Listeria monocytogenes wall teichoic acid decoration in virulence and cell-to-cell spread.

Wall teichoic acid (WTA) comprises a class of glycopolymers covalently attached to the peptidoglycan of gram positive bacteria. In Listeria monocytogenes, mutations that prevent addition of certain WTA decorating sugars are attenuating. However, the steps required for decoration and the pathogenic process interrupted are not well described.

In vitro properties of a Listeria monocytogenes bacteriophage-resistant mutant predict its efficacy as a live oral vaccine strain.

A Listeria monocytogenes glcV mutation precludes the binding of certain listerial phages and produces a profound attenuation characterized by the absence of detectable mutants in the livers and spleens of orally inoculated mice. In vitro, we found that the mutant formed plaques on mouse enterocyte monolayers as efficiently as the parent but the plaques formed were smaller. Intracellular growth rate determinations and examination of infected enterocytes by light and fluorescence microscopy established that the mutant was impaired not in intracellular growth rate but in cell-to-cell spreading.

Factors associated with the acquisition and severity of gestational listeriosis.

Gravid mammals are more prone to listeriosis than their nongravid counterparts. However, many features of the disease in gravid animals are not well defined. We determined, in mice, that increased susceptibility to lethal infection following oral inoculation begins surprisingly early in pregnancy and extends through embryonic development.

A Listeria monocytogenes mutant defective in bacteriophage attachment is attenuated in orally inoculated mice and impaired in enterocyte intracellular growth.

A Listeria monocytogenes bacteriophage was used to identify a phage-resistant Tn917 insertion mutant of the mouse-virulent listerial strain F6214-1. The mutant was attenuated when it was inoculated orally into female A/J mice and failed to replicate efficiently in cultured mouse enterocytes. Phage binding studies indicated that the mutant had a cell surface alteration that precluded phage attachment.

Host and bacterial factors in listeriosis pathogenesis.

Members of the Genus Listeria are ubiquitous environmental saprophytic microorganisms. If ingested they can cause a severe disseminated disease (listeriosis) that has a high mortality rate, the highest of any food-borne pathogen, even with antibiotic therapy. Central to the high mortality rate is the hallmark characteristic of the microorganism to grow intracellularly.

A specific gene expression program triggered by Gram-positive bacteria in the cytosol.

Innate and adaptive immunity depends critically on host recognition of pathogen-associated molecules. Toll-like receptors (TLRs) are key mediators of pathogen surveillance at the cell or phagocytic vacuole surface. However, mechanisms underlying recognition of pathogens in other cellular compartments remain unclear, and responses elicited by cytosolic challenge are poorly characterized.

Influence of pregnancy on the pathogenesis of listeriosis in mice inoculated intragastrically.

Pregnancy increases the risk of listeriosis, a systemic disease caused by Listeria monocytogenes. However, there is incomplete agreement on the reasons for this increased risk. We examined two features of listeriosis in gravid and nongravid female mice following intragastric (gavage) inoculation, namely, (i) disease severity (measured by lethality) and (ii) listerial infectivity (measured by liver and spleen colonization levels up to 120 h postinoculation).

Influence of extracellular bactericidal agents on bacteria within macrophages.

We employed gentamicin-sensitive and -resistant derivatives of Escherichia coli in a macrophage phagocytosis assay that compared lambda bacteriophage and gentamicin as extracellular bactericidal agents. Colony counts and direct microscopic examination of phagocytized E. coli supported the conclusion that gentamicin entered macrophages, even at low concentrations, and contributed to their bactericidal activity.

Characterization of Escherichia coli type 1 pilus mutants with altered binding specificities.

PCR mutagenesis and a unique enrichment scheme were used to obtain two mutants, each with a single lesion in fimH, the chromosomal gene that encodes the adhesin protein (FimH) of Escherichia coli type 1 pili. These mutants were noteworthy in part because both were altered in the normal range of cell types bound by FimH. One mutation altered an amino acid at a site previously shown to be involved in temperature-dependent binding, and the other altered an amino acid lining the predicted FimH binding pocket.

Genetic characterization of Escherichia coli type 1 pilus adhesin mutants and identification of a novel binding phenotype.

Five Escherichia coli type 1 pilus mutants that had point mutations in fimH, the gene encoding the type 1 pilus adhesin FimH, were characterized. FimH is a minor component of type 1 pili that is required for the pili to bind and agglutinate guinea pig erythrocytes in a mannose-inhibitable manner. Point mutations were located by DNA sequencing and deletion mapping.

Woodchuck lymphotoxin-alpha, -beta and tumor necrosis factor genes: structure, characterization and biological activity.

We cloned and characterized the woodchuck tumor necrosis factor (TNF) and lymphotoxin-alpha, -beta (LT-alpha, -beta) cDNAs, genes and proteins to facilitate study of the functions of these cytokines during the course of woodchuck hepatitis virus (WHV) infection. Woodchuck cDNA and genomic DNA libraries were screened with woodchuck-specific DNA probes to isolate the cDNA and gene clones for TNF, LT-alpha and LT-beta. The cDNAs for woodchuck TNF, LT-alpha and LT-beta code for proteins of 233, 205 and 310 amino acids respectively.

The mucosal phase of Listeria infection.

