bluuebunny/crossref_metadata_2025_split huggingface dataset:
Attenuated
Shigella flexneri
2a Δ
guaBA
Strain CVD 1204 Expressing Enterotoxigenic
Escherichia coli
(ETEC) CS2 and CS3 Fimbriae as a Live Mucosal Vaccine against
Shigella
and ETEC Infection
Authors: Altboum Zeev , Barry Eileen M. , Losonsky Genevieve , Galen James E. , Levine Myron M. 
Published: July 2002

ABSTRACT

To construct a prototype hybrid vaccine against
Shigella
and enterotoxigenic
Escherichia coli
(ETEC), the genes encoding the production of ETEC CS2 and CS3 fimbriae were isolated and expressed in attenuated
Shigella flexneri
2a
guaBA
strain CVD 1204. The CS2
cotA
to -
D
genes, isolated from ETEC strain C91F, and the CS3
cstA
to -
H
genes, subcloned from plasmid pCS100, were cloned into ∼15-copy-number-stabilized pGA1 behind the osmotically regulated
ompC
promoter, resulting in high expression of both fimbriae. Under nonselective in vitro growth conditions, pGA1-CS2 and pGA1-CS3 were stable in CVD 1204, exhibiting a plasmid loss of only approximately 1% per duplication. Expression of CS2 and CS3 reduced the invasiveness of
Shigella
for HeLa cells and slowed the intracellular growth rate. Guinea pigs immunized intranasally with CVD 1204(pGA1-CS2) or CVD 1204(pGA1-CS3), or with a mixture of these strains, developed secretory immunoglobulin A (IgA) in tears and serum IgG antibodies against
Shigella
lipopolysaccharide, CS2, and CS3 antigens. Moreover, the animals were protected against keratoconjunctivitis following conjunctival challenge with virulent
S. flexneri
2a strain 2457T. Animals immunized with
Shigella
expressing CS2 or CS3 developed serum antibodies that agglutinated
Shigella
as well as an ETEC strain bearing the homologous fimbriae, whereas animals immunized with combined CVD 1204(pGA1-CS2) and CVD 1204(pGA1-CS3) developed antibodies that agglutinated all three test strains. These observations support the feasibility of a multivalent vaccine against shigellosis and ETEC diarrhea consisting of multiple
Shigella
live vectors expressing relevant ETEC antigens.
Calprotectin Expression by Gingival Epithelial Cells
Authors: Ross Karen F. , Herzberg Mark C. 
Published: July 2002

ABSTRACTCalprotectin, a heterodimer of MRP8 and MRP14 with antimicrobial properties, is found in the cytosol of neutrophils, monocytes, and human gingival keratinocytes. During inflammation of the oral mucosa, the expression of immunoreactive calprotectin appears upregulated. Given the possible cell sources, we sought to learn if epithelial cells upregulate calprotectin in response to proinflammmatory agents. First, human gingival keratinocytes were maintained in primary culture until senescence. At each passage, cells were harvested and analyzed for quantitative expression of MRP8 and MRP14 subunit mRNA by RNase protection assays and calprotectin complex by enzyme-linked immunosorbent assay. Calprotectin expression was constitutive in the primary gingival keratinocytes, but calprotectin-specific mRNA and protein tended to increase as the cells neared senescence. To test whether calprotectin expression was inducible, immortalized gingival keratinocyte cultures were treated for 2 to 4 h with lipopolysaccharide (LPS) or interleukin-1β (IL-1β). As a positive control for inducible expression, immortalized keratinocytes were incubated with phorbol myristate acetate (PMA) (50 ng/ml) for 24 h. Incubation with PMA stimulated increased expression of MRP8 and MRP14 mRNA within 2 h, peaking within 5 h. MRP8- and MRP14-specific mRNA expression by immortalized keratinocytes appeared to be unaffected by LPS or IL-1β. In contrast, LPS, IL-1β, and PMA each upregulated IL-8. These data show that calprotectin mRNA is expressed constitutively in cultured keratinocytes, while expression by immortalized cells appears to be independent of the exogenous proinflammatory agents LPS and IL-1β.
Recruitment of Cytoskeletal and Signaling Proteins to Enteropathogenic and Enterohemorrhagic
Escherichia coli
Pedestals
Authors: Goosney Danika L. , DeVinney Rebekah , Finlay B. Brett 
Published: July 2002

