bluuebunny/crossref_metadata_2025_split huggingface dataset:
Use of Inverse PCR for Analysis of Class 1 Integrons Carrying an Unusual 3′ Conserved Segment Structure
Authors: Lee Mei-Feng , Peng Chien-Fang , Hsu Hui-Jine , Toh Han-Siong 
Published: November 2010

ABSTRACT

By using inverse PCR and DNA sequencing, 13
sul3
-associated mutational integrons, 2 defective class 1 integrons, and 1
qnrB2
-associated complex
sul1
-type class 1 integrons were identified in
Salmonella enterica
serovar Choleraesuis,
Pseudomonas aeruginosa
, and
Enterobacter cloacae
, respectively. In addition, conjugation and Southern hybridization demonstrated that unusual class 1 integrons were located on plasmids or integrated into chromosomal DNA. Thus, an inverse PCR assay can be a valuable tool for the analysis of unusual structures of the 3′ conserved region of class 1 integrons.
Expression of Multidrug Efflux Pump Genes
acrAB-tolC
,
mdfA
, and
norE
in
Escherichia coli
Clinical Isolates as a Function of Fluoroquinolone and Multidrug Resistance
Authors: Swick Michelle C. , Morgan-Linnell Sonia K. , Carlson Kimberly M. , Zechiedrich Lynn 
Published: November 2010

ABSTRACT

In a single quantitative study, we measured
acrA
,
acrB
,
tolC
,
mdfA
, and
norE
expression in
Escherichia coli
clinical isolates by using real-time PCR.
acrA
and
acrB
overexpression strongly correlated with fluoroquinolone and multidrug resistance;
tolC
,
mdfA
, and
norE
expression did not. The order of abundance of efflux pump transcripts in all fluoroquinolone-susceptible isolates was
tolC
(highest), then
acrA
and
acrB
, and then
mdfA
and
norE.
Our findings suggest
acrAB
overexpression is an indicator of multidrug resistance.
Combinatorial Phenotypic Signatures Distinguish Persistent from Resolving Methicillin-Resistant
Staphylococcus aureus
Bacteremia Isolates
Authors: Seidl Kati , Bayer Arnold S. , Fowler Vance G. , McKinnell James A. , Abdel Hady Wessam , Sakoulas George , Yeaman Michael R. , Xiong Yan Q. 
Published: November 2010

ABSTRACT

Persistent methicillin-resistant
Staphylococcus aureus
(MRSA) bacteremia (PB) (positive blood cultures after ≥7 days of therapy) represents a clinically challenging subset of invasive MRSA infections. In this investigation, we examined the potential correlation of specific virulence signatures with PB versus resolving MRSA bacteremia (RB) (negative blood cultures within 2 to 4 days of therapy) strains. Thirty-six MRSA isolates from patients enrolled in a recent multinational clinical trial were studied for (i) susceptibility to host defense cationic peptides (HDPs) (i.e., thrombin-induced platelet microbicidal proteins [tPMPs] and human neutrophil peptide 1 [hNP-1]); (ii) adherence to host endovascular ligands (fibronectin) and cells (endothelial cells); and (iii) biofilm formation. We found that PB isolates exhibited significantly reduced susceptibilities to tPMPs and hNP-1 (
P
< 0.001 and
P
= 0.023, respectively). There was no significant association between the PB outcome and fibronectin binding, endothelial cell binding, or biofilm formation (
P
= 0.25, 0.97, and 0.064 versus RB strains, respectively). However, multiple logistic regression analysis revealed that the PB outcome was significantly associated with the combination of reduced susceptibilities to HDPs and extent of biofilm formation (
P
< 0.0001). Similar results were obtained in a second analysis using days of bacteremia as a continuous outcome, showing that reduced HDP susceptibilities and increased biofilm formation cocontributed to predict the duration of bacteremia. Our data indicate that PB isolates have specific pathogenic signatures independent of conventional antimicrobial susceptibility. These combinatorial mosaics can be defined and used to prospectively distinguish PB from RB strains in advance and potentially to predict ultimate clinical outcomes.
The Antibiotic Monensin Causes Cell Cycle Disruption ofToxoplasma gondiiMediated through the DNA Repair Enzyme TgMSH-1
Authors: Lavine Mark D. , Arrizabalaga Gustavo 
Published: November 2010

