Serum-Mediated Resistance Induced with Immunogenic Preparations of Salmonella typhimurium

Serum obtained from animals immunized with attenuated Salmonella typhimurium strain RIA, heat-killed bacterial suspensions, or immunogenic ribosomal preparations was capable of passively conferring protection on normal recipient mice to challenge infection with virulent S. typhimurium strain SR-11. Protection was measured by the ability of a recipient animal to reduce the total number of challenge organisms significantly below that found in challenged control mice. Intraperitoneal administration of 0.1 ml of pooled sera was more effective than 0.05 ml in transferring resistance. The transfer of equivalent amounts of immune serum subcutaneously did not result in demonstrable resistance. In all cases in which serum transfer was effective, resistance was maximal at 5 days, greatly diminished by 10 days, and gone by 15 days post-transfer. Pooled serum obtained from normal donor mice or 0.2 ml of serum from mice hyperimmunized with immunogenic ribonucleic acid preparations did not possess the capacity to confer demonstrable resistance on normal recipients.     

Immunogenicity of Ribonucleic Acid Preparations Obtained from Salmonella typhimurium

Mice immunized with purified whole-cell ribonucleic acid (RNA), RNA from the bacterial “particulate” fraction, and ribosome-associated RNA obtained from Salmonella typhimurium were found to be resistant to subsequent challenge infection with virulent salmonellae. Chemically, the immunogenic nucleic acid fractions contained from 1 to 3% “contaminant” material defined (based on the mean of 19 different preparations) as protein (0.24%), deoxyribonucleic acid (0.43%), methyl pentose (0.64%), hexose (1.58%), and undefined carbohydrate (0.76%). Heptoses and lipoidal material were not detectable in any of the immunogenic preparations examined. Physically, the nucleic acid preparations, after analytical ultracentrifugation, exhibited three boundaries similar to those reported elsewhere in comparable systems: 4 to 5S, 16S, and 23S. An evaluation of the immunity induced by the ribosome-associated RNA established that the immune response was (i) comparable to that induced 15 days postimmunization with live salmonellae and by ribosomal vaccines, but greater at 30 days postimmunization than that in mice immunized with attenuated salmonellae; (ii) dependent on the quantity of immunogen administered; (iii) dependent on the size of the infective inocula; (iv) inhibited at 15 but not at 30 days postimmunization when the immunogenic nucleic acid preparations were incorporated into Freund's incomplete adjuvant, (v) reduced or lost by dialysis in relatively high or low immunizing doses, respectively; and (vi) unaffected by enzymatic treatment of the preparations with trypsin, deoxyribonuclease, Pronase plus pancreatic ribonuclease, or pancreatic ribonuclease alone. The possible mode of action of ribosome-associated RNA in inducing an immune response to subsequent challenge infection with the homologous organism is discussed.     

Antibiotic Resistance Patterns and Plasmid Profiles of Salmonella typhimurium Isolates in Turkey

 To understand the resistance mechanisms present in 75 isolates of Salmonella typhimurium derived from clinical infections in Turkey, antimicrobial resistance patterns and associated plasmids were investigated. Among the 22 strains that produced extended-spectrum β-lactamase (ESBL), 20 were resistant to aminoglycosides and 12 to trimethoprim-sulfamethoxazole. Strains that did not produce ESBL did not express aminoglycoside or trimethoprim-sulfamethoxazole resistance, although 27 of them were ampicillin resistant. None of the strains were resistant to imipenem or fluoroquinolones. Nineteen strains producing ESBL carried a plasmid of >100 MDa. Seven ESBL-producing strains conjugally transferred their ESBLs and trimethoprim-sulfamethoxazole resistance. No correlation was found between the resistance patterns and plasmids in non-ESBL-producing strains.     

Immunity in Experimental Salmonellosis I. Protection Induced by Rough Mutants of Salmonella typhimurium

Different rough mutants of Salmonella typhimurium were tested to determine their virulence and immunizing capacity when used as live vaccines for mice. All uridine diphosphate-galactose-4-epimeraseless mutants tested were much more potent immunizing agents than any other mutants. This capacity was not correlated with virulence or complexity of cell wall polysaccharide. For good protection, persistence of the rough strains in vivo was essential, but the protection lasted longer than the period during which bacteria were demonstrable in the liver and spleen of the mice. The outstanding immunizing capacity of the “epimeraseless” mutants is not dependent on the persistence of viable bacteria in the mouse.     

Virulence of R-Factor-Bearing Salmonella typhimurium

Antibiotic-resistant recombinants obtained from mating antibiotic-resistant Escherichia coli with virulent, antibiotic-sensitive Salmonella typhimurium are generally avirulent. After 32 consecutive transfers, two of four avirulent recombinants regained their virulence without loss of episome-mediated resistance.     

Position on Mouse Chromosome 1 of a Gene that Controls Resistance to Salmonella typhimurium

Ity is a gene which regulates the magnitude of Salmonella typhimurium growth in murine tissues and, hence, the innate salmonella resistance of mice. The results of a five-point backcross clearly showed that the correct gene order on chromosome 1 is fz-Idh-1-Ity-ln-Pep-3.     

