bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Screening of
Enterococcus
Virulence Determinants and Potential for Genetic Exchange between Food and Medical Isolates
Authors: Eaton Tracy J. , Gasson Michael J. 
Published: July 2002

ABSTRACT

Enterococci are used as starter and probiotic cultures in foods, and they occur as natural food contaminants. The genus
Enterococcus
is of increased significance as a cause of nosocomial infections, and this trend is exacerbated by the development of antibiotic resistance. In this study, we investigated the incidence of known virulence determinants in starter, food, and medical strains of
Enterococcus faecalis
,
E. faecium
, and
E. durans
. PCR and gene probe strategies were used to screen enterococcal isolates from both food and medical sources. Different and distinct patterns of incidence of virulence determinants were found for the
E. faecalis
and
E. faecium
strains. Medical
E. faecalis
strains had more virulence determinants than did food strains, which, in turn, had more than did starter strains. All of the
E. faecalis
strains tested possessed multiple determinants (between 6 and 11).
E. faecium
strains were generally free of virulence determinants, with notable exceptions. Significantly,
esp
and
gelE
determinants were identified in
E. faecium
medical strains. These virulence determinants have not previously been identified in
E. faecium
strains and may result from regional differences or the evolution of pathogenic
E. faecium
. Phenotypic testing revealed the existence of apparently silent
gelE
and
cyl
genes. In
E. faecalis
, the trend in these silent genes mirrors that of the expressed determinants. The potential for starter strains to acquire virulence determinants by natural conjugation mechanisms was investigated. Transconjugation in which starter strains acquired additional virulence determinants from medical strains was demonstrated. In addition, multiple pheromone-encoding genes were identified in both food and starter strains, indicating their potential to acquire other sex pheromone plasmids. These results suggest that the use of
Enterococcus
spp. in foods requires careful safety evaluation.
Enterocin P Selectively Dissipates the Membrane Potential of
Enterococcus faecium
T136
Authors: Herranz C. , Chen Y. , Chung H.-J. , Cintas L. M. , Hernández P. E. , Montville T. J. , Chikindas M. L. 
Published: July 2002

ABSTRACT

Enterocin P is a pediocin-like, broad-spectrum bacteriocin which displays a strong inhibitory activity against
Listeria monocytogenes
. The bacteriocin was purified from the culture supernatant of
Enterococcus faecium
P13, and its molecular mechanism of action against the sensitive strain
E. faecium
T136 was evaluated. Although enterocin P caused significant reduction of the membrane potential (ΔΨ) and the intracellular ATP pool of the indicator organism, the pH gradient (ΔpH) component of the proton motive force (Δp) was not dissipated. By contrast, enterocin P caused carboxyfluorescein efflux from
E. faecium
T136-derived liposomes.
Methods for Integrated Air Sampling and DNA Analysis for Detection of Airborne Fungal Spores
Authors: Williams Roger H. , Ward Elaine , McCartney H. Alastair 
Published: July 2002

ABSTRACTIntegrated air sampling and PCR-based methods for detecting airborne fungal spores, usingPenicillium roquefortias a model fungus, are described.P. roquefortispores were collected directly into Eppendorf tubes using a miniature cyclone-type air sampler. They were then suspended in 0.1% Nonidet P-40, and counted using microscopy. Serial dilutions of the spores were made. Three methods were used to produce DNA for PCR tests: adding untreated spores to PCRs, disrupting spores (fracturing of spore walls to release the contents) using Ballotini beads, and disrupting spores followed by DNA purification. ThreeP. roqueforti-specific assays were tested: single-step PCR, nested PCR, and PCR followed by Southern blotting and probing. Disrupting the spores was found to be essential for achieving maximum sensitivity of the assay. Adding untreated spores to the PCR did allow the detection ofP. roqueforti, but this was never achieved when fewer than 1,000 spores were added to the PCR. By disrupting the spores, with or without subsequent DNA purification, it was possible to detect DNA from a single spore. When known quantities ofP. roquefortispores were added to air samples consisting of high concentrations of unidentified fungal spores, pollen, and dust, detection sensitivity was reduced.P. roquefortiDNA could not be detected using untreated or disrupted spore suspensions added to the PCRs. However, using purified DNA, it was possible to detect 10P. roquefortispores in a background of 4,500 other spores. For all DNA extraction methods, nested PCR was more sensitive than single-step PCR or PCR followed by Southern blotting.
Development of Aflatoxin B
1
-Lysine Adduct Monoclonal Antibody for Human Exposure Studies
Authors: Wang Jia-Sheng , Abubaker Salahaddin , He Xia , Sun Guiju , Strickland Paul T. , Groopman John D. 
Published: July 2002

