bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Epidemiology of Antimicrobial-Resistant CommensalEscherichia coliStrains in a Cohort of Newborn Calves
Authors: Hoyle Deborah V. , Yates Catherine M. , Chase-Topping Margo E. , Turner Esther J. , Davies Sarah E. , Low J. Chris , Gunn George J. , Woolhouse Mark E. J. , Amyes Sebastian G. B. 
Published: November 2005

ABSTRACTPulsed-field gel electrophoresis (PFGE) was used to investigate the dissemination and diversity of ampicillin-resistant (Ampr) and nalidixic acid-resistant (Nalr) commensalEscherichia colistrains in a cohort of 48 newborn calves. Calves were sampled weekly from birth for up to 21 weeks and a single resistant isolate selected from positive samples for genotyping and further phenotypic characterization. The Amprpopulation showed the greatest diversity, with a total of 56 different genotype patterns identified, of which 5 predominated, while the Nalrpopulation appeared to be largely clonal, with over 97% of isolates belonging to just two different PFGE patterns. Distinct temporal trends were identified in the distribution of several Amprgenotypes across the cohort, with certain patterns predominating at different points in the study. Cumulative recognition of new Amprgenotypes within the cohort was biphasic, with a turning point coinciding with the housing of the cohort midway through the study, suggesting that colonizing strains were from an environmental source on the farm. Multiply resistant isolates dominated the collection, with >95% of isolates showing resistance to at least two additional antimicrobials. Carriage of resistance to streptomycin, sulfamethoxazole, and tetracycline was the most common combination, found across several different genotypes, suggesting the possible spread of a common resistance element across multiple strains. The proportion of Amprisolates carrying sulfamethoxazole resistance increased significantly over the study period (P< 0.05), coinciding with a decline in the most common genotype pattern. These data indicate that calves were colonized by a succession of multiply resistant strains, with a probable environmental source, that disseminated through the cohort over time.
Antimicrobial Resistance and Virulence Genes of
Escherichia coli
Isolates from Swine in Ontario
Authors: Boerlin Patrick , Travis Rebeccah , Gyles Carlton L. , Reid-Smith Richard , Heather Lim Nicol Janecko , Nicholson Vivian , McEwen Scott A. , Friendship Robert , Archambault Marie 
Published: November 2005

ABSTRACT

A total of 318
Escherichia coli
isolates obtained from diarrheic and healthy pigs in Ontario from 2001 to 2003 were examined for their susceptibility to 19 antimicrobial agents. They were tested by PCR for the presence of resistance genes for tetracycline, streptomycin, sulfonamides, and apramycin and of 12 common virulence genes of porcine
E. coli
. Antimicrobial resistance frequency among
E. coli
isolates from swine in Ontario was moderate in comparison with other countries and was higher in isolates from pigs with diarrhea than in isolates from healthy finisher pigs. Resistance profiles suggest that cephamycinases may be produced by ≥8% of enterotoxigenic
E. coli
(ETEC). Resistance to quinolones was detected only in enterotoxigenic
E. coli
(≤3%). The presence of
sul3
was demonstrated for the first time in Canada in porcine
E. coli
isolates. Associations were observed among
tetA
,
sul1
,
aadA
, and
aac(3)IV
and among
tetB
,
sul2
, and
strA
/
strB
, with a strong negative association between
tetA
and
tetB
. The
paa
and
sepA
genes were detected in 92% of porcine ETEC, and strong statistical associations due to colocation on a large plasmid were observed between
tetA
,
estA
,
paa
, and
sepA
. Due at least in part to gene linkages, the distribution of resistance genes was very different between ETEC isolates and other porcine
E. coli
isolates. This demonstrates that antimicrobial resistance epidemiology differs significantly between pathogenic and commensal
E. coli
isolates. These results may have important implications with regards to the spread and persistence of resistance and virulence genes in bacterial populations and to the prudent use of antimicrobial agents.
Rapid Identification and Enumeration of
Saccharomyces cerevisiae
Cells in Wine by Real-Time PCR
Authors: Martorell P. , Querol A. , Fernández-Espinar M. T. 
Published: November 2005

