bluuebunny/crossref_metadata_2025_split huggingface dataset:
Extraintestinal PathogenicEscherichia coliSurvives within Neutrophils
Authors: Nazareth Helen , Genagon Stacy A. , Russo Thomas A. 
Published: February 2007

ABSTRACTExtracellular pathogenicEscherichia coli(ExPEC) strains are common causes of a variety of clinical syndromes, including urinary tract infections, abdominal infections, nosocomial pneumonia, neonatal meningitis, and sepsis. ExPEC strains are extracellular bacterial pathogens; therefore, the innate immune response (e.g., professional phagocytes) plays a crucial role in the host defense against them. Studies using the model ExPEC strain CP9 demonstrated that it is relatively resistant to neutrophil-mediated bactericidal activity. Although this could be due to resistance to phagocytosis, the ability of CP9 to survive the intracellular killing mechanisms of neutrophils is another possibility. Using a variation of the intracellular invasion assay, we studied the survival of CP9 within peripheral blood-derived human neutrophils. Our results indicated that CP9 did survive within human neutrophils, but we were unable to demonstrate that intracellular replication occurred. This finding was not unique to CP9, since when a conservative assessment of survival was used, four of six additional ExPEC strains, but not anE. colilaboratory strain, were also capable of survival within neutrophils. Initial studies in which we began to decipher the mechanisms by which CP9 is able to successfully survive intracellular neutrophil-mediated bactericidal activity demonstrated that CP9 was at least partially susceptible to the neutrophil oxidative burst. Therefore, absolute resistance to the oxidative burst is not a mechanism by which ExPEC survives within neutrophils. In addition, electron microscopy studies showed that CP9 appeared to be present in phagosomes within neutrophils. Therefore, avoidance of phagosomal uptake or subsequent escape from the phagosome does not appear to be a mechanism that contributes to CP9's survival. These findings suggest that survival of ExPEC within neutrophils may be an important virulence mechanism.
Candidate Live, Attenuated
Salmonella enterica
Serotype Typhimurium Vaccines with Reduced Fecal Shedding Are Immunogenic and Effective Oral Vaccines
Authors: Abd El Ghany Moataz , Jansen Angela , Clare Simon , Hall Lindsay , Pickard Derek , Kingsley Robert A. , Dougan Gordon 
Published: February 2007

ABSTRACT

Environmental shedding of genetically manipulated microorganisms is an issue impeding the development of new live vaccines. We have investigated the immunogenicity of a number of novel
Salmonella enterica
serotype Typhimurium oral vaccine candidates that express the fragment C (TetC) component of tetanus toxin and harbor combinations of additional mutations in genes
shdA
,
misL
, and
ratB
that contribute to the persistence of serotype Typhimurium's colonization of the intestine. Serotype Typhimurium
aroA
(TetC) derivatives harboring additional mutations in either
shdA
or
misL
or combinations of these mutations exhibited a marked decrease in shedding of the vaccine strain in the feces of orally vaccinated mice. However, equivalent levels of anti-TetC and anti-
Salmonella
lipopolysaccharide immunoglobulin G (IgG), IgG1, IgG2a, and IgA were detected in sera of the vaccinated but not of the control mice. Cellular immune responses to TetC were detected in all vaccinated mice, regardless of the presence of the additional mutations in
shdA
or
misL
. Further, immunization with serotype Typhimurium
aroA
candidate vaccines harboring
shdA
and
misL
afforded complete protection against challenge with a virulent strain of serotype Typhimurium.
Molecular Damage and Induction of Proinflammatory Cytokines in Human Endothelial Cells Exposed to Shiga Toxin 1, Shiga Toxin 2, and α-Sarcin
Authors: Brigotti Maurizio , Carnicelli Domenica , Ravanelli Elisa , Vara Antonio González , Martinelli Chiara , Alfieri Roberta R. , Petronini Pier Giorgio , Sestili Piero 
Published: February 2007