ABSTRACT

Mouse monoclonal antibodies were developed against a synthetic aflatoxin B
1
(AFB)-lysine–cationized bovine serum albumin conjugate. The isotype of one of these antibodies, IIA4B3, has been classified as immunoglobulin G1(λ). The affinity and specificity of IIA4B3 were further characterized by a competitive radioimmunoassay. The affinities of IIA4B3 for AFB and its associated adducts and metabolites are ranked as follows: AFB-lysine > 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl formamido)-9-hydroxy-AFB > AFB = 8,9-dihydro-8-(
N
7
-guanyl)-9-hydroxy-AFB > aflatoxin M
1
> aflatoxin Q
1
. IIA4B3 had about a 10-fold higher affinity for binding to AFB-lysine adduct than to AFB when
3
H-AFB–lysine was used as the tracer. The concentration for 50% inhibition for AFB-lysine was 0.610 pmol; that for AFB was 6.85 pmol. IIA4B3 had affinities at least sevenfold and twofold higher than those of 2B11, a previously developed antibody against parent AFB, for the major aflatoxin-DNA adducts 8,9-dihydro-8-(
N
7
-guanyl)-9-hydroxy-AFB and 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl formamido)-9-hydroxy-AFB, respectively. An analytical method based on a competitive radioimmunoassay with IIA4B3 and
3
H-AFB–lysine was validated with a limit of detection of 10 fmol of AFB-lysine adduct. The method has been applied to the measurement of AFB-albumin adduct levels in human serum samples collected from the residents of areas at high risk for liver cancer.
Diversity and Characterization of Sulfate-Reducing Bacteria in Groundwater at a Uranium Mill Tailings Site
Authors: Chang Yun-Juan , Peacock Aaron D. , Long Philip E. , Stephen John R. , McKinley James P. , Macnaughton Sarah J. , Hussain A. K. M. Anwar , Saxton Arnold M. , White David C. 
Published: July 2002

ABSTRACT

Microbially mediated reduction and immobilization of U(VI) to U(IV) plays a role in both natural attenuation and accelerated bioremediation of uranium-contaminated sites. To realize bioremediation potential and accurately predict natural attenuation, it is important to first understand the microbial diversity of such sites. In this paper, the distribution of sulfate-reducing bacteria (SRB) in contaminated groundwater associated with a uranium mill tailings disposal site at Shiprock, N.Mex., was investigated. Two culture-independent analyses were employed: sequencing of clone libraries of PCR-amplified dissimilatory sulfite reductase (DSR) gene fragments and phospholipid fatty acid (PLFA) biomarker analysis. A remarkable diversity among the DSR sequences was revealed, including sequences from δ-Proteobacteria, gram-positive organisms, and the
Nitrospira
division. PLFA analysis detected at least 52 different mid-chain-branched saturate PLFA and included a high proportion of 10me16:0.
Desulfotomaculum
and
Desulfotomaculum
-like sequences were the most dominant DSR genes detected. Those belonging to SRB within δ-Proteobacteria were mainly recovered from low-uranium (≤302 ppb) samples. One
Desulfotomaculum
-like sequence cluster overwhelmingly dominated high-U (>1,500 ppb) sites. Logistic regression showed a significant influence of uranium concentration over the dominance of this cluster of sequences (
P
= 0.0001). This strong association indicates that
Desulfotomaculum
has remarkable tolerance and adaptation to high levels of uranium and suggests the organism's possible involvement in natural attenuation of uranium. The in situ activity level of
Desulfotomaculum
in uranium-contaminated environments and its comparison to the activities of other SRB and other functional groups should be an important area for future research.
Rapid Approach To Determine
rrn
Arrangement in
Salmonella
Serovars
Authors: Helm R. Allen , Maloy Stanley 
Published: July 2002

ABSTRACT

A PCR method was developed by which to rapidly and accurately determine the
rrn
arrangement of
Salmonella enterica
serovars. Primers were designed to the genomic regions flanking each of the seven
rrn
operons
.
PCR analysis using combinations of these primers will distinguish each of the possible arrangements of the
rrn
skeleton.
Construction and Environmental Release of a
Sinorhizobium meliloti
Strain Genetically Modified To Be More Competitive for Alfalfa Nodulation
Authors: van Dillewijn Pieter , Soto Marı́a José , Villadas Pablo J. , Toro Nicolás 
Published: July 2002