ABSTRACT

Enteropathogenic
Escherichia coli
(EPEC) is a human pathogen that attaches to intestinal epithelial cells and causes chronic watery diarrhea. A close relative, enterohemorrhagic
E. coli
(EHEC), causes severe bloody diarrhea and hemolytic-uremic syndrome. Both pathogens insert a protein, Tir, into the host cell plasma membrane where it binds intimin, the outer membrane ligand of EPEC and EHEC. This interaction triggers a cascade of signaling events within the host cell and ultimately leads to the formation of an actin-rich pedestal upon which the pathogen resides. Pedestal formation is critical in mediating EPEC- and EHEC-induced diarrhea, yet very little is known about its composition and organization. In EPEC, pedestal formation requires Tir tyrosine 474 phosphorylation. In EHEC Tir is not tyrosine phosphorylated, yet the pedestals appear similar. The composition of the EPEC and EHEC pedestals was analyzed by examining numerous cytoskeletal, signaling, and adapter proteins. Of the 25 proteins examined, only two, calpactin and CD44, were recruited to the site of bacterial attachment independently of Tir. Several others, including ezrin, talin, gelsolin, and tropomyosin, were recruited to the site of EPEC attachment independently of Tir tyrosine 474 phosphorylation but required Tir in the host membrane. The remaining proteins were recruited to the pedestal in a manner dependent on Tir tyrosine phosphorylation or were not recruited at all. Differences were also found between the EPEC and EHEC pedestals: the adapter proteins Grb2 and CrkII were recruited to the EPEC pedestal but were absent in the EHEC pedestal. These results demonstrate that although EPEC and EHEC recruit similar cytoskeletal proteins, there are also significant differences in pedestal composition.
Molecular and Genomic Analysis of Genes Encoding Surface-Anchored Proteins from
Clostridium difficile
Authors: Karjalainen Tuomo , Waligora-Dupriet Anne-Judith , Cerquetti Marina , Spigaglia Patrizia , Maggioni Andrea , Mauri Pierluigi , Mastrantonio Paola 
Published: July 2002

ABSTRACT

The gene
slpA
, encoding the S-layer precursor protein in the virulent
Clostridium difficile
strains C253 and 79–685, was identified. The precursor protein carries a C-terminal highly conserved anchoring domain, similar to the one found in the Cwp66 adhesin (previously characterized in strain 79–685), an SLH domain, and a variable N-terminal domain mediating cell adherence. The genes encoding the S-layer precursor proteins and the Cwp66 adhesin are present in a genetic locus carrying 17 open reading frames, 11 of which encode a similar two-domain architecture, likely to include surface-anchored proteins.
Influence of Cultivation Media on Genetic Regulatory Patterns in
Borrelia burgdorferi
Authors: Yang Xiaofeng , Popova Taissia G. , Goldberg Martin S. , Norgard Michael V. 
Published: July 2002

ABSTRACT

Barbour-Stoenner-Kelly II (BSKII) medium and BSKH medium both are routinely used for the cultivation of
Borrelia burgdorferi
. However, heretofore there have been no studies to compare how these two media affect gene expression patterns in virulent
B. burgdorferi
. In the present study, we found that some
B. burgdorferi
strain 297 genes (e.g.,
ospA
,
mlp-7A
,
mlp-8
,
p22
, and
lp6.6
) that typically are regulated by temperature or pH displayed their predicted pattern of expression when
B. burgdorferi
was cultivated in BSKH medium; this was not true when spirochetes were cultivated in conventional BSKII medium. The results suggest that BSKH medium is superior to BSKII medium for gene expression studies with
B. burgdorferi
.
Malnutrition Alters the Innate Immune Response and Increases Early Visceralization followingLeishmania donovaniInfection
Authors: Anstead Gregory M. , Chandrasekar Bysani , Zhao Weiguo , Yang Jue , Perez Luis E. , Melby Peter C. 
Published: July 2002