ABSTRACT
Treatment of human endothelial cells with Shiga toxin 1 and 2 leads to the upregulation of genes encoding proinflammatory molecules involved in the pathogenesis of hemolytic-uremic syndrome. The paradoxical effect of inhibitors of mRNA translation, such as Shiga toxins, that at the same time induce protein expression was investigated by studying the relationship between their enzymatic activity (abstraction of adenine from nucleic acids) and the induction of interleukin-8 and granulocyte-macrophage colony-stimulating factor in human endothelial cells. As a positive control, the fungal toxin α-sarcin, acting on the same rRNA sequence targeted by Shiga toxins with a different mechanism (RNase activity), was used. The three toxins caused ribosomal lesions that, in turn, induced the activation of p38 stress kinase with kinetics that paralleled the inhibition of translation. α-Sarcin was devoid of activity on DNA. Shiga toxin 2 targeted nuclear DNA with more rapid kinetics than did Shiga toxin 1. Since the fungal ribotoxin was fully effective in the induction of proinflammatory proteins, we conclude that damage to ribosomes is indispensable and sufficient to activate protein expression via induction of the stress-kinase cascade. However, gene upregulation events induced by Shiga toxin 2 were much more efficient than those triggered by Shiga toxin 1, although the two toxins impaired translation to the same extent and had overlapping time courses of stress kinase activation. Regulations independent of the ribotoxic stress were assumed to operate in intoxicated cells. We hypothesized that the two bacterial toxins recognize different DNA sequences inducing different regulating effects on gene expression.
Modulation of Naive CD4+T-Cell Responses to an Airway Antigen during Pulmonary Mycobacterial Infection
Authors: Anis Mursalin M. , Fulton Scott A. , Reba Scott M. , Harding Clifford V. , Boom W. Henry 
Published: February 2007

ABSTRACTDuring pulmonary mycobacterial infection, there is increased trafficking of dendritic cells from the lungs to the draining lymph nodes. We hypothesized that ongoing mycobacterial infection would modulate recruitment and activation of antigen-specific naive CD4+T cells after airway antigen challenge. BALB/c mice were infected by aerosol withMycobacterium bovisBCG. At peak bacterial burden in the lungs (4 to 6 weeks postinfection), carboxy-fluorescein diacetate succinimidyl ester-labeled naive ovalbumin-specific DO11.10 T cells were adoptively transferred into infected and uninfected mice. Recipient mice were challenged intranasally with soluble ovalbumin (OVA), and OVA-specific T-cell responses were measured in the lungs, draining mediastinal lymph nodes (MLN), and spleens. OVA challenge resulted in increased activation and proliferation of OVA-specific T cells in the draining MLN of both infected and uninfected mice. However, only BCG-infected mice had prominent OVA-specific T-cell activation, proliferation, and Th1 differentiation in the lungs. BCG infection caused greater distribution of airway OVA to pulmonary dendritic cells and enhanced presentation of OVA peptide by lung CD11c+cells. Together, these data suggest that an existing pulmonary mycobacterial infection alters the phenotype of lung dendritic cells so that they can activate antigen-specific naive CD4+T cells in the lungs in response to airway antigen challenge.
Invasion and Persistence ofMycobacterium aviumsubsp.paratuberculosisduring Early Stages of Johne's Disease in Calves
Authors: Wu Chia-wei , Livesey Michael , Schmoller Shelly K. , Manning Elizabeth J. B. , Steinberg Howard , Davis William C. , Hamilton Mary Jo , Talaat Adel M. 
Published: February 2007

ABSTRACTInfection withMycobacterium aviumsubsp.paratuberculosiscauses Johne's disease in cattle and is a serious problem for the dairy industry worldwide. Development of models to mimic aspects of Johne's disease remains an elusive goal because of the chronic nature of the disease. In this report, we describe a surgical approach employed to characterize the very early stages of infection of calves withM. aviumsubsp.paratuberculosis. To our surprise, strains ofM. aviumsubsp.paratuberculosiswere able to traverse the intestinal tissues within 1 h of infection in order to colonize distant organs, such as the liver and lymph nodes. Both the ileum and the mesenteric lymph nodes were persistently infected for months following intestinal deposition ofM. aviumsubsp.paratuberculosisdespite a lack of fecal shedding of mycobacteria. During the first 9 months of infection, humoral immune responses were not detected. Nonetheless, using flow cytometric analysis, we detected a significant change in the cells participating in the inflammatory responses of infected calves compared to cells in a control animal. Additionally, the levels of cytokines detected in both the ileum and the lymph nodes indicated that there were TH1-type-associated cellular responses but not TH2-type-associated humoral responses. Finally, surgical inoculation of a wild-type strain and a mutantM. aviumsubsp.paratuberculosisstrain (with an inactivatedgcpEgene) demonstrated the ability of the model which we developed to differentiate between the wild-type strain and a mutant strain ofM. aviumsubsp.paratuberculosisdeficient in tissue colonization and invasion. Overall, novel insights into the early stages of Johne's disease were obtained, and a practical model of mycobacterial invasiveness was developed. A similar approach can be used for other enteric bacteria.
A Defined O-Antigen Polysaccharide Mutant ofFrancisella tularensisLive Vaccine Strain Has Attenuated Virulence while Retaining Its Protective Capacity
Authors: Sebastian Shite , Dillon Simon T. , Lynch Jillian G. , Blalock LeeAnn T. , Balon Emmy , Lee Kristin T. , Comstock Laurie E. , Conlan J. Wayne , Rubin Eric J. , Tzianabos Arthur O. , Kasper Dennis L. 
Published: February 2007

