bluuebunny/crossref_metadata_2025_split huggingface dataset:
Concentrations, Viability, and Distribution of
Cryptosporidium
Genotypes in Lagoons of Swine Facilities in the Southern Piedmont and in Coastal Plain Watersheds of Georgia
Authors: Jenkins Michael B. , Liotta Janice L. , Lucio-Forster Araceli , Bowman Dwight D. 
Published: July 2010

ABSTRACT

Waste lagoons of swine operations are a source of
Cryptosporidium
oocysts. Few studies, however, have reported on oocyst concentrations in swine waste lagoons; none have reported on oocyst viability status, nor has there been a systematic assessment of species/genotype distributions across different types of swine facilities. Ten swine waste lagoons associated with farrowing, nursery, finishing, and gestation operations were each sampled once a month for a year. Oocysts were extracted from triplicate 900-ml effluent samples, enumerated by microscopy, and assessed for viability by dye exclusion/vital stain assay. DNA was extracted from processed samples, and 18S ribosomal DNA (rDNA) genes were amplified by PCR and sequenced for species and genotype identification. Oocysts were observed at each sampling time at each lagoon. Annual mean concentrations of total oocysts and viable oocysts ranged between 24 and 51 and between 0.6 and 12 oocysts ml
−1
effluent, respectively. The species and genotype distributions were dominated (95 to 100%) by
Cryptosporidium suis
and
Cryptosporidium
pig genotype II, the latter of which was found at eight of the lagoons. The lagoon at the gestation facility was dominated by
Cryptosporidium muris
(90%), and one farrowing facility showed a mix of pig genotypes,
Cryptosporidium muris
, and various genotypes of
C. parvum
. The zoonotic
C. parvum
bovine genotype was observed five times out of 407 18S rDNA sequences analyzed. Our results indicate that pigs can have mixed
Cryptosporidium
infections, but infection with
C. suis
is likely to be dominant.
Characterization of Novel
Brucella
Strains Originating from Wild Native Rodent Species in North Queensland, Australia
Authors: Tiller Rebekah V. , Gee Jay E. , Frace Michael A. , Taylor Trevor K. , Setubal Joao C. , Hoffmaster Alex R. , De Barun K. 
Published: July 2010

ABSTRACT

We report on the characterization of a group of seven novel
Brucella
strains isolated in 1964 from three native rodent species in North Queensland, Australia, during a survey of wild animals. The strains were initially reported to be
Brucella suis
biovar 3 on the basis of microbiological test results. Our results indicated that the rodent strains had microbiological traits distinct from those of
B. suis
biovar 3 and all other
Brucella
spp. To reinvestigate these rodent strains, we sequenced the 16S rRNA,
recA
, and
rpoB
genes and nine housekeeping genes and also performed multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA). The rodent strains have a unique 16S rRNA gene sequence compared to the sequences of the classical
Brucella
spp. Sequence analysis of the
recA
,
rpoB
, and nine housekeeping genes reveals that the rodent strains are genetically identical to each other at these loci and divergent from any of the currently described
Brucella
sequence types. However, all seven of the rodent strains do exhibit distinctive allelic MLVA profiles, although none demonstrated an amplicon for VNTR 07, whereas the other
Brucella
spp. did. Phylogenetic analysis of the MLVA data reveals that the rodent strains form a distinct clade separate from the classical
Brucella
spp. Furthermore, whole-genome sequence comparison using the maximal unique exact matches index (MUMi) demonstrated a high degree of relatedness of one of the seven rodent
Brucella
strains (strain NF 2653) to another Australian rodent
Brucella
strain (strain 83-13). Our findings strongly suggest that this group of
Brucella
strains isolated from wild Australian rodents defines a new species in the
Brucella
genus.
Presence of
Bacteroidales
as a Predictor of Pathogens in Surface Waters of the Central California Coast
Authors: Schriewer Alexander , Miller Woutrina A. , Byrne Barbara A. , Miller Melissa A. , Oates Stori , Conrad Patricia A. , Hardin Dane , Yang Hsuan-Hui , Chouicha Nadira , Melli Ann , Jessup Dave , Dominik Clare , Wuertz Stefan 
Published: July 2010