ABSTRACT

Despite the beneficial role of
Saccharomyces cerevisiae
in the food industry for food and beverage production, it is able to cause spoilage in wines. We have developed a real-time PCR method to directly detect and quantify this yeast species in wine samples to provide winemakers with a rapid and sensitive method to detect and prevent wine spoilage. Specific primers were designed for
S. cerevisiae
using the sequence information obtained from a cloned random amplified polymorphic DNA band that differentiated
S. cerevisiae
from its sibling species
Saccharomyces bayanus
,
Saccharomyces pastorianus
, and
Saccharomyces paradoxus
. The specificity of the primers was demonstrated for typical wine spoilage yeast species. The method was useful for estimating the level of
S. cerevisiae
directly in sweet wines and red wines without preenrichment when yeast is present in concentrations as low as 3.8 and 5 CFU per ml. This detection limit is in the same order as that obtained from glucose-peptone-yeast growth medium (GPY). Moreover, it was possible to quantify
S. cerevisiae
in artificially contaminated samples accurately. Limits for accurate quantification in wine were established, from 3.8 × 10
5
to 3.8 CFU/ml in sweet wine and from 5 × 10
6
to 50 CFU/ml in red wine.
Development and Application of Real-Time PCR Assays for Quantification of Genes Encoding Tetracycline Resistance
Authors: Yu Zhongtang , Michel Frederick C. , Hansen Glenn , Wittum Thomas , Morrison Mark 
Published: November 2005

ABSTRACT

We report here the development, validation, and use of three real-time PCR assays to quantify the abundance of the following three groups of tetracycline resistance genes:
tet
(A) and
tet
(C);
tet
(G); and
tet
genes encoding ribosomal protection proteins, including
tet
(M),
tet
(O),
tetB
(P),
tet
(Q),
tet
(S),
tet
(T), and
tet
(W). The assays were validated using known numbers of sample-derived
tet
gene templates added to microbiome DNA. These assays are both precise and accurate over at least 6 log
tet
gene copies. New
tet
gene variants were also identified from cloned
tet
amplicons as part of this study. The utility of these real-time PCR assays was demonstrated by quantifying the three
tet
gene groups present in bovine and swine manures, composts of swine manure, lagoons of hog house effluent, and samples from an Ekokan upflow biofilter system treating hog house effluent. The bovine manures were found to contain fewer copies of all three groups of
tet
genes than the swine manures. The composts of swine manures had substantially reduced
tet
gene abundance (up to 6 log), while lagoon storage or the upflow biofilter had little effect on
tet
gene abundance. These results suggest that the method of manure storage and treatment may have a substantial impact on the persistence and dissemination of
tet
genes in agricultural environments. These real-time PCR assays provide rapid, quantitative, cultivation-independent measurements of 10 major classes of
tet
genes, which should be useful for ecological studies of antibiotic resistance.
A Method Adapting Microarray Technology for Signature-Tagged Mutagenesis of
Desulfovibrio desulfuricans
G20 and
Shewanella oneidensis
MR-1 in Anaerobic Sediment Survival Experiments
Authors: Groh Jennifer L. , Luo Qingwei , Ballard Jimmy D. , Krumholz Lee R. 
Published: November 2005