ABSTRACTMalnutrition is a risk factor for the development of visceral leishmaniasis. However, the immunological basis for this susceptibility is unknown. We have developed a mouse model to study the effect of malnutrition on innate immunity and early visceralization followingLeishmania donovaniinfection. Three deficient diets were studied, including 6, 3, or 1% protein; these diets were also deficient in iron, zinc, and calories. The control diet contained 17% protein, was zinc and iron sufficient, and was provided ab libitum. Three days after infection withL. donovanipromastigotes, the total extradermal (lymph nodes, liver, and spleen) and skin parasite burdens were equivalent in the malnourished (3% protein) and control mice, but in the malnourished group, a greater percentage (39.8 and 4.0%, respectively;P= 0.009) of the extradermal parasite burden was contained in the spleen and liver. The comparable levels of parasites in the footpads in the two diet groups and the higher lymph node parasite burdens in the well-nourished mice indicated that the higher visceral parasite burdens in the malnourished mice were not due to a deficit in local parasite killing but to a failure of lymph node barrier function. Lymph node cells from the malnourished, infected mice produced increased levels of prostaglandin E2(PGE2) and decreased levels of interleukin-10. Inducible nitric oxide synthase activity was significantly lower in the spleen and liver of the malnourished mice. Thus, malnutrition causes a failure of lymph node barrier function afterL. donovaniinfection, which may be related to excessive production of PGE2and decreased levels of IL-10 and nitric oxide.
Chlamydia trachomatisPersistence in the Female Mouse Genital Tract: Inducible Nitric Oxide Synthase and Infection Outcome
Authors: Ramsey Kyle H. , Miranpuri Gurwattan S. , Sigar Ira M. , Ouellette Scott , Byrne Gerald I. 
Published: July 2002

ABSTRACTIt was previously reported that female mice resolve a primaryChlamydia trachomatisurogenital infection independent of inducible nitric oxide synthase (iNOS). We now report that although iNOS-deficient (NOS2−/−) mice resolve culture-apparent infection in a fashion similar to that of normal control (NOS2+/+) mice, they sustain significantly increased rates of disease, as assessed by hydrosalpinx formation. PCR amplification ofompAfollowed by Southern blot detection of amplicands revealed the presence of chlamydial DNA in the lower genital tracts of both NOS2−/−and NOS2+/+mice at ≥120 days postinfection and in upper genital tract tissues at >120 days postinfection. However, only NOS2−/−mice shed low numbers of viable chlamydiae from the lower genital tract after immunosuppressive treatment at 120 days postinfection. When cultured primary murine lung fibroblasts were activated in the presence of gamma interferon (IFN-γ), inhibition of chlamydial growth occurred in both NOS2+/+and NOS2−/−cells, but the inhibition was reversible after removal of the cytokine in the NOS2−/−primary cell culture only. The iNOS-independent inhibition was microbistatic but was independent of 2,3-indoleamine dioxygenase activity. We conclude that chlamydial DNA and antigens persist in mice subsequent to culture-apparent resolution. In addition, IFN-γ induces in vivo inhibition of chlamydial growth through microbistatic mechanisms in the absence of iNOS activity, but in the presence of iNOS activity, IFN-γ is microbicidal and effects eradication.
Enteropathogenic
Escherichia coli
Activates Ezrin, Which Participates in Disruption of Tight Junction Barrier Function
Authors: Simonovic Ivana , Arpin Monique , Koutsouris Athanasia , Falk-Krzesinski Holly J. , Hecht Gail 
Published: July 2002

