bluuebunny/crossref_metadata_2025 huggingface dataset:
Isolation and Characterization of a Cyclohexane-Metabolizing
Xanthobacter
sp
Authors: Trower Michael K. , Buckland R. Martin , Higgins Raymond , Griffin Martin
Published: January 2020

An unusual
Xanthobacter
sp., capable of independent growth on cyclohexane as the sole source of carbon and energy, has been isolated from soil by using classical enrichment techniques. The mean generation time for growth on cyclohexane was 6 h. The microorganism showed a limited ability to utilize hydrocarbons, with only alicyclic hydrocarbons closely related to cyclohexane supporting growth. Ultrastructural studies indicated the presence of electron-transparent vesicles in the cyclohexane-grown
Xanthobacter
sp., but the presence of complex intracytoplasmic membranes could not be identified. A soluble inducible enzyme capable of oxidizing cyclohexane was identified in cell extracts. This enzyme had a pH optimum of 6.5, an absolute specificity for NADPH, and a stoichiometric requirement for molecular O
2
which was consistent with the formation of cyclohexanol. The enzyme showed no activity towards straight chain alkanes and only a limited activity towards unsaturated ring compounds. Enzymatic studies with cell extracts have indicated the main route of metabolism of cyclohexane by this
Xanthobacter
sp. to proceed via cyclohexane → cyclohexanol → cyclohexanone → 1-oxa-2-oxocycloheptane (ε-caprolactone) → 6-hydroxyhexanoate (6-hydroxycaproate) → → adipic acid. Alternative routes involving initial double hydroxylation of the cyclohexane ring may operate fortuituously but are unlikely to represent major pathways for the dissimilation of cyclohexane by this microorganism.
Some goodness-of-fit methods for the Poisson plus added zeros distribution
Authors: el-Shaarawi A H
Published: January 2020

Methods for making inferences about the Poisson plus added zeros distribution and the truncated Poisson distribution are presented and illustrated with bacteriological data. Some of the methods are designed for testing the compatibility of the zero frequency with the Poisson distribution, whereas others are given for testing the goodness of fit for the truncated Poisson. In particular, a modified form of the Fisher index of dispersion is presented which is suitable for the truncated case. It is shown that the use of the usual expression of the index of dispersion for testing the adequacy of the truncated Poisson is not correct and leads to accepting inadequate fits more frequently than expected on the basis of test of significance. Furthermore, three test statistics are presented for testing the compatability of the zero frequency with the Poisson distribution. The results of the simulation show that two test statistics, one due to Cochran (W. G. Cochran, Biometrics 10:417-451, 1954) and the other to Rao and Chakravarti (C. R. Rao and I. M. Chakravarti, Biometrics 12:264-282, 1956), are preferable to those from the likelihood ratio test.
A survey for viruses from fresh water that infect a eucaryotic chlorella-like green alga
Authors: Van Etten J L , Van Etten C H , Johnson J K , Burbank D E
Published: January 2020

Viruses which formed plaques on lawns of a eucaryotic, chlorella-like green alga were detected in 37% of the 35 freshwater samples surveyed. Virus populations, monitored in seven locations, fluctuated both qualitatively and quantitatively over an 8-month period.
Simple screening method for identification of nonpleiotropic mutants for exoenzyme production
Authors: Rose K E , Heath H E , Sloan G L
Published: January 2020

A differential medium that distinguishes between pleiotropic and nonpleiotropic mutants for exoenzyme production has been developed for Staphylococcus simulans biovar staphylolyticus. The medium will facilitate genetic analysis of exoenzyme production by this organism. Generally useful strategies for increasing the sensitivity of indicator plates for detection of exoenzyme activities are presented.
Isotherm for Adsorption of
Agrobacterium tumefaciens
to Susceptible Potato (
Solanum tuberosum
L.) Tissues
Authors: Kluepfel Daniel A. , Pueppke Steven G.
Published: January 2020

Potato tuber disks were submerged in suspensions containing 10
1
to 10
9
cells of
Agrobacterium tumefaciens
B6 per ml. After 60 min, the disks were rinsed and homogenized, and portions of the homogenates were plated to measure the number of adsorbed bacteria. At low initial bacterial concentrations (10
5
/ml), 5 to 23% of the bacteria adsorbed. At higher bacterial concentrations, the corresponding value was approximately 1.2%. Adsorption was a reversible equilibrium process. Binding saturation was not achieved, and adsorbed bacteria were confined to monolayers on the surfaces of tissue prepared for scanning electron microscopy. Adsorption of strain B6 to potato tuber tissues is described accurately by the Freundlich adsorption isotherm and may be a nonspecific phenomenon.
Effect of Cultural Practices, Soil Phosphorus, Potassium, and pH on the Incidence of Fusarium Head Blight and Deoxynivalenol Levels in Wheat
Authors: Teich A. H. , Hamilton J. R.
Published: January 2020

In a survey conducted in 1984 in Essex, Lambton, and Middlesex Counties of Ontario, Canada, greater incidences of head blight and greater concentrations of deoxynivalenol in grain were observed in fields of winter wheat planted after corn than in those planted after soybeans, barley, and mixed grains. Neither head blight nor deoxynivalenol level were correlated significantly with soil P, K, or pH. Head blight was reduced in wheat planted after corn where the residues from a preceding corn crop were plowed down or where seed was treated with Vitaflo 280. However, head blight and deoxynivalenol levels were not affected significantly by the level of N application, cultivar, other diseases or herbicides.
Conserved Nodulation Genes in
Rhizobium meliloti
and
Rhizobium trifolii
Authors: Fisher Robert F. , Tu Janice K. , Long Sharon R.
Published: January 2020

