bluuebunny/crossref_metadata_2025_split huggingface dataset:
Population Turnover in a
Microcystis
Bloom Results in Predominantly Nontoxigenic Variants Late in the Season
Authors: Bozarth Connie S. , Schwartz Andrew D. , Shepardson Jonathan W. , Colwell Frederick S. , Dreher Theo W. 
Published: June 2010

ABSTRACT

Surface samples of the 2007
Microcystis
bloom occurring in Copco Reservoir on the Klamath River in Northern California were analyzed genetically by sequencing clone libraries made with amplicons at three loci: the internal transcribed spacer of the rRNA operon (ITS),
cpcBA
, and
mcyA
. Samples were taken between June and October, during which time two cell count peaks occurred, in mid-July and early September. The ITS and
cpcBA
loci could be classified into four or five allele groups, which provided a convenient means for describing the
Microcystis
population and its changes over time. Each group was numerically dominated by a single, highly represented sequence. Other members of each group varied by changes at 1 to 3 nucleotide positions, while groups were separated by up to 30 nucleotide differences. As deduced by a partial sampling of the clone libraries, there were marked population turnovers during the season, indicated by changes in allele composition at both the ITS and
cpcBA
loci. Different ITS and
cpcBA
genotypes appeared to be dominant at the two population peaks. Toxicity (amount of microcystin per cell) and toxigenic potential (
mcyB
copy number) were lower during the second peak, and the
mcyB
copy number fell further as the bloom declined.
Targeted Chromosomal Knockouts in
Mycoplasma pneumoniae
Authors: Krishnakumar Radha , Assad-Garcia Nacyra , Benders Gwynedd A. , Phan Quang , Montague Michael G. , Glass John I. 
Published: June 2010

ABSTRACT

Most gene knockouts in mycoplasmas are achieved through labor-intensive transposon mutagenesis. Here, we describe a method for making targeted deletions in
Mycoplasma pneumoniae
by use of homologous recombination. In this method,
M. pneumoniae
is transformed with a plasmid carrying an antibiotic resistance marker flanked by 1-kb regions surrounding the target gene. Following selection for the antibiotic resistance, colonies are screened for double crossovers which indicate complete deletion of the target open reading frame.
Hydroxyapatite-Mediated Separation of Double-Stranded DNA, Single-Stranded DNA, and RNA Genomes from Natural Viral Assemblages
Authors: Andrews-Pfannkoch Cynthia , Fadrosh Douglas W. , Thorpe Joyce , Williamson Shannon J. 
Published: June 2010

ABSTRACT
Metagenomics can be used to determine the diversity of complex, often unculturable, viral communities with various nucleic acid compositions. Here, we report the use of hydroxyapatite chromatography to efficiently fractionate double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), dsRNA, and ssRNA genomes from known bacteriophages. Linker-amplified shotgun libraries were constructed to generate sequencing reads from each hydroxyapatite fraction. Greater than 90% of the reads displayed significant similarity to the expected genomes at the nucleotide level. These methods were applied to marine viruses collected from the Chesapeake Bay and the Dry Tortugas National Park. Isolated nucleic acids were fractionated using hydroxyapatite chromatography followed by linker-amplified shotgun library construction and sequencing. Taxonomic analysis demonstrated that the majority of environmental sequences, regardless of their source nucleic acid, were most similar to dsDNA viruses, reflecting the bias of viral metagenomic sequence databases.
Alteration of Microbial Communities Colonizing Leaf Litter in a Temperate Woodland Stream by Growth of Trees under Conditions of Elevated Atmospheric CO
2
Authors: Kelly John J. , Bansal Amit , Winkelman Jonathan , Janus Lori R. , Hell Shannon , Wencel Marie , Belt Patricia , Kuehn Kevin A. , Rier Steven T. , Tuchman Nancy C. 
Published: June 2010