ABSTRACT

Highly efficient nitrogen-fixing strains selected in the laboratory often fail to increase legume production in agricultural soils containing indigenous rhizobial populations because they cannot compete against these populations for nodule formation. We have previously demonstrated, with a
Sinorhizobium meliloti
PutA
−
mutant strain, that proline dehydrogenase activity is required for colonization and therefore for the nodulation efficiency and competitiveness of
S. meliloti
on alfalfa roots (J. I. Jiménez-Zurdo, P. van Dillewijn, M. J. Soto, M. R. de Felipe, J. Olivares, and N. Toro, Mol. Plant-Microbe Interact. 8:492–498, 1995). In this work, we investigated whether the
putA
gene could be used as a means of increasing the competitiveness of
S. meliloti
strains. We produced a construct in which a constitutive promoter was placed 190 nucleotides upstream from the start codon of the
putA
gene. This resulted in an increase in the basal expression of this gene, with this increase being even greater in the presence of the substrate proline. We found that the presence of multicopy plasmids containing this
putA
gene construct increased the competitiveness of
S. meliloti
in microcosm experiments in nonsterile soil planted with alfalfa plants subjected to drought stress only during the first month. We investigated whether this construct also increased the competitiveness of
S. meliloti
strains under agricultural conditions by using it as the inoculum in a contained field experiment at León, Spain. We found that the frequency of nodule occupancy was higher with inoculum containing the modified
putA
gene for samples that were analyzed after 34 days but not for samples that were analyzed later.
nifH
Sequences and Nitrogen Fixation in Type I and Type II Methanotrophs
Authors: Auman Ann J. , Speake Catherine C. , Lidstrom Mary E. 
Published: July 2002

ABSTRACT

Some methane-oxidizing bacteria (methanotrophs) are known to be capable of expressing nitrogenase and utilizing N
2
as a nitrogen source. However, no sequences are available for
nif
genes in these strains, and the known nitrogen-fixing methanotrophs are confined mainly to a few genera. The purpose of this work was to assess the nitrogen-fixing capabilities of a variety of methanotroph strains.
nifH
gene fragments from four type I methanotrophs and seven type II methanotrophs were PCR amplified and sequenced. Nitrogenase activity was confirmed in selected type I and type II strains by acetylene reduction. Activities ranged from 0.4 to 3.3 nmol/min/mg of protein. Sequence analysis shows that the
nifH
sequences from the type I and type II strains cluster with
nifH
sequences from other gamma proteobacteria and alpha proteobacteria, respectively. The translated
nifH
sequences from three
Methylomonas
strains show high identity (95 to 99%) to several published translated environmental
nifH
sequences PCR amplified from rice roots and a freshwater lake. The translated
nifH
sequences from the type II strains show high identity (94 to 99%) to published translated
nifH
sequences from a variety of environments, including rice roots, a freshwater lake, an oligotrophic ocean, and forest soil. These results provide evidence for nitrogen fixation in a broad range of methanotrophs and suggest that nitrogen-fixing methanotrophs may be widespread and important in the nitrogen cycling of many environments.
Survival of Clinical and Poultry-Derived Isolates of
Campylobacter jejuni
at a Low Temperature (4°C)
Authors: Chan Kam Fai , Le Tran Huyen , Kanenaka R. Y. , Kathariou S. 
Published: July 2002

ABSTRACT

Campylobacter jejuni
is a leading cause of bacterial gastroenteritis in humans, and contamination of poultry has been implicated in illness. The bacteria are fastidious in terms of their temperature requirements, being unable to grow below ca. 31°C, but have been found to be physiologically active at lower temperatures and to tolerate exposure to low temperatures in a strain-dependent manner. In this study, 19 field isolates of
C. jejuni
(10 of clinical and 9 of poultry origin) were studied for their ability to tolerate prolonged exposure to low temperature (4°C). Although substantial variability was found among different strains, clinical isolates tended to be significantly more likely to remain viable following cold exposure than poultry-derived strains. In contrast, the relative degree of tolerance of the bacteria to freezing at −20°C and freeze-thawing was strain specific but independent of strain source (poultry versus clinical) and degree of cold (4°C) tolerance.
Production of Recombinant α-Galactosidases in
Thermus thermophilus
Authors: Fridjonsson Olafur , Mattes Ralf 
Published: July 2002