ABSTRACT

Enteropathogenic
Escherichia coli
(EPEC) is an important human intestinal pathogen, especially in infants. EPEC adherence to intestinal epithelial cells induces the accumulation of a number of cytoskeletal proteins beneath the bacteria, including the membrane-cytoskeleton linker ezrin. Evidence suggests that ezrin can participate in signal transduction. The aim of this study was to determine whether ezrin is activated following EPEC infection and if it is involved in the cross talk with host intestinal epithelial cells. We show here that following EPEC attachment to intestinal epithelial cells there was significant phosphorylation of ezrin, first on threonine and later on tyrosine residues. A significant increase in cytoskeleton-associated ezrin occurred following phosphorylation, suggesting activation of this molecule. Nonpathogenic
E. coli
and EPEC strains harboring mutations in type III secretion failed to elicit this response. Expression of dominant-negative ezrin significantly decreased the EPEC-elicited association of ezrin with the cytoskeleton and attenuated the disruption of intestinal epithelial tight junctions. These results suggest that ezrin is involved in transducing EPEC-initiated signals that ultimately affect host physiological functions.
Delayed Invasion of the Kidney and Brain byBorrelia crociduraein Plasminogen-Deficient Mice
Authors: Nordstrand Annika , Shamaei-Tousi Alireza , Ny Annelii , Bergström Sven 
Published: July 2002

ABSTRACTBorrelia crociduraeis an etiologic agent of relapsing fever in Africa and is transmitted to humans by the bite of soft ticks of the genusOrnithodoros. The role of the plasminogen (Plg) activation system for the pathogenicity ofB. crociduraewas investigated by infection of Plg-deficient (plg−/−) and Plg wild-type (plg+/+) mice. No differences in spirochetemia were observed between theplg−/−andplg+/+mice. However, signs indicative of brain invasion, such as neurological symptoms and/or histopathological changes, were more common inplg+/+mice. Quantitative immunohistochemical analysis demonstrated infection of spirochetes in kidney interstitium and brain as soon as 2 days postinoculation. Lower numbers of extravascular spirochetes inplg−/−mice during the first days of infection suggested a less efficient invasion mechanism in these mice than in theplg+/+mice. The invasion of the kidneys inplg−/−mice produced no significant inflammation, as seen by quantitative immunohistochemistry of the CD45 common leukocyte marker. However, significant kidney inflammation was observed with infection in theplg+/+mice. In brain, inflammation was more severe inplg+/+mice than inplg−/−mice, and the numbers of CD45+cells increased significantly with duration of infection in theplg+/+mice. The results show that invasion of brain and kidney occurs as early as 2 days after inoculation. Also, Plg is not required for establishment of spirochetemia by the organism, whereas it is involved in the invasion of organs.
Molting, Ecdysis, and Reproduction of
Trichinella spiralis
Are Supported In Vitro by Intestinal Epithelial Cells
Authors: Gagliardo L. F. , McVay C. S. , Appleton J. A. 
Published: July 2002

ABSTRACT

Trichinella spiralis
is an obligate parasite of animals that has an unusual intracellular life cycle. Investigation of parasitism at the cellular and molecular levels has been challenging because of a shortage of tools for in vitro cultivation of
T. spiralis.
We have found that
T. spiralis
larvae molt, ecdyse, develop to adulthood, and reproduce when they are inoculated onto cultured intestinal epithelial cells. Initially, larvae invade and migrate through cells in a monolayer (T. ManWarren, L. Gagliardo, J. Geyer, C. McVay, S. Pearce-Kelling, and J. Appleton, Infect. Immun. 65:4806-4812, 1997). During prolonged culture in Caco-2 epithelial cells, L1 larvae molted and ecdysed with efficiencies as high as 50%. Molting and ecdysis in vitro required entry of the parasite into cells; conditions that prevented entry into cells also prevented ecdysis. When larvae were inoculated at a low density and cultured for 5 to 9 days, as many as 50% of the larvae developed to adult stages. Low numbers of mature male worms with copulatory appendages were observed in these cultures. The majority of worms that survived for five or more days were unfertilized females. Low-density cultures supported development of female worms with embryos at rates of 4 to 5%. These results show that the intestinal life cycle of
T. spiralis
can be supported entirely by host epithelial cells. Our model should allow more detailed investigation of intracellular parasitism by
T. spiralis
.
Helicobacter pylori
Uses Motility for Initial Colonization and To Attain Robust Infection
Authors: Ottemann Karen M. , Lowenthal Andrew C. 
Published: July 2002