Plasmids which contained wild-type or mutated
Rhizobium meliloti
nodulation (
nod
) genes were introduced into Nod
−
R. trifolii
mutants ANU453 and ANU851 and tested for their ability to nodulate clover. Cloned wild-type and mutated
R. meliloti nod
gene segments restored ANU851 to Nod
+
, with the exception of
nodD
mutants. Similarly, wild-type and mutant
R. meliloti nod
genes complemented ANU453 to Nod
+
, except for
nodCII
mutants. Thus, ANU851 identifies the equivalent of the
R. meliloti nodD
genes, and ANU453 specifies the equivalent of the
R. meliloti nodCII
genes. In addition, cloned wild-type
R. trifolii nod
genes were introduced into seven
R. meliloti
Nod
−
mutants. All seven mutants were restored to Nod
+
on alfalfa. Our results indicate that these genes represent common nodulation functions and argue for an allelic relationship between
nod
genes in
R. meliloti
and
R. trifolii
.
Mixed carbon source utilization of meat-spoiling Pseudomonas fragi 72 in relation to oxygen limitation and carbon dioxide inhibition
Authors: Molin G
Published: January 2020

The growth of meat-spoiling Pseudomonas fragi 72 was studied on a defined salt medium supplemented with L-aspartate, citrate, creatine, creatinine, D-glucose, L-glutamate, and L-lactate. The utilization of the different carbon sources was followed in batch and continuous culture and under the influence of oxygen limitation and carbon dioxide inhibition (50% CO2 in air). Under nonrestricted atmospheric conditions in batch culture, the organism showed a preference in the utilization of the carbon sources in the order glucose greater than lactate greater than citrate greater than aspartate-glutamate greater than creatine greater than creatinine. The first five sources were utilized simultaneously. The order of preference was changed in continuous culture to lactate-citrate-glutamate-aspartate greater than glucose greater than creatine greater than creatinine. All carbon sources were utilized at lower dilution rates, but as the rate was increased the concentration of the carbon sources started to increase in the effluent and the preference could be seen. Under conditions of oxygen limitation the preference for glucose was weakened, but for lactate it was slightly enhanced (batch and continuous culture). Under conditions of CO2 inhibition, the preference for glucose was enhanced. However, lactate and amino acids were still preferred to glucose in the continuous culture. The utilization of creatine and creatinine was blocked by CO2 in batch culture, and only a slight utilization of creatine was noticed in a chemostat at lower dilution rates.
Isolation and Some Properties of the Enzyme That Transforms Eremofortin C to PR Toxin
Authors: Chang Shenq-Chyi , Wei Yau-Huei , Liu Mei-Ling , Wei Ru-Dong
Published: January 2020

PR toxin and eremofortin C are secondary metabolites of
Penicillium roqueforti.
The chemical structures of these two compounds are closely related to each other and differ only by an aldehyde and an alcohol group at the C-12 position. In an effort to better understand the biosynthesis of PR toxin, we discovered the enzyme of
P. roqueforti
that is responsible for the transformation of eremofortin C to PR toxin. The maximum activity of the enzyme in the culture medium was found to occur on day 13, which corresponded to the maximal production of PR toxin in the medium. The enzyme was isolated and purified from the culture medium and the mycelium of the fungus, respectively, through a procedure involving ammonium sulfate fractionation and DEAE-cellulose chromatography. The specific activity increased 20- and 8-fold, respectively, and the yield was 33.3 and 21.6%, respectively, for the enzyme from the medium and mycelium. The optimal pH for the enzyme reaction was ca. pH 5.6. The enzyme reaction was temperature dependent. The rates followed a linear time course when it catalyzed the transformation at 30°C and decayed with time when reacted at higher temperatures. At 100°C, the enzyme activity was completely lost. The

K
m

and

V
max

of the enzyme as determined at 30°C were 0.02 mM and 4.0 μmol/min per mg, respectively. The molecular weight of the enzyme was estimated by gel filtration on a high-pressure liquid chromatography I-250 protein column to be ca. 40,000.
Isolation of a Butyrate-Utilizing Bacterium in Coculture with
Methanobacterium thermoautotrophicum
from a Thermophilic Digester
Authors: Henson J. Michael , Smith P. H.
Published: January 2020

Sludge from a thermophilic, 55°C digester produced methane without a lag period when enriched with butyrate. The sludge was found by most-probable-number enumeration to have ca. 5 × 10
6
butyrate-utilizing bacteria per ml. A thermophilic butyrate-utilizing bacterium was isolated in coculture with
Methanobacterium thermoautotrophicum.
This bacterium was a gram-negative, slightly curved rod, occurred singly, was nonmotile, and did not appear to produce spores. When this coculture was incubated with
Methanospirillum hungatei
at 37°C, the quantity of methane produced was less than 5% of the methane produced when the coculture was incubated at 55°C, the routine incubation temperature. The coculture required clarified digester fluid. The addition of yeast extract to medium containing 5% clarified digester fluid stimulated methane production when a
Methanosarcina
sp. was present. Hydrogen in the gas phase prevented butyrate utilization. However, when the hydrogen was removed, butyrate utilization began. Penicillin G and
d
-cycloserine caused the complete inhibition of butyrate utilization by the coculture. The ability of various ecosystems to convert butyrate to methane was studied. Marine sediments enriched with butyrate required a 2-week incubation period before methanogenesis began. Hypersaline sediments did not produce methane after 3 months when enriched with butyrate.
Growth of
Agaricus campestris
NRRL 2334 in the Form of Pellets
Authors: Martin Antonio M. , Bailey Valerie I.
Published: January 2020

