bluuebunny/crossref_metadata_2025_split huggingface dataset:
Prevalence of Virulence Genes and Clonality in
Escherichia coli
Strains That Cause Bacteremia in Cancer Patients
Authors: Hilali Farida , Ruimy Raymond , Saulnier Patrick , Barnabé Christian , Lebouguénec Chantal , Tibayrenc Michel , Andremont Antoine 
Published: July 2002

ABSTRACT

Phenotypic analysis of
Escherichia coli
strains causing bacteremia in cancer patients suggests that they possess specific virulence properties. To investigate this hypothesis, we compared the frequency of the virulence-related genes
cnf1
,
cnf2
,
papC
,
hlyC
, and
iut
in 155
E. coli
strains isolated from hospitalized cancer patients with epidemiologically unrelated cases of bacteremia to their frequency in 70
E. coli
strains isolated from the feces of healthy unrelated volunteers. Of the blood isolates, 24, 37, and 26% were positive for
cnf1
,
papC
, and
hlyC
, respectively, versus only 6, 17, and 6% of the fecal isolates (
P
< 0.05 in all instances). By contrast, 47% of both isolates carried the
iut
gene. The patients' clinical characteristics did not significantly influence these frequencies. The presence on various pathogenicity islands (PAIs) of a combination of the
cnf1
,
papC
, and
hlyC
genes on the chromosome was strongly suggested by Southern blotting of pulsed-field gel electrophoresis (PFGE) patterns with specific DNA probes. The phylogenetic relatedness among 60 strains carrying three, two, one, or no virulence genes and 6 ECOR strains included as references was determined by neighbor joining, the unweighted pair-group method with arithmetic mean, and Wagner analysis of the randomly amplified polymorphic DNA (RAPD) patterns generated by 11 primers. Identification of a major cluster including 96.4% of the strains carrying the
cnf1
,
papC
, and
hlyC
genes and ECOR subgroup B2 strains suggested that the virulent
E. coli
strains causing bacteremia in cancer patients are closely related to ECOR B2 strains. The presence in the
E. coli
population surveyed of a strong linkage disequilibrium, and especially of a highly significant correlation between PFGE and RAPD genetic distances, confirms that clonal propagation has a major impact on the
E. coli
population structure. Nevertheless, low bootstrap values in the phylogenetic tree suggested that frequent genetic exchange inhibits the individualization of discrete genetic lineages, which are stable on an evolutionary scale.
Granulocyte-Macrophage Colony-Stimulating Factor-Deficient Mice Have Impaired Resistance to Blood-Stage Malaria
Authors: Riopel Julie , Tam MiFong , Mohan Karkada , Marino Michael W. , Stevenson Mary M. 
Published: July 2002

ABSTRACTThe contribution of granulocyte-macrophage colony-stimulating factor (GM-CSF), a hematopoietic and immunoregulatory cytokine, to resistance to blood-stage malaria was investigated by infecting GM-CSF-deficient (knockout [KO]) mice withPlasmodium chabaudiAS. KO mice were more susceptible to infection than wild-type (WT) mice, as evidenced by higher peak parasitemia, recurrent recrudescent parasitemia, and high mortality.P. chabaudiAS-infected KO mice had impaired splenomegaly and lower leukocytosis but equivalent levels of anemia compared to infected WT mice. Both bone marrow and splenic erythropoiesis were normal in infected KO mice. However, granulocyte-macrophage colony formation was significantly decreased in these tissues of uninfected and infected KO mice, and the numbers of macrophages in the spleen and peritoneal cavity were significantly lower than in infected WT mice. Serum levels of gamma interferon (IFN-γ) were found to be significantly higher in uninfected KO mice, and the level of this cytokine was not increased during infection. In contrast, IFN-γ levels were significantly above normal levels in infected WT mice. During infection, tumor necrosis factor alpha (TNF-α) levels were significantly increased in KO mice and were significantly higher than TNF-α levels in infected WT mice. Our results indicate that GM-CSF contributes to resistance toP. chabaudiAS infection and that it is involved in the development of splenomegaly, leukocytosis, and granulocyte-macrophage hematopoiesis. GM-CSF may also regulate IFN-γ and TNF-α production and activity in response to infection. The abnormal responses seen in infected KO mice may be due to the lack of GM-CSF during development, to the lack of GM-CSF in the infected mature mice, or to both.
Impaired Pulmonary NF-κB Activation in Response to Lipopolysaccharide in NADPH Oxidase-Deficient Mice
Authors: Koay M. Audrey , Christman John W. , Segal Brahm H. , Venkatakrishnan Annapurna , Blackwell Thomas R. , Holland Steven M. , Blackwell Timothy S. 
Published: July 2002