ABSTRACT

Signature-tagged mutagenesis (STM) is a powerful technique that can be used to identify genes expressed by bacteria during exposure to conditions in their natural environments. To date, there have been no reports of studies in which this approach was used to study organisms of environmental, rather than pathogenic, significance. We used a mini-Tn
10
transposon-bearing plasmid, pBSL180, that efficiently and randomly mutagenized
Desulfovibrio desulfuricans
G20 in addition to
Shewanella oneidensis
MR-1. Using these organisms as model sediment-dwelling anaerobic bacteria, we developed a new screening system, modified from former STM procedures, to identify genes that are critical for sediment survival. The screening system uses microarray technology to visualize tags from input and output pools, allowing us to identify those lost during sediment incubations. While the majority of data on survival genes identified will be presented in future papers, we report here on chemotaxis-related genes identified by our STM method in both bacteria in order to validate our method. This system may be applicable to the study of numerous environmental bacteria, allowing us to identify functions and roles of survival genes in various habitats.
Metabolic Engineering of
Corynebacterium glutamicum
for
l
-Serine Production
Authors: Peters-Wendisch Petra , Stolz Michael , Etterich Helga , Kennerknecht Nicole , Sahm Hermann , Eggeling Lothar 
Published: November 2005

ABSTRACT

Although
l
-serine proceeds in just three steps from the glycolytic intermediate 3-phosphoglycerate, and as much as 8% of the carbon assimilated from glucose is directed via
l
-serine formation, previous attempts to obtain a strain producing
l
-serine from glucose have not been successful. We functionally identified the genes
serC
and
serB
from
Corynebacterium glutamicum
, coding for phosphoserine aminotransferase and phosphoserine phosphatase, respectively. The overexpression of these genes, together with the third biosynthetic
serA
gene,
serA
Δ
197
, encoding an
l
-serine-insensitive 3-phosphoglycerate dehydrogenase, yielded only traces of
l
-serine, as did the overexpression of these genes in a strain with the
l
-serine dehydratase gene
sdaA
deleted. However, reduced expression of the serine hydroxymethyltransferase gene
glyA
, in combination with the overexpression of
serA
Δ
197
,
serC
, and
serB
, resulted in a transient accumulation of up to 16 mM
l
-serine in the culture medium. When
sdaA
was also deleted, the resulting strain,
C. glutamicum
Δ
sdaA
::pK18mob
glyA
′(pEC-T18mob2
serA
Δ197
CB
), accumulated up to 86 mM
l
-serine with a maximal specific productivity of 1.2 mmol h
−1
g (dry weight)
−1
. This illustrates a high rate of
l
-serine formation and also utilization in the
C. glutamicum
wild type. Therefore, metabolic engineering of
l
-serine production from glucose can be achieved only by addressing the apparent key position of this amino acid in the central metabolism.
16S rRNA Gene-Based Identification of Midgut Bacteria from Field-Caught
Anopheles gambiae
Sensu Lato and
A. funestus
Mosquitoes Reveals New Species Related to Known Insect Symbionts
Authors: Lindh Jenny M. , Terenius Olle , Faye Ingrid 
Published: November 2005

ABSTRACT

Field-collected mosquitoes of the two main malaria vectors in Africa,
Anopheles gambiae
sensu lato and
Anopheles funestus
, were screened for their midgut bacterial contents. The midgut from each blood-fed mosquito was screened with two different detection pathways, one culture independent and one culture dependent. Bacterial species determination was achieved by sequence analysis of 16S rRNA genes. Altogether, 16 species from 14 genera were identified, 8 by each method. Interestingly, several of the bacteria identified are related to bacteria known to be symbionts in other insects. One isolate,
Nocardia corynebacterioides
, is a relative of the symbiont found in the vector for Chagas' disease that has been proven useful as a paratransgenic bacterium. Another isolate is a novel species within the γ-proteobacteria that could not be phylogenetically placed within any of the known orders in the class but is close to a group of insect symbionts. Bacteria representing three intracellular genera were identified, among them the first identifications of
Anaplasma
species from mosquitoes and a new mosquito-
Spiroplasma
association. The isolates will be further investigated for their suitability for a paratransgenic
Anopheles
mosquito.
Recognition of Individual Genes in Diverse Microorganisms by Cycling Primed In Situ Amplification
Authors: Kenzaka Takehiko , Tamaki Shigeru , Yamaguchi Nobuyasu , Tani Katsuji , Nasu Masao 
Published: November 2005