ABSTRACTMonensin is a polyether ionophore antibiotic that is widely used in the control of coccidia in animals. Despite its significance in veterinary medicine, little is known about its mode of action and potential mechanisms of resistance in coccidian parasites. Here we show that monensin causes accumulation of the coccidianToxoplasma gondiiat an apparent late-S-phase cell cycle checkpoint. In addition, experiments utilizing a monensin-resistantT. gondiimutant show that this effect of monensin is dependent on the function of a mitochondrial homologue of the MutS DNA damage repair enzyme (TgMSH-1). Furthermore, the same TgMSH-1-dependent cell cycle disruption is observed with the antiparasitic ionophore salinomycin and the DNA alkylating agent methyl nitrosourea. Our results suggest a novel mechanism for the mode of action of monensin and salinomycin on coccidial parasites, in which the drug activates an MSH-1-dependent cell cycle checkpoint by an unknown mechanism, ultimately leading to the death of the parasite. This model would indicate that cell cycle disruption is an important mediator of drug susceptibility and resistance to ionophoric antibiotics in coccidian parasites.
Development and Validation of a High-Performance Liquid Chromatography Method for Determination of Cefquinome Concentrations in Sheep Plasma and Its Application to Pharmacokinetic Studies
Authors: Uney Kamil , Altan Feray , Elmas Muammer 
Published: November 2010

ABSTRACT

Cefquinome has a broad spectrum of antibacterial activity and was developed especially for use in animals. A simple and sensitive high-performance liquid chromatography (HPLC) method with UV-visible detection for quantification of cefquinome concentrations in sheep plasma was developed and validated. Separation of cefquinome from plasma components was achieved on a Phenomenex Gemini C
18
column (250 mm by 4.6 mm; internal diameter [i.d.], 5 μm). The mobile phase consisted of acetonitrile and 0.1% trifluoroacetic acid in water and was delivered at a rate of 0.9 ml/min. A simple and rapid sample preparation involved the addition of methanol to 200 μl of plasma to precipitate plasma proteins followed by direct injection of 50 μl of supernatant into the high-performance liquid chromatography system. The linearity range of the proposed method was 0.02 to 12 μg/ml. The intraday and interday coefficients of variation obtained from cefquinome were less than 5%, and biases ranged from −3.76% to 1.24%. Mean recovery based on low-, medium-, and high-quality control standards ranged between 92.0 and 93.9%. Plasma samples were found to be stable in various storage conditions (freeze-thaw, postpreparative, short-term, and long-term stability). The method described was found to be readily available, practicable, cheap, rapid, sensitive, precise, and accurate. It was successfully applied to the study of the pharmacokinetics of cefquinome in sheep. This method can be very useful and an alternate to performing pharmacokinetic studies in the determination of cefquinome for clinical use.
Whole-Genome Analysis ofSalmonella entericaSerovar Typhimurium T000240 Reveals the Acquisition of a Genomic Island Involved in Multidrug Resistance via IS1Derivatives on the Chromosome
Authors: Izumiya Hidemasa , Sekizuka Tsuyoshi , Nakaya Hideo , Taguchi Masumi , Oguchi Akio , Ichikawa Natsuko , Nishiko Rika , Yamazaki Shuji , Fujita Nobuyuki , Watanabe Haruo , Ohnishi Makoto , Kuroda Makoto 
Published: November 2010

ABSTRACTSalmonella entericaserovar Typhimurium is frequently associated with life-threatening systemic infections, and the recent global emergence of multidrug resistance inS. entericaisolates from agricultural and clinical settings has raised concerns. In this study, we determined the whole-genome sequence of fluoroquinolone-resistantS. entericaserovar Typhimurium T000240 strain (DT12) isolated from human gastroenteritis in 2000. Comparative genome analysis revealed that T000240 displays high sequence similarity to strain LT2, which was originally isolated in 1940, indicating that progeny of LT2 might be reemerging. T000240 possesses a unique 82-kb genomic island, designated as GI-DT12, which is composed of multidrug resistance determinants, including a Tn2670-like composite transposon (class 1 integron [intI1,blaoxa-30,aadA1,qacEΔ1, andsul1], mercury resistance proteins, and chloramphenicol acetyltransferase), a Tn10-like tetracycline resistance protein (tetA), the aerobactin iron-acquisition siderophore system (lutAandlucABC), and an iron transporter (sitABCD). Since GI-DT12 is flanked by IS1derivatives, IS1-mediated recombination likely played a role in the acquisition of this genomic island through horizontal gene transfer. The aminoglycoside-(3)-N-acetyltransferase (aac(3)) gene and a class 1 integron harboring thedfrA1gene cassette responsible for gentamicin and trimethoprim resistance, respectively, were identified on plasmid pSTMDT12_L and appeared to have been acquired through homologous recombination with IS26. This study represents the first characterization of the unique genomic island GI-DT12 that appears to be associated with possible IS1-mediated recombination inS. entericaserovar Typhimurium. It is expected that future whole-genome studies will aid in the characterization of the horizontal gene transfer events for the emergingS. entericaserovar Typhimurium strains.
Complementarity-Determining Region 3 Size Spectratypes of T Cell Receptor β Chains in CD8+T Cells following Antiviral Treatment of Chronic Hepatitis B
Authors: Ma Shi-Wu , Li Yong-Yin , Zhang Guang-Wen , Huang Xuan , Sun Jian , Li Chris , Abbott William G. H. , Hou Jin-Lin 
Published: November 2010