ABSTRACTReactive oxygen species (ROS) are thought to be involved in intracellular signaling, including activation of the transcription factor NF-κB. We investigated the role of NADPH oxidase in the NF-κB activation pathway by utilizing knockout mice (p47phox−/−) lacking the p47phoxcomponent of NADPH oxidase. Wild-type (WT) controls and p47phox−/−mice were treated with intraperitoneal (i.p.)Escherichia colilipopolysaccharide (LPS) (5 or 20 μg/g of body weight). LPS-induced NF-κB binding activity and accumulation of RelA in nuclear protein extracts of lung tissue were markedly increased in WT compared to p47phox−/−mice 90 min after treatment with 20 but not 5 μg of i.p. LPS per g. In another model of lung inflammation, RelA nuclear translocation was reduced in p47phox−/−mice compared to WT mice following treatment with aerosolized LPS. In contrast to NF-κB activation in p47phox−/−mice, LPS-induced production of macrophage inflammatory protein 2 in the lungs and neutrophilic lung inflammation were not diminished in these mice compared to WT mice. We conclude that LPS-induced NF-κB activation is deficient in the lungs of p47phox−/−mice compared to WT mice, but this abnormality does not result in overt alteration in the acute inflammatory response.
Immunization with theChlamydia trachomatisMouse Pneumonitis Major Outer Membrane Protein Can Elicit a Protective Immune Response against a Genital Challenge
Authors: Pal Sukumar , Theodor Ida , Peterson Ellena M. , de la Maza Luis M. 
Published: July 2002

ABSTRACTInfertility, ectopic pregnancy, and chronic abdominal pain are frequent complications of genital infections withChlamydia trachomatis. In an attempt to produce a vaccine to protect against this pathogen we purified and refolded theC. trachomatismouse pneumonitis (MoPn) major outer membrane protein (MOMP). This preparation, mixed with Freund's adjuvant using vortexing or sonication, was used to immunize BALB/c mice that were subsequently challenged in the upper genital tract. Vaginal cultures were taken on a weekly basis, and mice were mated 6 weeks after the challenge. Gels of the vortexed MOMP showed a predominant band with a molecular size of 62 kDa and weaker bands at 42 and 132 kDa, while the sonicated MOMP had a single band with a molecular size of 42 kDa. Following immunization with these two preparations, strong humoral and cell-mediated immune responses were detected in the mice inoculated with the vortexed MOMP. On the other hand, mice immunized with the sonicated MOMP had a strong humoral immune response but a relatively weak cell-mediated immune response, as determined by a T-cell lymphoproliferative assay and level of cytokine production by splenocytes. Vaginal cultures showed that the mice immunized with the vortexed MOMP were significantly protected, as determined by a decrease in the number of animals with positive cultures, the length of time the mice shed viable organisms, and the number of inclusion-forming units recovered per mouse. Animals immunized with the sonicated MOMP, on the other hand, showed a weaker level of protection based on the same three parameters. After mating, the number of fertile animals and number of embryos per mouse were significantly higher for the mice immunized with vortexed MOMP, but not for the mice immunized with sonicated MOMP, compared to those of the control groups. In conclusion, immunization with a purified and refolded preparation of theC. trachomatisMoPn MOMP confers a significant level of protection in mice against a genital challenge.
Heme Utilization in
Bordetella avium
Is Regulated by RhuI, a Heme-Responsive Extracytoplasmic Function Sigma Factor
Authors: Kirby Amy E. , Metzger Daniel J. , Murphy Erin R. , Connell Terry D. 
Published: July 2002