Listeria monocytogenes is an enteroinvasive bacterial pathogen of man and animals. Listeriae have been shown capable of infecting the host by translocating from the intestinal lumen through Peyer's Patches (PP), however, results of experiments now indicate that these facultative intracellular parasites may also translocate through PP-independent routes. With regards to this, on occasion we observed that listeriae were absent from the PP of mice inoculated intragastrically with L.

Host and bacterial factors involved in the innate ability of mouse macrophages to eliminate internalized unopsonized Escherichia coli.

In an effort to better understand genetic and cellular factors that influence innate immunity, we examined host and bacterial factors involved in the nonopsonic phagocytosis and killing of Escherichia coli K-12 by mouse macrophages. Unelicited (resident) peritoneal macrophages from five different mouse strains, BALB/c, C57BL/6, CD-1, C3H/HeJ, and C3H/HeN, were employed. Additional macrophage populations were obtained from CD-1 mice (bone marrow-derived macrophages).

Roles for tumor necrosis factor and gamma interferon in resistance to enteric listeriosis.

Listeria monocytogenes normally infects the host by translocating from the intestinal lumen. Experiments were carried out to determine if, when, and where tumor necrosis factor (TNF) and gamma interferon (IFN-gamma) function in antibacterial resistance during enteric listeriosis. Groups of normal mice and severe combined immunodeficient (SCID) mice were injected with neutralizing monoclonal antibodies (MAb) specific for each cytokine and then inoculated intragastrically with L.

Role of tumor necrosis factor and gamma interferon in acquired resistance to Cryptococcus neoformans in the central nervous system of mice.

Although naive C.B-17 and BALB/cBy mice die of meningoencephalitis within 5 weeks of intravenous infection with an opportunistic strain of Cryptococcus neoformans, immunized mice express an acquired, CD4+ T-cell-dependent immunity and survive an intravenous infection. Infusion of lymphocytes from immune mice into severe combined immunodeficiency (SCID) mice renders these mice more resistant to cryptococcal brain infection than uninfused controls.

Gamma interferon-dependent temporary resistance to acute Toxoplasma gondii infection independent of CD4+ or CD8+ lymphocytes.

It is well established that resistance to acute primary Toxoplasma gondii infection is mediated by a gamma interferon (IFN-gamma)-dependent mechanism. The present in vivo experiments were undertaken to investigate the cellular basis for this resistance. We show here that immunocompetent T.

Mice that lack the interferon-gamma receptor have profoundly altered responses to infection with Bacillus Calmette-Guérin and subsequent challenge with lipopolysaccharide.

Mice with a targeted disruption of the interferon gamma receptor gene (IFN-gamma R0/0 mice) and control wild-type mice were inoculated with the Bacillus Calmette-Guérin (BCG) strain of Mycobacterium bovis. BCG infection was not lethal for wild-type mice whereas all IFN-gamma R0/0 mice died approximately 7-9 wk after inoculation. Histological examination at 2 and 6 wk after BCG inoculation showed that livers of IFN-gamma R0/0 mice had higher numbers of acid-fast bacteria than wild-type mice, especially at 6 wk.

Gamma interferon functions in resistance to Cryptosporidium parvum infection in severe combined immunodeficient mice.

Severe combined immunodeficient (SCID) adult mice are relatively resistant to Cryptosporidium parvum infection, even though they are deficient in both T- and B-cell function. The requirement for gamma interferon (IFN-gamma) in this resistance was examined by treatment of these mice with monoclonal antibody to IFN-gamma. SCID mice injected intraperitoneally with monoclonal anti-IFN-gamma 4 h before and three times weekly after challenge with C.

Listeria monocytogenes-induced interferon-gamma primes the host for production of tumor necrosis factor and interferon-alpha/beta.

Mice acquired an enhanced capacity for the production of tumor necrosis factor (TNF) and the interferons (IFN)-alpha, and -beta shortly after intravenous injection of viable Listeria monocytogenes. By the end of the first day of a sublethal infection, mice were primed to produce 100-1000 times more endotoxin-induced serum TNF than is produced by normal mice. Acquisition of the augmented capacity for TNF production was due to L.

Endotoxin-induced tumor necrosis factor alpha synthesis in murine embryo fibroblasts.

Murine embryo fibroblasts (MEF) were found to secrete tumor necrosis factor (TNF) in response to stimulation with endotoxin. Endotoxin-induced TNF production by MEF was inhibited by cycloheximide. However, reversal of the effect of this inhibitor on protein synthesis results in TNF being secreted in amounts equivalent to those produced by endotoxin-induced MEF not treated with cycloheximide.

Interleukin-6 production in a murine model of Pneumocystis carinii pneumonia: relation to resistance and inflammatory response.

The production of interleukin-6 (IL-6) and its possible relationship to host resistance and inflammatory response to Pneumocystis carinii infection were examined in mice with severe combined immunodeficiency (SCID mice). IL-6 activity was detected in the serum and lungs of P. carinii-infected mice but not in mice free of P.

Interleukin 1: an important mediator of host resistance against Pneumocystis carinii.

The importance of endogenous interleukin 1 (IL-1) in resistance to Pneumocystis carinii infection was examined in a SCID mouse model. Naturally acquired pulmonary infection of P. carinii in SCID mice was completely cleared by reconstitution of the infected mice with immunocompetent spleen cells.