The production of pellets of the fungus
Agaricus campestris
NRRL 2334 was studied in submerged fermentation with peat extract as the main substrate source. Pellets up to 6 mm in diameter were obtained when the peat extract was diluted to reduce the concentration of growth inhibitors. Yeast extract and yeast extract plus glucose were the most effective nutrient supplements in the diluted peat extract media and stimulated the formation of large pellets which contained 44.4% crude protein, 2.8% fat, and 9% ash (dry weight basis). No solid supports were required for the growth of the pellets. The effects on the growth morphology of several dilution ratios of the peat extract, rates of agitation and aeration, and time were investigated.
Association of
Rhizobium
Strains with Roots of
Trifolium repens
Authors: Badenoch-Jones Jane , Flanders David J. , Rolfe Barry G.
Published: January 2020

Two techniques were used to assess the binding of rhizobia to clover roots: indirect counting after radiolabeling the bacteria and direct counting by using phase-contrast microscopy. Microscopic observations revealed a large variability in the number of bacteria associated with individual root hairs. This variability made unbiased counting by microscopy difficult. Systematic examination of all visible root hairs and “blind” counting of coded strains and treatments were adopted to minimize observer bias. The validity of the radiolabeling method was also examined in some detail. The reproducibility of results from this method was satisfactory. However, drawbacks of this method included its lack of sensitivity and its failure to distinguish between bacteria attached to mature root hairs, emerging root hairs, and undifferentiated epidermal cells. The method also failed to distinguish between individual bacteria and any aggregates that may be present. The ability of a number of chosen mutant strains of
Rhizobium trifolii
and their corresponding parent strains, as well as a number of nonhomologous strains, to bind to clover roots was assessed by using both of these methods. Our results gave no indication of specificity of
R. trifolii
binding to clover roots. 2-Deoxy-
d
-glucose did not appear to have a major inhibitory effect on the attachment of rhizobia to the host root, which suggests that lectin cross-bridging is not an obligatory step in the initiation of infection even though it may occur under some conditions. The presence or absence of the symbiotic plasmid was not correlated with bacterial adherence to the host plant root. Since host specificity functions are carried on this plasmid, our results suggest that binding of rhizobia to the legume root is not the basis of host specificity.
Inhibition of mitogen-induced blastogenesis in human lymphocytes by T-2 toxin and its metabolites
Authors: Forsell J H , Kateley J R , Yoshizawa T , Pestka J J
Published: January 2020

Concentrations of T-2, HT-2, 3'-OH T-2, 3'-OH HT-2, T-2 triol, and T-2 tetraol toxins which inhibited [3H]thymidine uptake in mitogen-stimulated human peripheral lymphocytes by 50% were 1.5, 3.5, 4.0, 50, 150, and 150 ng/ml, respectively. The results suggested that the initial hydrolysis of T-2 toxin and the hydroxylation of T-2 toxin to 3'-OH T-2 toxin did not significantly decrease the immunotoxicity of the parent molecule, whereas further hydrolysis to T-2 triol and T-2 tetraol toxins or hydroxylation to 3'-OH HT-2 toxin decreased in vitro toxicity for human lymphocytes.
High Production of Thermostable β-Galactosidase of
Bacillus stearothermophilus
in
Bacillus subtilis
Authors: Hirata Haruhisa , Negoro Seiji , Okada Hirosuke
Published: January 2020

By cloning the β-galactosidase gene of
Bacillus stearothermophilus
IAM11001 (ATCC 8005) into
Bacillus subtilis
, enzyme production was enhanced 50 times. β-Galactosidase could be purified to 80% homogeneity by incubating the cell extract of
B. subtilis
at 70°C for 15 min, followed by centrifugation to remove the denatured proteins. Because of its heat stability and ease of production, β-galactosidase is suitable for application in industrial processes.
Survival of Bacteria and Fungi in Relation to Water Activity and the Solvent Properties of Water in Biopolymer Gels
Authors: Mugnier J. , Jung G.
Published: January 2020

Survival of bacteria (
Rhizobium, Agrobacterium
, and
Arthrobacter
spp.), fungal spores (
Penicillium
sp.), and yeasts (
Saccharomyces
sp.) was studied in relation to water activity (a
w
) and the presence of nutritive solutes. The cells were entrapped in polysaccharide gels, as is done to immobilize cells or enzymes, and then dehydrated. The number of living cells (10
10
g of dry polymer
−1
) remained constant for periods of storage of >3 years at 28°C when the inocula were kept at an a
w
of <0.069. At a
w
values between 0.069 and 0.83 the number of survivors diminished more and more rapidly as the a
w
was raised. For a given a
w
and organism, there were large differences in survival rate as a function of the nutritive solutes used to culture the microorganisms. Low-molecular-weight compounds (with three or five carbon atoms) had a deleterious effect on survival, whereas compounds of higher molecular weight (C
6
to C
12
) had a protecting effect. Thus, the a
w
alone was not a sufficient explanation for the deterioration of the inocula. Survival seemed to be more directly related to some properties of the water in the biopolymer. New concepts such as the discontinuity of properties of water and the point of mobilization of solutes, already proposed by Duckworth and Kelly (J. Food Technol. 8:105-113, 1973) and Seow (J. Sci. Food Agric., 26:535-536, 1975), have been taken into consideration to explain the interactions of water with the biopolymer and their specific effects on the microorganisms.
Inhibition of Clostridium botulinum 52A toxicity and protease activity by sodium acid pyrophosphate in media systems
Authors: Wagner M K , Busta F F
Published: January 2020