ABSTRACT

Elevated atmospheric CO
2
can cause increased carbon fixation and altered foliar chemical composition in a variety of plants, which has the potential to impact forested headwater streams because they are detritus-based ecosystems that rely on leaf litter as their primary source of organic carbon. Fungi and bacteria play key roles in the entry of terrestrial carbon into aquatic food webs, as they decompose leaf litter and serve as a source of nutrition for invertebrate consumers. This study tested the hypothesis that changes in leaf chemistry caused by elevated atmospheric CO
2
would result in changes in the size and composition of microbial communities colonizing leaves in a woodland stream. Three tree species,
Populus tremuloides
,
Salix alba
, and
Acer saccharum
, were grown under ambient (360 ppm) or elevated (720 ppm) CO
2
, and their leaves were incubated in a woodland stream. Elevated-CO
2
treatment resulted in significant increases in the phenolic and tannin contents and C/N ratios of leaves. Microbial effects, which occurred only for
P. tremuloides
leaves, included decreased fungal biomass and decreased bacterial counts. Analysis of fungal and bacterial communities on
P. tremuloides
leaves via terminal restriction fragment length polymorphism (T-RFLP) and clone library sequencing revealed that fungal community composition was mostly unchanged by the elevated-CO
2
treatment, whereas bacterial communities showed a significant shift in composition and a significant increase in diversity. Specific changes in bacterial communities included increased numbers of alphaproteobacterial and cytophaga-flavobacter-bacteroides (CFB) group sequences and decreased numbers of betaproteobacterial and firmicutes sequences, as well as a pronounced decrease in overall Gram-positive bacterial sequences.
Association of the Melioidosis AgentBurkholderia pseudomalleiwith Water Parameters in Rural Water Supplies in Northern Australia
Authors: Draper A. D. K. , Mayo M. , Harrington G. , Karp D. , Yinfoo D. , Ward L. , Haslem A. , Currie B. J. , Kaestli M. 
Published: June 2010

ABSTRACTWe analyzed water parameters and the occurrence of the melioidosis agentBurkholderia pseudomalleiin 47 water bores in Northern Australia.B. pseudomalleiwas associated with soft, acidic bore water of low salinity but high iron levels. This finding aids in identifying water supplies at risk of contamination with this pathogenic bacterium.
Distribution of Shiga-Toxigenic
Escherichia coli
O157 in the Gastrointestinal Tract of Naturally O157-Shedding Cattle at Necropsy
Authors: Keen James E. , Laegreid William W. , Chitko-McKown Carol G. , Durso Lisa M. , Bono James L. 
Published: June 2010

ABSTRACT

Shiga-toxigenic
Escherichia coli
(STEC) O157 occurrence was determined along the entire gastrointestinal tract (GIT) of each of four naturally shedding cattle and at three sites in 61 slaughter cattle. STEC O157 was distributed along the entire GIT, though interanimal distribution was variable. Neither feces nor rectoanal-junction samples accurately predicted the STEC O157-negative status of any particular animal.
Improved Molecular Detection ofAngiostrongylus cantonensisin Mollusks and Other Environmental Samples with a Species-Specific Internal Transcribed Spacer 1-Based TaqMan Assay
Authors: Qvarnstrom Yvonne , da Silva Ana Cristina Aramburu , Teem John L. , Hollingsworth Robert , Bishop Henry , Graeff-Teixeira Carlos , da Silva Alexandre J. 
Published: June 2010

ABSTRACTAngiostrongylus cantonensisis the most common cause of human eosinophilic meningitis. Humans become infected by ingesting food items contaminated with third-stage larvae that develop in mollusks. We report the development of a real-time PCR assay for the species-specific identification ofA. cantonensisin mollusk tissue.
Decreasing Trend of Overlapping Multilocus Sequence Types between Human and Chicken
Campylobacter jejuni
Isolates over a Decade in Finland
Authors: de Haan C. P. A. , Kivistö R. , Hakkinen M. , Rautelin H. , Hänninen M. L. 
Published: June 2010

ABSTRACT

We describe the long-term multilocus sequence typing (MLST) analysis of the population structure and dynamics of 454 Finnish human
Campylobacter jejuni
isolates, as well as 208 chicken isolates, collected during the mid-1990s to 2007. The sequence type clonal complexes (ST CC) ST-45 CC, ST-21 CC, and ST-677 CC were the most common ones found among all isolates, and they covered 73.9% of all isolates. The ST-283 CC also was found frequently among chicken isolates (8.2%). The predominant STs among all isolates were ST-45, ST-50, and ST-677. ST-137 and ST-230 were common among human isolates, and ST-267 was found more frequently among chicken isolates than human isolates. The ST-45 CC was significantly associated with chicken isolates (
P
< 0.01), whereas the ST-21 CC was associated with human isolates (
P
< 0.001). The ST-677 CC was not associated with any host (
P
= 0.5), and an opposite temporary trend of this complex was seen among chicken and human isolates, with an increase in the former and a decrease in the latter during the study period. Furthermore, the ST-22 and ST-48 CCs were significantly associated with human isolates (
P
< 0.01), but neither of the CCs was found in chicken isolates. The annual overlap between STs from human and chicken isolates decreased from 76% at the beginning of the study to 58% at the end. Our results suggest that the importance of chicken as a reservoir for strains associated with human infections has declined despite the consumption of domestic chicken meat increasing during the follow-up period by 83%.
Biochemical Properties of Pectate Lyases Produced by Three Different
Bacillus
Strains Isolated from Fermenting Cocoa Beans and Characterization of Their Cloned Genes
Authors: Ouattara Honoré G. , Reverchon Sylvie , Niamke Sébastien L. , Nasser William 
Published: June 2010