ABSTRACT

The value of
Bacteroidales
genetic markers and fecal indicator bacteria (FIB) to predict the occurrence of waterborne pathogens was evaluated in ambient waters along the central California coast.
Bacteroidales
host-specific quantitative PCR (qPCR) was used to quantify fecal bacteria in water and provide insights into contributing host fecal sources. Over 140 surface water samples from 10 major rivers and estuaries within the Monterey Bay region were tested over 14 months with four
Bacteroidales
-specific assays (universal, human, dog, and cow), three FIB (total coliforms, fecal coliforms, and enterococci), two protozoal pathogens (
Cryptosporidium
and
Giardia
spp.), and four bacterial pathogens (
Campylobacter
spp.,
Escherichia coli
O157:H7,
Salmonella
spp., and
Vibrio
spp.). Indicator and pathogen distribution was widespread, and detection was not highly seasonal.
Vibrio cholerae
was detected most frequently, followed by
Giardia
,
Cryptosporidium
,
Salmonella
, and
Campylobacter
spp. Bayesian conditional probability analysis was used to characterize the
Bacteroidales
performance assays, and the ratios of concentrations determined using host-specific and universal assays were used to show that fecal contamination from human sources was more common than livestock or dog sources in coastal study sites. Correlations were seen between some, but not all, indicator-pathogen combinations. The ability to predict pathogen occurrence in relation to indicator threshold cutoff levels was evaluated using a weighted measure that showed the universal
Bacteroidales
genetic marker to have a comparable or higher mean predictive potential than standard FIB. This predictive ability, in addition to the
Bacteroidales
assays providing information on contributing host fecal sources, supports using
Bacteroidales
assays in water quality monitoring programs.
Eugenol Reduces the Expression of Virulence-Related Exoproteins in
Staphylococcus aureus
Authors: Qiu Jiazhang , Feng Haihua , Lu Jing , Xiang Hua , Wang Dacheng , Dong Jing , Wang Jianfeng , Wang Xiaoliang , Liu Juxiong , Deng Xuming 
Published: July 2010

ABSTRACT

Eugenol, an essential oil component in plants, has been demonstrated to possess activity against both Gram-positive and Gram-negative bacteria. This study examined the influence that subinhibitory concentrations of eugenol may have on the expression of the major exotoxins produced by
Staphylococcus aureus
. The results from a tumor necrosis factor (TNF) release assay and a hemolysin assay indicated that
S. aureus
cultured with graded subinhibitory concentrations of eugenol (16 to 128 μg/ml) dose dependently decreased the TNF-inducing and hemolytic activities of culture supernatants. Western blot analysis showed that eugenol significantly reduced the production of staphylococcal enterotoxin A (SEA), SEB, and toxic shock syndrome toxin 1 (the key exotoxins to induce TNF release), as well as the expression of α-hemolysin (the major hemolysin to cause hemolysis). In addition, this suppression was also evaluated at the transcriptional level via real-time reverse transcription (RT)-PCR analysis. The transcriptional analysis indicated that 128 μg/ml of eugenol remarkably repressed the transcription of the
S. aureus sea
,
seb
,
tst
, and
hla
genes. According to these results, eugenol has the potential to be rationally applied on food products as a novel food antimicrobial agent both to inhibit the growth of bacteria and to suppress the production of exotoxins by
S. aureus
.
High Diversity of Extended-Spectrum Beta-Lactamase-Producing Bacteria in an Urban River Sediment Habitat
Authors: Lu Su-Ying , Zhang Ya-Li , Geng Sui-Na , Li Tian-Yu , Ye Zhuo-Ming , Zhang Dong-Sheng , Zou Fei , Zhou Hong-Wei 
Published: July 2010

ABSTRACTAntibiotic-resistant bacteria (ARB) have been surveyed widely in water bodies, but few studies have determined the diversity of ARB in sediment, which is the most taxon-abundant habitat in aquatic environments. We isolated 56 extended-spectrum β-lactamase (ESBL)-producing bacteria from a single sediment sample taken from an urban river in China. All strains were confirmed for ESBL-producing capability by both the clavulanic acid combination disc method and MIC determination. Of the isolated strains, 39 were classified asEnterobacteriaceae(consisting of the generaEscherichia,Klebsiella,Serratia, andAeromonas) by 16S rRNA gene sequencing and biochemical analysis. The present study identifies, for the first time, ESBL-producing strains from the familiesBrucellaceaeandMoraxellaceae. TheblaCTX-Mgene was the most dominant of the ESBL genes (45 strains), while theblaTEMgene was the second-most dominant (22 strains). A total of five types ofblaCTX-Mfragments were identified, with both known and novel sequences. A library ofblaCTX-Mcloned from the sediment DNA showed an even higher diversity ofblaCTX-Msequences. The discovery of highly diverse ESBL-producing bacteria and ESBL genes, particularlyblaCTX, in urban river sediment raises alarms for potential dissemination of ARB in communities through river environments.
Estimation of Uptake of Humic Substances from Different Sources by
Escherichia coli
Cells under Optimum and Salt Stress Conditions by Use of Tritium-Labeled Humic Materials
Authors: Kulikova Natalia A. , Perminova Irina V. , Badun Gennady A. , Chernysheva Maria G. , Koroleva Olga V. , Tsvetkova Eugenia A. 
Published: July 2010

