Typhimurium-infected Mice Research Articles

Th1 and Th2 cell involvement in immune response to Salmonella typhimurium porins.

In understanding the regulation of the specific immune response to Salmonella typhimurium, the role of a surface major component (porins) was studied. In this study we demonstrate that purified porins are able to induce a different response to that induced by the porins present on the S. typhimurium cell surface. Porin-treated or orally infected mice show anti-porin antibodies with bactericidal activity. The complete adoptive transfer of resistance to S. typhimurium is achieved only using splenic T cells from survivor mice after experimental infection. After stimulation with specific antigen in vitro CD4+ cells from porin-immunized mice released large amounts of interleukin-4 (IL-4), at a time when CD4+ cells from S. typhimurium-infected mice predominantly secreted interferon-gamma (IFN-gamma). Limiting dilution analysis showed that infection resulted in a higher precursor frequency of IFN-gamma-producing CD4+ T cells and a lower precursor frequency of IL-4-producing CD4+ T cells, while immunization with porins resulted in a higher precursor frequency of IL-4-producing cells and a low frequency of IFN-gamma-producing cells. Analysis of polymerase chain reaction-amplified cDNA from the spleens of infected mice revealed that IFN-gamma, IL-2 and IL-12 p40 mRNA were found 5 days after in vitro challenge and increased after 15 days; IL-10 expression was barely present after both 5 and 15 days, while IL-4 mRNA expression was not detected. In immunized mice, the IL-4 mRNA expression increased after 15 days, IFN-gamma mRNA expression disappeared entirely after 15 days, while IL-2, IL-10 and IL-12 mRNA remained relatively unchanged.

Th1 response in Salmonella typhimurium-infected mice with a high or low rate of bacterial clearance.

Previous studies have shown that the capacity to clear an attenuated strain of Salmonella typhimurium after the second week of infection varies widely among mouse strains. Bacterial clearance is mediated by CD4+ T cells and is regulated in part by the H-2 complex. The aim of the present study was to compare the patterns of cytokine mRNA expression in the spleens of C57BL/6 (H-2b) and CBA (H-2k) mice, which exhibit a low and a high rate of bacterial clearance, respectively. A transient increase in interleukin-12 (IL-12) mRNA levels was found in both mouse strains. Gamma interferon (IFN-gamma) gene expression was higher and more sustained in C57BL/6 than in CBA mice. No increase in IL-4 mRNA was detected. A transient increase in IL-10 mRNA was found in C57BL/6 mice. Separation of spleen cells into CD4+ and CD4- fractions showed that CD4+ T cells produced the bulk of IFN-gamma in both mouse strains and of IL-10 in C57BL/6 mice. Infection of H-2 congenic mice induced a higher level of IFN-gamma mRNA expression by CD4+ T cells in mice with a low rate of clearance (H-2b) than in mice with a high rate of clearance (H-2q). Treatment of infected C57BL/6 mice with anti-IFN-gamma or anti-CD4 monoclonal antibodies indicated that IFN-gamma participates in resistance in the early phase of infection, but not in bacterial clearance, and that CD4+ T cells mediate bacterial clearance during the 3rd week of infection. Taken together, these results suggest that defective bacterial clearance in H-2b mice is not linked to defective IFN-gamma production and that CD4+ T cells mediate bacterial clearance by an IFN-gamma-independent mechanism.

Detection of porin antigen in serum for early diagnosis of mouse infections with Salmonella typhimurium.

The monoclonal antibodies to porin, an outer membrane protein isolated from Salmonella typhimurium and sandwich enzyme linked immunosorbent assay (ELISA) has made possible the detection of porin from sera of S. typhimurium-infected mice. The specificity of the monoclonal antibodies was ascertained based on their cross-reactivity with porins isolated from S. typhi, Shigella flexneri and Escherichia coli and lipopolysaccharide (LPS) of S. typhimurium and E. coli. Serum samples were found to be positive for porin as early as 3 days after intravenous and 5 days after oral infection. In addition, a positive correlation was observed between the bacterial load and the concentration of porin detected in the sera. On the other hand, analysis of sera for anti-porin antibody showed diametrically opposite time kinetics with antigenaemia. These results indicate that porin accumulates in the serum of infected mice much earlier than the appearance of antibodies to porin. Thus detection of porin holds promise for early diagnosis of typhoid.

Formation of Interferon-Gamma and Tumor Necrosis Factor in Mice during <i>Salmonella typhimurium </i>Infection

Formation of interferon-gamma (IFN-gamma) and tumor necrosis factor (TNF) during Salmonella typhimurium infection was investigated in lipopolysaccharide (LPS)-sensitive C3H/HeN and C57BL/10ScSn(B10ScSn), and LPS-resistant (lpsd mutant) C3H/HeJ and C57BL/10ScCr(B10ScCr) mice. When infected with 50 colony-forming units (CFU) of S. typhimurium C5, C3H/HeN and B10ScSn mice became hypersensitive to the lethal effect of LPS. In the case of lpsd mutants, only C3H/HeJ mice became hypersensitive to LPS, while B10ScCr mice remained resistant. C3H/HeJ as well as B10ScSn mice produced significant amounts of plasma IFN-gamma on day 3 after infection. By this time bacterial CFU in the liver of B10ScSn and C3H/HeJ mice were 10(6.7) and 10(7.1), respectively. In B10ScCr mice, however, IFN-gamma was not detectable although bacteria present in the liver exceeded 10(8) CFU. On the other hand, plasma TNF was not detectable in any of the mouse strains during S. typhimurium infection. When S. typhimurium-infected mice were challenged with LPS on day 3, significant amounts of plasma TNF were measured in C3H/HeN and B10ScSn mice, while in the lpsd mutant C3H/HeJ and B10ScCr mice plasma TNF was undetectable.

