bluuebunny/crossref_metadata_2025 huggingface dataset:
Seroprevalence of Antibodies to Microorganisms Known To Cause Arterial and Myocardial Damage in Patients with or without Coronary Stenosis
Authors: Stöllberger C. , Mölzer G. , Finsterer J.
Published: July 2002

Infections are assumed to play a role in coronary artery disease (CAD) and cardiomyopathies. It is unknown whether the seroprevalence of antibodies to these microorganisms is higher in patients with than without CAD. The seroprevalence of antibodies to
Bartonella henselae, Borrelia burgdorferi, Chlamydia pneumoniae, Coxiella burnetii, Helicobacter pylori
, human granulocytic
Ehrlichia, Leptospira, Rickettsia conorii
, and
Treponema pallidum
was assessed prospectively in patients with exertional dyspnea or anginal chest pain who underwent coronary angiography because of suspected CAD. Patients with normal angiograms (NA) were those in whom no more than 50% stenosis of any coronary artery was found. Patients with CAD were patients who underwent percutaneous transluminal coronary angioplasty. There were 50 patients with CAD (9 female) and 62 with NA (25 female), with a mean age of 62 years. All patients had antibodies to at least one microorganism: to
B. henselae
, 8% of CAD patients and 5% of NA patients; to
B. burgdorferi
IgG, 14% CAD and 6% NA; to
B. burgdorferi
IgM, 6% CAD and 3% NA; to
C. pneumoniae
lipopolysaccharide (LPS) IgA, 76% CAD and 77% NA; to
C. pneumoniae
LPS IgG, 80% CAD and 90% NA; to
C. burnetii
, 0% CAD and 5% NA; to
H. pylori
, 92% CAD and 68% NA; to human granulocytic
Ehrlichia
, 8% CAD and 3% NA; to
Leptospira
IgG, 4% CAD and 2% NA; to
R. conorii
, 10% in both groups; and to
T. pallidum
, 2% CAD and 0% NA. The seroprevalence of antibodies to micro-organisms known to induce arterial and myocardial damage does not differ between patients with CAD and NA.
Inhibition of Ganciclovir-Susceptible and -Resistant Human Cytomegalovirus Clinical Isolates by the Benzimidazole
l
-Riboside 1263W94
Authors: McSharry James J. , McDonough Ann , Olson Betty , Talarico Christine , Davis Michele , Biron Karen K.
Published: July 2002

The average 50% inhibitory concentration (IC
50
) values for AD169 were 0.22 ± 0.09 μM 1263W94 and 5.36 ± 0.12 μM ganciclovir. For 35 human cytomegalovirus (HCMV) clinical isolates the average IC
50
was 0.42 ± 0.09 μM 1263W94, and for 26 ganciclovir-susceptible HCMV clinical isolates the average IC
50
was 3.78 ± 1.62 μM ganciclovir. Nine HCMV clinical isolates that were resistant to ganciclovir were completely susceptible to 1263W94.
Quantitation of Immunoglobulin to Hepatitis E Virus by Enzyme Immunoassay
Authors: Innis Bruce L. , Seriwatana Jitvimol , Robinson Robin A. , Shrestha Mrigendra P. , Yarbough Patrice O. , Longer Charles F. , Scott Robert M. , Vaughn David W. , Myint Khin Saw Aye
Published: July 2002

We developed a quantitative enzyme immunoassay (EIA) for antibody to hepatitis E virus (HEV) by using truncated HEV capsid protein expressed in the baculovirus system to improve seroepidemiology, to contribute to hepatitis E diagnosis, and to enable vaccine evaluations. Five antigen lots were characterized; we used a reference antiserum to standardize antigen potency. We defined Walter Reed antibody units (WR U) with a reference antiserum by using the four-parameter logistic model, established other reference pools as assay standards, and determined the conversion factor: 1 WR U/ml = 0.125 World Health Organization unit (WHO U) per ml. The EIA performed consistently; median intra- and intertest coefficients of variation were 9 and 12%, respectively. The accurate minimum detection limit with serum diluted 1:1,000 was 5.6 WR U/ml; the test could detect reliably a fourfold antibody change. In six people followed from health to onset of hepatitis E, the geometric mean antibody level rose from 7.1 WR U/ml to 1,924.6 WR U/ml. We used the presence of 56- and 180-kDa bands by Western blotting as a confirmatory test and to define true-negative and -positive serum specimens. A receiver-operating characteristics plot identified 30 WR U/ml as an optimum cut-point (sensitivity, 86%; specificity, 89%). The EIA detected antibody more sensitively than a commercially available test. The EIA was transferred to another laboratory, where four operators matched reference laboratory results for a panel of unknowns. Quantitation of antibody to HEV and confirmation of its specificity by Western blotting make HEV serology more meaningful.
Bacterial Immunoglobulin Superantigen Proteins A and L Activate Human Heart Mast Cells by Interacting with Immunoglobulin E
Authors: Genovese Arturo , Bouvet Jean-Pierre , Florio Giovanni , Lamparter-Schummert Bärbel , Björck Lars , Marone Gianni
Published: July 2002

