bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genetic Diversity of
Campylobacter jejuni
Isolates from Farm Animals and the FarmEnvironment
Authors: Colles F. M. , Jones K. , Harding R. M. , Maiden M. C. J. 
Published: December 2003

ABSTRACT

The
genetic diversity of
Campylobacter jejuni
isolates from farm
animals and their environment was investigated by multilocus sequence
typing (MLST). A total of 30 genotypes, defined by allelic profiles
(assigned to sequence types [STs]), were found in 112
C.
jejuni
isolates originating in poultry, cattle, sheep, starlings,
and slurry. All but two of these genotypes belonged to one of nine
C. jejuni
clonal complexes previously identified in isolates
from human disease and retail food samples and one clonal complex
previously associated with an environmental source. There was some
evidence for the association of certain clonal complexes with
particular farm animals: isolates belonging to the ST-45 complex
predominated among poultry isolates but were absent among sheep
isolates, while isolates belonging to the ST-61 and ST-42 complexes
were predominant among sheep isolates but were absent from the poultry
isolates. In contrast, ST-21 complex isolates were distributed among
the different isolation sources. Comparison with MLST data from 91
human disease isolates showed small but significant genetic
differentiation between the farm and human isolates; however,
representatives of six clonal complexes were found in both samples.
These data demonstrate that MLST and the clonal complex model can be
used to identify and compare the genotypes of
C. jejuni
isolates from farm animals and the environment with those from retail
food and human
disease.
Predominance of
vanA
Genotype amongVancomycin-Resistant
Enterococcus
Isolates from Poultry andSwine in CostaRica
Authors: Bustamante Warner , Alpízar Angélica , Hernández Silvia , Pacheco Alexandra , Vargas Natalia , Herrera Marco Luis , Vargas Álvaro , Caballero Magaly , García Fernando 
Published: December 2003

ABSTRACT

The
use of avoparcin as a growth promoter is considered to have selected
for vancomycin-resistant enterococci (VRE). In Costa Rica, the use of
avoparcin for poultry and swine was intensive until the product was
withdrawn from the market in 2000. We evaluated the presence of VRE in
poultry, swine, and cattle fecal samples obtained during 1998 and 1999.
A total of 185 VRE isolates were recovered from 116 out of 893 samples.
Enterococcus faecium
was the most frequently isolated species
(50.8%), being predominant among poultry (71.6%) and
swine (37.7%) isolates, but it was not recovered from the bovine
samples. The second-most-frequently-isolated species from poultry and
swine, respectively, were
E. durans
(23.2%)
and
E. faecalis
(21.7%).
E. casseliflavus
was the only species obtained from bovine
samples, but it was not found among the avian isolates. An evident
predominance of the
vanA
determinant among
vancomycin-resistant enterococcal species from poultry and swine, but
not from cattle, was observed and was similar to the situation in
European countries before avoparcin was forbidden. The diversity of the
vanA
determinant in the isolates was assessed by detection of
the IS
1251
insertion in the
vanSH
intergenic region
and of the IS
1476
insertion in the
vanXY
intergenic
region. However, in none of the 154
vanA
+
isolates recovered in this study were those insertions
detected.
Spatial Analysis of Archaeal Community Structure inGrasslandSoil
Authors: Nicol Graeme W. , Glover L. Anne , Prosser James I. 
Published: December 2003

ABSTRACT

The
complex structure of soil and the heterogeneity of resources available
to microorganisms have implications for sampling regimens when the
structure and diversity of microbial communities are analyzed. To
assess the heterogeneity in community structure, archaeal communities,
which typically contain sequences belonging to the nonthermophilic
Crenarchaeota
, were examined at two contrasting spatial scales
by using PCR-denaturing gradient gel electrophoresis (DGGE) analysis
followed by unweighted pair group method with arithmetic mean analysis
of 16S rRNA- and ribosomal DNA-derived profiles. A
macroscale analysis was carried out with soil cores taken
at 2-m intervals along triplicate 8-m transects from both managed
(improved) and natural (unimproved) grassland rhizosphere soils. A
microscale analysis was carried out with a single soil core by
assessing the effects of both sample size (10, 1, and 0.1 g)
and distance between samples. The much reduced complexity of archaeal
profiles compared to the complexity typical of the bacterial community
facilitated visual comparison of profiles based on band presence and
revealed different levels of heterogeneity between sets of samples. At
the macroscale level, heterogeneity over the transect could not be
related to grassland type. Substantial heterogeneity was observed
across both improved and unimproved transects, except for one improved
transect that exhibited substantial homogeneity, so that profiles for a
single core were largely representative of the entire transect. At the
smaller scale, the heterogeneity of the archaeal community structure
varied with sample size within a single 8- by 8-cm core. The archaeal
DGGE profiles for replicate 10-g soil samples were similar, while those
for 1-g samples and 0.1-g samples showed greater heterogeneity. In
addition, there was no relationship between the archaeal profiles and
the distance between 1- or 0.1-g samples, although relationships
between community structure and distance of separation may occur at a
smaller scale. Our findings demonstrate the care required when workers
attempt to obtain a representative picture of microbial community
structure in the soil
environment.
Real-Time PCR Assay for Detection and Enumeration ofDekkera bruxellensisin Wine
Authors: Phister Trevor G. , Mills David A. 
Published: December 2003