The effects of two pH levels (5.55 or 5.85) in combination with 0.4% sodium acid pyrophosphate (SAPP), NaH2PO4 X H2O, Na2HPO4 X 7H2O, or NaCl on the growth and toxicity of Clostridium botulinum 52A were studied. Absorbancy measurements at 630 nm, microscopic observations, and the mouse bioassay procedure were used to observe the effects. At pH 5.55 and 5.85 most control cultures exhibited toxicity when cell lysis began. Vegetative cell development was normal (4 micron long; 1 micron wide). SAPP-containing (0.4%) treatment cultures displayed similar growth and lysis but no or delayed (48 h) toxicity. Cells grown in the SAPP treatment culture were longer and wider (6 micron long; 1.5 micron wide) than in most other treatment cultures. Trypsinization of nontoxic supernatants from 0.4% SAPP resulted in toxicity. Addition of 0.4% SAPP to toxic C. botulinum supernatant delayed but did not prevent death of mice. The addition of various levels of SAPP to toxic supernatants resulted in a decrease in zone size with an increase in the level of SAPP (9 mm with 0.4% SAPP to 7 mm with 1.0% SAPP), using a dual substrate protease assay. A decrease in the zone size also occurred with the supernatant from cultures grown in the presence of SAPP and with Bacillus polymyxa protease dilutions containing 0.4% SAPP. Results suggest that the actual production or function of the protease responsible for toxin activation may have been inhibited by the presence of SAPP.
Nutritional Requirements of
Methanosarcina
sp. Strain TM-1
Authors: Murray Patti A. , Zinder Stephen H.
Published: January 2020

Methanosarcina
sp. strain TM-1, an acetotrophic, thermophilic methanogen isolated from an anaerobic sludge digestor, was originally reported to require an anaerobic sludge supernatant for growth. It was found that the sludge supernatant could be replaced with yeast extract (1 g/liter), 6 mM bicarbonate-30% CO
2
, and trace metals, with a doubling time on methanol of 14 h. For growth on either methanol or acetate, yeast extract could be replaced with CaCl
2
· 2H
2
O (13.6 μM minimum) and the vitamin
p
-aminobenzoic acid (PABA, ca. 3 nM minimum), with a doubling time on methanol of 8 to 9 h. Filter-sterilized folic acid at 0.3 μM could not replace PABA. The antimetabolite sulfanilamide (20 mM) inhibited growth of and methanogenesis by
Methanosarcina
sp. strain TM-1, and this inhibition was reversed by the addition of 0.3 μM PABA. When a defined medium buffered with 20 mM
N,N
-bis(2-hydroxyethyl)-2-aminoethanesulfonic acid was used, it was shown that
Methanosarcina
sp. strain TM-1 required 6 mM bicarbonate-30% CO
2
for optimal growth and methanogenesis from methanol. Cells growing on acetate were less dependent on bicarbonate-CO
2
. When we used a defined medium in which the only organic compounds present were methanol or acetate, nitrilotriacetic acid (0.2 mM), and PABA, it was possible to limit batch cultures of
Methanosarcina
sp. strain TM-1 for nitrogen at NH
4
+
concentrations at or below 2.0 mM, in marked contrast with
Methanosarcina barkeri
227, which fixes dinitrogen when grown under NH
4
+
limitation.
Monoclonal antibody-based immunoassay for type A Clostridium botulinum toxin is comparable to the mouse bioassay
Authors: Shone C , Wilton-Smith P , Appleton N , Hambleton P , Modi N , Gatley S , Melling J
Published: January 2020

A monoclonal antibody (BA11) has been produced against Clostridium botulinum type A neurotoxin by the fusion of myeloma cells (P3 NS1/1-Ag4-1) with spleen cells from BALB/c mice immunized with botulinum type A neurotoxoid. The antibody bound specifically to botulinum type A neurotoxin, showing no cross-reactivity with types B and E botulinum toxins or with any of several other bacterial toxins tested. The monoclonal antibody did not bind to botulinum type A neurotoxin which had been denatured with sodium dodecyl sulfate and bound only weakly to each of the separated heavy and light subunits of the neurotoxin, suggesting a conformational requirement for the antigenic determinant of the antibody. A sensitive immunoassay for C. botulinum type A toxin with monoclonal antibody BA11 in conjunction with an enzyme amplication system has been developed which allows detection of 5 to 10 mouse 50% lethal doses ml-1 of purified neurotoxin. The assay was equally sensitive when applied to the detection of crude toxin in food stuffs; the average value for the minimum level of detectable toxin in extracts of tinned salmon or corned beef was 9 +/- 3.1 mouse 50% lethal doses ml-1.
Intestinal microbial flora after feeding phytohemagglutinin lectins (Phaseolus vulgaris) to rats
Authors: Banwell J G , Howard R , Cooper D , Costerton J W
Published: January 2020

Incorporation of purified phytohemagglutinin (PHA) lectins derived from red kidney beans (Phaseolus vulgaris) in the diet of weanling rats will cause growth failure, malabsorption of nutrients, and bacterial overgrowth in the small intestine. These effects are not caused by feeding a similar quantity of PHA to germfree rats. To define the morphological and bacterial changes on the mucosal surfaces of the jejunum, ileum, and cecum in greater detail, we pair fed two groups of weanling rats isocaloric, isonitrogenous diets with or without 0.5% PHA protein. On the jejunal surfaces of control rats, the mucous layer was a confluent covering with sparsely scattered bacteria and protozoa. In PHA-treated rats, the mucous layer was thin and discontinuous, and the microvillous surface of the tissue was extensively populated by bacterial cells of two distinct morphotypes--a gram-negative rod and a gram-positive coccobacillus. In all PHA-treated animals, these bacteria formed adherent monospecific or mixed adherent microcolonies on the tissue surface. Tissue damage was observed in PHA-exposed jejunal tissue as evidenced by vesiculation of the microvillous plasma membrane and by damage to the brush border membrane. On the ileal surfaces of control rats, there was a thick mucous layer within which small numbers of bacteria and protozoa were seen. Segmented filamentous bacteria were anchored in the tissue surface. In PHA-treated rats, the ileal surface was only incompletely covered by a mucous layer, and the overlying mucosal surface was extensively covered by large numbers of protozoan cells (predominantly Hexamita muris). Most of the ileal surfaces not covered by the mucous layer were occupied and virtually occluded by an overgrowth of these protozoan cells with occasional cells of Giardia muris and the tissue-associated segmented bacillus. In the ceca of control rats, the mucosa was incompletely covered by a discontinuous mucous layer and colonized by an unnamed Spirillum sp., other bacteria, and occasional protozoa. The cecal surfaces of PHA-treated rats retained most of their incomplete overlying mucous layer, which was heavily colonized by the same type of Spirillum sp. seen in untreated animals; intestinal crypts were colonized. These descriptive morphological studies demonstrate that exposure to purified PHA in the diet caused characteristic changes in the microbial ecology of the small intestine. The changes in microbial flora contributed to the malabsorption of nutrients in the small intestines of PHA-fed animals.
Potential Importance of Fish Predation and Zooplankton Grazing on Natural Populations of Freshwater Bacteria
Authors: Riemann Bo
Published: January 2020