ABSTRACT

Pectinolytic enzymes play an important role in cocoa fermentation. In this study, we characterized three extracellular pectate lyases (Pels) produced by bacilli isolated from fermenting cocoa beans. These enzymes, named Pel-22, Pel-66, and Pel-90, were synthesized by
Bacillus pumilus
BS22,
Bacillus subtilis
BS66, and
Bacillus fusiformis
BS90, respectively. The three Pels were produced under their natural conditions and purified from the supernatants using a one-step chromatography method. The purified enzymes exhibited optimum activity at 60°C, and the half-time of thermoinactivation at this temperature was approximately 30 min. Pel-22 had a low specific activity compared with the other two enzymes. However, it displayed high affinity for the substrate, about 2.5-fold higher than those of Pel-66 and Pel-90. The optimum pHs were 7.5 for Pel-22 and 8.0 for Pel-66 and Pel-90. The three enzymes
trans
-eliminated polygalacturonate in a random manner to generate two long oligogalacturonides, as well as trimers and dimers. A synergistic effect was observed between Pel-22 and Pel-66 and between Pel-22 and Pel-90, but not between Pel-90 and Pel-66. The Pels were also strongly active on highly methylated pectins (up to 60% for Pel-66 and Pel-90 and up to 75% for Pel-22). Fe
2+
was found to be a better cofactor than Ca
2+
for Pel-22 activity, while Ca
2+
was the best cofactor for Pel-66 and Pel-90. The amino acid sequences deduced from the cloned genes showed the characteristics of Pels belonging to Family 1. The
pel
-
66
and
pel
-
90
genes appear to be very similar, but they are different from the
pe
l-
22
gene. The characterized enzymes form two groups, Pel-66/Pel-90 and Pel-22; members of the different groups might cooperate to depolymerize pectin during the fermentation of cocoa beans.
Characterization of a
gne
::IS
629
O Rough:H7
Escherichia coli
Strain from a Hemorrhagic Colitis Patient
Authors: Rump Lydia V. , Beutin Lothar , Fischer Markus , Feng Peter C. H. 
Published: June 2010

ABSTRACT

Shiga-toxigenic
Escherichia coli
strains that are O rough:H7 due to
gne
::IS
629
were thought to be rare and to have unknown pathogenic potential. Recently, an O rough:H7 strain caused by
gne
::IS
629
was isolated from a hemorrhagic colitis patient, suggesting that these strains are pathogenic and may not be as rare as anticipated.
Redirecting Reductant Flux into Hydrogen Production via Metabolic Engineering of Fermentative Carbon Metabolism in a Cyanobacterium
Authors: McNeely Kelsey , Xu Yu , Bennette Nick , Bryant Donald A. , Dismukes G. Charles 
Published: June 2010

ABSTRACT

Some aquatic microbial oxygenic photoautotrophs (AMOPs) make hydrogen (H
2
), a carbon-neutral, renewable product derived from water, in low yields during autofermentation (anaerobic metabolism) of intracellular carbohydrates previously stored during aerobic photosynthesis. We have constructed a mutant (the
ldhA
mutant) of the cyanobacterium
Synechococcus
sp. strain PCC 7002 lacking the enzyme for the NADH-dependent reduction of pyruvate to
d
-lactate, the major fermentative reductant sink in this AMOP. Both nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS) metabolomic methods have shown that autofermentation by the
ldhA
mutant resulted in no
d
-lactate production and higher concentrations of excreted acetate, alanine, succinate, and hydrogen (up to 5-fold) compared to that by the wild type. The measured intracellular NAD(P)(H) concentrations demonstrated that the NAD(P)H/NAD(P)
+
ratio increased appreciably during autofermentation in the
ldhA
strain; we propose this to be the principal source of the observed increase in H
2
production via an NADH-dependent, bidirectional [NiFe] hydrogenase. Despite the elevated NAD(P)H/NAD(P)
+
ratio, no decrease was found in the rate of anaerobic conversion of stored carbohydrates. The measured energy conversion efficiency (ECE) from biomass (as glucose equivalents) converted to hydrogen in the
ldhA
mutant is 12%. Together with the unimpaired photoautotrophic growth of the
ldhA
mutant, these attributes reveal that metabolic engineering is an effective strategy to enhance H
2
production in AMOPs without compromising viability.
Control of the Type 3 Secretion System in
Vibrio harveyi
by Quorum Sensing through Repression of ExsA
Authors: Waters Christopher M. , Wu Julie T. , Ramsey Meghan E. , Harris Rebecca C. , Bassler Bonnie L. 
Published: June 2010