ABSTRACT

The primary goal of this paper is to demonstrate potential strengths of the use of tritium-labeled humic substances (HS) to quantify their interaction with living cells under various conditions. A novel approach was taken to study the interaction between a model microorganism and the labeled humic material. The bacterium
Escherichia coli
was used as a model microorganism. Salt stress was used to study interactions of HS with living cells under nonoptimum conditions. Six tritium-labeled samples of HS originating from coal, peat, and soil were examined. To quantify their interaction with
E. coli
cells, bioconcentration factors (BCF) were calculated and the amount of HS that penetrated into the cell interior was determined, and the liquid scintillation counting technique was used as well. The BCF values under optimum conditions varied from 0.9 to 13.1 liters kg
−1
of cell biomass, whereas under salt stress conditions the range of corresponding values increased substantially and accounted for 0.2 to 130 liters kg
−1
. The measured amounts of HS that penetrated into the cells were 23 to 167 mg and 25 to 465 mg HS per kg of cell biomass under optimum and salt stress conditions, respectively. This finding indicated increased penetration of HS into
E. coli
cells under salt stress.
Simple Genetic Selection Protocol for Isolation of Overexpressed Genes That Enhance Accumulation of Membrane-Integrated Human G Protein-Coupled Receptors inEscherichia coli
Authors: Skretas Georgios , Georgiou George 
Published: July 2010

ABSTRACTThe efficient production of membrane proteins in bacteria remains a major challenge. In this work, we sought to identify overexpressed genes that enhance the yields of recombinant membrane proteins inEscherichia coli. We developed a genetic selection system for bacterial membrane protein production, consisting of membrane protein fusions with the enzyme β-lactamase and facile selection of high-production strains on ampicillin-containing media. This system was used to screen the ASKA library, an ordered library of plasmids encoding all the knownE. coliopen reading frames (ORFs), and several clones with the ability to accumulate enhanced amounts of recombinant membrane proteins were selected. Notably, coexpression ofybaB, a gene encoding a putative DNA-binding protein of unknown function, was found to enhance the accumulation of a variety of membrane-integrated human G protein-coupled receptors and other integral membrane proteins inE. coliby up to 10-fold. The results of this study highlight the power of genetic approaches for identifying factors that impact membrane protein biogenesis and for generating engineered microbial hosts for membrane protein production.
Survival of
Campylobacter jejuni
under Conditions of Atmospheric Oxygen Tension with the Support of
Pseudomonas
spp
Authors: Hilbert Friederike , Scherwitzel Manuela , Paulsen Peter , Szostak Michael P. 
Published: July 2010

ABSTRACT

Campylobacter jejuni
is a major food-borne pathogen. Despite causing enteritis in humans, it is a well-adapted intestinal microorganism in animals, hardly ever generating disease symptoms. Nevertheless, as a true microaerophilic microorganism it is still puzzling how
Campylobacter
cells can survive on chicken meat, the main source of human infection. In this study, we demonstrate that
C. jejuni
is able to withstand conditions of atmospheric oxygen tension when cocultured with
Pseudomonas
species, major food-spoiling bacteria that are frequently found on chicken meat in rather high numbers. Using an
in vitro
survival assay, interactions of 145
C. jejuni
wild-type strains and field isolates from chicken meat, broiler feces, and human clinical samples with type strains and food isolates of
Pseudomonas
spp.,
Proteus mirabilis
,
Citrobacter freundii
,
Micrococcus luteus
, and
Enterococcus faecalis
were studied. When inoculated alone or in coculture with
Proteus mirabilis
,
Citrobacter freundii
,
Micrococcus luteus
, or
Enterococcus faecalis
type strains,
Campylobacter
cells were able to survive ambient oxygen levels for no more than 18 h. In contrast,
Campylobacter
bacteria inoculated with type strains or wild-type isolates of
Pseudomonas
showed a prolonged aerobic survival of up to >48 h. This microbial commensalism was diverse in
C. jejuni
isolates from different sources; isolates from chicken meat and humans in coculture with
P
seudomonas
putida
were able to use this survival support better than fecal isolates from broilers. Scanning electron microscopy revealed the development of fiberlike structures braiding
P. putida
and
C. jejuni
cells. Hence, it seems that microaerophilic
C. jejuni
is able to survive ambient atmospheric oxygen tension by metabolic commensalism with
Pseudomonas
spp. This bacterium-bacterium interaction might set the basis for survival of
C. jejuni
on chicken meat and thus be the prerequisite step in the pathway toward human infection.
Differential Filamentation of Candida albicans and Candida dubliniensis Is Governed by Nutrient Regulation of
UME6
Expression
Authors: O'Connor Leanne , Caplice Nicole , Coleman David C. , Sullivan Derek J. , Moran Gary P. 
Published: July 2010