ABSTRACT

A
Thermus thermophilus
selector strain for production of thermostable and thermoactive α-galactosidase was constructed. For this purpose, the native α-galactosidase gene (
agaT
) of
T. thermophilus
TH125 was inactivated to prevent background activity. In our first attempt, insertional mutagenesis of
agaT
by using a cassette carrying a kanamycin resistance gene led to bacterial inability to utilize melibiose (α-galactoside) and galactose as sole carbohydrate sources due to a polar effect of the insertional inactivation. A Gal
+
phenotype was assumed to be essential for growth on melibiose. In a Gal
−
background, accumulation of galactose or its metabolite derivatives produced from melibiose hydrolysis could interfere with the growth of the host strain harboring recombinant α-galactosidase. Moreover, the AgaT
−
strain had to be Km
s
for establishment of the plasmids containing α-galactosidase genes and the kanamycin resistance marker. Therefore, a suitable selector strain (AgaT
−
Gal
+
Km
s
) was generated by applying integration mutagenesis in combination with phenotypic selection. To produce heterologous α-galactosidase in
T. thermophilus
, the isogenes
agaA
and
agaB
of
Bacillus stearothermophilus
KVE36 were cloned into an
Escherichia coli-Thermus
shuttle vector. The region containing the
E. coli
plasmid sequence (pUC-derived vector) was deleted before transformation of
T. thermophilus
with the recombinant plasmids. As a result, transformation efficiency and plasmid stability were improved. However, growth on minimal agar medium containing melibiose was achieved only following random selection of the clones carrying a plasmid-based mutation that had promoted a higher copy number and greater stability of the plasmid.
Phylogeny of Culturable Estuarine Bacteria Catabolizing Riverine Organic Matter in the Northern Baltic Sea
Authors: Kisand Veljo , Cuadros Rocio , Wikner Johan 
Published: July 2002

ABSTRACT

The objective of our study was to isolate and determine the phylogenetic affiliation of culturable estuarine bacteria capable of catabolizing riverine dissolved organic matter (RDOM) under laboratory conditions. Additions of RDOM consistently promoted the growth of estuarine bacteria in carbon-limited dilution cultures, with seasonal variation in growth rates and yields. At least 42 different taxa were culturable on solid agar media and, according to quantitative DNA-DNA hybridizations, constituted 32 to 89% of the total bacterial number in the enriched treatments. Five species in the
Cytophaga-Flexibacter-Bacteroides
group and one in the γ-proteobacteria phylogenetic group (
Marinomonas
sp.) were numerically dominant during the stationary phase of the RDOM-enriched dilution cultures but not in the control cultures. Four of the isolates in
Cytophaga-Flexibacter-Bacteroides
group were putatively affiliated with the genus
Flavobacterium
. All dominating isolates were determined to be new species based on comparison to the current databases. The same group of species dominated independently of the season investigated, suggesting a low diversity of bacteria catabolizing RDOM in the estuary. It also suggested a broad tolerance of the dominating species to seasonal variation in hydrography, chemistry, and competition with other species. Taken together, our results suggest that a limited group of bacteria, mainly in the
Flavobacterium
genus, played an important role in introducing new energy and carbon to the marine system in the northern Baltic Sea.
Production of New Unsaturated Lipids during Wood Decay by Ligninolytic Basidiomycetes
Authors: Gutiérrez Ana , del Río José C. , Martínez-Íñigo María J. , Martínez María J. , Martínez Ángel T. 
Published: July 2002

ABSTRACT

Lipids were analyzed by gas chromatography-mass spectrometry for a 7-week in vitro decay of eucalypt wood by four ligninolytic basidiomycetes. The sound wood contained up to 75 mg of lipophilic compounds per 100 g of wood. Hydrolysis of sterol esters, which represented 38% of total wood lipids, occurred during the fungal decay. The initial increase of linoleic and other free unsaturated fatty acids paralleled the decrease of sterol esters. Moreover, new lipid compounds were found at advanced stages of wood decay that were identified from their mass spectra as unsaturated dicarboxylic acids consisting of a long aliphatic chain attached to the C-3 position of itaconic acid. These dicarboxylic acids were especially abundant in the wood treated with
Ceriporiopsis subvermispora
(up to 24 mg per 100 g of wood) but also were produced by
Phlebia radiata
,
Pleurotus pulmonarius
, and
Bjerkandera adusta
. We hypothesize that three main alkylitaconic acids (tetradecylitaconic,
cis
-7-hexadecenylitaconic, and hexadecylitaconic acids) are synthesized by fungi in condensation reactions involving palmitic, oleic, and stearic acids. We suggest that both wood unsaturated fatty acids (present in free form or released from esters during natural decay) and unsaturated metabolites synthesized by fungi could serve as a source for peroxidizable lipids in lignin degradation by white rot basidiomycetes.
Molecular Biological Detection and Characterization of
Clostridium
Populations in Municipal Landfill Sites
Authors: Van Dyke M. I. , McCarthy A. J. 
Published: July 2002