ABSTRACT

Efficient utilization of heme as an iron (Fe) source by
Bordetella avium
requires
bhuR
, an Fe-regulated gene which encodes an outer membrane heme receptor. Upstream of
bhuR
is a 507-bp open reading frame, hereby designated
rhuI
(for regulator of heme uptake), which codes for a 19-kDa polypeptide. Whereas the 19-kDa polypeptide had homology to a subfamily of alternative sigma factors known as the extracytoplasmic function (ECF) sigma factors, it was hypothesized that
rhuI
encoded a potential in-
trans
regulator of the heme receptor gene in
trans
. Support for the model was strengthened by the identification of nucleotide sequences common to ECF sigma-dependent promoters in the region immediately upstream of
bhuR
. Experimental evidence for the regulatory activities of
rhuI
was first revealed by recombinant experiments in which overproduction of
rhuI
was correlated with a dramatically increased expression of BhuR. A putative
rhuI
-dependent
bhuR
promoter was identified in the 199-bp region located proximal to
bhuR
. When a transcriptional fusion of the 199-bp region and a promoterless
lacZ
gene was introduced into
Escherichia coli
, promoter activity was evident, but only when
rhuI
was coexpressed in the cell. Sigma competition experiments in
E
.
coli
demonstrated that
rhuI
conferred biological properties on the cell that were consistent with RhuI having sigma factor activity. Heme, hemoglobin, and several other heme-containing proteins were shown to be the extracellular inducers of the
rhuI
-dependent regulatory system. Fur titration assays indicated that expression of
rhuI
was probably Fur dependent.
Characterization of Auxotrophic Mutants of
Mycobacterium tuberculosis
and Their Potential as Vaccine Candidates
Authors: Smith Debbie A. , Parish Tanya , Stoker Neil G. , Bancroft Gregory J. 
Published: July 2002

ABSTRACT

Auxotrophic mutants of
Mycobacterium tuberculosis
have been proposed as new vaccine candidates. We have analyzed the virulence and vaccine potential of
M. tuberculosis
strains containing defined mutations in genes involved in methionine (
metB
), proline (
proC
), or tryptophan (
trpD
) amino acid biosynthesis. The
metB
mutant was a prototrophic strain, whereas the
proC
and
trpD
mutants were auxotrophic for proline and tryptophan, respectively. Following infection of murine bone marrow-derived macrophages, H37Rv and the
metB
mutant strain survived intracellularly for over 10 days, whereas over 90% of
proC
and
trpD
mutants were killed during this time. In SCID mice, both H37Rv and the
metB
mutant were highly virulent, with mouse median survival times (MST) of 28.5 and 42 days, respectively. The
proC
mutant was significantly attenuated (MST, 130 days), whereas the
trpD
mutant was essentially avirulent in an immunocompromised host. Following infection of immunocompetent DBA mice with H37Rv, mice survived for a median of 83.5 days and the
metB
mutant now showed a clear reduction in virulence, with two of five infected mice surviving for 360 days. Both
proC
and
trpD
mutants were avirulent (MST of >360 days). In vaccination studies, prior infection with either the
proC
or
trpD
mutant gave protection equivalent (
proC
mutant) to or better (
trpD
mutant) than BCG against challenge with
M. tuberculosis
H37Rv. In summary,
proC
and
trpD
genes are essential for the virulence of
M. tuberculosis
, and mutants with disruptions in either of these genes show strong potential as vaccine candidates.
Comparison of
Vibrio cholerae
Pathogenicity Islands in Sixth and Seventh Pandemic Strains
Authors: Karaolis David K. R. , Lan Ruiting , Kaper James B. , Reeves Peter R. 
Published: July 2002

ABSTRACT

Epidemic
Vibrio cholerae
strains possess a large cluster of essential virulence genes on the chromosome called the
Vibrio
pathogenicity island (VPI). The VPI contains the
tcp
gene cluster encoding the type IV pilus toxin-coregulated pilus colonization factor which can act as the cholera toxin bacteriophage (CTXΦ) receptor. The VPI also contains genes that regulate virulence factor expression. We have fully sequenced and compared the VPI of the seventh-pandemic (El Tor biotype) strain N16961 and the sixth-pandemic (classical biotype) strain 395 and found that the N16961 VPI is 41,272 bp and encodes 29 predicted proteins, whereas the 395 VPI is 41,290 bp. In addition to various nucleotide and amino acid polymorphisms, there were several proteins whose predicted size differed greatly between the strains as a result of frameshift mutations. We hypothesize that these VPI sequence differences provide preliminary evidence to help explain the differences in virulence factor expression between epidemic strains (i.e., the biotypes) of
V. cholerae
.
Local and Systemic Immune Responses to Rectal Administration of Recombinant Cholera Toxin B Subunit in Humans
Authors: Jertborn Marianne , Nordström Inger , Kilander Anders , Czerkinsky Cecil , Holmgren Jan 
Published: July 2002

