2,4-dinitrophenylhydrazones indicated that the products were conjugated carbonyls and carboxylic acids. Six of the photoproducts were readily consumed by bacterial strain CC9M. In illuminated aquatic environments, coupled bio- and photodegradative mechanisms probably contribute to the degradation of PDCAs.
Effect of Indoleacetic Acid and Related Indoles on
Lactobacillus
sp. Strain 11201 Growth, Indoleacetic Acid Catabolism, and 3-Methylindole Formation
Authors: Honeyfield D. C. , Carlson J. R.
Published: January 2020

A study was conducted to determine the activity of the 3-methylindole (3MI)-forming enzyme in
Lactobacillus
sp. strain 11201. Cells were incubated anaerobically with 17 different indolic and aromatic compounds. Indoleacetic acid (IAA), 5-hydroxyindoleacetic acid, 5-methoxy-3-indoleacetic acid, indole-3-pyruvate, or indole-3-propionic acid induced 3MI-forming activity. The highest total enzyme activity induced by IAA was observed in cells incubated with an initial concentration of 1.14 mM IAA. Peak activity of the 3MI-forming enzyme occurred 4 h after bacteria were incubated with either 0.114 or 1.14 mM IAA. Enzyme activity peaked earlier (2 h) and disappeared more rapidly at 5.7 mM IAA than at other concentrations of IAA. The effects of IAA and 3MI on the growth of
Lactobacillus
sp. strain 11201 and formation of 3MI from IAA also were determined. Bacterial growth and 3MI formation from IAA were reduced in medium containing exogenous 3MI. The growth depression observed in medium containing 5.7 mM IAA appears to be due to the toxicity of 3MI rather than IAA. The formation of 3MI in this ruminal
Lactobacillus
sp. is mediated by an inducible enzyme, and as 3MI accumulates, bacterial growth and rates of 3MI formation from IAA are reduced.
New Method for Extraction of Streptomycete Spores from Soil and Application to the Study of Lysogeny in Sterile Amended and Nonsterile Soil
Authors: Herron Paul R. , Wellington Elizabeth M. H.
Published: January 2020

A new method for the isolation and enumeration of streptomycete spores from soil was developed. This method makes use of a cation-exchange resin to disperse soil particles. It allowed the detection of 10 spores in 100 g of sterile soil, while ca. 10
3
could be accurately enumerated in 100 g. This method was applied to studying the fate of a marked actinophage in soil. In sterile amended and nonsterile soil, relatively high numbers of actinophages were only found during the first few days of the experiment when the host streptomycete was in the mycelial form. Later, after sporulation, lysogens could be detected in sterile amended soil and could still be found 60 days after inoculation. Although no lysogens were found in nonsterile soil, the introduced phage could still be detected in the free state after 60 days, albeit at a low titer.
Interconversion of aflatoxin B1 and aflatoxicol by several fungi
Authors: Nakazato M , Morozumi S , Saito K , Fujinuma K , Nishima T , Kasai N
Published: January 2020

Four fungal strains, namely, Aspergillus niger, Eurotium herbariorum, a Rhizopus sp., and non-aflatoxin (AF)-producing Aspergillus flavus, which could convert AF-B1 to aflatoxicol (AFL), could also reconvert AFL to AF-B1. The interconversion of AF-B1 to AFL and of AFL to AF-B1 was ascertained to occur during proliferation of the fungi. These reactions were distinctly observed in cell-free systems obtained from disrupted mycelia of A. flavus and the Rhizopus sp., but they were not observed in culture filtrates from intact (nondisrupted) mycelia of the same strains. The interconversion activities of AF-B1 and AFL were not observed when the cell-free systems were preheated at 100 degrees C. These findings strongly suggest that the interconversion of AF-B1 and AFL is mediated by intracellular enzymes of A. flavus and the Rhizopus sp. In addition, the isomerization of AFL-A to AFL-B observed in culture medium was also found to occur by the lowering of the culture pH.
Improved fluorogenic assay for rapid detection of Vibrio parahaemolyticus in foods
Authors: Miyamoto T , Miwa H , Hatano S
Published: January 2020

An improved fluorogenic assay for the rapid detection of Vibrio parahaemolyticus was developed. In the improved assay, the enrichment of V. parahaemolyticus was carried out in arabinose-glucuronate medium (0.5% arabinose, 0.25% glucuronate, 0.1% polypeptone, 0.1% yeast extract, 0.1% ammonium sulfate, 2% NaCl, 2 micrograms of polymyxin B sulfate per ml, pH 8.5) at 37 degrees C. After the cultivation, the trypsinlike activity of the bacteria was measured by fluorescence with the fluorogenic substrate benzoyl-L-arginine-7-aminomethylcoumarin. Even in the presence of 3 x 10(5) cells of Vibrio alginolyticus, 20 cells of V. parahaemolyticus were clearly detected after a 6-h enrichment cultivation by the assay. Fifty contaminated samples of 14 seafoods were examined for V. parahaemolyticus by the fluorogenic assay after enrichment cultivation for 6 or 8 h. The results were then compared with those obtained by the conventional bromothymol blue Teepol agar assay and the most-probable-number method. There was a linear relationship between trypsinlike activity measured by the assay and the number of V. parahaemolyticus cells in seafood as determined by the bromothymol blue Teepol agar and most-probable-number methods. Correlation coefficients were 0.95 and 0.93 after a 6-h cultivation and an 8-h cultivation, respectively. The presence of 10 cells of V. parahaemolyticus per gram of seafood sample was detected after a 10-h total detection time by the fluorogenic assay.
Chemotaxis of Pseudomonas putida toward chlorinated benzoates
Authors: Harwood C S , Parales R E , Dispensa M
Published: January 2020

The chlorinated aromatic acids 3-chlorobenzoate and 4-chlorobenzoate are chemoattractants for Pseudomonas putida PRS2000. These compounds are detected by a chromosomally encoded chemotactic response to benzoate which is inducible by beta-ketoadipate, an intermediate of benzoate catabolism. Plasmid pAC27, encoding enzymes for 3-chlorobenzoate degradation, does not appear to carry genes for chemotaxis toward chlorinated compounds.
Adherence of Bordetella bronchiseptica 276 to porcine trachea maintained in organ culture
Authors: Dugal F , Girard C , Jacques M
Published: January 2020

Two organ culture models have been adapted for porcine tracheae in order to study colonization by Bordetella bronchiseptica. Rings or segments excised from tracheae of newborn piglets were incubated overnight at 37 degrees C in a nutrient medium under 5% CO2-95% air conditions. Tracheal segments were infected with B bronchiseptica 276, and after different incubation times, bacterial counts were done. B. bronchiseptica adhered well to tracheae maintained in culture, and no statistically significant differences between the two models were observed. Noninfected tracheal mucosae maintained a normal appearance for several days, whereas infected mucosae showed typical damage caused by B. bronchiseptica, namely, loss of ciliary activity and cilia and sloughing of ciliated cells. Our data indicated that porcine tracheal organ culture could be advantageously used to study in vitro colonization by B. bronchiseptica.