bluuebunny/crossref_metadata_2025_split huggingface dataset:
Escherichia coli
Heat Shock Protein DnaK: Production and Consequences in Terms of Monitoring Cooking
Authors: Seyer Karine , Lessard Martin , Piette Gabriel , Lacroix Monique , Saucier Linda 
Published: June 2003

ABSTRACT

Through use of commercially available DnaK proteins and anti-DnaK monoclonal antibodies, a competitive enzyme-linked immunosorbent assay was developed to quantify this heat shock protein in
Escherichia coli
ATCC 25922 subjected to various heating regimens. For a given process lethality (
F
70
10
of 1, 3, and 5 min), the intracellular concentration of DnaK in
E. coli
varied with the heating temperature (50 or 55°C). In fact, the highest DnaK concentrations were found after treatments at the lower temperature (50°C) applied for a longer time. Residual DnaK after heating was found to be necessary for cell recovery, and additional DnaK was produced during the recovery process. Overall, higher intracellular concentrations of DnaK tended to enhance cell resistance to a subsequent lethal stress. Indeed,
E. coli
cells that had undergone a sublethal heat shock (105 min at 55°C,
F
70
10
= 3 min) accompanied by a 12-h recovery (containing 76,786 ± 25,230 molecules/cell) resisted better than exponentially growing cells (38,500 ± 6,056 molecules/cell) when later heated to 60°C for 50 min (
F
70
10
= 5 min). Results reported here suggest that using stress protein to determine cell adaptation and survival, rather than cell counts alone, may lead to more efficient heat treatment.
The Bacterium
Thermus thermophilus
, Like Hyperthermophilic Archaea, Uses a Two-Step Pathway for the Synthesis of Mannosylglycerate
Authors: Empadinhas Nuno , Albuquerque Luciana , Henne Anke , Santos Helena , Costa Milton S. da 
Published: June 2003

ABSTRACT

The biosynthetic pathway for the synthesis of the compatible solute α-mannosylglycerate (MG) in the thermophilic bacterium
Thermus thermophilus
HB27 was identified based on the activities of recombinant mannosyl-3-phosphoglycerate synthase (MPGS) (EC 2.4.1.217) and mannosyl-3-phosphoglycerate phosphatase (MPGP) (EC 3.1.3.70). The sequences of homologous genes from the archaeon
Pyrococcus horikoshii
were used to identify MPGS and MPGP genes in
T. thermophilus
HB27 genome. Both genes were separately cloned and overexpressed in
Escherichia coli
, yielding 3 to 4 mg of pure recombinant protein per liter of culture. The molecular masses were 43.6 and 28.1 kDa for MPGS and MPGP, respectively. The recombinant MPGS catalyzed the synthesis of α-mannosyl-3-phosphoglycerate (MPG) from GDP-mannose and
d
-3-phosphoglycerate, while the recombinant MPGP catalyzed the dephosphorylation of MPG to MG. The recombinant MPGS had optimal activity at 80 to 90°C and a pH optimum near 7.0; MPGP had maximal activity between 90 and 95°C and at pH 6.0. The activities of both enzymes were strictly dependent on divalent cations; Mn
2+
was most effective for MPGS, while Mn
2+
, Co
2+
, Mg
2+
, and to a lesser extent Ni
2+
activated MPGP. The organization of MG biosynthetic genes in
T. thermophilus
HB27 is different from the
P. horikoshii
operon-like structure, since the genes involved in the conversion of fructose-6-phosphate to GDP-mannose are not found immediately downstream of the contiguous MPGS and MPGP genes. The biosynthesis of MG in the thermophilic bacterium
T. thermophilus
HB27, proceeding through a phosphorylated intermediate, is similar to the system found in hyperthermophilic archaea.
Isolation of a Methanogen from Deep Marine Sediments That Contain Methane Hydrates, and Description of
Methanoculleus submarinus
sp. nov
Authors: Mikucki Jill A. , Liu Yitai , Delwiche Mark , Colwell Frederick S. , Boone David R. 
Published: June 2003