ABSTRACTTraditional methods to detect the spoilage yeastDekkera bruxellensisfrom wine involve lengthy enrichments. To overcome this difficulty, we developed a quantitative real-time PCR method to directly detect and enumerateD. bruxellensisin wine. Specific PCR primers toD. bruxellensiswere designed to the 26S rRNA gene, and nontarget yeast and bacteria common to the winery environment were not amplified. The assay was linear over a range of cell concentrations (6 log units) and could detect as little as 1 cell per ml in wine. The addition of large amounts of nontarget yeasts did not impact the efficiency of the assay. This method will be helpful to identify possible routes ofD. bruxellensisinfection in winery environments. Moreover, the time involved in performing the assay (3 h) should enable winemakers to more quickly make wine processing decisions in order to reduce the threat of spoilage byD. bruxellensis.
Characterization of
Vibrio fluvialis
-LikeStrains Implicated in Limp LobsterDisease
Authors: Tall B. D. , Fall S. , Pereira M. R. , Ramos-Valle M. , Curtis S. K. , Kothary M. H. , Chu D. M. T. , Monday S. R. , Kornegay L. , Donkar T. , Prince D. , Thunberg R. L. , Shangraw K. A. , Hanes D. E. , Khambaty F. M. , Lampel K. A. , Bier J. W. , Bayer R. C. 
Published: December 2003

ABSTRACT

Studies
were undertaken to characterize and determine the pathogenic mechanisms
involved in a newly described systemic disease in
Homarus
americanus
(American lobster) caused by a
Vibrio
fluvialis
-like microorganism. Nineteen isolates were obtained from
eight of nine lobsters sampled. Biochemically, the isolates resembled
V. fluvialis
, and the isolates grew optimally at 20°C;
none could grow at temperatures above 23°C. The type strain
(1AMA) displayed a thermal reduction time (D value) of 5.77 min at
37°C. All of the isolates required at least 1% NaCl for
growth. Collectively, the data suggest that these isolates may embody a
new biotype. Pulsed-field gel electrophoresis (PFGE) analysis of the
isolates revealed five closely related subgroups. Some isolates
produced a sheep hemagglutinin that was neither an outer membrane
protein nor a metalloprotease. Several isolates possessed
capsules. The isolates were highly susceptible to a variety of
antibiotics tested. However, six isolates were resistant to
erythromycin. Seventeen isolates harbored plasmids. Lobster challenge
studies revealed that the 50% lethal dose of a plasmid-positive
strain was 100-fold lower than that of a plasmid-negative strain,
suggesting that the plasmid may enhance the pathogenicity of these
microorganisms in lobsters. Microorganisms that were recovered from
experimentally infected lobsters exhibited biochemical and PFGE
profiles that were indistinguishable from those of the challenge
strain. Tissue affinity studies demonstrated that the challenge
microorganisms accumulated in heart and midgut tissues as well as in
the hemolymph. Culture supernatants and polymyxin B lysates of the
strains caused elongation of CHO cells in tissue culture, suggesting
the presence of a hitherto unknown enterotoxin. Both plasmid-positive
and plasmid-negative strains caused significant dose-related intestinal
fluid accumulations in suckling mice. Absence of viable organisms in
the intestinal contents of mice suggests that these microorganisms
cause diarrhea in mice by intoxication rather than by an infectious
process. Further, these results support the thermal reduction data at
37°C and suggest that the mechanism(s) that led to fluid
accumulation in mice differs from the disease process observed in
lobsters by requiring neither the persistence of viable microorganisms
nor the presence of plasmids. In summary, results of lobster studies
satisfy Koch's postulates at the organismal and molecular levels;
the findings support the hypothesis that these
V.
fluvialis
-like organisms were responsible for the originally
described systemic disease, which is now called limp lobster
disease.
BiologicalTraits ofXylella fastidiosaStrains from Grapes andAlmonds
Authors: Almeida Rodrigo P. P. , Purcell Alexander H. 
Published: December 2003