ABSTRACT

Primer sets specific for 16S rRNA genes were designed for four phylogenetic groups of clostridia known to contain mesophilic cellulolytic species. Specific amplification of these groups from landfill leachate DNA extracts demonstrated the widespread occurrence of clostridia from the
Clostridium thermocellum
and
C. leptum
groups. In contrast, the
C. botulinum
group was never detected, and the
C. coccoides
-
C. lentocellum
group was only occasionally detected. Amplification products were analyzed by temporal thermal gel electrophoresis to generate profiles of the clostridial groups and to identify dominant bands. Sequence analysis of 17 landfill clones confirmed that the primers were specific for the clostridial subgroups and that the cloned sequences had a close relationship with known cellulose-degrading clostridia. The primers have therefore been authenticated for use in the rapid identification of clostridia in anaerobic environments.
Ochratoxin Production by the
Aspergillus ochraceus
Group and
Aspergillus alliaceus
Authors: Bayman Paul , Baker James L. , Doster Mark A. , Michailides Themis J. , Mahoney Noreen E. 
Published: July 2002

ABSTRACT

Ochratoxin A is a toxic and carcinogenic fungal secondary metabolite; its presence in foods is increasingly regulated. Various fungi are known to produce ochratoxins, but it is not known which species produce ochratoxins consistently and which species cause ochratoxin contamination of various crops. We isolated fungi in the
Aspergillus ochraceus
group (section
Circumdati
) and
Aspergillus alliaceus
from tree nut orchards, nuts, and figs in California. A total of 72 isolates were grown in potato dextrose broth and yeast extract-sucrose broth for 10 days at 30°C and tested for production of ochratoxin A in vitro by high-pressure liquid chromatography. Among isolates from California figs, tree nuts, and orchards,
A. ochraceus
and
Aspergillus melleus
were the most common species. No field isolates of
A. ochraceus
or
A. melleus
produced ochratoxin A above the level of detection (0.01 μg/ml). All
A. alliaceus
isolates produced ochratoxin A, up to 30 μg/ml. We examined 50,000 figs for fungal infections and measured ochratoxin content in figs with visible fungal colonies. Pooled figs infected with
A. alliaceus
contained ochratoxin A, figs infected with the
A. ochraceus
group had little or none, and figs infected with
Penicillium
had none. These results suggest that the little-known species
A. alliaceus
is an important ochratoxin-producing fungus in California and that it may be responsible for the ochratoxin contamination occasionally observed in figs.
Involvement of
Rhodocyclus
-Related Organisms in Phosphorus Removal in Full-Scale Wastewater Treatment Plants
Authors: Zilles Julie L. , Peccia Jordan , Kim Myeong-Woon , Hung Chun-Hsiung , Noguera Daniel R. 
Published: July 2002

ABSTRACT

The participation of organisms related to
Rhodocyclus
in full-scale enhanced biological phosphorus removal (EBPR) was investigated. By using fluorescent in situ hybridization techniques, the communities of
Rhodocyclus
-related organisms in two full-scale wastewater treatment plants were estimated to represent between 13 and 18% of the total bacterial population. However, the fractions of these communities that participated in polyphosphate accumulation depended on the type of treatment process evaluated. In a University of Cape Town EBPR process, the percentage of
Rhodocyclus
-related cells that contained polyphosphate was about 20% of the total bacterial population, but these cells represented as much as 73% of the polyphosphate-accumulating organisms (PAOs). In an aerated-anoxic EBPR process,
Rhodocyclus
-related PAOs were less numerous, accounting for 6% of the total bacterial population and 26% of the total PAO population. In addition, 16S ribosomal DNA sequences 99.9% similar to the sequences of
Rhodocyclus
-related organisms enriched in acetate-fed bench-scale EBPR reactors were recovered from both full-scale plants. These results confirmed the involvement of
Rhodocyclus
-related organisms in EBPR and demonstrated their importance in full-scale processes. In addition, the results revealed a significant correlation between the type of EBPR process and the PAO community.