The rates of ingestion of natural bacterial assemblages by natural populations of zooplankton (>50 μm in size) were measured during a 19-day period in eutrophic Frederiksborg Slotssø, Denmark, as well as in experimental enclosures (containing 5.3 m
3
of lake water). The fish and nutrients of the enclosures were manipulated. In enclosures without fish, large increases in ingestion by zooplankton >140 μm in size were found (up to 3 μg of C liter
−1
h
−1
), compared with values less than 0.3 μg of C liter
−1
h
−1
in the enclosures with fish and in the open lake.
Daphnia cucullata
and
D. galeata
dominated the community of zooplankton of >140 μm. Ingestion rates for zooplankton between 50 and 140 μm decreased after a period of about 8 days, in all enclosures and in the lake, to values below 0.1 μg of C liter
−1
h
−1
. On the last 2 sampling days, somewhat higher values were observed in the enclosures with fish present. The >50-μm zooplankton ingested 48 to 51% of the bacterial net secondary production in enclosures without fish, compared to 4% in the enclosures with added fish. Considering the sum of bacterial secondary production plus biomass change, 35 to 41% of the available bacteria were ingested by zooplankton of >50 μm in the enclosures without fish, compared with 4 to 6% in the enclosures with added fish and 21% in the open lake. Fish predation reduced the occurrence of zookplankton sized >50 μm and thus left a large proportion of the available bacteria to zooplankton sized <50 μm. In fact, there were 4.6 × 10
3
to 5.0 × 10
3
flagellates (4 to 8 μm in size) ml
−1
in the enclosures with fish added as well as in the lake, compared with 0.5 × 10
2
to 2.3 × 10
2
ml
−1
in the enclosures without fish. This link in the food chain was reduced when fish predation on zooplankton was eliminated and a direct route of dissolved organic matter, via the bacteria to the zooplankton, was established.
Muramic Acid Measurements for Bacterial Investigations in Marine Environments by High-Pressure Liquid Chromatography
Authors: Mimura Toru , Romano Jean-Claude
Published: January 2020

Muramic acid, a constituent of procaryotic cell walls, was assayed by high-pressure liquid chromatography in samples from several marine environments (water column, surface microlayer, and sediment) and a bacterial culture. It is used as a microbial biomass indicator. The method gave a good separation of muramic acid from interfering compounds with satisfactory reproducibility. A pseudomonad culture had a muramic acid content of 4.7 � 10
−10
to 5.3 � 10
−10
μg per cell during growth. In natural water samples, highly significant relationships were found between muramic acid concentrations and bacterial numbers for populations of 10
8
to 10
11
cells per liter. The muramic acid content in natural marine water decreased from 5.3 � 10
−10
to 1.6 � 10
−10
μg per cell with increasing depth. In coastal sediments exposed to sewage pollution, concentrations of muramic acid, ATP, organic carbon, and total amino acids displayed a parallel decrease with increasing distance from the sewage outlet. Advantages of muramic acid measurement by high-pressure liquid chromatography are its high sensitivity and reduction of preparation steps, allowing a short time analysis.
Isolation and characterization of two new fusaric acid analogs from Fusarium moniliforme NRRL 13,163
Authors: Burmeister H R , Grove M D , Peterson R E , Weisleder D , Plattner R D
Published: January 2020

Fusarium moniliforme NRRL 13,163 produced two new fusaric acid analogs, a 10,11-dihydroxyfusaric acid and a diacid of fusaric acid in which the C-11 methyl was oxidized to a carboxyl. Several hundred milligrams of the 10,11-dihydroxyfusaric acid were routinely recovered from a kilogram of corn grit medium. It crystallized as white, irregularly shaped rectangles that melted at 153 to 154 degrees C. The diacid analog of fusaric acid crystallized as white rods that melted at 210 to 211 degrees C. Unlike the consistent recovery experienced with the 10,11-dihydroxyfusaric acid, the diacid analog proved difficult to purify after the initial discovery and was detectable in subsequent fermentations only by mass spectrometry.
Models for the kinetics of biodegradation of organic compounds not supporting growth
Authors: Schmidt S K , Simkins S , Alexander M
Published: January 2020