ABSTRACTAn increased CD8+T cell response to hepatitis B virus (HBV) peptides occurs between 12 and 24 weeks after starting antiviral therapy for chronic hepatitis B. It is not known whether these cells have antiviral function. The aim of this study was to determine whether clonal expansions of CD8+T cells at these time points predict the virological response to therapy. Peripheral blood CD8+T cells were obtained from 20 patients treated with lamivudine or telbivudine for chronic hepatitis B at baseline, 12 weeks, and 24 weeks. The CDR3 spectratype of each T cell receptor (TCR) β chain variable region (Vβ) gene family was analyzed, and the changes in the numbers of Vβ families with clonal expansions were compared in subjects with (n= 12) and without (n= 8) a virological response (52 week HBV DNA < 300 copies/ml). The number of CD8+TCR Vβ families with clonal expansions at 12 weeks relative to baseline (median [10th to 90th percentile], +2.5 [0 to +7] versus +1 [0 to +2],P= 0.03) and at 24 weeks relative to 12 weeks (+1 [0 to +2] versus −1 [−3 to +4],P= 0.006) was higher in subjects with a virological response versus subjects without a virological response, as were interleukin-2 (IL-2) but not IL-21 mRNA levels in peripheral blood mononuclear cells. The duration of new expansions at 12 weeks was higher (P< 0.0001) in responders. Increased numbers of CD8+T cell expansions after antiviral therapy are associated with a virological response to treatment. These CD8+T cells are a potential target for a therapeutic vaccine for chronic hepatitis B.
Diversity of the Early Step of the Futalosine Pathway
Authors: Arakawa Chisato , Kuratsu Masahiro , Furihata Kazuo , Hiratsuka Tomoshige , Itoh Nobuya , Seto Haruo , Dairi Tohru 
Published: November 2010

ABSTRACT

We recently demonstrated that the futalosine pathway was operating in some bacteria for the biosynthesis of menaquinone and that futalosine was converted into dehypoxanthinyl futalosine (DHFL) by an MqnB of
Thermus thermophilus
. In this study, we found that aminodeoxyfutalosine, which has adenine instead of hypoxanthine in futalosine, was directly converted into DHFL by an MqnB of
Helicobacter pylori
. Therefore, this step is potentially an attractive target for the development of specific anti-
H. pylori
drugs.
NALP1 Influences Susceptibility to Human Congenital Toxoplasmosis, Proinflammatory Cytokine Response, and Fate ofToxoplasma gondii-Infected Monocytic Cells
Authors: Witola William H. , Mui Ernest , Hargrave Aubrey , Liu Susan , Hypolite Magali , Montpetit Alexandre , Cavailles Pierre , Bisanz Cordelia , Cesbron-Delauw Marie-France , Fournié Gilbert J. , McLeod Rima 
Published: November 2010

ABSTRACTNALP1 is a member of the NOD-like receptor (NLR) family of proteins that form inflammasomes. Upon cellular infection or stress, inflammasomes are activated, triggering maturation of proinflammatory cytokines and downstream cellular signaling mediated through the MyD88 adaptor.Toxoplasma gondiiis an obligate intracellular parasite that stimulates production of high levels of proinflammatory cytokines that are important in innate immunity. In this study, susceptibility alleles for human congenital toxoplasmosis were identified in the NALP1 gene. To investigate the role of the NALP1 inflammasome during infection withT. gondii, we genetically engineered a human monocytic cell line for NALP1 gene knockdown by RNA interference. NALP1 silencing attenuated progression ofT. gondiiinfection, with accelerated host cell death and eventual cell disintegration. In line with this observation, upregulation of the proinflammatory cytokines interleukin-1β (IL-1β), IL-18, and IL-12 uponT. gondiiinfection was not observed in monocytic cells with NALP1 knockdown. These findings suggest that the NALP1 inflammasome is critical for mediating innate immune responses toT. gondiiinfection and pathogenesis. Although there have been recent advances in understanding the potent activity of inflammasomes in directing innate immune responses to disease, this is the first report, to our knowledge, on the crucial role of the NALP1 inflammasome in the pathogenesis ofT. gondiiinfections in humans.
Enhanced Virulence of
Salmonella enterica
Serovar Typhimurium after Passage through Mice
Authors: Mastroeni Pietro , Morgan Fiona J. E. , McKinley Trevelyan J. , Shawcroft Ewan , Clare Simon , Maskell Duncan J. , Grant Andrew J. 
Published: November 2010