ABSTRACTXylella fastidiosais a xylem-limited bacterium that causes various diseases, among them Pierce's disease of grapevine (PD) and almond leaf scorch (ALS). PD and ALS have long been considered to be caused by the same strain of this pathogen, but recent genetic studies have revealed differences amongX. fastidiosaisolated from these host plants. We tested the hypothesis that ALS is caused by PD and ALS strains in the field and found that both groups ofX. fastidiosacaused ALS and overwintered within almonds after mechanical inoculation. Under greenhouse conditions, all isolates caused ALS and all isolates from grapes caused PD. However, isolates belonging to almond genetic groupings did not cause PD in inoculated grapes but systemically infected grapes with lower frequency and populations than those belonging to grape strains. Isolates able to cause both PD and ALS developed 10-fold-higher concentrations ofX. fastidiosain grapes than in almonds. In the laboratory, isolates from grapes overwintered with higher efficiency in grapes than in almonds and isolates from almonds overwintered better in almonds than in grapes. We assigned strains from almonds into groups I and II on the basis of their genetic characteristics, growth on PD3 solid medium, and bacterial populations within inoculated grapevines. Our results show that genetically distinct strains from grapes and almonds differ in population behavior and pathogenicity in grapes and in the ability to grow on two different media.
Identification and Population Dynamics of Yeasts inSourdough Fermentation Processes by PCR-Denaturing GradientGelElectrophoresis
Authors: Meroth Christiane B. , Hammes Walter P. , Hertel Christian 
Published: December 2003

ABSTRACTFour sourdoughs (A to D) were produced under practical conditions, using a starter obtained from a mixture of three commercially available sourdough starters and baker's yeast. The doughs were continuously propagated until the composition of the microbiota remained stable. A fungi-specific PCR-denaturing gradient gel electrophoresis (DGGE) system was established to monitor the development of the yeast biota. The analysis of the starter mixture revealed the presence ofCandida humilis,Debaryomyces hansenii,Saccharomyces cerevisiae, andSaccharomyces uvarum. In sourdough A (traditional process with rye flour),C. humilisdominated under the prevailing fermentation conditions. In rye flour sourdoughs B and C, fermented at 30 and 40°C, respectively,S. cerevisiaebecame predominant in sourdough B, whereas in sourdough C the yeast counts decreased within a few propagation steps below the detection limit. In sourdough D, which corresponded to sourdough C in temperature but was produced with rye bran,Candida kruseibecame dominant. Isolates identified asC. humilisandS. cerevisiaewere shown by randomly amplified polymorphic DNA-PCR analysis to originate from the commercial starters and the baker's yeast, respectively. The yeast species isolated from the sourdoughs were also detected by PCR-DGGE. However, in the gel, additional bands were visible. Because sequencing of these PCR fragments from the gel failed, cloning experiments with 28S rRNA amplicons obtained from rye flour were performed, which revealedCladosporiumsp.,Saccharomyces servazii,S. uvarum, an unculturable ascomycete,Dekkera bruxellensis,Epicoccum nigrum, andS. cerevisiae.The last four species were also detected in sourdoughs A, B, and C.
Survival of
Helicobacter pylori
in a Natural Freshwater Environment
Authors: Adams B. L. , Bates T. C. , Oliver J. D. 
Published: December 2003