Role of H-2 and non-H-2 genes in control of bacterial clearance from the spleen in Salmonella typhimurium-infected mice.

The ability of mice to clear Salmonella typhimurium from their spleens in the late phase of infection was studied after inoculation with a temperature-sensitive mutant. Clearance of bacteria was delayed in C57BL/6 mice compared with BALB/c, C3H/HeJ, DBA/2, A/J, and CBA mice. The responses of F1 hybrids, backcrosses, and recombinant inbred strains derived from C57BL/6 and BALB/c (both Itys) and of H-2 congenic mice were analyzed. The results showed that the low rate of bacterial clearance was recessive, that the rate of clearance was under polygenic control, and that an H-2-linked gene(s) plays a major role. Among H-2 congenic mice with a C57BL/10 background, three phenotypes of bacterial clearance could be distinguished: high (H-2j, H-2q, and H-2u), intermediate (H-2d, H-2f, H-2k, H-2p, H-2r, H-2s, and H-2v), and low (H-2b) rates. The effect of the H-2 complex was apparent with different genetic backgrounds (Itys and Ityr). In recombinant inbred strains derived from C57BL/6 (Itys) and A/J (Ityr) mice, the effect of the H-2b haplotype on bacterial clearance appeared to be fully expressed only in strains carrying the Itys allele.

T-2 toxin effects on the serum amyloid P-component (SAP) response of Listeria monocytogenes- and Salmonella typhimurium-infected mice

T-2 mycotoxin, given to mice 4 days prior to an intraperitoneal inoculation with Listeria monocytogenes EGD, increases the acute phase response as determined by measurements of serum amyloid protein-P (SAP), and decreases the severity of the infection. Conversely, when T-2 toxin is given simultaneously with L. monocytogenes the mice become more susceptible to the infection, and the SAP levels attained are diminished relative to the non-toxin-treated Listeria-infected controls. T-2 toxin given 4 days prior to intraperitoneal inoculation with Salmonella typhimurium had no effect on either the resultant infection or SAP levels. These results indicate that T-2 toxin modulates the acute phase response to infection, and are consistent with an in vivo role for SAP as a nonspecific host resistance factor.

Cellular response in Salmonella typhimurium-infected mice: evaluation of Salmonella receptors of B lymphocytes

The cellular response in the course of experimental infection with Salmonella typhimurium was studied in mice. T cells were detected by the presence of theta-antigen, B cells by the binding of fluorescent immunoglobulins, and cells with receptors by labeled Salmonella binding. Lymphocytes were from spleen and lymph nodes. Results have been divided into three groups: group A, including mice with slight symptomatology; group B, including those with serious infection symptomatology; and group C, including mice that died in the course of the experiment. In spleen and lymph nodes of group A mice, an increase in the percentage of T and B lymphocytes was observed. This increase reached a peak 10 days after experimental infection. In lymph nodes, the B-cell percentage was equal to the percentage of T cells, whereas in spleen lymphocytes the B-cell percentage was higher. In spleens of group B mice we observed the same response as in mice of group A, whereas in lymph nodes there was a low response of T and B lymphocytes. In group C mice, there was no significant response of T and B lymphocytes in either spleen or lymph nodes. In B lymphocytes prepared from spleens of surviving mice, a small number of Salmonella receptors was detected: 200 bacterial cells per 10(9) lymphocytes.

Mechanisms of acquired resistance in mouse typhoid.

Experiments in vitro comparing normal mouse peritoneal macrophages with cells from Salmonella typhimurium-infected mice have shown that the "immune" macrophages have conspicuously enhanced microbicidal properties. Whereas normal macrophages could inactivate only 50 to 60% of intracellular S. typhimurium pretreated with immune serum, cells from infected animals killed virtually all ingested organisms and did so at an accelerated rate. Macrophages from Listeria monocytogenes-infected mice were shown to possess similarly enhanced microbicidal activity against S. typhimurium. Furthermore, the growth of S. typhimurium in the liver and spleen was more effectively restricted in Listeria-infected mice than in animals vaccinated with heat-killed S. typhimurium, even though the Listeria-infected animals possessed no demonstrable cross-reacting antibody to S. typhimurium. The lack of resistance in the mice vaccinated with heat-killed organisms could not be attributed to any deficiency of humoral factors, since the serum from these animals was as effective at promoting phagocytosis and killing by macrophages as serum from actively infected (and demonstrably resistant) mice. Conversely, Salmonella-infected mice were totally resistant to intravenous challenge with L. monocytogenes. The level of resistance in individual animals was related to the numbers of residual Salmonellae remaining in the tissues; mice with heavier residual infections being the more resistant. Specific antiserum from mice vaccinated with heat-killed S. typhimurium was found to be significantly protective only when the intraperitoneal route of challenge was employed. The foregoing studies have been interpreted to mean that enhancement of the microbicidal ability of macrophages is the mechanism of major importance in acquired resistance to S. typhimurium infection in mice.