ABSTRACT

We isolated a methanogen from deep in the sediments of the Nankai Trough off the eastern coast of Japan. At the sampling site, the water was 950 m deep and the sediment core was collected at 247 m below the sediment surface. The isolated methanogen was named Nankai-1. Cells of Nankai-1 were nonmotile and highly irregular coccoids (average diameter, 0.8 to 2 μm) and grew with hydrogen or formate as a catabolic substrate. Cells required acetate as a carbon source. Yeast extract and peptones were not required but increased the growth rate. The cells were mesophilic, growing most rapidly at 45°C (no growth at ≤10°C or ≥55°C). Cells grew with a maximum specific growth rate of 2.43 day
−1
at 45°C. Cells grew at pH values between 5.0 and 8.7 but did not grow at pH 4.7 or 9.0. Strain Nankai-1 grew in a wide range of salinities, from 0.1 to 1.5 M Na
+
. The described phenotypic characteristics of this novel isolate were consistent with the in situ environment of the Nankai Trough. This is the first report of a methanogenic isolate from methane hydrate-bearing sediments. Phylogenetic analysis of its 16S rRNA gene sequence indicated that it is most closely related to
Methanoculleus marisnigri
(99.1% sequence similarity), but DNA hybridization experiments indicated a DNA sequence similarity of only 49%. Strain Nankai-1 was also found to be phenotypically similar to
M. marisnigri
, but two major phenotypic differences were found: strain Nankai-1 does not require peptones, and it grows fastest at a much higher temperature. We propose a new species,
Methanoculleus submarinus
, with strain Nankai-1 as the type strain.
Fluorescent Heteroduplex Assay for Monitoring
Bacillus anthracis
and Close Relatives in Environmental Samples
Authors: Merrill Lori , Richardson Jennifer , Kuske Cheryl R. , Dunbar John 
Published: June 2003

ABSTRACT

A fluorescent heteroduplex method was developed to assess the presence of 16S rRNA gene (rDNA) sequences from
Bacillus anthracis
and close relatives in PCR-amplified 16S rDNA sequence mixtures from environmental samples. The method uses a single-stranded, fluorescent DNA probe, 464 nucleotides in length, derived from a
B. anthracis
16S rRNA gene. The probe contains a unique, engineered deletion such that all probe-target duplexes are heteroduplexes with an unpaired G at position 343 (ΔG343). Heteroduplex profiles of sequences ≥85% similar to the probe were produced using an ABI 377 sequencer in less than 3 h. The method divides strains of the
Bacillus cereus
-
Bacillus thuringiensis
-
B. anthracis
group into two subgroups. Each subgroup is defined by a specific 16S rRNA gene sequence type. Sequence type A, containing one mismatch with the probe, occurs in
B. anthracis
and a small number of closely related clonal lineages represented mostly by food-borne pathogenic isolates of
B. cereus
and
B. thuringiensis
. Sequence type B, containing two mismatches with the probe, is found in the majority of
B. cereus
and
B. thuringiensis
strains examined to date. Sequence types A and B, when hybridized to the probe, generate two easily differentiated heteroduplexes. Thus, from heteroduplex profiles, the presence of
B. cereus
-
B. thuringiensis
-
B. anthracis
subgroups in environmental samples can be inferred unambiguously. The results show that fluorescent heteroduplex analysis is an effective profiling technique for detection and differentiation of sequences representing small phylogenetic or functional groups in environmental samples.
Identification and Manipulation of Soil Properties To Improve the Biological Control Performance of Phenazine-ProducingPseudomonas fluorescens
Authors: Ownley Bonnie H. , Duffy Brion K. , Weller David M. 
Published: June 2003

ABSTRACTPseudomonas fluorescens2-79RN10protects wheat against take-all disease caused byGaeumannomyces graminisvar.tritici; however, the level of protection in the field varies from site to site. Identification of soil factors that exert the greatest influence on disease suppression is essential to improving biocontrol. In order to assess the relative importance of 28 soil properties on take-all suppression, seeds were treated with strain 2-79RN10(which produces phenazine-1-carboxylate [PCA+]) or a series of mutants with PCA+and PCA−phenotypes. Bacterized seeds were planted in 10 soils, representative of the wheat-growing region in the Pacific Northwest. Sixteen soil properties were correlated with disease suppression. Biocontrol activity of PCA+strains was positively correlated with ammonium-nitrogen, percent sand, soil pH, sodium (extractable and soluble), sulfate-sulfur, and zinc. In contrast, biocontrol was negatively correlated with cation-exchange capacity (CEC), exchangeable acidity, iron, manganese, percent clay, percent organic matter (OM), percent silt, total carbon, and total nitrogen. Principal component factor analysis of the 16 soil properties identified a three-component solution that accounted for 87 percent of the variance in disease rating (biocontrol). A model was identified with step-wise regression analysis (R2= 0.96; Cpstatistic = 6.17) that included six key soil properties: ammonium-nitrogen, CEC, iron, percent silt, soil pH, and zinc. As predicted by our regression model, the biocontrol activity of 2-79RN10was improved by amending a soil low in Zn with 50 μg of zinc-EDTA/g of soil. We then investigated the negative correlation of OM with disease suppression and found that addition of OM (as wheat straw) at rates typical of high-OM soils significantly reduced biocontrol activity of 2-79RN10.
A Stable Bioluminescent Construct of
Escherichia coli
O157:H7 for Hazard Assessments of Long-Term Survival in the Environment
Authors: Ritchie Jennifer M. , Campbell Graeme R. , Shepherd Jill , Beaton Yvonne , Jones Davey , Killham Ken , Artz Rebekka R. E. 
Published: June 2003