ABSTRACT

The interaction between
Salmonella enterica
and the host immune system is complex. The outcome of an infection is the result of a balance between the
in vivo
environment where the bacteria survive and grow and the regulation of fitness genes at a level sufficient for the bacteria to retain their characteristic rate of growth in a given host. Using bacteriological counts from tissue homogenates and fluorescence microscopy to determine the spread, localization, and distribution of
S. enterica
in the tissues, we show that, during a systemic infection,
S. enterica
adapts to the
in vivo
environment. The adaptation becomes a measurable phenotype when bacteria that have resided in a donor animal are introduced into a recipient naïve animal. This adaptation does not confer increased resistance to early host killing mechanisms but can be detected as an enhancement in the bacterial net growth rate later in the infection. The enhanced growth rate is lost upon a single passage
in vitro
, and it is therefore transient and not due to selection of mutants. The adapted bacteria on average reach higher intracellular numbers in individual infected cells and therefore have patterns of organ spread different from those of nonadapted bacteria. These experiments help in developing an understanding of the influence of passage in a host on the fitness and virulence of
S. enterica
.
The Vi Capsular Polysaccharide Prevents Complement Receptor 3-Mediated Clearance of
Salmonella enterica
Serotype Typhi
Authors: Wilson R. Paul , Winter Sebastian E. , Spees Alanna M. , Winter Maria G. , Nishimori Jessalyn H. , Sanchez Jesus F. , Nuccio Sean-Paul , Crawford Robert W. , Tükel Çagla , Bäumler Andreas J. 
Published: November 2010

ABSTRACT

Capsular polysaccharides are important virulence factors of invasive bacterial pathogens. Here we studied the role of the virulence (Vi) capsular polysaccharide of
Salmonella enterica
serotype Typhi (
S.
Typhi) in preventing innate immune recognition by complement. Comparison of capsulated
S.
Typhi with a noncapsulated mutant (Δ
tviBCDE vexABCDE
mutant) revealed that the Vi capsule interfered with complement component 3 (C3) deposition. Decreased complement fixation resulted in reduced bacterial binding to complement receptor 3 (CR3) on the surface of murine macrophages
in vitro
and decreased CR3-dependent clearance of Vi capsulated
S.
Typhi from the livers and spleens of mice. Opsonization of bacteria with immune serum prior to intraperitoneal infection increased clearance of capsulated
S.
Typhi from the liver. Our data suggest that the Vi capsule prevents CR3-dependent clearance, which can be overcome in part by a specific antibody response.
Adsorption of Components of the Plasma Kinin-Forming System on the Surface ofPorphyromonas gingivalisInvolves Gingipains as the Major Docking Platforms
Authors: Rapala-Kozik Maria , Bras Grazyna , Chruscicka Barbara , Karkowska-Kuleta Justyna , Sroka Aneta , Herwald Heiko , Nguyen Ky-Anh , Eick Sigrun , Potempa Jan , Kozik Andrzej 
Published: November 2010