ABSTRACTCycling primed in situ amplification-fluorescent in situ hybridization (CPRINS-FISH) was developed to recognize individual genes in a single bacterial cell. In CPRINS, the amplicon was long single-stranded DNA and thus retained within the permeabilized microbial cells. FISH with a multiply labeled fluorescent probe set enabled significant reduction in nonspecific background while maintaining high fluorescence signals of target bacteria. The ampicillin resistance gene inEscherichia coli, chloramphenicol acetyltransferase gene in different gram-negative strains, and RNA polymerase sigma factor (rpoD) gene inAeromonasspp. could be detected under identical permeabilization conditions. After concentration of environmental freshwater samples onto polycarbonate filters and subsequent coating of filters in gelatin, no decrease in bacterial cell numbers was observed with extensive permeabilization. The detection rates of bacterioplankton in river and pond water samples by CPRINS-FISH with a universal 16S rRNA gene primer and probe set ranged from 65 to 76% of total cell counts (mean, 71%). The concentrations of cells detected by CPRINS-FISH targeting of therpoDgenes ofAeromonas sobriaandA. hydrophilain the water samples varied between 2.1 × 103and 9.0 × 103cells ml−1and between undetectable and 5.1 × 102cells ml−1, respectively. These results demonstrate that CPRINS-FISH provides a high sensitivity for microscopic detection of bacteria carrying a specific gene in natural aquatic samples.
Methylotrophic Metabolism Is Advantageous for
Methylobacterium extorquens
during Colonization of
Medicago truncatula
under Competitive Conditions
Authors: Sy Abdoulaye , Timmers Antonius C. J. , Knief Claudia , Vorholt Julia A. 
Published: November 2005

ABSTRACT

Facultative methylotrophic bacteria of the genus
Methylobacterium
are commonly found in association with plants. Inoculation experiments were performed to study the importance of methylotrophic metabolism for colonization of the model legume
Medicago truncatula
. Competition experiments with
Methylobacterium extorquens
wild-type strain AM1 and methylotrophy mutants revealed that the
ability to use methanol as a carbon and energy source provides a
selective advantage during colonization of
M. truncatula
.
Differences in the fitness of mutants defective in different stages of
methylotrophic metabolism were found; whereas approximately 25% of the
mutant incapable of oxidizing methanol to formaldehyde (deficient in
methanol dehydrogenase) was recovered, 10% or less of the mutants
incapable of oxidizing formaldehyde to CO
2
(defective in
biosynthesis of the cofactor tetrahydromethanopterin) was
recovered. Interestingly, impaired fitness of the mutant
strains compared with the wild type was found on leaves and roots.
Single-inoculation experiments showed, however, that mutants with
defects in methylotrophy were capable of plant colonization at the
wild-type level, indicating that methanol is not the only carbon source
that is accessible to
Methylobacterium
while it is associated
with plants. Fluorescence microscopy with a green fluorescent
protein-labeled derivative of
M. extorquens
AM1 revealed that
the majority of the bacterial cells on leaves were on the surface and
that the cells were most abundant on the lower, abaxial side. However,
bacterial cells were also found in the intercellular spaces inside the
leaves, especially in the epidermal cell layer and immediately
underneath this
layer.
Anaerobic Microbial Communities in Lake Pavin, a Unique Meromictic Lake in France
Authors: Lehours Anne-C. , Bardot Corinne , Thenot Aurelie , Debroas Didier , Fonty Gerard 
Published: November 2005