ABSTRACT

A chromosomally
lux-
marked (Tn
5 luxCDABE
) strain of nontoxigenic
Escherichia coli
O157:H7 was constructed by transposon mutagenesis and shown to have retained the O157, H7, and intimin phenotypes
.
The survival characteristics of this strain in the experiments performed (soil at −5, −100, and −1,500 kPa matric potential and artificial groundwater) were indistinguishable from the wild-type strain. Evaluation of potential luminescence was found to be a rapid, cheap, and quantitative measure of viable
E. coli
O157:H7 Tn
5 luxCDABE
populations in environmental samples. In the survival studies, bioluminescence of the starved populations of
E. coli
O157:H7 Tn
5 luxCDABE
could be reactivated to the original levels of light emission, suggesting that these populations remain viable and potentially infective to humans. The attributes of the construct offer a cheap and low-risk substitute to the use of verocytotoxin-producing
E. coli
O157:H7 in long-term survival studies.
Development of a Thermally Regulated Broad-Spectrum Promoter System for Use in Pathogenic Gram-Positive Species
Authors: Schofield David A. , Westwater Caroline , Hoel Brian D. , Werner Phillip A. , Norris James S. , Schmidt Michael G. 
Published: June 2003

ABSTRACT

Selectively regulating gene expression is an essential molecular tool that is lacking for many pathogenic gram-positive bacteria. In this report, we describe the evaluation of a series of promoters regulated by the bacteriophage P1 temperature-sensitive C1 repressor in
Enterococcus faecium
,
Enterococcus faecalis
, and
Staphylococcus aureus
. Using the
lacZ
gene to monitor gene expression, we examined the strength, basal expression, and induced expression of synthetic promoters carrying C1 operator sites. The promoters exhibited extremely low basal expression and, under inducing conditions, gave high levels of expression (100- to 1,000-fold induction). We demonstrate that the promoter system could be modulated by temperature and showed rapid induction and that the mechanism of regulation occurred at the level of transcription. Controlled expression with the same constructs was also demonstrated in the gram-negative bacterium
Escherichia coli
. However, low basal expression and the ability to achieve derepression were dependent on both the number of mismatches in the C1 operator sites and the promoter driving
c
1 expression. Since the promoters were designed to contain conserved promoter elements from gram-positive species and were constructed in a broad-host-range plasmid, this system will provide a new opportunity for controlled gene expression in a variety of gram-positive bacteria.
Giardia
Cysts in Wastewater Treatment Plants in Italy
Authors: Cacciò Simone M. , De Giacomo Marzia , Aulicino Francesca A. , Pozio Edoardo 
Published: June 2003