ABSTRACT

Candida dubliniensis
is closely related to
Candida albicans
; however, it is responsible for fewer infections in humans and is less virulent in animal models of infection.
C. dubliniensis
forms fewer hyphae
in vivo
, and this may contribute to its reduced virulence. In this study we show that, unlike
C. albicans
,
C. dubliniensis
fails to form hyphae in yeast extract-peptone-dextrose (YPD) medium supplemented with 10% (vol/vol) fetal calf serum (YPDS medium). However,
C. dubliniensis
filaments in water plus 10% (vol/vol) fetal calf serum (WS), and this filamentation is inhibited by the addition of peptone and glucose. Repression of filamentation in YPDS medium could be partly overcome by preculture in synthetic Lee's medium. Unlike
C. albicans
, inoculation of
C. dubliniensis
in YPDS medium did not result in increased
UME6
transcription. However, >100-fold induction of
UME6
was observed when
C. dubliniensis
was inoculated in nutrient-poor WS medium. The addition of increasing concentrations of peptone to WS medium had a dose-dependent effect on reducing
UME6
expression. Transcript profiling of
C. dubliniensis
hyphae in WS medium identified a starvation response involving expression of genes in the glyoxylate cycle and fatty acid oxidation. In addition, a core, shared transcriptional response with
C. albicans
could be identified, including expression of virulence-associated genes including
SAP456
,
SAP7
,
HWP1
, and
SOD5
. Preculture in nutrient-limiting medium enhanced adherence of
C. dubliniensis
, epithelial invasion, and survival following coculture with murine macrophages. In conclusion,
C. albicans
, unlike
C. dubliniensis
, appears to form hyphae in liquid medium regardless of nutrient availability, which may account for its increased capacity to cause disease in humans.
Roles of Minor Pilin Subunits Spy0125 and Spy0130 in the Serotype M1
Streptococcus pyogenes
Strain SF370
Authors: Smith Wendy D. , Pointon Jonathan A. , Abbot Emily , Kang Hae Joo , Baker Edward N. , Hirst Barry H. , Wilson Janet A. , Banfield Mark J. , Kehoe Michael A. 
Published: July 2010

ABSTRACT

Adhesive pili on the surface of the serotype M1
Streptococcus pyogenes
strain SF370 are composed of a major backbone subunit (Spy0128) and two minor subunits (Spy0125 and Spy0130), joined covalently by a pilin polymerase (Spy0129). Previous studies using recombinant proteins showed that both minor subunits bind to human pharyngeal (Detroit) cells (A. G. Manetti et al., Mol. Microbiol. 64:968-983, 2007), suggesting both may act as pilus-presented adhesins. While confirming these binding properties, studies described here indicate that Spy0125 is the pilus-presented adhesin and that Spy0130 has a distinct role as a wall linker. Pili were localized predominantly to cell wall fractions of the wild-type
S. pyogenes
parent strain and a
spy0125
deletion mutant. In contrast, they were found almost exclusively in culture supernatants in both
spy0130
and
srtA
deletion mutants, indicating that the housekeeping sortase (SrtA) attaches pili to the cell wall by using Spy0130 as a linker protein. Adhesion assays with antisera specific for individual subunits showed that only anti-rSpy0125 serum inhibited adhesion of wild-type
S. pyogenes
to human keratinocytes and tonsil epithelium to a significant extent. Spy0125 was localized to the tip of pili, based on a combination of mutant analysis and liquid chromatography-tandem mass spectrometry analysis of purified pili. Assays comparing parent and mutant strains confirmed its role as the adhesin. Unexpectedly, apparent spontaneous cleavage of a labile, proline-rich (8 of 14 residues) sequence separating the N-terminal ∼1/3 and C-terminal ∼2/3 of Spy0125 leads to loss of the N-terminal region, but analysis of internal
spy0125
deletion mutants confirmed that this has no significant effect on adhesion.
A Putrescine-Inducible Pathway Comprising PuuE-YneI in Which γ-Aminobutyrate Is Degraded into Succinate in
Escherichia coli
K-12
Authors: Kurihara Shin , Kato Kenji , Asada Kei , Kumagai Hidehiko , Suzuki Hideyuki 
Published: July 2010