ABSTRACTFrancisella tularensis, the causative agent of tularemia, has been designated a CDC category A select agent because of its low infective dose (<10 CFU), its ready transmission by aerosol, and its ability to produce severe morbidity and high mortality. The identification and characterization of this organism's virulence determinants will facilitate the development of a safe and effective vaccine. We report that inactivation of thewbtA-encoded dehydratase of the O-antigen polysaccharide (O-PS) locus of the still-unlicensed live vaccine strain ofF. tularensis(LVS) results in a mutant (the LVSwbtAmutant) with remarkably attenuated virulence. Western blot analysis and immune electron microscopy studies associate this loss of virulence with a complete lack of surface O-PS expression. A likely mechanism for attenuation is shown to be the transformation from serum resistance in the wild-type strain to serum sensitivity in the mutant. Despite this significant attenuation in virulence, the LVSwbtAmutant remains immunogenic and confers protective immunity on mice against challenge with an otherwise lethal dose of eitherF. tularensisLVS or a fully virulent clinical isolate ofF. tularensistype B. Recognition and characterization of the pivotal role of O-PS in the virulence of this intracellular bacterial pathogen may have broad implications for the creation of a safe and efficacious vaccine.
Mucosal and Systemic Immune Responses in Patients with Diarrhea Due to CS6-Expressing Enterotoxigenic
Escherichia coli
Authors: Qadri Firdausi , Ahmed Tanvir , Ahmed Firoz , Bhuiyan M. Saruar , Mostofa Mohammad Golam , Cassels Frederick J. , Helander Anna , Svennerholm Ann-Mari 
Published: February 2007

ABSTRACT

Colonization factor CS6 expressed by enterotoxigenic
Escherichia coli
(ETEC) is a nonfimbrial polymeric protein. A substantial proportion of ETEC strains isolated from patients in endemic settings and in people who travel to regions where ETEC is endemic are ETEC strains expressing CS6, either alone or in combination with fimbrial colonization factor CS5 or CS4. However, relatively little is known about the natural immune responses elicited against CS6 expressed by ETEC strains causing disease. We studied patients who were hospitalized with diarrhea (
n
= 46) caused by CS6-expressing ETEC (ETEC expressing CS6 or CS5 plus CS6) and had a disease spectrum ranging from severe dehydration (27%) to moderate or mild dehydration (73%). Using recombinant CS6 antigen, we found that more than 90% of the patients had mucosal immune responses to CS6 expressed as immunoglobulin (IgA) antibody-secreting cells (ASC) or antibody in lymphocyte supernatant (ALS) and that about 57% responded with CS6-specific IgA antibodies in feces. More than 80% of the patients showed IgA seroconversion to CS6. Significant increases in the levels of anti-CS6 antibodies of the IgG isotype were also observed in assays for ASC (75%), ALS (100%), and serum (70%). These studies demonstrated that patients hospitalized with the noninvasive enteric pathogen CS6-expressing ETEC responded with both mucosal and systemic antibodies against CS6. Studies are needed to determine if the anti-CS6 responses protect against reinfection and if protective levels of CS6 immunity are induced by vaccination.
Contributions of [4Fe-4S]-FNR and Integration Host Factor to
fnr
Transcriptional Regulation
Authors: Mettert Erin L. , Kiley Patricia J. 
Published: February 2007