Human heart mast cells (HHMC) have been identified in heart tissue, perivascularly, and in the intima of coronary arteries. In vitro activation of isolated HHMC induces the release of vasoactive and proinflammatory mediators (histamine, tryptase, and cysteinyl leukotriene C4[LTC4]). We investigated the effects of several bacterial proteins on HHMC activation in vitro. HHMC released histamine, tryptase, and LTC4in response toStaphylococcus aureusCowan 1 and the immunoglobulin (Ig)-binding protein A, but not toS. aureusWood 46, which does not synthesize protein A. The effect of protein A was inhibited by preincubation with monoclonal IgM VH3+. Some strains ofPeptostreptococcus magnusexpress an Ig light chain-binding surface protein called protein L. Such bacteria and soluble protein L stimulated the release of preformed and newly synthesized mediators from HHMC. Preincubation of HHMC with either protein A or protein L resulted in complete cross-desensitization to a subsequent challenge with the heterologous stimulus or anti-IgE. Monoclonal IgE (κ chains) blocked protein L-induced release, whereas IgE (λ chains) had no effect. Streptococcal protein G, formyl-containing tripeptide, and pepstatin A did not activate HHMC. Bacterial products protein A and protein L and intact bacteria (S. aureusandP. magnus) activate HHMC by acting as Ig superantigens.
A Region ofPlasmodium falciparumAntigen Pfs25 That Is the Target of Highly Potent Transmission-Blocking Antibodies
Authors: Stowers Anthony W. , Keister David B. , Muratova Olga , Kaslow David C.
Published: July 2002

Each of the four epidermal growth factor (EGF)-like domains of thePlasmodium falciparumsexual-stage antigen Pfs25 has been individually expressed as a yeast-secreted recombinant protein (yEGF1 through yEGF4). All four are recognized by the immune sera of animals and humans vaccinated with TBV25H (the corresponding yeast-secreted full-length recombinant form of Pfs25), with antibody titers to yEGF1 and yEGF2 weakly correlating with the ability of the sera to block the transmission of parasites to the mosquito host. All four proteins are poorly immunogenic in mice vaccinated with aluminum hydroxide-absorbed formulations. However, all four successfully primed the mice to mount an effective secondary antibody response after a single boost with TBV25H. Sera from mice vaccinated with yEGF2-TBV25H completely block the development of oocysts in mosquito midguts in membrane-feeding assays. Further, of the four proteins, only the depletion of antibodies to yEGF2 from the sera of rabbits vaccinated with TBV25H consistently abolished the ability of those sera to block oocyst development. Thus, antibodies to the second EGF-like domain of Pfs25 appear to mediate a very potent blocking activity, even at low titers. Vaccination strategies that target antibody response towards this domain may improve the efficacy of future transmission-blocking vaccines.
Characterization of Bactericidal Immune Responses following Vaccination with Acellular Pertussis Vaccines in Adults
Authors: Weingart Christine L. , Keitel Wendy A. , Edwards Kathryn M. , Weiss Alison A.
Published: July 2002

Sera from six adults, collected before and after acellular pertussis vaccination, and from a placebo control were examined for the ability to elicit two bactericidal immune defenses, (i) antibody-dependent complement-mediated bacterial lysis and (ii) opsonization and phagocytosis by human neutrophils. The samples were chosen based on low preimmunization titers and strong postimmunization responses to various combinations of vaccine antigens. All but two prevaccination samples demonstrated activity indicative of complement-mediated lysis. Preimmunization activity could have been due to prior infection or childhood immunization. Immunization did not result in improved bactericidal activity for any of the individuals, and in two cases immunization caused a statistically significant decrease in complement-mediated lysis. Similarly, opsonization with the postimmunization sera failed to enhance attachment or phagocytosis of bacteria by neutrophils, and one postimmunization sample with a strong response to filamentous hemagglutinin caused an inhibition of phagocytosis that was statistically significant compared to that observed for the no-serum control. In summary, booster immunization of adults with acellular pertussis vaccines was not found to increase bactericidal activity over preimmunization levels. Identifying ways to promote bactericidal immune responses might improve the efficacy of acellular pertussis vaccines.
Metalloprotease Activity in a Small Heat Shock Protein of the Human Malaria Parasite
Plasmodium vivax
Authors: Fakruddin J. Mohamed , Biswas Sukla , Sharma Yagya D.
Published: July 2002

The malaria parasite affects millions of people each year, lives and multiplies in two different hosts, and synthesizes a large number of proteases and heat shock proteins (HSPs) for its survival. We describe here the characterization of a metalloprotease activity which resides in the small HSP (PVHSP28) of the common but noncultivable human malaria parasite
Plasmodium vivax
. The protein is expressed by erythrocytic stages of the parasite. It is expressed as a ∼55-kDa polypeptide which is then processed to the 28-kDa mature protein. The latter was found to be an active protease in gelatin zymography. This protease showed its optimal activity at 37°C (pH 7.6). It also retained its proteolytic activity at higher temperatures of up to 55°C. The enzyme belongs to the metalloprotease class, as its proteolytic activity was most effectively blocked by 1,10-phenanthroline and was restored to a maximal level by the addition of zinc metal ions. Inhibitors for the cysteine, serine, and aspartate classes of proteases were ineffective against this enzyme. A homology search indicates that PVHSP28 probably belongs to a new class of HSPs which possess the metalloprotease signature sequence.
Pathogenic Mycobacteria Disrupt the Macrophage Actin Filament Network
Authors: Guérin Isabelle , de Chastellier Chantal
Published: July 2002

Phagosomes with pathogenic mycobacteria retain fusion and intermingling characteristics of early endosomes indefinitely. The time course of acquisition of newly endocytosed tracers becomes, however, atypical (lag instead of immediate acquisition) starting from day 1 postinfection (p.i.), thereby suggesting that additional factors affect this process. Disruption of the actin filament (F-actin) network by cytochalasin D perturbs the movement of early endosomes and probably fusion events among early endosomes and phagosomes. Here we compare, by immunofluorescence microscopy, the morphology and distribution of F-actin in macrophages infected with virulentMycobacterium avium, in uninfected macrophages, or in macrophages after phagocytosis of nonpathogenic bacteria (Mycobacterium smegmatisorBacillus subtilis) or hydrophobic latex particles. In uninfected cells, F-actin appeared as a network of small filaments distributed throughout the cell; about 80% of the cells also displayed one or two small patches of F-actin at the cell periphery. VirulentM. aviumcaused a marked disorganization of the F-actin network starting from day 1 p.i. The most salient features were the formation of several large patches, the progressive disappearance of the small filaments, and the appearance of large numbers of tiny punctate structures starting from day 2 p.i. With the three other particles, the F-actin network was unmodified compared to that in uninfected cells. The atypical lag in acquisition of newly endocytosed tracers byM. avium-containing phagosomes, therefore, seems to coincide with the disorganization of the F-actin network.
Serum Immunoglobulin G (IgG) and IgG Subclass Responses to the RgpA-Kgp Proteinase-Adhesin Complex ofPorphyromonas gingivalisin Adult Periodontitis
Authors: O'Brien-Simpson Neil M. , Black Catherine L. , Bhogal Peter S. , Cleal Steven M. , Slakeski Nada , Higgins Thomas J. , Reynolds Eric C.
Published: July 2002