We developed 12 models of kinetics to describe the metabolism of organic substrates that are not supporting bacterial growth. These models can be used to describe the biodegradation of organic compounds that are not supporting growth when the responsible populations are growing logistically, logarithmically, or linearly or are not increasing in numbers. Nonlinear regression analysis was used to fit patterns of mineralization by two bacteria to these kinetic models. Pseudomonas acidovorans mineralized 1 ng of phenol per ml while growing exponentially at the expense of uncharacterized organic carbon in a synthetic medium. Phenol at a concentration of 1 ng/ml did not affect the growth of P. acidovorans. These data were best fit by the model that incorporates the equation for logarithmic growth and assumes a concentration of test substrate well below its Km value. In the absence of a second substrate, glucose at concentrations below those supporting growth was mineralized by Salmonella typhimurium in a manner best described by pseudo first-order kinetics. In the presence of different concentrations of arabinose, however, the kinetics of glucose mineralization by S. typhimurium reflected linear, logistic, or logarithmic growth of the population on arabinose. We conclude that the kinetics of mineralization of organic compounds at concentrations too low to support growth are best described either by the first-order model or by models that incorporate expressions for the kinetics of growth of the metabolizing population on other substrates. When growth is at the expense of other substrates, the kinetics observed reflect such growth, as well as the concentration of the substrate of interest.(ABSTRACT TRUNCATED AT 250 WORDS)
Indirect enzyme-linked immunosorbent assay for the mycotoxin zearalenone
Authors: Liu M T , Ram B P , Hart L P , Pestka J J
Published: January 2020

A competitive indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of zearalenone, an estrogenic mycotoxin. Zearalenone was converted to zearalenone-6'-carboxymethyloxime and conjugated to bovine serum albumin and poly-L-lysine for use as immunogen and solid-phase marker, respectively. Immunization of rabbits with the bovine serum albumin conjugate resulted in zearalenone antibody titers of 20,480 in 11 weeks. A competitive indirect ELISA was conducted by simultaneously incubating zearalenone with zearalenone antiserum over zearalenone-6'-carboxymethyloxime poly-L-lysine solid phase and then determining the bound rabbit immunoglobulin with goat anti-rabbit peroxidase conjugate. Response range for zearalenone in the resulting competition curve was between 1 and 50 ng/ml. Reactivities of this antiserum for alpha-zearalenol, beta-zearalenol, alpha-zearalanol, and beta-zearalanol were, respectively, 50, 12, 6, and 3% of that found for zearalenone. By using the competitive indirect ELISA, zearalenone was detectable in methanol-water extracts of corn, wheat, and pig feed samples.
Sensitivity to bile salts of Shigella flexneri sublethally heat stressed in buffer or broth
Authors: Tollison S B , Johnson M G
Published: January 2020

Batch cultures of Shigella flexneri M4243 were grown at 37 degrees C in broth to early stationary phase, washed, and heated at 50 degrees C in 0.1 M phosphate buffer (pH 7.0). Cells were surface plated on a tryptic phytone glucose agar (TPGA), TPGA with 0.15 or 0.85% bile salts no. 3 (TPGA-BS 0.15 or TPGA-BS 0.85), or TPGA with 0.25 or 0.50% sodium deoxycholate (TPGA-DC 0.25 or TPGA-DC 0.50). Cells sampled after no heating produced colony counts on TPGA-BS 0.85 or on TPGA-DC 0.50 that were no more than about 0.5 log lower than for unheated cell samples plated on TPGA. Cells heated at 50 degrees C for 30 min produced colony counts on TPGA-DC 0.50 or on TPGA-BS 0.85 that were about 1.5 logs lower than on TPGA. Cells heated for 30 min and shifted to TPG broth at 37 degrees C to allow resuscitation required about 2 h to regain tolerance to 0.85% BS. However, heated cells resuscitated on solid TPGA at 35 degrees C before being challenged with overlays of TPGA-BS 0.85 or TPGA-DC 0.50 required 6 to 8 h on TPGA to regain tolerance to 0.85% BS or 0.50% DC. To regain tolerance to overlays of 0.15% BS or 0.25% DC, heated cells required resuscitation periods on TPGA of about 2 or 2 to 6 h, respectively. Cells heated in TPG broth and sampled after no heating produced colony counts on TPGA that were about 1.5 logs lower than for unheated cell suspensions, suggesting greater apparent injury when heat stressed in broth than in buffer.(ABSTRACT TRUNCATED AT 250 WORDS)
Microbiology and ration digestibility in the hindgut of the ovine
Authors: Lewis S M , Dehority B A
Published: January 2020

Contents of the terminal ilea, ceca-proximal colons, and terminal recta were obtained from nine sheep, three of which were fed 100% orchardgrass hay, three of which were fed 60% cracked corn-40% orchardgrass hay, and three of which were fed 80% cracked corn-20% orchardgrass hay. The digestibility of dry matter in the ceca was greatest when the all-hay diet was fed, whereas the percentage of cellulose digestion in the ceca increased with increasing levels of concentrate. For all diets, the total volatile fatty acid concentrations were higher in the ceca than in the other two sites. The cecal pH levels decreased with increased corn levels in the diet. The total microbiol numbers per gram of ileal and cecal contents increased in response to feeding of concentrate; however, across all diets, the ileal counts were 8% or less of the cecal counts. In contrast, the cellulolytic microbial numbers in the ilea were 50% or more of those in the ceca and were highest with the all-hay diet. Both bacterial and end product concentrations in the ceca were equivalent to those occurring in rumina. A total of 16 cellulolytic cultures were isolated and characterized from ileal and cecal contents of animals fed all three diets. Seven gram-negative-rod-shaped organisms were identified as Butyrivibrio fibrisolvens, whose capacity to digest cellulose exceeded that of several rumen strains. The guanine-plus-cytosine content of the DNA of one strain of B. fibrisolvens was 38.8 mol%, compared with the only previously reported value for this species of 41.2 mol%.(ABSTRACT TRUNCATED AT 250 WORDS)
Reduced virulence of Yersinia enterocolitica by copper-induced injury
Authors: Singh A , LeChevallier M W , McFeters G A
Published: January 2020