ABSTRACT

Reductions in annual rainfall in some regions and increased human consumption have caused a shortage of water resources at the global level. The recycling of treated wastewaters has been suggested for certain domestic, industrial, and agricultural activities. The importance of microbiological and parasitological criteria for recycled water has been repeatedly emphasized. Among water-borne pathogens, protozoa of the genera
Giardia
and
Cryptosporidium
are known to be highly resistant to water treatment procedures and to cause outbreaks through contaminated raw or treated water. We conducted an investigation in four wastewater treatment plants in Italy by sampling wastewater at each stage of the treatment process over the course of 1 year. The presence of the parasites was assessed by immunofluorescence with monoclonal antibodies. While
Cryptosporidium
oocysts were rarely observed,
Giardia
cysts were detected in all samples throughout the year, with peaks observed in autumn and winter. The overall removal efficiency of cysts in the treatment plants ranged from 87.0 to 98.4%. The removal efficiency in the number of cysts was significantly higher when the secondary treatment consisted of active oxidation with O
2
and sedimentation instead of activated sludge and sedimentation (94.5% versus 72.1 to 88.0%;
P
= 0.05, analysis of variance). To characterize the cysts at the molecular level, the β
-
giardin gene was PCR amplified, and the products were sequenced or analyzed by restriction. Cysts were typed as assemblage A or B, both of which are human pathogens, stressing the potential risk associated with the reuse of wastewater.
Use of Antibiotic Resistance Analysis for Representativeness Testing of Multiwatershed Libraries
Authors: Wiggins Bruce A. , Cash Philip W. , Creamer Wes S. , Dart Scott E. , Garcia Preston P. , Gerecke Todd M. , Han Jennifer , Henry Brian L. , Hoover Kylie B. , Johnson Erika L. , Jones K. C. , McCarthy Jacquie G. , McDonough Justin A. , Mercer Sarah A. , Noto Michael J. , Park Haewon , Phillips Matthew S. , Purner Stephanie M. , Smith Brian M. , Stevens Erin N. , Varner Amy K. 
Published: June 2003

ABSTRACT
The use of antibiotic resistance analysis (ARA) for microbial source tracking requires the generation of a library of isolates collected from known sources in the watershed. The size and composition of the library are critical in determining if it represents the diversity of patterns found in the watershed. This study was performed to determine the size that an ARA library needs to be to be representative of the watersheds for which it will be used and to determine if libraries from different watersheds can be merged to create multiwatershed libraries. Fecal samples from known human, domesticated, and wild animal sources were collected from six Virginia watersheds. From these samples, enterococci were isolated and tested by ARA. Based on cross-validation discriminant analysis, only the largest of the libraries (2,931 isolates) were found to be able to classify nonlibrary isolates as well as library isolates (i.e., were representative). Small libraries tended to have higher average rates of correct classification, but were much less able to correctly classify nonlibrary isolates. A merged multiwatershed library (6,587 isolates) was created and was found to be large enough to be representative of the isolates from the contributing watersheds. When isolates that were collected from the contributing watersheds approximately 1 year later were analyzed with the multiwatershed library, they were classified as well as the isolates in the library, suggesting that the resistance patterns are temporally stable for at least 1 year. The ability to obtain a representative, temporally stable library demonstrates that ARA can be used to identify sources of fecal pollution in natural waters.
Glucose Metabolism in
Lactococcus lactis
MG1363 under Different Aeration Conditions: Requirement of Acetate To Sustain Growth under Microaerobic Conditions
Authors: Nordkvist Mikkel , Jensen Niels Bang Siemsen , Villadsen John 
Published: June 2003

ABSTRACT

Lactococcus lactis
subsp.
lactis
MG1363 was grown in batch cultures on a defined medium with glucose as the energy source under different aeration conditions, namely, anaerobic conditions, aerobic conditions, and microaerobic conditions with a dissolved oxygen tension of 5% (when saturation with air was used as the reference). The maximum specific growth rate was high (0.78 to 0.91 h
−1
) under all aeration conditions but decreased with increasing aeration, and more than 90% of the glucose was converted to lactate. However, a shift in by-product formation was observed. Increasing aeration resulted in acetate, CO
2
, and acetoin replacing formate and ethanol as end products. Under microaerobic conditions, growth came to a gradual halt, although more than 60% of the glucose was still left. A decline in growth was not observed during microaerobic cultivation when acetate was added to the medium. We hypothesize that the decline in growth was due to a lack of acetyl coenzyme A (acetyl-CoA) needed for fatty acid synthesis since acetyl-CoA can be synthesized from acetate by means of acetate kinase and phosphotransacetylase activities.
Rapid Detection of
Campylobacter coli
,
C. jejuni
, and
Salmonella enterica
on Poultry Carcasses by Using PCR-Enzyme-Linked Immunosorbent Assay
Authors: Hong Yang , Berrang Mark E. , Liu Tongrui , Hofacre Charles L. , Sanchez Susan , Wang Lihua , Maurer John J. 
Published: June 2003