Serum immunoglobulin G (IgG), IgM, and IgG subclass responses to the RgpA-Kgp proteinase-adhesin complex ofPorphyromonas gingivaliswere examined by enzyme-linked immunosorbent assay using adult periodontitis patients and age- and sex-matched controls. Twenty-five sera from subjects with adult periodontitis (diseased group) and 25 sera from healthy subjects (control group) were used for the study. Sera and subgingival plaque samples from 10 sites were collected from each patient at the time of clinical examination. The level ofP. gingivalisin the plaque samples was determined using a DNA probe. Highly significant positive associations between the percentage of sites positive forP. gingivalisand measures of disease severity (mean pocket depth, mean attachment loss, and percentage of sites that bled on probing) were found. The diseased group had significantly higher specific IgG responses to the RgpA-Kgp complex than did the control group, and the responses were significantly associated with mean probing depths and percentage of sites positive forP. gingivalis. Analysis of the IgG subclass responses to the RgpA-Kgp complex revealed that the subclass distribution for both the diseased and control groups was IgG4 > IgG2 > IgG3 = IgG1. The IgG2 response to the complex was positively correlated with mean probing depth, whereas the IgG4 response was negatively correlated with this measure of disease severity. Immunoblot analysis of the RgpA-Kgp complex showed that sera from healthy subjects and those with low levels of disease, with high IgG4 and low IgG2 responses, reacted with the RgpA27, Kgp39, and RgpA44 adhesins; however, sera from diseased subjects with low IgG4 and high IgG2 responses reacted only with the RgpA44 and/or Kgp44 adhesins. Epitope mapping of the RgpA27 adhesin localized a major epitope recognized by IgG4 antibodies in sera from subjects with high IgG4 and low IgG2 responses to the RgpA-Kgp complex which was not recognized by sera from diseased subjects with low IgG4 and high IgG2 responses.
Passive Transfer of Antiserum Specific for Immunogens Derived from a Nontypeable
Haemophilus influenzae
Adhesin and Lipoprotein D Prevents Otitis Media after Heterologous Challenge
Authors: Kennedy Bobbie-Jo , Novotny Laura A. , Jurcisek Joseph A. , Lobet Yves , Bakaletz Lauren O.
Published: July 2002

We recently determined that passive transfer of serum directed against a synthetic peptide called LB1 or a recombinant fusion protein immunogen [LPD-LB1(f)
2,1,3
] could prevent otitis media after challenge with a homologous nontypeable
Haemophilus influenzae
(NTHI) isolate. NTHI residing in the nasopharynx was rapidly cleared from this site, thus preventing it from ascending the eustachian tube and inducing otitis media in chinchillas compromised by an ongoing viral upper respiratory tract infection. While LB1 is based solely on one NTHI adhesin, the latter immunogen, LPD-LB1(f)
2,1,3
, was designed to incorporate two NTHI antigens shown to play a role in the pathogenesis of otitis media; lipoprotein D (LPD) and the P5-homologous fimbrin adhesin. The design of LPD-LB1(f)
2,1,3
also accommodated for the recently demonstrated existence of three major groupings, based on amino acid sequence diversity, in the third surface-exposed region of P5-fimbrin. LPD-LB1(f)
2,1,3
was thus designed to potentially confer broader protection against challenge by diverse strains of NTHI. Chinchillas were passively immunized here with serum specific for either LB1 or for LPD-LB1(f)
2,1,3
prior to challenge with a member of all three groups of NTHI relative to diversity in region 3. The transferred serum pools were also analyzed for titer, specificity, and several functional activities. We found that both serum pools had equivalent ability to mediate C′-dependent killing and to inhibit adherence of NTHI strains to human oropharyngeal cells. When passively transferred, both serum pools significantly inhibited the signs and incidence of otitis media (
P
≤ 0.01) induced by any of the three challenge isolates. Despite providing protection against disease, the ability of these antisera to induce total eradication of NTHI from the nasopharynx was not equivalent among NTHI groups. These data thus suggested that while early, complete eradication of NTHI from the nasopharynx was highly protective, reduction of the bacterial load to below a critical threshold level appeared to be similarly effective.
Identification of a Gene within a Pathogenicity Island of Enterotoxigenic
Escherichia coli
H10407 Required for Maximal Secretion of the Heat-Labile Enterotoxin
Authors: Fleckenstein James M. , Lindler Luther E. , Elsinghorst Eric A. , Dale James B.
Published: July 2002