ABSTRACTEnhanced production of proinflammatory bradykinin-related peptides, the kinins, has been suggested to contribute to the pathogenesis of periodontitis, a common inflammatory disease of human gingival tissues. In this report, we describe a plausible mechanism of activation of the kinin-generating system, also known as the contact system or kininogen-kallikrein-kinin system, by the adsorption of its plasma-derived components such as high-molecular-mass kininogen (HK), prekallikrein (PK), and Hageman factor (FXII) to the cell surface of periodontal pathogenPorphyromonas gingivalis. The adsorption characteristics of mutant strains deficient in selected proteins of the cell envelope suggested that the surface-associated cysteine proteinases, gingipains, bearing hemagglutinin/adhesin domains (RgpA and Kgp) serve as the major platforms for HK and FXII adhesion. These interactions were confirmed by direct binding tests using microplate-immobilized gingipains and biotinylated contact factors. Other bacterial cell surface components such as fimbriae and lipopolysaccharide were also found to contribute to the binding of contact factors, particularly PK. Analysis of kinin release in plasma upon contact withP. gingivalisshowed that the bacterial surface-dependent mechanism is complementary to the previously described kinin generation system dependent on HK and PK proteolytic activation by the gingipains. We also found that severalP. gingivalisclinical isolates differed in the relative significance of these two mechanisms of kinin production. Taken together, these data show the importance of this specific type of bacterial surface-host homeostatic system interaction in periodontal infections.
TheN-Ethyl-N-Nitrosourea-InducedGoldenticketMouse Mutant Reveals an Essential Function ofStingin theIn VivoInterferon Response toListeria monocytogenesand Cyclic Dinucleotides
Authors: Sauer John-Demian , Sotelo-Troha Katia , von Moltke Jakob , Monroe Kathryn M. , Rae Chris S. , Brubaker Sky W. , Hyodo Mamoru , Hayakawa Yoshihiro , Woodward Joshua J. , Portnoy Daniel A. , Vance Russell E. 
Published: November 2010

ABSTRACTType I interferons (IFNs) are central regulators of the innate and adaptive immune responses to viral and bacterial infections. Type I IFNs are induced upon cytosolic detection of microbial nucleic acids, including DNA, RNA, and the bacterial second messenger cyclic-di-GMP (c-di-GMP). In addition, a recent study demonstrated that the intracellular bacterial pathogenListeria monocytogenesstimulates a type I IFN response due to cytosolic detection of bacterially secreted c-di-AMP. The transmembrane signaling adaptor Sting (Tmem173, Mita, Mpys, Eris) has recently been implicated in the induction of type I IFNs in response to cytosolic DNA and/or RNA. However, the role of Sting in response to purified cyclic dinucleotides or duringin vivo L. monocytogenesinfection has not been addressed. In order to identify genes important in the innate immune response, we have been conducting a forward genetic mutagenesis screen in C57BL/6 mice using the mutagenN-ethyl-N-nitrosourea (ENU). Here we describe a novel mutant mouse strain,Goldenticket(Gt), that fails to produce type I IFNs uponL. monocytogenesinfection. By genetic mapping and complementation experiments, we found thatGtmice harbor a single nucleotide variant (T596A) ofStingthat functions as a null allele and fails to produce detectable protein. Analysis of macrophages isolated fromGtmice revealed thatStingis absolutely required for the type I interferon response to both c-di-GMP and c-di-AMP. Additionally,Stingis required for the response to c-di-GMP andL. monocytogenes in vivo. Our results provide new functions forStingin the innate interferon response to pathogens.
Clostridium perfringens
TpeL Glycosylates the Rac and Ras Subfamily Proteins
Authors: Nagahama Masahiro , Ohkubo Akiko , Oda Masataka , Kobayashi Keiko , Amimoto Katsuhiko , Miyamoto Kazuaki , Sakurai Jun 
Published: November 2010

ABSTRACT

Clostridium perfringens
TpeL belongs to a family of large clostridial cytotoxins that encompasses
Clostridium difficile
toxin A (TcdA) and B (TcdB) and
Clostridium sordellii
lethal toxin (TcsL). We report here the identification of the TpeL-catalyzed modification of small GTPases. A recombinant protein (TpeL1-525) derived from the TpeL N-terminal catalytic domain in the presence of streptolysin O (SLO) induced the rounding of Vero cells and the glycosylation of cellular Rac1. Among several hexoses tested, UDP-
N
-acetyl-glucosamine (UDP-GlcNAc) and UDP-glucose (UDP-Glc) served as cosubstrates for TpeL1-525-catalyzed modifications. TpeL1-525 catalyzed the incorporation of UDP-Glc into Ha-Ras, Rap1B, and RalA and of UDP-GlcNAc into Rac1, Ha-Ras, Rap1B, and RalA. In Rac1, TpeL and TcdB share the same acceptor amino acid for glycosylation, Thr-35. In Vero cells treated with TpeL1-525 in the presence of SLO, glycosylation leads to a translocation of the majority of Rac1 and Ha-Ras to the membrane. We demonstrate for first time that TpeL uses both UDP-GlcNAc and UDP-Glc as donor cosubstrates and modifies the Rac1 and Ras subfamily by glycosylation to mediate its cytotoxic effects.