ABSTRACT

Maintaining appropriate levels of the global regulator FNR is critical to its function as an O
2
sensor. In this study, we examined the mechanisms that control transcription of
fnr
to increase our understanding of how FNR protein levels are regulated. Under anaerobic conditions, one mechanism that controls
fnr
expression is negative autoregulation by the active [4Fe-4S] form of FNR. Through DNase I footprinting and in vitro transcription experiments, we observed that direct binding of [4Fe-4S]-FNR to the predicted downstream FNR binding site is sufficient for repression of the
fnr
promoter in vitro. In addition, the downstream FNR binding site was required for repression of transcription from
fnr
′-
lacZ
fusions in vivo. No repression of
fnr
was observed in vivo or in vitro with the apoprotein form of FNR, indicating that repression requires the dimeric, Fe-S cluster-containing protein. Furthermore, our in vitro and in vivo data suggest that [4Fe-4S]-FNR does not bind to the predicted upstream FNR binding site within the
fnr
promoter. Rather, we provide evidence that integration host factor binds to this upstream region and increases in vivo expression of P
fnr
under both aerobic and anaerobic conditions.
Assembly of Fimbrial Structures inPseudomonas aeruginosa: Functionality and Specificity of Chaperone-Usher Machineries
Authors: Ruer Ségolène , Stender Silke , Filloux Alain , de Bentzmann Sophie 
Published: February 2007

ABSTRACTFimbrial or nonfimbrial adhesins assembled by the bacterial chaperone-usher pathway have been demonstrated to play a key role in pathogenesis. Such an assembly mechanism has been exemplified in uropathogenicEscherichia colistrains with the Pap and the Fim systems. InPseudomonas aeruginosa, three gene clusters (cupA,cupB, andcupC) encodingchaperone-usherpathway components have been identified in the genome sequence of the PAO1 strain. The Cup systems differ from the Pap or Fim systems, since they obviously lack numbers of genes encoding fimbrial subunits. Nevertheless, the CupA system has been demonstrated to be involved in biofilm formation on solid surfaces, whereas the role of the CupB and CupC systems in biofilm formation could not be clearly elucidated. Moreover, these gene clusters were described as poorly expressed under standard laboratory conditions. ThecupBandcupCclusters are directly under the control of a two-component regulatory system designated RocA1/S1/R. In this study, we revealed that Roc1-dependent induction of thecupBandcupCgenes resulted in a high level of biofilm formation, with CupB and CupC acting with synergy in clustering bacteria for microcolony formation. Very importantly, this phenotype was associated with the assembly of cell surface fimbriae visualized by electron microscopy. Finally, we observed that the CupB and CupC systems are specialized in the assembly of their own fimbrial subunits and are not exchangeable.
Characterization of the Variants, Flanking Genes, and Promoter Activity of the
Leifsonia xyli
subsp.
cynodontis
Insertion Sequence IS
1237
Authors: Lin Hui , Li Tai-Yuan , Xie Mao-Hua , Zhang Yi 
Published: February 2007

ABSTRACT

We performed a comprehensive study of the distribution and function of an insertion sequence (IS) element, IS
1237
, in the genome of
Leifsonia xyli
subsp.
cynodontis
, a useful genetic carrier for expressing beneficial foreign genes in plants. Two shorter IS
1237
isoforms, IS
1237
d1 and IS
1237
d2 resulting from precise deletion between two nonperfect repeats, were found in the bacterial genome at a level that was one-fifth the level of wild-type IS
1237
. Both the genome and native plasmid pCXC100 harbor a truncated toxin-antitoxin cassette that is precisely fused with a 5′-truncated IS
1237
sequence at one nonperfect repeat, indicating that it is a hot site for DNA rearrangement. Nevertheless, no transposition activity was detected when the putative transposase of IS
1237
was overexpressed in
Escherichia coli
. Using thermal asymmetric interlaced PCR, we identified 13 upstream and 10 downstream unique flanking sequences, and two pairs of these sequences were from the same loci, suggesting that IS
1237
has up to 65 unique loci in the
L. xyli
subsp.
cynodontis
chromosome. The presence of TAA or TTA direct repeat sequences at most insertion sites indicated that IS
1237
inserts into the loci by active transposition. IS
1237
showed a high propensity for insertion into other IS elements, such as IS
Lxc1
and IS
Lxc2
, which could offer IS
1237
a nonautonomous transposition pathway through the host IS elements. Interestingly, we showed that IS
1237
has a strong promoter at the 3′ end and a weak promoter at the 5′ end, and both promoters promote the transcription of adjacent genes in different gram-positive bacteria. The high-copy-number nature of IS
1237
and its promoter activity may contribute to bacterial fitness.
Characterization of an Exceedingly Active NADH Oxidase from the Anaerobic Hyperthermophilic Bacterium
Thermotoga maritima
Authors: Yang Xianqin , Ma Kesen 
Published: February 2007