Studies of the pathogenesis of enterotoxigenic
Escherichia coli
(ETEC) have largely centered on extrachromosomal determinants of virulence, in particular the plasmid-encoded heat-labile (LT) and heat-stable enterotoxins and the colonization factor antigens. ETEC causes illnesses that range from mild diarrhea to severe cholera-like disease. These differences in disease severity are not readily accounted for by our current understanding of ETEC pathogenesis. Here we demonstrate that Tia, a putative adhesin of ETEC
H10407
, is encoded on a large chromosomal element of approximately 46 kb that shares multiple features with previously described
E. coli
pathogenicity islands. Further analysis of the region downstream from
tia
revealed the presence of several candidate open reading frames (ORFs) in the same transcriptional orientation as
tia
. The putative proteins encoded by these ORFs bear multiple motifs associated with bacterial secretion apparatuses. An in-frame deletion in one candidate gene identified here as
leoA
(labile enterotoxin output) resulted in marked diminution of secretion of the LT enterotoxin and lack of fluid accumulation in a rabbit ileal loop model of infection. Although previous studies have suggested that
E. coli
lacks the capacity to secrete LT, our studies show that maximal release of LT from the periplasm of
H10407
is dependent on one or more elements encoded on a pathogenicity island.
Antigenic Equivalence of Human T-Cell Responses to
Mycobacterium tuberculosis
-Specific RD1-Encoded Protein Antigens ESAT-6 and Culture Filtrate Protein 10 and to Mixtures of Synthetic Peptides
Authors: Arend Sandra M. , Geluk Annemieke , van Meijgaarden Krista E. , van Dissel Jaap T. , Theisen Michael , Andersen Peter , Ottenhoff Tom H. M.
Published: July 2002

The early secreted antigenic target 6-kDa protein (ESAT-6) and culture filtrate protein 10 (CFP-10) are promising antigens for reliable immunodiagnosis of tuberculosis. Both antigens are encoded by RD1, a genomic region present in all strains of
Mycobacterium tuberculosis
and
M. bovis
but lacking in all
M. bovis
bacillus Calmette-Guérin vaccine strains. Production and purification of recombinant antigens are laborious and costly, precluding rapid and large-scale testing. Aiming to develop alternative diagnostic reagents, we have investigated whether recombinant ESAT-6 (rESAT-6) and recombinant CFP-10 (rCFP-10) can be replaced with corresponding mixtures of overlapping peptides spanning the complete amino acid sequence of each antigen. Proliferation of
M. tuberculosis
-specific human T-cell lines in response to rESAT-6 and rCFP-10 and that in response to the corresponding peptide mixtures were almost completely correlated (
r
= 0.96,
P
< 0.0001 for ESAT-6;
r
= 0.98,
P
< 0.0001 for CFP-10). More importantly, the same was found when gamma interferon production by peripheral blood mononuclear cells in response to these stimuli was analyzed (
r
= 0.89,
P
< 0.0001 for ESAT-6;
r
= 0.89,
P
< 0.0001 for CFP-10). Whole protein antigens and the peptide mixtures resulted in identical sensitivity and specificity for detection of infection with
M. tuberculosis
. The peptides in each mixture contributing to the overall response varied between individuals with different HLA-DR types. Interestingly, responses to CFP-10 were significantly higher in the presence of HLA-DR15, which is the major subtype of DR2. These results show that mixtures of synthetic overlapping peptides have potency equivalent to that of whole ESAT-6 and CFP-10 for sensitive and specific detection of infection with
M. tuberculosis
, and peptides have the advantage of faster production at lower cost.
Investigation of the Role of Type IV
Aeromonas
Pilus (Tap) in the Pathogenesis of
Aeromonas
Gastrointestinal Infection
Authors: Kirov Sylvia M. , Barnett Timothy C. , Pepe Cynthia M. , Strom Mark S. , Albert M. John
Published: July 2002

Although there is substantial evidence that type IV pili purified from diarrhea-associated
Aeromonas
species (designated Bfp for bundle-forming pilus) are intestinal colonization factors (S. M. Kirov, L. A. O'Donovan, and K. Sanderson, Infect. Immun. 67:5447–5454, 1999), nothing is known regarding the function of a second family of
Aeromonas
type IV pili (designated Tap for type IV
Aeromonas
pilus), identified following the cloning of a pilus biogenesis gene cluster
tapABCD
. Related pilus gene clusters are widely conserved among gram-negative bacteria, but their significance for virulence has been controversial. To investigate the role of Tap pili in
Aeromonas
pathogenesis, mutants of
Aeromonas
strains (a fish isolate of
A. hydrophila
and a human dysenteric isolate of
A. veronii
bv. sobria) were prepared by insertional inactivation of the
tapA
gene which encodes the type IV pilus subunit protein, TapA. Exotoxic activities were unaffected by the mutation in
tapA
. Inactivation of
tapA
had no effect on the bacterial adherence of these two isolates to HEp-2 cells. For the
A. veronii
bv. sobria isolate, adhesion to Henle 407 intestinal cells and to human intestinal tissue was also unaffected. There was no significant effect on the duration of colonization or incidence of diarrhea when the
A. veronii
bv. sobria strain was tested in the removable intestinal tie adult rabbit diarrhea model or on its ability to colonize infant mice. Evidence was obtained that demonstrated that TapA was expressed by both
Aeromonas
species and was present on the cell surface, although if assembled into pili this pilus type appears to be an uncommon one under standard bacterial growth conditions. Further studies into factors which may influence Tap expression are required, but the present study suggests that Tap pili may not be as significant as Bfp pili for
Aeromonas
intestinal colonization.
Comparative Genomics ofHelicobacter pylori: Analysis of the Outer Membrane Protein Families
Authors: Alm Richard A. , Bina James , Andrews Beth M. , Doig Peter , Hancock Robert E. W. , Trust Trevor J.
Published: July 2002