A sublethal concentration of copper (0.75 mg/liter) caused substantial injury (87 to 95%) of Yersinia enterocolitica serotype O:8 cells in 72 h at 4 degrees C without producing extensive cell death. Copper-injured cells had a higher 50% lethal dose in mice (2,700 CFU) than uninjured cells (150 CFU). This reduced virulence correlated with more rapid clearance of the injured cells from the blood of mice after intravenous inoculation. A possible role of the liver in this process was shown by significant cell accumulation in mouse livers when copper-injured Y. enterocolitica cells were administered, compared with uninjured bacteria. In vitro studies with isolated mouse liver membranes showed higher titers of aggregation with copper-injured cells than control cells. The in vitro aggregation reaction and blood clearance activity in vivo were abolished by sugars that are known to interact with a hepatic lectin. Our data suggest that copper-induced injury reduces the virulence of Y. enterocolitica and that the liver may be involved in nonimmune rapid clearance of the injured cells, probably by interaction with a hepatic lectin(s).
Changes in virulence of waterborne enteropathogens with chlorine injury
Authors: LeChevallier M W , Singh A , Schiemann D A , McFeters G A
Published: January 2020

We designed experiments to assess the effect of chlorine injury on the virulence of waterborne enteropathogens. Higher chlorine doses (0.9 to 1.5 mg/liter) were necessary to produce injured Yersinia enterocolitica, Salmonella typhimurium, and Shigella spp. than to produce injured enterotoxigenic Escherichia coli or coliform bacteria (0.25 to 0.5 mg/liter) in the test system used; 50% lethal dose experiments in which mice were used showed that injured Y. enterocolitica cells were 20 times less virulent than uninjured control cells (3,300 and 160 CFU, respectively). This decrease in virulence was not related to reduced attachment to Henle 407 intestinal epithelial cells, but could be related to a loss of HeLa cell invasiveness. In contrast, injured S. typhimurium and enterotoxigenic E. coli cells lost their ability to attach to Henle cells. These data show that some enteropathogens and coliform bacteria differ in their sensitivities to chlorine injury and that the virulence determinants affected by chlorine may vary from one pathogen to another.
Recombinant plasmid conferring proline overproduction and osmotic tolerance
Authors: Jakowec M W , Smith L T , Dandekar A M
Published: January 2020

A recombinant plasmid carrying the proBA (pro-74) mutant allele which governs osmotic tolerance and proline overproduction was constructed by using the broad-host-range plasmid vector pQSR49. The physiological, biochemical, and genetic properties of strains carrying the pQSR49 derivatives pMJ101 and pMJ1, mutant and wild type, respectively, were investigated. pMJ101 conferred enhanced osmotolerance compared with strains carrying the wild type, pMJ1. These results are in contrast to those obtained previously with strains carrying recombinant plasmids based on pBR322 that failed to confer the osmotic tolerance phenotype. gamma-Glutamyl kinase (first step in proline biosynthesis) from strains carrying pMJ101 was 200-fold less sensitive to feedback inhibition than was the wild-type enzyme. As expected, the intracellular proline levels of strains carrying pMJ101 were more than an order of magnitude higher than those of the wild type. An analysis of copy number revealed that the pQSR49 constructs were present in the cell at a level six- to eightfold lower than those of the pBR322 recombinants, which may account for the difference in phenotype. We found that the genetic stability of the pQSR49 derivative in a variety of gram-negative bacteria was dependent on the insert orientation and the presence of foreign DNA on the plasmid. These factors may be significant in future studies aimed at expanding the osmotolerance phenotype to a broad range of gram-negative bacteria.
Survival and enumeration of the fecal indicators Bifidobacterium adolescentis and Escherichia coli in a tropical rain forest watershed
Authors: Carrillo M , Estrada E , Hazen T C
Published: January 2020

The density of Bifidobacterium spp., fecal coliforms, Escherichia coli, and total anaerobic bacteria, acridine orange direct counts, percentages of total bacterial community activity and respiration, and 12 physical and chemical parameters were measured simultaneously at six sites for 12 months in the Mameyes River rain forest watershed, Puerto Rico. The densities of all bacteria were higher than those reported for uncontaminated temperate rivers, even though other water quality parameters would indicate that all uncontaminated sites were oligotrophic. The highest densities for all indicator bacteria were at the site receiving sewage effluent; however, the highest elevation site in the watershed had the next highest densities. Correlations between bacterial densities, nitrates, temperature, phosphates, and total phosphorus indicated that all viable counts were related to nutrient levels, regardless of the site sampled. In situ diffusion chamber studies at two different sites indicated that E. coli could survive, remain physiologically active, and regrow at rates that were dependent on nutrient levels of the ambient waters. Bifidobacterium adolescentis did not survive at either site but did show different rates of decline and physiological activity at the two sites. Bifidobacteria show promise as a better indicator of recent fecal contamination in tropical freshwaters than E. coli or fecal coliforms; however, the YN-6 medium did not prove to be effective for enumeration of bifidobacteria. The coliform maximum contaminant levels for assessing water usability for drinking and recreation appear to be unworkable in tropical freshwaters.
Plasmid DNA in Strains of
Pediococcus cerevisiae
and
Pediococcus pentosaceus
Authors: Graham Donald C. , McKay Larry L.
Published: January 2020

Five parental strains of
Pediococcus
were examined for plasmid content. Each strain contained three to six resident plasmids, ranging in size from 4.5 to 39.5 megadaltons. A bacteriocin-like substance produced by
Pediococcus cerevisiae
FBB63 was tentatively linked to a 10.5-megadalton plasmid after being cured with novobiocin.