ABSTRACT

The
Bacteria
and
Archaea
from the meromictic Lake Pavin were analyzed in samples collected along a vertical profile in the anoxic monimolimnion and were compared to those in samples from the oxic mixolimnion. Nine targeted 16S rRNA oligonucleotide probes were used to assess the distribution of
Bacteria
and
Archaea
and to investigate the in situ occurrence of sulfate-reducing bacteria and methane-producing
Archaea
involved in the terminal steps of the anaerobic degradation of organic material. The diversity of the complex microbial communities was assessed from the 16S rRNA polymorphisms present in terminal restriction fragment (TRF) depth patterns. The densities of the microbial community increased in the anoxic layer, and
Archaea
detected with probe ARCH915 represented the largest microbial group in the water column, with a mean
Archaea/Eubacteria
ratio of 1.5. Terminal restriction fragment length polymorphism (TRFLP) analysis revealed an elevated archaeal and bacterial phylotype richness in anoxic bottom-water samples. The structure of the
Archaea
community remained rather homogeneous, while TRFLP patterns for the eubacterial community revealed a heterogeneous distribution of eubacterial TRFs.
Proof that
Burkholderia
Strains Form Effective Symbioses with Legumes: a Study of Novel
Mimosa
-Nodulating Strains from South America
Authors: Chen Wen-Ming , de Faria Sergio M. , Straliotto Rosângela , Pitard Rosa M. , Simões-Araùjo Jean L. , Chou Jui-Hsing , Chou Yi-Ju , Barrios Edmundo , Prescott Alan R. , Elliott Geoffrey N. , Sprent Janet I. , Young J. Peter W. , James Euan K. 
Published: November 2005

ABSTRACT

Twenty
Mimosa
-nodulating bacterial strains from Brazil and Venezuela, together with eight reference
Mimosa
-nodulating rhizobial strains and two other β-rhizobial strains, were examined by amplified rRNA gene restriction analysis. They fell into 16 patterns and formed a single cluster together with the known β-rhizobia,
Burkholderia caribensis
,
Burkholderia phymatum
, and
Burkholderia tuberum
. The 16S rRNA gene sequences of 15 of the 20 strains were determined, and all were shown to belong to the genus
Burkholderia
; four distinct clusters could be discerned, with strains isolated from the same host species usually clustering very closely. Five of the strains (MAP3-5, Br3407, Br3454, Br3461, and Br3469) were selected for further studies of the symbiosis-related genes
nodA
, the NodD-dependent regulatory consensus sequences (
nod
box), and
nifH
. The
nodA
and
nifH
sequences were very close to each other and to those of
B. phymatum
STM815,
B. caribensis
TJ182, and
Cupriavidus taiwanensis
LMG19424 but were relatively distant from those of
B. tuberum
STM678. In addition to nodulating their original hosts, all five strains could also nodulate other
Mimosa
spp., and all produced nodules on
Mimosa pudica
that had nitrogenase (acetylene reduction) activities and structures typical of effective N
2
-fixing symbioses. Finally, both wild-type and green fluorescent protein-expressing transconjugant strains of Br3461 and MAP3-5 produced N
2
-fixing nodules on their original hosts,
Mimosa bimucronata
(Br3461) and
Mimosa pigra
(MAP3-5), and hence this confirms strongly that
Burkholderia
strains can form effective symbioses with legumes.
Construction of an Alpha Toxin Gene Knockout Mutant of
Clostridium perfringens
Type A by Use of a Mobile Group II Intron
Authors: Chen Yue , McClane Bruce A. , Fisher Derek J. , Rood Julian I. , Gupta Phalguni 
Published: November 2005

ABSTRACT

In developing
Clostridium perfringens
as a safe vaccine vector, the alpha toxin gene (
plc
) in the bacterial chromosome must be permanently inactivated. Disrupting genes in
C. perfringens
by traditional mutagenesis methods is very difficult. Therefore, we developed a new strategy using group II intron-based Target-Tron technology to inactivate the
plc
gene in
C. perfringens
ATCC 3624. Western blot analysis showed no production of alpha toxin protein in the culture supernatant of the
plc
mutant. Advantages of this technology, such as site specificity, relatively high frequency of insertion, and introduction of no antibiotic resistance genes into the chromosome, could facilitate construction of other
C. perfringens
mutants.
Factors Influencing the Ability of
Listeria monocytogenes
To Pass through a Membrane Filter by Active Infiltration
Authors: Nakazawa Kentaro , Hasegawa Hiroyuki , Nakagawa Yoji , Terao Michinori , Matsuyama Tohey 
Published: November 2005