ABSTRACT

Helicobacter pylori
has been shown to require flagella for infection of the stomach. To analyze whether flagella themselves or motility is needed by these pathogens, we constructed flagellated nonmotile mutants. This was accomplished by using both an insertion mutant and an in-frame deletion of the
motB
gene. In vitro, these mutants retain flagella (Fla
+
) but are nonmotile (Mot
−
). By using FVB/N mice, we found that these mutants had reduced ability to infect mice in comparison to that of their isogenic wild-type counterparts. When these mutants were coinfected with wild type, we were unable to detect any
motB
mutant. Finally, by analyzing the 50% infectious dose, we found that motility is needed for initial colonization of the stomach mucosa. These results support a model in which motility is used for the initial colonization of the stomach and also to attain full infection levels.
Specific Entry ofHelicobacter pyloriinto Cultured Gastric Epithelial Cells via a Zipper-Like Mechanism
Authors: Kwok Terry , Backert Steffen , Schwarz Heinz , Berger Jürgen , Meyer Thomas F. 
Published: July 2002

ABSTRACTAlthoughHelicobacter pylorihas generally been considered an extracellular pathogen, a number of in vitro infection experiments and biopsy examinations have shown that it is capable of occasionally entering mammalian host cells. Here, we characterized this entry process by using AGS cells as a host cell model. In gentamicin protection-invasion assays, the number ofH. pyloricolonies recovered was lower than that forSalmonella entericaserovar Typhimurium X22,Escherichia coliexpressing InvA, andYersinia enterocoliticaYO:9 grown at 25°C but higher than that forNeisseria gonorrhoeaeVP1 andY. enterocoliticaYO:9 grown at 37°C. At the ultrastructural level, the entry process was observed to occur via a zipper-like mechanism. InternalizedH. pyloriwas bound in tight LAMP-1-containing vacuoles in close association with condensed filamentous actin and tyrosine phosphorylation signals. Wortmannin, a potent inhibitor of phosphatidylinositol 3-kinase, and calphostin C, an inhibitor of protein kinase C, both inhibited the entry ofH. pyloriin a sensitive and dose-dependent manner; however, the level of entry was enhanced by sodium vanadate, an inhibitor of tyrosine phosphatases and ATPases. Furthermore, the cytokine tumor necrosis factor alpha antagonized the entry ofH. pyloriinto AGS cells. Collectively, these results demonstrate that the entry ofH. pyloriinto AGS cells occurs via a zipper-like mechanism which involves various host signal transduction events.
Afa, a Diffuse Adherence Fibrillar Adhesin Associated with Enteropathogenic
Escherichia coli
Authors: Keller Rogéria , Ordoñez Juana G. , de Oliveira Rosana R. , Trabulsi Luiz R. , Baldwin Thomas J. , Knutton Stuart 
Published: July 2002

ABSTRACT

O55 is one of the most frequent enteropathogenic
Escherichia coli
(EPEC) O serogroups implicated in infantile diarrhea in developing countries. Multilocus enzyme electrophoresis analysis showed that this serogroup includes two major electrophoretic types (ET), designated ET1 and ET5. ET1 corresponds to typical EPEC, whilst ET5 comprises strains with different combinations of virulence genes, including those for localized adherence (LA) and diffuse adherence (DA). Here we report that ET5 DA strains possess a DA adhesin, designated EPEC Afa. An 11.6-kb chromosomal region including the DA adhesin operon from one O55:H
−
ET5 EPEC strain was sequenced and found to encode a protein with 98% identity to AfaE-1, an adhesin associated with uropathogenic
E. coli
. Although described as an afimbrial adhesin, we show that both AfaE-1 and EPEC Afa possess fine fibrillar structures. This is the first characterization and demonstration of an Afa adhesin associated with EPEC.
Fc-Dependent and Fc-Independent Opsonization of
Cryptococcus neoformans
by Anticapsular Monoclonal Antibodies: Importance of Epitope Specificity
Authors: Netski Dale , Kozel Thomas R. 
Published: July 2002