The two complete genomic sequences ofHelicobacter pyloriJ99 and 26695 were used to compare the paralogous families (related genes within one genome, likely to have related function) of genes predicted to encode outer membrane proteins which were present in each strain. We identified five paralogous gene families ranging in size from 3 to 33 members; two of these families contained members specific for eitherH. pyloriJ99 orH. pylori26695. Most orthologous protein pairs (equivalent genes between two genomes, same function) shared considerable identity between the two strains. The unusual set of outer membrane proteins and the specialized outer membrane may be a reflection of the adaptation ofH. pylorito the unique gastric environment where it is found. One subfamily of proteins, which contains both channel-forming and adhesin molecules, is extremely highly related at the sequence level and has likely arisen due to ancestral gene duplication. In addition, the largest paralogous family contained two essentially identical pairs of genes in both strains. The presence and genomic organization of these two pairs of duplicated genes were analyzed in a panel of independentH. pyloriisolates. While one pair was present in every strain examined, one allele of the other pair appeared partially deleted in several isolates.
Purification of Protease from a Mixture of Exfoliative Toxin and Newborn-Mouse Epidermis
Authors: Ninomiya Junya , Ito Yayoi , Takiuchi Iwao
Published: July 2002

Although the role of exfoliative toxin in staphylococcal scalded-skin syndrome has been suggested to be that of a serine protease, it has not been demonstrated to show proteolytic activity. Our purpose was to purify a proteolytic enzyme from a mixture of exfoliative toxin and newborn-mouse epidermis. We used gel filtration and ion-exchange and hydroxyapatite chromatography with a high-pressure liquid chromatography system. A casein-hydrolyzing enzyme was isolated from the mixture. The molecular mass of the enzyme was confirmed to be 20 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Subcutaneous injection of the purified enzyme into newborn mice reproduced the epidermal splitting that is seen in staphylococcal scalded-skin syndrome. These results suggest that exfoliative toxin does not work as a protease itself but that some reaction between exfoliative toxin and an epidermal component(s) first produces a protease, after which epidermal splitting occurs.
Soluble CD14 Enhances Membrane CD14-Mediated Responses to Peptidoglycan: Structural Requirements Differ from Those for Responses to Lipopolysaccharide
Authors: Dziarski Roman , Viriyakosol Suganya , Kirkland Theo N. , Gupta Dipika
Published: July 2002

The purpose of this study was to identify the functional significance of the binding of soluble CD14 (sCD14) to bacterial peptidoglycan (PGN) and to compare the structural requirements of sCD14 for the binding to PGN and lipopolysaccharide (LPS) and for sCD14-mediated enhancement of PGN- and LPS-induced cell responses. sCD14 did not facilitate the responses of membrane CD14 (mCD14)-negative pre-B 70Z/3 cells to PGN, although it facilitated the responses of these cells to LPS and although mCD14 facilitated the responses of 70Z/3 cells to PGN. sCD14 enhanced mCD14-mediated cell activation by both PGN and LPS, but only the responses to LPS, and not to PGN, were enhanced by LPS-binding protein. Four 4- or 5-amino-acid-long sequences within the 65-amino-acid N-terminal region of sCD14 were needed for binding to both PGN and LPS and for enhancement of cell activation by both PGN and LPS. However, deletions of individual sequences had different effects on the ability of sCD14 to bind to PGN and to LPS and on the ability to enhance the responses to PGN and to LPS. Thus, there are different structural requirements of sCD14 for binding to PGN and to LPS and for the enhancement of PGN- and LPS-induced cell activation.
Cytokine Responses toTreponema pectinovorumandTreponema denticolain Human Gingival Fibroblasts
Authors: Nixon Connie S. , Steffen Michelle J. , Ebersole Jeffrey L.
Published: July 2002

Human gingival fibroblasts were challenged withTreponema pectinovorumandTreponema denticolato test three specific hypotheses: (i) these treponemes induce different cytokine profiles from the fibroblasts, (ii) differences in cytokine profiles are observed after challenge with live versus killed treponemes, and (iii) differences in cytokine profiles are noted from different gingival fibroblast cell lines when challenged with these treponemes. Three normal gingival fibroblast cell cultures were challenged withT. pectinovorumandT. denticolastrains, and the supernatants were analyzed for cytokine production (i.e., interleukin-1α [IL-1α], IL-1β, IL-6, IL-8, IL-10, gamma interferon, macrophage chemotactic protein 1 [MCP-1], platelet-derived growth factor, tumor necrosis factor alpha, and granulocyte-macrophage colony-stimulating factor). Unstimulated fibroblast cell lines produced IL-6, IL-8, and MCP-1.T. pectinovorumroutinely elicited the greatest production of these cytokines from the fibroblast cell lines, increasing 10- to 50-fold over basal production. WhileT. denticolaalso induced IL-6 and IL-8 production, these levels were generally lower than those elicited by challenge withT. pectinovorum. MCP-1 levels were significantly lower afterT. denticolachallenge, and the kinetics suggested that this microorganism actually inhibited basal production by the fibroblasts. No basal or stimulated production of the other cytokines was observed. Significant differences were noted in the responsiveness of the various cell lines with respect to the two species of treponemes and the individual cytokines produced. Finally, deadT. pectinovorumgenerally induced a twofold-greater level of IL-6 and IL-8 than the live bacteria. These results supported the idea that different species of oral treponemes can elicit proinflammatory cytokine production by gingival cells and that this stimulation did not require live microorganisms. Importantly, a unique difference was noted in the ability ofT. pectinovorumto induce a robust MCP-1 production, whileT. denticolaappeared to inhibit this activity of the fibroblasts. While the general cytokine profiles of the fibroblast cell cultures were similar, significant differences were noted in the quantity of individual cytokines produced, which could relate to individual patient variation in local inflammatory responses in the periodontium.
Immunity againstHelicobacter pylori: Significance of Interleukin-4 Receptor α Chain Status and Gender of Infected Mice
Authors: Aebischer Toni , Laforsch Stephanie , Hurwitz Robert , Brombacher Frank , Meyer Thomas F.
Published: July 2002