ABSTRACT

Listeria monocytogenes
infiltrated the reticulate structure of a membrane filter and passed through a filter with pore sizes of 0.45 μm and 0.2 μm in 6 to 24 h and 5 to 6 days, respectively. Flagellar motility and expansive pressure generated by the growing bacterial population were indicated as the driving forces of infiltration.
Impact of Bacterial NO
3
−
Transport on Sediment Biogeochemistry
Authors: Sayama Mikio , Risgaard-Petersen Nils , Nielsen Lars Peter , Fossing Henrik , Christensen Peter Bondo 
Published: November 2005

ABSTRACT

Experiments demonstrated that
Beggiatoa
could induce a H
2
S-depleted suboxic zone of more than 10 mm in marine sediments and cause a divergence in sediment NO
3
−
reduction from denitrification to dissimilatory NO
3
−
reduction to ammonium. pH, O
2
, and H
2
S profiles indicated that the bacteria oxidized H
2
S with NO
3
−
and transported S
0
to the sediment surface for aerobic oxidation.
Bactericidal Actions of a Silver Ion Solution on
Escherichia coli
, Studied by Energy-Filtering Transmission Electron Microscopy and Proteomic Analysis
Authors: Yamanaka Mikihiro , Hara Keita , Kudo Jun 
Published: November 2005

ABSTRACT

Bactericidal actions of the silver ion on
Escherichia coli
as a model microorganism were studied using energy-filtering transmission electron microscopy (EFTEM), two-dimensional electrophoresis (2-DE), and matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS). EFTEM observations demonstrated that the silver ion readily infiltrates the interior of
E. coli
, contrary to the early hypothesis that it resides initially in the cell membrane area. Furthermore, 2-DE and MALDI-TOF MS indicated that the expression of a ribosomal subunit protein as well as that of some other enzymes and proteins is affected by the silver ion. The present results demonstrate for the first time that one of the major bactericidal functions of the silver ion is its interaction with the ribosome and the ensuing inhibition in expression of the enzymes and proteins essential to ATP production.
Discrimination between
Francisella tularensis
and
Francisella
-Like Endosymbionts when Screening Ticks by PCR
Authors: Kugeler Kiersten J. , Gurfield Nikos , Creek Jean G. , Mahoney Kerry S. , Versage Jessica L. , Petersen Jeannine M. 
Published: November 2005

ABSTRACT

The presence of
Francisella
-like endosymbionts in tick species known to transmit tularemia poses a potential diagnostic problem for laboratories that screen tick samples by PCR for
Francisella tularensis.
Tick samples initially considered positive for
F. tularensis
based on standard 16S rRNA gene PCR were found to be positive only for
Francisella-
like endosymbionts using a multitarget
F. tularensis
TaqMan assay (IS
Ftu2
,
tul4
, and
iglC
) and 16S rRNA gene sequencing. Specificity of PCR-based diagnostics for
F. tularensis
should be carefully evaluated with appropriate specimen types prior to diagnostic use.
Creating New Genes by Plasmid Recombination in
Escherichia coli
and
Bacillus subtilis
Authors: Gomez Ana , Galic Tatjana , Mariet Jean-François , Matic Ivan , Radman Miroslav , Petit Marie-Agnès 
Published: November 2005

ABSTRACT

Gene shuffling is a way of creating proteins with interesting new characteristics, starting from diverged sequences. We tested an alternative to gene shuffling based on plasmid recombination and found that
Bacillus subtilis
efficiently recombines sequences with 4% divergence, and
Escherichia coli mutS
is more appropriate for sequences with 22% divergence.