ABSTRACT

Monoclonal antibodies (MAbs) reactive with glucuronoxylomannan (GXM), the major capsular polysaccharide of the yeast
Cryptococcus neoformans
, produce distinct capsular reactions when viewed by differential interference contrast microscopy. These reactions depend on the epitope specificity of the antibody. Opsonic activities of immunoglobulin G1 (IgG1) MAbs that produce patterns termed rim and puffy were examined. Rim-pattern MAbs are reactive with an epitope shared by GXM serotypes A, B, C, and D. Puffy-pattern MAbs are reactive only with serotypes A and D. In phagocytosis assays, using serotype A cells and resident murine peritoneal macrophages, rim-pattern MAbs were markedly more opsonic than puffy-pattern MAbs. F(ab′)
2
fragments of rim-pattern MAbs were synergistic with heat-labile factors in normal human serum for opsonization of the yeast. F(ab′)
2
fragments of puffy-pattern MAbs were also synergistic with normal serum in opsonization but at a much lower level than fragments of rim-pattern MAbs. Normal serum alone was not opsonic. F(ab′)
2
fragments of rim-pattern MAbs, but not puffy-pattern MAbs, stimulated phagocytosis of encapsulated cryptococci in the absence of serum. This serum-independent opsonic action of F(ab′)
2
fragments was abrogated by pretreatment of macrophages with purified GXM, suggesting the involvement of a phagocyte GXM receptor. The results indicate that (i) there are multiple mechanisms by which anticapsular IgG MAbs facilitate phagocytosis of encapsulated cryptococci, (ii) some anti-GXM antibodies are opsonic in an Fc-independent manner, and (iii) opsonic activity correlates with the capsular reaction and occurs in an epitope-specific manner.
Induction of Interleukin-12 and Gamma Interferon Requires Tumor Necrosis Factor Alpha for Protective T1-Cell-Mediated Immunity to PulmonaryCryptococcus neoformansInfection
Authors: Herring Amy C. , Lee John , McDonald Roderick A. , Toews Galen B. , Huffnagle Gary B. 
Published: July 2002

ABSTRACTThe development of T1-cell-mediated immunity is required to clear a pulmonaryCryptococcus neoformansinfection. The objective of these studies was to determine the mechanism by which tumor necrosis factor alpha (TNF-α) augments the development of pulmonary T1 immunity toC. neoformansinfection. TNF-α expression was detected in lavage sample cells at days 2, 3, and 7 followingC. neoformansinfection. The numbers of CFU in the lung were not different between control and anti-TNF-α-treated mice at any time point examined during the afferent phase of the response (days 0 to 7). However, neutralization of TNF-α prevented the initiation of pulmonary clearance during the efferent phase of the response (day 14). Administration of anti-TNF-α monoclonal antibody (day 0) diminished the lung levels of TNF-α, interleukin-12 (IL-12), and gamma interferon (IFN-γ) induced byC. neoformansat day 7 postinfection. Neutralization of TNF-α (day 0) also altered the IFN-γ/IL-4 ratio in the lung-associated lymph nodes at day 7 followingC. neoformansinfection. Anti-TNF-α-treated mice developed a pulmonary eosinophilia at day 14 postinfection. Consistent with the pulmonary eosinophilia, anti-TNF-α-treated mice exhibited elevated serum immunoglobulin E and inhibition of the anticryptococcal delayed-type hypersensitivity response, indicating a shift toward a T2 response. Neutralization of IL-12 also prevented lung leukocyte production of IFN-γ in response to the infection. These findings demonstrate that afferent-phase TNF-α production is essential for the induction of IL-12 and IFN-γ and neutralization of early TNF-α results in a T2 shift of the T1/T2 balance of antifungal immunity.