ABSTRACT

The type 3 secretion system (T3SS) genes of
Vibrio harveyi
are activated at low cell density and repressed at high cell density by quorum sensing (QS). Repression requires LuxR, the master transcriptional regulator of QS-controlled genes. Here, we determine the mechanism underlying the LuxR repression of the T3SS system. Using a fluorescence-based cell sorting approach, we isolated
V. harveyi
mutants that are unable to express T3SS genes at low cell density and identified two mutations in the
V. harveyi exsBA
operon. While LuxR directly represses the expression of
exsBA
, complementation and epistasis analyses reveal that it is the repression of
exsA
expression, but not
exsB
expression, that is responsible for the QS-mediated repression of T3SS genes at high cell density. The present work further defines the genes in the
V. harveyi
QS regulon and elucidates a mechanism demonstrating how multiple regulators can be linked in series to direct the expression of QS target genes specifically at low or high cell density.
Recombineering Using RecTE from
Pseudomonas syringae
Authors: Swingle Bryan , Bao Zhongmeng , Markel Eric , Chambers Alan , Cartinhour Samuel 
Published: June 2010

ABSTRACT

In this report, we describe the identification of functions that promote genomic recombination of linear DNA introduced into
Pseudomonas
cells by electroporation. The genes encoding these functions were identified in
Pseudomonas syringae
pv. syringae B728a based on similarity to the lambda Red Exo/Beta and RecET proteins encoded by the lambda and Rac bacteriophages of
Escherichia coli
. The ability of the pseudomonad-encoded proteins to promote recombination was tested in
P. syringae
pv. tomato DC3000 using a quantitative assay based on recombination frequency. The results show that the
Pseudomonas
RecT homolog is sufficient to promote recombination of single-stranded DNA oligonucleotides and that efficient recombination of double-stranded DNA requires the expression of both the RecT and RecE homologs. Additionally, we illustrate the utility of this recombineering system to make targeted gene disruptions in the
P. syringae
chromosome.
Occurrence and Molecular Characteristics of Methicillin-Resistant
Staphylococcus aureus
and Methicillin-Resistant
Staphylococcus pseudintermedius
in an Academic Veterinary Hospital
Authors: Ishihara Kanako , Shimokubo Natsumi , Sakagami Akie , Ueno Hiroshi , Muramatsu Yasukazu , Kadosawa Tsuyoshi , Yanagisawa Chie , Hanaki Hideaki , Nakajima Chie , Suzuki Yasuhiko , Tamura Yutaka 
Published: June 2010

ABSTRACT

Recently, methicillin-resistant
Staphylococcus aureus
(MRSA) and methicillin-resistant
Staphylococcus pseudintermedius
(MRSP) have been increasingly isolated from veterinarians and companion animals. With a view to preventing the spread of MRSA and MRSP, we evaluated the occurrence and molecular characteristics of each in a veterinary college. MRSA and MRSP were isolated from nasal samples from veterinarians, staff members, and veterinary students affiliated with a veterinary hospital. Using stepwise logistic regression, we identified two factors associated with MRSA carriage: (i) contact with an identified animal MRSA case (odds ratio [OR], 6.9; 95% confidence interval [95% CI], 2.2 to 21.6) and (ii) being an employee (OR, 6.2; 95% CI, 2.0 to 19.4). The majority of MRSA isolates obtained from individuals affiliated with the veterinary hospital and dog patients harbored
spa
type t002 and a type II staphylococcal cassette chromosome
mec
(SCC
mec
), similar to the hospital-acquired MRSA isolates in Japan. MRSA isolates harboring
spa
type t008 and a type IV SCC
mec
were obtained from one veterinarian on three different sampling occasions and also from dog patients. MRSA carriers can also be a source of MRSA infection in animals. The majority of MRSP isolates (85.2%) carried hybrid SCC
mec
type II-III, and almost all the remaining MRSP isolates (11.1%) carried SCC
mec
type V. MRSA and MRSP were also isolated from environmental samples collected from the veterinary hospital (5.1% and 6.4%, respectively). The application of certain disinfection procedures is important for the prevention of nosocomial infection, and MRSA and MRSP infection control strategies should be adopted in veterinary medical practice.
Specific and Rapid Enumeration of Viable but Nonculturable and Viable-Culturable Gram-Negative Bacteria by Using Flow Cytometry
Authors: Khan Mohiuddin M. Taimur , Pyle Barry H. , Camper Anne K. 
Published: June 2010