ABSTRACT

γ-Aminobutyrate (GABA) is metabolized to succinic semialdehyde by GABA aminotransferase (GABA-AT), and the succinic semialdehyde is subsequently oxidized to succinate by succinic semialdehyde dehydrogenase (SSADH). In
E
scherichia
coli
, there are duplicate GABA-ATs (GabT and PuuE) and duplicate SSADHs (GabD and YneI). While GabT and GabD have been well studied previously, the characterization and expression analysis of PuuE and YneI are yet to be investigated. By analyzing the amino acid profiles in cells of Δ
puuE
and/or Δ
gabT
mutants, this study demonstrated that PuuE plays an important role in GABA metabolism in
E. coli
cells. The similarity of the amino acid sequences of PuuE and GabT is 67.4%, and it was biochemically demonstrated that the catalytic center of GabT is conserved as an amino acid residue important for the enzymatic activity in PuuE as Lys-247. However, the regulation of expression of PuuE is significantly different from that of GabT. PuuE is induced by the addition of putrescine to the medium and is repressed by succinate and low aeration conditions; in contrast, GabT is almost constitutive. Similarly, YneI is induced by putrescine, while GabD is not. For
E. coli
, PuuE is important for utilization of putrescine as a sole nitrogen source and both PuuE and YneI are important for utilization of putrescine as a sole carbon source. The results demonstrate that the PuuE-YneI pathway was a putrescine-inducible GABA degradation pathway for utilizing putrescine as a nutrient source.
Genetic and Functional Analyses of the
mob
Operon on Conjugative Transposon CTn
341
from
Bacteroides
spp
Authors: Peed Lindsay , Parker Anita C. , Smith C. Jeffrey 
Published: July 2010

ABSTRACT

Bacteroides
are Gram-negative anaerobes indigenous to the intestinal tract of humans, and they are important opportunistic pathogens. Mobile genetic elements, such as conjugative transposons (CTns), have contributed to an increase in antibiotic resistance in these organisms. CTns are self-transmissible elements that belong to the superfamily of
i
ntegrative and
c
onjugative
e
lements (ICEs). CTn
341
is 52 kb; it encodes tetracycline resistance and its transfer is induced by tetracycline. The mobilization region of CTn
341
was shown to be comprised of a three-gene operon,
mobABC
, and the transfer origin,
oriT
. The three genes code for a nicking accessory protein, a relaxase, and a VirD4-like coupling protein, respectively. The Mob proteins were predicted to mediate the formation of the relaxosome complex, nick DNA at the
oriT
, and shuttle the DNA/protein complex to the mating-pore apparatus. The results of mutational studies indicated that the three genes are required for maximal transfer of CTn
341
. Mob gene transcription was induced by tetracycline, and this regulation was mediated through the two-component regulatory system, RteAB. The
oriT
region of CTn
341
was located within 100 bp of
mobA
, and a putative
Bacteroides
consensus nicking site was observed within this region. Mutation of the putative nick site resulted in a loss of transfer. This study demonstrated a role of the mobilization region for transfer of
Bacteroides
CTns and that tetracycline induction occurs for the
mob
gene operon, as for the
tra
gene operon(s), as shown previously.
The Paralogous MarR/DUF24-Family Repressors YodB and CatR Control Expression of the Catechol Dioxygenase CatE inBacillus subtilis
Authors: Chi Bui Khanh , Kobayashi Kazuo , Albrecht Dirk , Hecker Michael , Antelmann Haike 
Published: July 2010

ABSTRACTThe redox-sensing MarR/DUF24-type repressor YodB controls expression of the azoreductase AzoR1 and the nitroreductase YodC that are involved in detoxification of quinones and diamide inBacillus subtilis. In the present paper, we identified YodB and its paralog YvaP (CatR) as repressors of theyfiDE(catDE) operon encoding a catechol-2,3-dioxygenase that also contributes to quinone resistance. Inactivation of both CatR and YodB is required for full derepression ofcatDEtranscription. DNA-binding assays and promoter mutagenesis studies showed that CatR protects two inverted repeats with the consensus sequence TTAC-N5-GTAA overlapping the −35 promoter region (BS1) and the transcriptional start site (TSS) (BS2). The BS1 operator was required for binding of YodBin vitro. CatR and YodB share the conserved N-terminal Cys residue, which is required for redox sensing of CatRin vivoas shown by Cys-to-Ser mutagenesis. Our data suggest that CatR is modified by intermolecular disulfide formation in response to diamide and quinonesin vitroandin vivo. Redox regulation of CatR occurs independently of YodB, and no protein interaction was detected between CatR and YodBin vivousing protein cross-linking and mass spectrometry.