ABSTRACT

An NADH oxidase from the anaerobic hyperthermophilic bacterium
Thermotoga maritima
was purified. The enzyme was very active in catalyzing the reduction of oxygen to hydrogen peroxide with an optimal pH value of 7 at 80°C. The

V
max

was 230 ± 14 μmol/min/mg (
k
cat
/

K
m

= 548,000 min
−1
mM
−1
), and the

K
m

values for NADH and oxygen were 42 ± 3 and 43 ± 4 μM, respectively. The NADH oxidase was a heterodimeric flavoprotein with two subunits with molecular masses of 54 kDa and 46 kDa. Its gene sequences were identified, and the enzyme might represent a new type of NADH oxidase in anaerobes. An NADH-dependent peroxidase with a specific activity of 0.1 U/mg was also present in the cell extract of
T. maritima
.
The DeoR-Type Regulator SugR Represses Expression ofptsGinCorynebacterium glutamicum
Authors: Engels Verena , Wendisch Volker F. 
Published: February 2007

ABSTRACTCorynebacterium glutamicumgrows on a variety of carbohydrates and organic acids. Uptake of the preferred carbon source glucose via the phosphoenolpyruvate-dependent phosphotransferase system (PTS) is reduced during coutilization of glucose with acetate, sucrose, or fructose compared to growth on glucose as the sole carbon source. Here we show that the DeoR-type regulator SugR (NCgl1856) represses expression ofptsG, which encodes the glucose-specific PTS enzyme II. Overexpression ofsugRresulted in reducedptsGmRNA levels, decreased glucose utilization, and perturbed growth on media containing glucose. In mutants lackingsugR, expression of theptsG′-′catfusion was increased two- to sevenfold during growth on gluconeogenic carbon sources but remained similar during growth on glucose or other sugars. As shown by DNA microarray analysis, SugR also regulates expression of other genes, includingptsSand the putative NCgl1859-fruK-ptsFoperon. Purified SugR bound to DNA regions upstream ofptsG,ptsS, and NCgl1859, and a 75-bpptsGpromoter fragment was sufficient for SugR binding. Fructose-6-phosphate interfered with binding of SugR to theptsGpromoter DNA. Thus, while during growth on gluconeogenic carbon sources SugR repressesptsG,ptsGexpression is derepressed during growth on glucose or under other conditions characterized by high fructose-6-phosphate concentrations, representing one mechanism which allowsC. glutamicumto adapt glucose uptake to carbon source availability.
A Complete Set of Flagellar Genes Acquired by Horizontal Transfer Coexists with the Endogenous Flagellar System inRhodobacter sphaeroides
Authors: Poggio Sebastian , Abreu-Goodger Cei , Fabela Salvador , Osorio Aurora , Dreyfus Georges , Vinuesa Pablo , Camarena Laura 
Published: February 2007

ABSTRACTBacteria swim in liquid environments by means of a complex rotating structure known as the flagellum. Approximately 40 proteins are required for the assembly and functionality of this structure.Rhodobacter sphaeroideshas two flagellar systems. One of these systems has been shown to be functional and is required for the synthesis of the well-characterized single subpolar flagellum, while the other was found only after the genome sequence of this bacterium was completed. In this work we found that the second flagellar system ofR. sphaeroidescan be expressed and produces a functional flagellum. In many bacteria with two flagellar systems, one is required for swimming, while the other allows movement in denser environments by producing a large number of flagella over the entire cell surface. In contrast, the second flagellar system ofR. sphaeroidesproduces polar flagella that are required for swimming. Expression of the second set of flagellar genes seems to be positively regulated under anaerobic growth conditions. Phylogenic analysis suggests that the flagellar system that was initially characterized was in fact acquired by horizontal transfer from a γ-proteobacterium, while the second flagellar system contains the native genes. Interestingly, other α-proteobacteria closely related toR. sphaeroideshave also acquired a set of flagellar genes similar to the set found inR. sphaeroides, suggesting that a common ancestor received this gene cluster.