ABSTRACT

The
mode by which
Helicobacter pylori
, the causative agent of most
gastric ulcers, is transmitted remains undetermined. Epidemiological
evidence suggests these organisms are waterborne; however,
H.
pylori
has rarely been grown from potential water sources. This
may be due to the ability of this organism to rapidly enter the viable
but nonculturable (VBNC) state. Our investigation examines the entrance
of
H. pylori
into this state in laboratory cultures and a
natural freshwater environment as well as the relationship between
morphology and culturability. To this end, membrane diffusion chambers
were utilized to expose the cells to the natural fluctuations of a
freshwater stream. In both the laboratory and environment, samples were
assayed for culturability using plate counts and stained using a
LIVE/DEAD
Bac
Light assay for viability and morphological
determinations. Additionally, water samples were collected, six
environmental parameters were measured, and resuscitation conditions
were examined.
H. pylori
was observed to lose culturability in
the laboratory and stream, although viability was maintained. While the
results of our study agree with those of previous studies which
suggested that there is a transition in morphology from rods to cocci
as culturability is lost, the morphological distribution of cells did
not change as culturability was lost in the environment. The majority
of cells in the VBNC state in the laboratory are cocci; however, all
morphological forms were present in the environment. The results of
these studies suggest that
H. pylori
persists in laboratory
cultures and the environment in the VBNC state and that cells in this
state represent a public health
hazard.
Enumeration and Characterization of Iron(III)-ReducingMicrobial Communities from Acidic Subsurface Sediments ContaminatedwithUranium(VI)
Authors: Petrie Lainie , North Nadia N. , Dollhopf Sherry L. , Balkwill David L. , Kostka Joel E. 
Published: December 2003

ABSTRACTIron(III)-reducing bacteria have been demonstrated to rapidly catalyze the reduction and immobilization of uranium(VI) from contaminated subsurface sediments. Thus, these organisms may aid in the development of bioremediation strategies for uranium contamination, which is prevalent in acidic subsurface sediments at U.S. government facilities. Iron(III)-reducing enrichment cultures were initiated from pristine and contaminated (high in uranium, nitrate; low pH) subsurface sediments at pH 7 and pH 4 to 5. Enumeration of Fe(III)-reducing bacteria yielded cell counts of up to 240 cells ml−1for the contaminated and background sediments at both pHs with a range of different carbon sources (glycerol, acetate, lactate, and glucose). In enrichments where nitrate contamination was removed from the sediment by washing, MPN counts of Fe(III)-reducing bacteria increased substantially. Sediments of lower pH typically yielded lower counts of Fe(III)-reducing bacteria in lactate- and acetate-amended enrichments, but higher counts were observed when glucose was used as an electron donor in acidic enrichments. Phylogenetic analysis of 16S rRNA gene sequences extracted from the highest positive MPN dilutions revealed that the predominant members of Fe(III)-reducing consortia from background sediments were closely related to members of theGeobacteraceaefamily, whereas a recently characterized Fe(III) reducer (Anaeromyxobactersp.) and organisms not previously shown to reduce Fe(III) (PaenibacillusandBrevibacillusspp.) predominated in the Fe(III)-reducing consortia of contaminated sediments. Analysis of enrichment cultures by terminal restriction fragment length polymorphism (T-RFLP) strongly supported the cloning and sequencing results. Dominant members of the Fe(III)-reducing consortia were observed to be stable over several enrichment culture transfers by T-RFLP in conjunction with measurements of Fe(III) reduction activity and carbon substrate utilization. Enrichment cultures from contaminated sites were also shown to rapidly reduce millimolar amounts of U(VI) in comparison to killed controls. With DNA extracted directly from subsurface sediments, quantitative analysis of 16S rRNA gene sequences with MPN-PCR indicated thatGeobacteraceaesequences were more abundant in pristine compared to contaminated environments,whereasAnaeromyxobactersequences were more abundant in contaminated sediments. Thus, results from a combination of cultivation-based and cultivation-independent approaches indicate that the abundance/community composition of Fe(III)-reducing consortia in subsurface sediments is dependent upon geochemical parameters (pH, nitrate concentration) and that microorganisms capable of producing spores (gram positive) or spore-like bodies (Anaeromyxobacter) were representative of acidic subsurface environments.
Single-Copy Green Fluorescent Protein Gene FusionsAllow Accurate Measurement of
Salmonella
Gene Expression InVitro and during Infection of MammalianCells
Authors: Hautefort Isabelle , Proença Maria José , Hinton Jay C. D. 
Published: December 2003