ABSTRACT

Contamination of retail poultry by
Campylobacter
spp. and
Salmonella enterica
is a significant source of human diarrheal disease. Isolation and identification of these microorganisms require a series of biochemical and serological tests. In this study,
Campylobacter ceuE
and
Salmonella invA
genes were used to design probes in PCR-enzyme-linked immunosorbent assay (ELISA), as an alternative to conventional bacteriological methodology, for the rapid detection of
Campylobacter jejuni
,
Campylobacter coli
, and
S. enterica
from poultry samples. With PCR-ELISA (40 cycles), the detection limits for
Salmonella
and
Campylobacter
were 2 × 10
2
and 4 × 10
1
CFU/ml, respectively. ELISA increased the sensitivity of the conventional PCR method by 100- to 1,000-fold. DNA was extracted from carcass rinses and tetrathionate enrichments and used in PCR-ELISA for the detection of
Campylobacter
and
S. enterica
, respectively. With PCR-ELISA,
Salmonella
was detected in 20 of 120 (17%) chicken carcass rinses examined, without the inclusion of an enrichment step. Significant correlation was observed between PCR-ELISA and cultural methods (kappa = 0.83; chi-square test,
P
< 0.001) with only one false negative (1.67%) and four false positives (6.67%) when PCR-ELISA was used to screen 60 tetrathionate enrichment cultures for
Salmonella
. With PCR-ELISA, we observed a positive correlation between the ELISA absorbance (optical density at 405 nm) and the campylobacter cell number in carcass rinse, as determined by standard culture methods. Overall, PCR-ELISA is a rapid and cost-effective approach for the detection and enumeration of
Salmonella
and
Campylobacter
bacteria on poultry.
Bacterial Colonization of Particles: Growth and Interactions
Authors: Grossart Hans-Peter , Kiørboe Thomas , Tang Kam , Ploug Helle 
Published: June 2003

ABSTRACT
Marine particles in the ocean are exposed to diverse bacterial communities, and colonization and growth of attached bacteria are important processes in the degradation and transformation of the particles. In an earlier study, we showed that the initial colonization of model particles by individual bacterial strains isolated from marine aggregates was a function of attachment and detachment. In the present study, we have investigated how this colonization process was further affected by growth and interspecific interactions among the bacteria. Long-term incubation experiments showed that growth dominated over attachment and detachment after a few hours in controlling the bacterial population density on agar particles. In the absence of grazing mortality, this growth led to an equilibrium population density consistent with the theoretical limit due to oxygen diffusion. Interspecific interaction experiments showed that the presence of some bacterial strains (“residents”) on the agar particles either increased or decreased the colonization rate of other strains (“newcomers”). Comparison between an antibiotic-producing strain and its antibiotic-free mutant showed no inhibitory effect on the newcomers due to antibiotic production. On the contrary, hydrolytic activity of the antibiotic-producing strain appeared to benefit the newcomers and enhance their colonization rate. These results show that growth- and species-specific interactions have to be taken into account to adequately describe bacterial colonization of marine particles. Changes in colonization pattern due to such small-scale processes may have profound effects on the transformation and fluxes of particulate matter in the ocean.
Chemical and Biological Interactions during Nitrate and Goethite Reduction by
Shewanella putrefaciens
200
Authors: Cooper D. Craig , Picardal Flynn W. , Schimmelmann Arndt , Coby Aaron J. 
Published: June 2003

ABSTRACT

Although previous research has demonstrated that NO
3
−
inhibits microbial Fe(III) reduction in laboratory cultures and natural sediments, the mechanisms of this inhibition have not been fully studied in an environmentally relevant medium that utilizes solid-phase, iron oxide minerals as a Fe(III) source. To study the dynamics of Fe and NO
3
−
biogeochemistry when ferric (hydr)oxides are used as the Fe(III) source,
Shewanella putrefaciens
200 was incubated under anoxic conditions in a low-ionic-strength, artificial groundwater medium with various amounts of NO
3
−
and synthetic, high-surface-area goethite. Results showed that the presence of NO
3
−
inhibited microbial goethite reduction more severely than it inhibited microbial reduction of the aqueous or microcrystalline sources of Fe(III) used in other studies. More interestingly, the presence of goethite also resulted in a twofold decrease in the rate of NO
3
−
reduction, a 10-fold decrease in the rate of NO
2
−
reduction, and a 20-fold increase in the amounts of N
2
O produced. Nitrogen stable isotope experiments that utilized δ
15
N values of N
2
O to distinguish between chemical and biological reduction of NO
2
−
revealed that the N
2
O produced during NO
2
−
or NO
3
−
reduction in the presence of goethite was primarily of abiotic origin. These results indicate that concomitant microbial Fe(III) and NO
3
−
reduction produces NO
2
−
and Fe(II), which then abiotically react to reduce NO
2
−
to N
2
O with the subsequent oxidation of Fe(II) to Fe(III).