ABSTRACT
The induction of immune responses to rectally administered recombinant cholera toxin B subunit (CTB) in humans was studied. Three immunizations induced high levels of CTB-specific antibody-secreting cells, particular of the immunoglobulin A isotype, in both rectum and peripheral blood. Antitoxin antibody responses in rectal secretions and serum were also found.
Cloning and Expression of Two Novel Hemin Binding Protein Genes fromTreponema denticola
Authors: Xu Xiaoping , Holt Stanley C. , Kolodrubetz David 
Published: July 2002

ABSTRACTTreponema denticoladoes not appear to produce siderophores, so it must acquire iron by other pathways. Indeed,T. denticolahas been shown to have an iron-regulated 44-kDa outer membrane protein (HbpA) with hemin binding ability. To characterize the HbpA protein, its gene was cloned from genomic DNA libraries ofT. denticola. Sequence analysis of thehbpAopen reading frame indicated that it encoded a 42.8-kDa protein with a 23-amino-acid signal peptide. HbpA has no significant homology to any proteins in the databases. Southern blot analysis demonstrated thathbpAis present in severalT. denticolaATCC strains and clinical isolates, but not inTreponema pectinovorum, Treponema socranskii, orEscherichia coli. HbpA, expressed as a recombinant protein inE. coliand purified by antibody affinity chromatography, has hemin binding activity as determined by lithium dodecyl sulfate-polyacrylamide gel electrophoresis with tetramethylbenzidine staining. Northern blot analysis showed that there were twohbpA-containing transcripts, of approximately 1.3 and 2.6 kb, and that the RNA levels were low-iron induced. Interestingly, the 2.6-kb mRNA also encoded a second protein with significant homology tohbpA. This downstream gene, calledhbpB, was cloned and sequenced and its product was expressed as a fusion protein inE. coli. ThehbpBgene product is 49% identical to HbpA and binds hemin. Thus,T. denticolahas two novel hemin binding proteins which may be part of a previously unrecognized iron acquisition pathway.
Distinct Cytokine Regulation by Cholera Toxin and Type II Heat-Labile Toxins Involves Differential Regulation of CD40 Ligand on CD4+T Cells
Authors: Martin Michael , Metzger Daniel J. , Michalek Suzanne M. , Connell Terry D. , Russell Michael W. 
Published: July 2002

ABSTRACTCholera toxin (CT) and the type II heat-labile enterotoxins (HLT) LT-IIa and LT-IIb act as potent systemic and mucosal adjuvants and induce distinct T-helper (Th)-cell cytokine profiles. In the present study, CT and the type II HLT were found to differentially affect cytokine production by anti-CD3-stimulated human peripheral blood mononuclear cells (PBMC), and the cellular mechanisms responsible were investigated. CT suppressed interleukin-2 (IL-2), tumor necrosis factor alpha (TNF-α), and IL-12 production by PBMC cultures more than either LT-IIa or LT-IIb. CT but not LT-IIa or LT-IIb reduced the expression of CD4+T-cell surface activation markers (CD25 and CD69) and subsequent proliferative responses of anti-CD3-stimulated T cells. CT but not LT-IIa or LT-IIb significantly reduced the expression of CD40 ligand (CD40L) on CD4+T cells. In a coculture system, CT-treated CD4+T cells induced significantly less TNF-α and IL-12 p70 production by both autologous monocytes and monocyte-derived dendritic cells than either LT-IIa- or LT-IIb-treated CD4+T cells. These findings demonstrate that CT, LT-IIa, and LT-IIb differentially affect CD40-CD40L interactions between antigen-presenting cells and T cells and help explain the distinct cytokine profiles observed with type I and type II HLT when used as mucosal adjuvants.
Vibrio cholerae tolC
Is Required for Bile Resistance and Colonization
Authors: Bina James E. , Mekalanos John J. 
Published: July 2002