ABSTRACT

We
developed a reliable and flexible green fluorescent protein (GFP)-based
system for measuring gene expression in individual bacterial cells.
Until now, most systems have relied upon plasmid-borne
gfp
gene fusions, risking problems associated with plasmid instability. We
show that a recently developed GFP variant, GFP
+
, is
suitable for assessing bacterial gene expression. Various

gfp
+

transcriptional fusions were
constructed and integrated as single copies into the chromosome of
Salmonella enterica
serovar Typhimurium. A comparison of the
expression levels of
proU-lacZ
and

proU-gfp
+

fusions showed that
GFP
+
reported
proU
activity in individual
Salmonella
cells as accurately as β-galactosidase
reported activity for entire populations. The single-copy

gfp
+

fusions were ideal for monitoring up-
and downregulation of
Salmonella
virulence genes. We
discovered that in vitro induction of the
SPI1
gene
prgH
occurs only in a portion of the population and that
the proportion varies with the growth phase. We determined the level of
expression of the SPI2 gene
ssaG
in bacteria released from
murine macrophages. Our results demonstrate for the first time that
single-copy GFP
+
fusions reliably report gene
expression in simple and complex environments. This approach promises
to allow accurate measurement of gene expression in individual bacteria
during animal
infection.
Role of the Glycine Betaine and Carnitine Transporters in Adaptation ofListeria monocytogenesto Chill Stress in Defined Medium
Authors: Angelidis Apostolos S. , Smith Gary M. 
Published: December 2003

ABSTRACTThe food-borne pathogenListeria monocytogenesproliferates at refrigeration temperatures, rendering refrigeration ineffective in the preservation ofListeria-contaminated foods. The uptake and intracellular accumulation of the potent compatible solutes glycine betaine and carnitine has been shown to be a key mediator of the pathogen's cold-tolerant phenotype. To date, three compatible solute systems are known to operate inL. monocytogenes: glycine betaine porter I (BetL), glycine betaine porter II (Gbu), and the carnitine transporter OpuC. We investigated the specificity of each transporter towards each compatible solute at 4°C by examining mutant derivatives ofL. monocytogenes10403S that possess each of the transporters in isolation. Kinetic and steady-state compatible solute accumulation data together with growth rate experiments demonstrated that under cold stress glycine betaine transport is primarily mediated by Gbu and that Gbu-mediated betaine uptake results in significant growth stimulation of chill-stressed cells. BetL and OpuC can serve as minor porters for the uptake of betaine, and their action is capable of providing a small degree of cryotolerance. Under cold stress, carnitine transport occurs primarily through OpuC and results in a high level of cryoprotection. Weak carnitine transport occurs via Gbu and BetL, conferring correspondingly weak cryoprotection. No other transporter inL. monocytogenes10403S appears to be involved in transport of either compatible solute at 4°C, since a triple mutant strain yielded neither transport nor accumulation of glycine betaine or carnitine and could not be rescued by either osmolyte when grown at that temperature.
Experimental Examination of BacteriophageLatent-Period Evolution as a Response to BacterialAvailability
Authors: Abedon Stephen T. , Hyman Paul , Thomas Cameron 
Published: December 2003

ABSTRACTFor obligately lytic bacteriophage (phage) a trade-off exists between fecundity (burst size) and latent period (a component of generation time). This trade-off occurs because release of phage progeny from infected bacteria coincides with destruction of the machinery necessary to produce more phage progeny. Here we employ phage mutants to explore issues of phage latent-period evolution as a function of the density of phage-susceptible bacteria. Theory suggests that higher bacterial densities should select for shorter phage latent periods. Consistently, we have found that higher host densities (≥∼107bacteria/ml) can enrich stocks of phage RB69 for variants that display shorter latent periods than the wild type. One such variant, dubbed sta5, displays a latent period that is ∼70 to 80% of that of the wild type—which is nearly as short as the RB69 eclipse period—and which has a corresponding burst size that is ∼30% of that of the wild type. We show that at higher host densities (≥∼107bacteria/ml) the sta5 phage can outcompete the RB69 wild type, though only under conditions of direct (same-culture) competition. We interpret this advantage as corresponding to slightly faster sta5 population growth, resulting in multifold increases in mutant frequency during same-culture growth. The sta5 advantage is lost, however, given indirect (different-culture) competition between the wild type and mutant or given same-culture competition but at lower densities of phage-susceptible bacteria (≤∼106bacteria/ml). From these observations we suggest that phage displaying very short latent periods may be viewed as specialists for propagation when bacteria within cultures are highly prevalent and transmission between cultures is easily accomplished.