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Sulfate-Dependent Interspecies H
2
Transfer between
Methanosarcina barkeri
and
Desulfovibrio vulgaris
during Coculture Metabolism of Acetate or Methanol
Authors: Phelps T. J. , Conrad R. , Zeikus J. G.
Published: January 2020

We compared the metabolism of methanol and acetate when
Methanosarcina barkeri
was grown in the presence and absence of
Desulfovibrio vulgaris.
The sulfate reducer was not able to utilize methanol or acetate as the electron donor for energy metabolism in pure culture, but was able to grow in coculture. Pure cultures of
M. barkeri
produced up to 10 μmol of H
2
per liter in the culture headspace during growth on acetate or methanol. In coculture with
D. vulgaris
, the gaseous H
2
concentration was ≤2 μmol/liter. The fractions of
14
CO
2
produced from [
14
C]methanol and 2-[
14
C]acetate increased from 0.26 and 0.16, respectively, in pure culture to 0.59 and 0.33, respectively, in coculture. Under these conditions, approximately 42% of the available electron equivalents derived from methanol or acetate were transferred and were utilized by
D. vulgaris
to reduce approximately 33 μmol of sulfate per 100 μmol of substrate consumed. As a direct consequence, methane formation in cocultures was two-thirds that observed in pure cultures. The addition of 5.0 mM sodium molybdate or exogenous H
2
decreased the effects of
D. vulgaris
on the metabolism of
M. barkeri.
An analysis of growth and carbon and electron flow patterns demonstrated that sulfate-dependent interspecies H
2
transfer from
M. barkeri
to
D. vulgaris
resulted in less methane production, increased CO
2
formation, and sulfide formation from substrates not directly utilized by the sulfate reducer as electron donors for energy metabolism and growth.
Activity Stain for Rapid Characterization of Pectic Enzymes in Isoelectric Focusing and Sodium Dodecyl Sulfate-Polyacrylamide Gels
Authors: Ried J. L. , Collmer A.
Published: January 2020

A system was developed for the rapid characterization of microbial pectic enzyme complexes and then tested on
Erwinia chrysanthemi
and
Sclerotium rolfsii.
Pectic enzymes in minute samples of crude culture filtrates were resolved by ultrathin-layer polyacrylamide gel isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then assayed with an ultrathin pectate-agarose overlay stained with ruthenium red. The simple procedure can be completed within 30 min after isoelectric focusing, can detect extremely low levels of pectate lyase (6.4 × 10
−6
μmol of product per min), and is sufficiently sensitive to determine the pectate lyase isozyme profile of a single bacterial colony with a diameter of 4 mm. Pectate lyases and polygalacturonases can be distinguished by altering buffer conditions in the overlays. The assay system revealed additional isozymes not resolved by classical techniques and generally corroborated the previously published isoelectric points and molecular weights of the pectate lyase isozymes and exo-poly-α-
d
-galacturonosidase produced by
E. chrysanthemi
and the endopolygalacturonase and exopolygalacturonase produced by
S. rolfsii.
Fungal metabolism and detoxification of the nitropolycyclic aromatic hydrocarbon 1-nitropyrene
Authors: Cerniglia C E , Freeman J P , White G L , Heflich R H , Miller D W
Published: January 2020

Nitropolycyclic aromatic hydrocarbons are ubiquitous environmental pollutants, many of which are potent mutagens in bacterial and mammalian cells and carcinogenic to rodents. In this study, we investigated the fungal metabolism of 1-nitropyrene and determined the mutagenic activity of the metabolites toward Salmonella typhimurium TA98, TA98NR, and TA100. Cunninghamella elegans metabolized 1-nitropyrene to form glucoside conjugates of 6-hydroxy-1-nitropyrene and 8-hydroxy-1-nitropyrene. The metabolites were isolated by reversed-phase high-pressure liquid chromatography and characterized by application of UV absorption, 1H-nuclear magnetic resonance, and mass spectroscopy. Mutagenicity assays performed on samples extracted from incubations of C. elegans with 1-nitropyrene indicated that mutagenic activity decreased with time. Consistent with the loss in mutagenic activity, the glucoside conjugates of 6- and 8-hydroxy-1-nitropyrene were nonmutagenic in the Salmonella reversion assay. The results indicate that the fungus C. elegans metabolizes 1-nitropyrene to detoxified products.
Genetic Recombination and Transformation in Protoplasts of
Thermomonospora fusca
Authors: Pidcock Kenneth A. , Montenecourt Bland S. , Sands Jeffrey A.
Published: January 2020

Protoplasts were produced from the thermophilic actinomycete
Thermomonospora fusca
and were regenerated to 0.1% of the direct count on regeneration agar. Recombination after protoplast fusion was demonstrated with drug-resistant mutants of
T. fusca
YX. A single thiostrepton-resistant colony was isolated after transformation of
T. fusca
YX with the streptomycete vector pIJ702, providing the first evidence for transformation in the genus
Thermomonospora
and suggesting that some mesophilic streptomycete genes can be expressed in thermophilic actinomycetes. Of 20 thermophilic actinomycete strains isolated from self-heated composts, 3 were found to harbor native plasmid DNA, providing potential sequences for the development of
Thermomonospora-Streptomyces
shuttle vectors.
Statistical association of dietary components with Simonsiella species residing in normal human mouths
Authors: Gregory D A , Kuhn D A , Daly K R , Flygenring K
Published: January 2020

Members of the genus Simonsiella, aerobic, multicellular filamentous gliding bacteria, were detected in swabbings from the palates of 32% of 212 human subjects free of gross oral pathologies. Nutritional evaluations for 142 of the subjects showed a significantly greater daily intake among 53 Simonsiella carriers for 13 dietary variables, including four fat components, but there was no significantly greater daily intake for any of the carbohydrate components. Overall, there was a general excess dietary intake by Simonsiella carriers. The mean dietary intake of the carriers was numerically greater than that of the noncarriers for 70 of 74 dietary variables.