Effects of antiinflammatory agents on chronic Salmonella typhimurium infection in a mouse model.

Anti-inflammatory corticosteroids have been demonstrated to lower the resistance of the host to a range of infections in several different animal models. We have adapted the mouse model for Salmonella typhimurium infection, using a subcutaneous inoculation of 10(4) organisms in CBA/Ca mice. The chronic infection was potentiated by treatment with hydrocortisone given orally in the diet for up to 4 weeks, beginning on day 30 after infection. A dose of 25 mg/kg of body weight resulted in 100% mortality from Salmonella infection within 24 days of beginning hydrocortisone treatment. Nonsteroid drugs, indomethacin and a trial agent BW 755C, were compared with the corticosteroid in this model. No mortality was recorded during the 4 weeks of drug treatment in these groups. Fecal samples were monitored weekly, and drug-related effects on the normal fecal flora were observed. The kinetics of S. typhimurium infection were followed in both fecal and organ samples for the duration of the experiments. The model is useful for the determination of the lowered resistance of the host to infection after drug treatment and specifically demonstrated here the potential of the trial drug BW 755C as an active anti-inflammatory agent without one of the undesirable side effects of corticosteroids.

Genetic control of resistance to Salmonella typhimurium infection in high and low antibody responder mice.

Mice selected by Biozzi for high and low responses to sheep erythrocytes were investigated for resistance to subcutaneous Salmonella typhimurium infection. The resistance was measured by LD50 values, viable bacterial counts in liver and spleen at 10 days, and the kinetics of infection over 4 weeks. High responder mice were susceptible to S. typhimurium injected subcutaneously (LD50 less than 10) and low line resistant (LD50 3 x 10(6)). Control of natural resistance to S. typhimurium in inbred mice is primarily by a single gene. Ity, on chromosome 1. Results with hybrid generations of Biozzi mice with either BALB/c (sensitive) or CBA (resistant) inbred mice indicated additional genetic control of resistance in Biozzi mice. Analysis of resistance data of backcrosses of (high x low)F1 with either parental strain showed this genetic control to be at least one other gene in the Biozzi mice, not linked to Ity. The antibody responses in the hybrid generations and inbred and Biozzi parental strains were tested by haemagglutination assays and ELISA. After specific stimulation of the mice there was an inverse relationship between resistance to S. typhimurium and antibody levels.

The protein-lipopolysaccharide complex extracted with trichloracetic acid from Salmonella typhimurium effective in protection of mice against S. typhimurium infection.

A protein-lipopolysaccharide complex has previously been postulated as necessary to protect susceptible mice against Salmonella typhimurium infection. Lipopolysaccharide attached to non-specific proteins, bovine serum albumin or methylated BSA, gave a specific delayed-type hypersensitivity (DTH) reaction when injected into the footpad of mice sensitized with sublethal doses of S. typhimurium. However, immunization of BALB/c mice with the complex gave no survivors after challenge with 100 LD50 S. typhimurium. Trichloracetic acid extraction of bacterial cultures produced lipopolysaccharide with attached protein. This method gave simple and convenient production of an active factor, demonstrating few major protein bands after electrophoresis. The complex elicited specific DTH reactions in sensitized mice and protected 37% of BALB/c mice against 100 LD50 S. typhimurium. Combinations of protein:lipopolysaccharide were used in DTH experiments to determine the relative importance of the components. A minimum requirement for both was demonstrated, although the ratio was not critical. Use of O-antigenic mutant strains of Salmonella indicated a role for protein in the specificity of activity of the complex.

Early haemopoietic responses to Salmonella typhimurium infection in resistant and susceptible mice.

The response of colony forming cells in the bone marrow and spleen of resistant (CBA) and susceptible (C57BL) mice to Salmonella typhimurium infection was studied for 4 days after infection. The number and size of the colonies were assessed. The resistant strain exhibited an immediate response to challenge, sharply increasing the number of colonies to 2.5 times normal over 2-3 days after infection. In contrast the susceptible strain gave a slowly increased response to the same challenge, which never exceeded 1.2 times normal and fell to 0.8 times the normal. When mouse strains were immunized there was a clear distinction between the splenic and bone marrow cellularity. Immunization appeared to enhance the splenic cellularity in resistant mice but failed to in susceptible mice. In the bone marrow of susceptible mice, however, there was some evidence of an elevated response.

O-antigenic specificity of the protective supernatant factor from Salmonella typhimurium effective in S. typhimurium-infected mice.

A supernatant factor (SF) prepared from cultures of Salmonella typhimurium protected naturally susceptible inbred mice against challenge with S. typhimurium subcutaneously injected (s.c.i.), but not against Salmonella enteritidis, suggesting tha the relevant specificity involved lipopolysaccharide. Further experiments were performed with two transductant strains of S. typhimurium. Strain SH6701 has O-antigen 4 from S. typhimurium and SH6703 had O-antigen 9 from S. enteritidis. Immunization with SF from SH6701 protected 95% BALB/c mice challenged with 100 LD50 S. typhimurium s.c.i., whereas SH6703 immunization had no effect on survival or mean survival time. SH 6703 SF gave some protection against homologous challenge. Antibody titres and delayed-type hypersensitivity reactions were also tested in immunized mice. The SF was, therefore, specific in that O-antigen 4 was necessary to protect mice against S. typhimurium challenge. Since we have previously demonstrated protein to be necessary for protection by SF, the active factor may be in the form of a protein-lipopolysaccharide complex.