Vaccination of interleukin-4 (IL-4) receptor α (IL-4Rα) chain-deficient BALB/c mice withHelicobacter pyloriurease and cholera toxin or with urease-expressing, live attenuatedSalmonella entericaserovar Typhimurium cells revealed that protection againstH. pyloriinfection is independent of IL-4- or IL-13-mediated signals. A comparison of male and female mice suggests a sexual dimorphism in the extent of bacterial colonization that is particularly evident in the absence of the IL-4Rα chain.
Essential Role for Estrogen in Protection against
Vibrio vulnificus
-Induced Endotoxic Shock
Authors: Merkel Sandra M. , Alexander Sarah , Zufall Eric , Oliver James D. , Huet-Hudson Yvette M.
Published: July 2002

Little is known about the underlying mechanisms that result in a sexually dimorphic response to
Vibrio vulnificus
endotoxic shock.
V. vulnificus
is a gram-negative bacterium, considered one of the most invasive and rapidly fatal human pathogens known. However, 85% of individuals that develop endotoxic shock from
V. vulnificus
are males. Using the rat, we have developed a model for
V. vulnificus
endotoxic shock that mimics the sexually dimorphic response in humans. Gonadectomy in females results in increased mortality, and estrogen replacement results in decreased mortality in both gonadectomized males and females. These results demonstrate that estrogen is providing protection against
V. vulnificus
lipopolysaccharide-induced endotoxic shock.
Assessment of Internalization and Viability of
Porphyromonas gingivalis
in KB Epithelial Cells by Confocal Microscopy
Authors: Houalet-Jeanne S. , Pellen-Mussi P. , Tricot-Doleux S. , Apiou J. , Bonnaure-Mallet M.
Published: July 2002

Porphyromonas gingivalis (P. gingivalis)
is considered to be one of the main periodontal pathogens. The goal of this work was to confirm the ability of
P. gingivalis
to invade host cells. We detected
P. gingivalis
inside KB cells by confocal microscopy and analyzed the various aspects of the adherence and internalization process. Lysates of
P. gingivalis
-infected KB cells were also examined using anaerobic growth techniques. The results showed the viability and ability to replicate, inside the host cells, of the internalized pathogen. The production of vesicles was also tracked for the first time. Confocal microscopy revealed
P. gingivalis
in a perinuclear position.
Staphylococcus caprae
Strains Carry Determinants Known To Be Involved in Pathogenicity: a Gene Encoding an Autolysin-Binding Fibronectin and the
ica
Operon Involved in Biofilm Formation
Authors: Allignet Jeanine , Aubert Sylvie , Dyke Keith G. H. , El Solh Nevine
Published: July 2002

The
atlC
gene (1,485 bp), encoding an autolysin which binds fibronectin, and the
ica
operon, involved in biofilm formation, were isolated from the chromosome of an infectious isolate of
Staphylococcus caprae
and sequenced. AtlC (155 kDa) is similar to the staphylococcal autolysins Atl, AtlE, Aas (48 to 72% amino acid identity) and contains a putative signal peptide of 29 amino acids and two enzymatic centers (
N
-acetylmuramoyl-
l
-alanine amidase and endo-β-
N
-acetylglucosaminidase) interconnected by three imperfect fibronectin-binding repeats. The glycine-tryptophan (GW) motif found in the central and end part of each repeat may serve for cell surface anchoring of AtlC as they do in
Listeria monocytogenes
. The
S. caprae ica
operon contains four genes closely related to
S. epidermidis
and
S. aureus icaA
,
icaB
,
icaC
, and
icaD
genes (≥ 68% similarity) and is preceded by a gene similar to
icaR
(≥70% similarity). The polypeptides deduced from the
S. caprae ica
genes exhibit 67 to 88% amino acid identity to those of
S. epidermidis
and
S. aureus ica
genes. The
ica
operon and
icaR
gene were analyzed in 14
S. caprae
strains from human specimens or goats' milk. Some of the strains produced biofilm, and others did not. All strains carry the
ica
operon and
icaR
of the same sizes and in the same relative positions, suggesting that the absence of biofilm formation is not related to the insertion of a mobile element such as an insertion sequence or a transposon.
Immunochemical and Biological Characterization of Three Capsular Polysaccharides from a Single
Bacteroides fragilis
Strain
Authors: Kalka-Moll Wiltrud M. , Wang Ying , Comstock L. E. , Gonzalez Sylvia E. , Tzianabos Arthur O. , Kasper Dennis L.
Published: July 2002

Although
Bacteroides fragilis
accounts for only 0.5% of the normal human colonic flora, it is the anaerobic species most frequently isolated from intra-abdominal and other infections with an intestinal source. The capsular polysaccharides of
B. fragilis
are part of a complex of surface polysaccharides and are the organism's most important virulence factors in the formation of intra-abdominal abscesses. Two capsular polysaccharides from strain NCTC 9343, PS A1 and PS B1, have been characterized structurally. Their most striking feature is a zwitterionic charge motif consisting of both positively and negatively charged substituent groups on each repeating unit. This zwitterionic motif is essential for abscess formation. In this study, we sought to elucidate structural features of the capsular polysaccharide complex of a commonly studied
B. fragilis
strain, 638R, that is distinct from strain 9343. We sought a more general picture of the species to establish basic structure-activity and structure-biosynthesis relationships among abscess-inducing polysaccharides. Strain 638R was found to have a capsular polysaccharide complex from which three distinct carbohydrates could be isolated by a complex purification procedure. Compositional and immunochemical studies demonstrated a zwitterionic charge motif common to all of the capsular polysaccharides that correlated with their ability to induce experimental intra-abdominal abscesses. Of interest is the range of net charges of the isolated polysaccharides—from positive (PS C2) to balanced (PS A2) to negative (PS 3). Relationships among structural components of the zwitterionic polysaccharides and their molecular biosynthesis loci were identified.
In Vivo Dynamics of Type 1 Fimbria Regulation in Uropathogenic
Escherichia coli
during Experimental Urinary Tract Infection
Authors: Gunther Nereus W. , Lockatell Virginia , Johnson David E. , Mobley Harry L. T.
Published: July 2002