ABSTRACT

TolC and its homologues are outer membrane proteins that are essential for the transport of many molecules across the cell envelope. In this study we characterized the gene encoding
Vibrio cholerae
TolC.
V. cholerae tolC
mutants failed to secrete the RTX cytotoxin, were hypersensitive to antimicrobial agents, and were deficient in intestinal colonization.
Opsonized VirulentEdwardsiella tardaStrains Are Able To Adhere to and Survive and Replicate within Fish Phagocytes but Fail To Stimulate Reactive Oxygen Intermediates
Authors: Rao Putanae S. Srinivasa , Lim Tit Meng , Leung Ka Yin 
Published: July 2002

ABSTRACTEdwardsiella tardais responsible for hemorrhagic septicemia (edwardsiellosis) in fish and also causes diseases in higher vertebrates such as birds, reptiles, and mammals, including humans. Interactions ofE. tardawith blue gourami phagocytes were studied by light microscopy as well as by adherence, intracellular replication, and superoxide anion assays. Both nonopsonized virulent (PPD130/91 and AL9379) and avirulent (PPD125/87 and PPD76/87) bacteria could adhere to and survive and replicate within phagocytes, while only opsonized virulent strains replicated within the phagocytes. Furthermore, only avirulentE. tardaelicited a higher rate of production of reactive oxygen intermediates (ROIs) by phagocytes, indicating that they were unable to avoid and/or resist reactive oxygen radical-based killing by the fish phagocytes. TnphoAtransposon mutagenesis was used to construct a library of 200 alkaline phosphatase (PhoA+) fusion mutants from a total of 182,000 transconjugants derived fromE. tardaPPD130/91. Five of these mutants induced more ROI production in phagocytes than the wild-type strain. Two mutants had lower replication ability inside phagocytes and moderately higher 50% lethal dose values than the wild-type strain. Sequence analysis revealed that three of these mutants had insertions at sequences having homology to PhoS, dipeptidase, and a surface polymer ligase of lipid A core proteins of other pathogens. These three independent mutations might have changed the cell surface characteristics of the bacteria, which in turn induced phagocytes to produce increased ROIs. Sequences from two other mutants had no homology to known genes, indicating that they may be novel genes for antiphagocytic killing. The present study showed that there are differences in the interactions of virulent and avirulentE. tardaorganisms with fish phagocytes and PhoA+fusion mutants that could be used successfully to identify virulence genes. The information elucidated here would help in the development of suitable strategies to combat the disease caused byE. tarda.
Erysipelothrix rhusiopathiae
YS-1 as a Live Vaccine Vehicle for Heterologous Protein Expression and Intranasal Immunization of Pigs
Authors: Shimoji Yoshihiro , Oishi Eiji , Kitajima Takashi , Muneta Yoshihiro , Shimizu Shinya , Mori Yasuyuki 
Published: July 2002

ABSTRACT

We have developed a system in which a foreign antigen is delivered and expressed on the surface of an attenuated strain of
Erysipelothrix rhusiopathiae
YS-1 and have examined the ability of a such recombinant
E. rhusiopathiae
strain to function as a mucosal vaccine vector. The C-terminal portion, including two repeat regions, R1 and R2, of the P97 adhesin of
Mycoplasma hyopneumoniae
strain E-1 was successfully translocated and expressed on the
E. rhusiopathiae
YS-1 cell surface after it was fused to SpaA.1, a cell surface protective antigen of
E. rhusiopathiae.
BALB/c mice subcutaneously immunized with the
E. rhusiopathiae
recombinant strains developed specific antibodies against SpaA.1 protein and were protected from lethal challenge with the highly virulent homologous
E. rhusiopathiae
Fujisawa-SmR strain, showing the efficacy of this heterologous-antigen expression system as a vaccine against
E. rhusiopathiae
infection. To determine whether protective immune responses are induced in target species, newborn, specific-pathogen-free piglets were immunized intranasally with a recombinant strain designated YS-19. The immunized piglets developed specific anti-SpaA.1 immunoglobulin G (IgG) antibodies in their serum and were protected from death by erysipelas, showing that mucosal vaccination of piglets with YS-19 induces systemic immune responses. Furthermore, YS-19-immunized piglets showed higher levels of P97-specific IgA antibodies in the respiratory tract than did YS-1-immunized piglets. Thus,
E. rhusiopathiae
YS-1 appears to be a promising vaccine vector for mucosal delivery that can induce local and systemic immune responses.