ABSTRACT

An issue of critical concern in microbiology is the ability to detect viable but nonculturable (VBNC) and viable-culturable (VC) cells by methods other than existing approaches. Culture methods are selective and underestimate the real population, and other options (direct viable count and the double-staining method using epifluorescence microscopy and inhibitory substance-influenced molecular methods) are also biased and time-consuming. A rapid approach that reduces selectivity, decreases bias from sample storage and incubation, and reduces assay time is needed. Flow cytometry is a sensitive analytical technique that can rapidly monitor physiological states of bacteria. This report outlines a method to optimize staining protocols and the flow cytometer (FCM) instrument settings for the enumeration of VBNC and VC bacterial cells within 70 min. Experiments were performed using the FCM to quantify VBNC and VC
Escherichia coli
O157:H7,
Pseudomonas aeruginosa
,
Pseudomonas syringae
, and
Salmonella enterica
serovar Typhimurium cells after staining with different fluorescent probes: SYTO 9, SYTO 13, SYTO 17, SYTO 40, and propidium iodide (PI). The FCM data were compared with those for specific standard nutrient agar to enumerate the number of cells in different states. By comparing results from cultures at late log phase, 1 to 64% of cells were nonculturable, 40 to 98% were culturable, and 0.7 to 4.5% had damaged cell membranes and were therefore theoretically dead. Data obtained using four different Gram-negative bacteria exposed to heat and stained with PI also illustrate the usefulness of the approach for the rapid and unbiased detection of dead versus live organisms.
Using Chemobiosynthesis and Synthetic Mini-Polyketide Synthases To Produce Pharmaceutical Intermediates in
Escherichia coli
Authors: Menzella Hugo G. , Carney John R. , Li Yong , Santi Daniel V. 
Published: June 2010

ABSTRACT

Recombinant microbial whole-cell biocatalysis is a valuable approach for producing enantiomerically pure intermediates for the synthesis of complex molecules. Here, we describe a method to produce polyketide intermediates possessing multiple stereogenic centers by combining chemobiosynthesis and engineered mini-polyketide synthases (PKSs). Chemobiosynthesis allows the introduction of unnatural moieties, while a library of synthetic bimodular PKSs expressed from codon-optimized genes permits the introduction of a variety of ketide units. To validate the approach, intermediates for the synthesis of
trans
-9,10-dehydroepothilone D were generated. The designer molecules obtained have the potential to greatly reduce the manufacturing cost of epothilone analogues, thus facilitating their commercial development as therapeutic agents.
Complementation ofListeria monocytogenesNull Mutants with SelectedListeria seeligeriVirulence Genes Suggests Functional Adaptation of Hly and PrfA and Considerable Diversification ofprfARegulation inL. seeligeri
Authors: Lucas Stelling Courtney R. , Orsi Renato H. , Wiedmann Martin 
Published: June 2010

ABSTRACTWhileListeria seeligeriandL. monocytogenescontain the mainListeriavirulence gene cluster, onlyL. monocytogenesis considered an intracellular pathogen. Initial evolutionary analyses showed that the virulence genesprfA,hly, andplcAare conserved inL. seeligeri, with specific Hly and PrfA amino acid residues showing evidence for positive selection inL. seeligeri. Our data also show that temperature-dependent transcript patterns forprfA, which encodes a transcriptional regulator of virulence genes, differed betweenL. monocytogenesandL. seeligeri. To further investigate the divergence of virulence gene function and regulation,L. seeligeri prfA(prfALS),hly(hlyLS), andplcA(plcALS), as well asprfALSconstructs with differentprfApromoter regions, were introduced into appropriateL. monocytogenesnull mutants. Only whenprfALSwas under the control of theL. monocytogenes prfApromoters (P1- and P2prfA) (P1P2LMprfALS) wasprfALSable to fully complement the ΔprfALMdeletion.hlyLSintroduced into anL. monocytogenesbackground under its native promoter showed transcript levels similar to those ofhlyLMand was able to partially restoreL. monocytogeneswild-type-level hemolysis and intracellular growth, even though HlyLMand HlyLSshowed distinct patterns of cell- and supernatant-associated hemolytic activities. Our data indicate that (i) regulation ofprfAexpression differs betweenL. monocytogenesandL. seeligeri, althoughhlytranscription is temperature dependent in both species, and (ii) PrfA and Hly functions are largely, but not fully, conserved betweenL. seeligeriandL. monocytogenes. Virulence gene homologues and their expression thus appear to have adapted to distinct but possibly related functions in these two species.