Escherichia coli
is the primary cause of uncomplicated infections of the urinary tract including cystitis. More serious infections, characterized as acute pyelonephritis, can also develop. Type 1 fimbriae of
E. coli
contribute to virulence in the urinary tract; however, only recently has the expression of the type 1 fimbriae been investigated in vivo using molecular techniques. Transcription of type 1 fimbrial genes is controlled by a promoter that resides on a 314-bp invertible element capable of two orientations. One places the promoter in the ON orientation, allowing for transcription; the other places the promoter in the OFF orientation, preventing transcription. A PCR-based assay was developed to measure the orientation of the invertible element during an experimental urinary tract infection in mice. Using this assay, it was found that the percentage of the population ON in urine samples correlated with the respective CFU per gram of bladder (
P
= 0.0006) but not with CFU per gram of kidney (
P
> 0.069). Cystitis isolates present in the urine of mice during the course of infection had a higher percentage of their invertible elements in the ON orientation than did pyelonephritis isolates (85 and 34%, respectively, at 24 h;
P
< 0.0001). In general, cystitis isolates, unlike pyelonephritis isolates, were more likely to maintain their invertible elements in the ON orientation for the entire period of infection.
E. coli
cells expressing type 1 fimbriae, expelled in urine, were shown by scanning electron microscopy to be densely packed on the surface of uroepithelial cells. These results suggest that expression of type 1 fimbriae is more critical for cystitis strains than for pyelonephritis strains in the early stages of an infection during bladder colonization.
Gamma Interferon Is Not Required for Arthritis Resistance in the Murine Lyme Disease Model
Authors: Glickstein Lisa , Edelstein Meg , Dong Jay Zengjun
Published: July 2002

Lyme arthritis is the most common complication following infection of human individuals withBorrelia burgdorferisensu stricto. In mice,B. burgdorferiinfection leads to arthritis of the tibiotarsal joints. Arthritis severity in mice is under host genetic control, as BALB/c mice developed mild arthritis but C3H/He mice developed severe disease followingB. burgdorferiinfection. To study the role of gamma interferon (IFN-γ) in arthritogenesis, targeted mutant mice lacking the IFN-γ receptor (IFN-γR) were infected by inoculation withB. burgdorferi. IFN-γR−/−and parental 129/SvEv mice developed mild arthritis of similar severity, as determined both by weekly tibiotarsal joint measurements and histopathology at 2 and 5 weeks postinfection. Both strains of mice had the same spirochetal burden in the joints, suggesting that the IFN-γR−/−mice were not impaired in controlling spirochetal expansion in vivo. The wild-type mice mounted a Th1 response, with a predominance of CD4+IFN-γ+T cells observed by flow cytometry. In contrast, the IFN-γR−/−mice mounted a Th2 response, with a predominance of CD4+IL-4+T cells. As expected given their cytokine profile, the IFN-γR−/−mice produced fewer CD8+IFN-γ+and MAC-1+IL-12+cells and less immunoglobulin G2a (IgG2a) than their wild-type counterparts. These results strongly suggest that IFN-γ is not required for arthritis resistance or as part of an effective immune response againstB. burgdorferi.
Mapping the Ligand-Binding Region of
Borrelia burgdorferi
Fibronectin-Binding Protein BBK32
Authors: Probert William S. , Kim Jung Hwa , Höök Magnus , Johnson Barbara J. B.
Published: July 2002

The cellular attachment and entry of pathogenic microorganisms can be facilitated by the expression of microbial adhesins that bind fibronectin. We have previously described a
Borrelia burgdorferi
gene,
bbk32
, that encodes a 47-kDa fibronectin-binding protein. In this study, the ligand-binding region of BBK32 from
B. burgdorferi
isolate B31 was localized to 32 amino acids. The
bbk32
gene was cloned and sequenced from three additional
B. burgdorferi
isolates representing different genospecies of
B. burgdorferi
sensu lato. All four
bbk32
genes encoded proteins having fibronectin-binding activity when expressed in
Escherichia coli
, and the deduced proteins shared 81 to 91% amino acid sequence identity within the ligand-binding domain. In addition, the ligand-binding region of BBK32 was found to share sequence homology with a fibronectin-binding peptide defined for protein F1 of
Streptococcus pyogenes
. The structural and functional similarity between the ligand-binding region of BBK32 and the UR region of protein F1 suggests a common mechanism of cellular adhesion and entry for
B. burgdorferi
and
S. pyogenes
.
Establishing a Direct Role for the
Bartonella bacilliformis
Invasion-Associated Locus B (IalB) Protein in Human Erythrocyte Parasitism
Authors: Coleman Sherry A. , Minnick Michael F.
Published: July 2002

The invasion-associated locus A and B genes (
ialAB
) of
Bartonella bacilliformis
were previously shown to confer an erythrocyte-invasive phenotype upon
Escherichia coli
, indirectly implicating their role in virulence. We report the first direct demonstration of a role for
ialB
as a virulence factor in
B. bacilliformis
. The presence of a secretory signal sequence and amino acid sequence similarity to two known outer membrane proteins involved in virulence suggested that IalB was an outer membrane protein. To develop an antiserum for protein localization, the
ialB
gene was cloned in frame into an expression vector with a six-histidine tag and under control of the
lacZ
promoter. The IalB fusion protein was purified by nickel affinity chromatography and used to raise polyclonal antibodies. IalB was initially localized to the bacterial membrane fraction. To further localize IalB,
B. bacilliformis
inner and outer membranes were fractionated by sucrose density gradient centrifugation and identified by appearance, buoyant density (ρ), and cytochrome
b
content. Inner and outer membrane proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and IalB was positively identified by Western blot. Contrary to expectations, IalB was localized to the inner membrane of the pathogen. To directly demonstrate a role for IalB in erythrocyte parasitism, the
B. bacilliformis ialB
gene was disrupted by insertional mutagenesis. The resulting
ialB
mutant strain was complemented in
trans
with a replicative plasmid encoding the full-length
ialB
gene. PCR and high-stringency DNA hybridization confirmed mutagenesis and transcomplementation events. Abrogation and restoration of
ialB
expression was verified by SDS-PAGE and immunoblotting. In vitro virulence assays showed that mutagenesis of
ialB
decreased bacterial association and invasion of human erythrocytes by 47 to 53% relative to controls. Transcomplementation of
ialB
restored erythrocyte association and invasion rates to levels observed in the parental strain. These data provide direct evidence for IalB's role in erythrocyte parasitism and represent the first demonstration of molecular Koch's postulates for a
Bartonella
species.
Effect of Low-Dose Antigen Exposure on Development of Immunity to
Helicobacter pylori
Infection in Mice
Authors: Radcliff Fiona J. , Ferrero Richard L.
Published: July 2002

The effect of low-dose antigen exposure on the development of immunity to
Helicobacter pylori
infection was studied in outbred mice. Animals that were primed with a subinfectious number of
H. pylori
bacteria exhibited significantly lower bacterial loads after challenge with an infectious dose of pathogen (versus controls,
P
< 0.05).
Toxin Levels in Serum Correlate with the Development of Staphylococcal Scalded Skin Syndrome in a Murine Model
Authors: Plano Lisa R. W. , Adkins Becky , Woischnik Markus , Ewing Ruth , Collins Carleen M.
Published: July 2002

Staphylococcal scalded skin syndrome (SSSS) is an exfoliative dermatitis that results from infection with exfoliative toxin-producingStaphylococcus aureus. SSSS is seen primarily in infants and children. Here we ask if there is a specific maturation process that protects healthy adults from this syndrome. For these studies, an active recombinant exfoliative toxin A (rETA) was used in a neonatal mouse model. A time course generated on the susceptibility to the toxin as a function of mouse age indicated that BALB/c mice developed the characteristic symptoms of SSSS until day 7 of life. Between day 7 and day 8 of life there was a dramatic decrease in susceptibility, such that mice at day 9 of life were resistant to the effects of the toxin. This time course corresponds approximately to the time needed for maturation of the adaptive immune response, and SSSS in adults is often identified with immunocompromised states. Therefore, mice deficient in this response were examined. Adult mice thymectomized at birth and adult SCID mice did not develop the symptoms of SSSS after injection with the toxin, indicating that the adaptive immune response is not responsible for the lack of susceptibility observed in the older mice. SSSS in adults is also associated with renal disorders, suggesting that levels of toxin in serum are important in the development of the disease. rETA was not cleared as efficiently from the serum of 1-day-old mice compared to clearance from 10-day-old mice. Ten-day-old mice were given repeated injections of toxin so that the maximal level of toxin was maintained for a sustained period of time, and exfoliation occurred in these mice. Thus, whereas the adaptive immune response is not needed for protection of adult mice from SSSS, efficient clearance of the toxin from the bloodstream is a critical factor.
Characterization of the Secreted MPT53 Antigen of
Mycobacterium tuberculosis
Authors: Johnson Sadie , Brusasca PierNatale , Lyashchenko Konstantin , Spencer John S. , Wiker Harald G. , Bifani Pablo , Shashkina Elena , Kreiswirth Barry , Harboe Morten , Schluger Neil , Gomez Manuel , Gennaro Maria Laura
Published: July 2002

MPT53 is a secreted protein of
Mycobacterium tuberculosis
. Southern transfer and hybridization showed
mpt53
to be conserved in the
M. tuberculosis
complex and to have homology with DNA from
Mycobacterium avium
and other nontuberculous mycobacteria. However, anti-MPT53 polyclonal antibodies detected no antigen in the culture filtrates of
M. avium
and other nontuberculous mycobacteria. MPT53 of
M. tuberculosis
induced strong, tuberculosis-specific antibody responses in guinea pigs but induced no delayed-type hypersensitivity. Involvement in immune responses during human tuberculosis was very modest.
Primary Role for CD4+T Lymphocytes in Recovery from Oropharyngeal Candidiasis
Authors: Farah C. S. , Elahi S. , Drysdale K. , Pang G. , Gotjamanos T. , Seymour G. J. , Clancy R. L. , Ashman R. B.
Published: July 2002

Oropharyngeal candidiasis is associated with defects in cell-mediated immunity and is commonly seen in human immunodeficiency virus positive individuals and AIDS patients. A model for oral candidiasis in T-cell-deficient BALB/c and CBA/CaHnu/numice was established. After inoculation with 108Candida albicansyeasts, these mice displayed increased levels of oral colonization compared to euthymic control mice and developed a chronic oropharyngeal infection. Histopathological examination ofnu/nuoral tissues revealed extensive hyphae penetrating the epithelium, with polymorphonuclear leukocyte microabscess formation. Adoptive transfer of either naive or immune lymphocytes into immunodeficient mice resulted in the recovery of these animals from the oral infection. Reconstitution of immunodeficient mice with naive CD4+but not CD8+T cells significantly decreased oral colonization compared to controls. Interleukin-12 and gamma interferon were detected in the draining lymph nodes of immunodeficient mice following reconstitution with naive lymphocytes. This study demonstrates the direct requirement for T lymphocytes in recovery from oral candidiasis and suggests that this is associated with the production of cytokines by CD4+T helper cells.