Salmonella typhimurium Virulence Genes Are Induced upon Bacterial Invasion into Phagocytic and Nonphagocytic Cells

Survival and growth of salmonellae within host cells are important aspects of bacterial virulence. We have developed an assay to identify Salmonella typhimurium genes that are induced inside Salmonella-containing vacuoles within macrophage and epithelial cells. A promoterless luciferase gene cassette was inserted randomly into the Salmonella chromosome, and the resulting mutants were screened for genes upregulated in intracellular bacteria compared to extracellular bacteria. We identified four genes in S. typhimurium that were upregulated upon bacterial invasion of both phagocytic and nonphagocytic cells. Expression of these genes was not induced by factors secreted by host cells or media alone. All four genes were induced at early time points (2 to 4 h) postinvasion and continued to be upregulated within host cells at later times (5 to 7 h). One mutant contained an insertion in the ssaR gene, within Salmonella pathogenicity island 2 (SPI-2), which abolished bacterial virulence in a murine typhoid model. Two other mutants contained insertions within SPI-5, one in the sopB/sigD gene and the other in a downstream gene, pipB. The insertions within SPI-5 resulted in the attenuation of S. typhimurium in the mouse model. The fourth mutant contained an insertion within a previously undescribed region of the S. typhimurium chromosome, iicA (induced intracellularly A). We detected no effect on virulence as a result of this insertion. In conclusion, all but one of the genes identified in this study were virulence factors within pathogenicity islands, illustrating the requirement for specific gene expression inside mammalian cells and indicating the key role that virulence factor regulation plays in Salmonella pathogenesis.     

Interleukin 18 Contributes to Host Resistance and Gamma Interferon Production in Mice Infected with Virulent Salmonella typhimurium

Spleen and peritoneal macrophages obtained from innately resistant A/J mice released low levels of interleukin 18 (IL-18) upon infection with Salmonella typhimurium C5 RP4. Incubating the cells with recombinant gamma interferon (rIFN-γ) enhanced IL-18 production. A/J mice treated in vivo with anti-IL-18 antibodies showed impaired resistance to infection, with increased bacterial loads in the liver and spleen. Administration of rIL-18 could protect A/J mice from challenge with a lethal dose of virulent salmonellae, with a dramatic reduction in bacterial numbers in the tissues. rIL-18 administration did not ameliorate the disease in IFN-γ-R^−/− mice. IL-18 proved to be required for IFN-γ production by mouse splenocytes from conventional, scid, and rag-1^−/− mice; in vivo IL-18 neutralization caused a decrease in circulating IFN-γ levels. Thus, IL-18 is a key factor in early host resistance to Salmonella and probably acts via IFN-γ.     

Identification of Three Highly Attenuated Salmonella typhimurium Mutants That Are More Immunogenic and Protective in Mice than a Prototypical aroA Mutant

A panel of Salmonella typhimurium 14028s mutants, which were previously shown to be highly attenuated in the BALB/c mouse model of infection, were analyzed for their potential as live Salmonella oral-vaccine candidates. A prototypical aroA mutant was chosen as a basis of comparison. From the panel of mutants initially chosen for this study, three mutants with comparable levels of attenuation elicited higher Salmonella-specific serum immunoglobulin G (IgG) and/or mucosal secretory-IgA antibody titers than the aroA vaccine strain. The three mutants, CL288, CL401, and CL554, also elicited a better protective immune response than the aroA control strain, after a single oral dose of 1 × 10⁹ to 2 × 10⁹ bacteria.     

Oral Immunization of Mice with Killed Salmonella typhimurium Vaccine

A study was undertaken to assess the efficacy of oral, parenteral, and intraperitoneal immunization methods of administering killed Salmonella typhimurium vaccine to mice and to evaluate the effectiveness of single and multiple doses of the vaccine containing varied numbers of the killed bacteria. A further objective of this study was to evaluate the effect of adding substances to the vaccine to which have been ascribed "adjuvant" properties. The protection was estimated by isolation of bacteria from the spleen and feces after oral challenge of the mice with live S. typhimurium. The results showed that one or more doses of 10(10) organisms given orally led to significant protection. This rate of protection increased proportionately with the number of doses up to 10 doses, which offered 100% protection. Streptomycin, when added to multiple doses of 10(9) or more organisms given orally, increased the degree of protection, but beryllium sulfate and pertussis vaccine did not. Although multiple doses afforded similar systemic protection by all three routes of immunization, oral immunization yielded significantly greater local protection than that observed after subcutaneous or intraperitoneal immunization.     

Assay of typhoid vaccines with Salmonella typhosa-Salmonella typhimurium hybrids

Salmonella typhimurium hybrids expressing the S. typhosa antigens 9, d, and Vi were constructed by genetic crosses with an S. typhosa Hfr donor. The hybrids retained the same degree of mouse virulence as their S. typhimurium parent strain, the minimum lethal dose being less than 50 organisms when tested either in C₅₇ black mice or Swiss white mice. Vaccination of the Swiss white mice with S. typhosa Ty2 vaccines prepared by acetone treatment, alcohol treatment, or heat-killing conferred significant protection against challenge by the hybrid strains but not against their S. typhimurium parent. Both the acetone-treated and alcohol-treated typhoid vaccines were markedly more protective than the heat-killed, phenol-preserved vaccine.     

Salmonella typhimurium meningitis in infancy

We report a case of meningitis due to Salmonella typhimurium in a four-month-old female infant. The child was brought to the pediatric emergency department with complaints of fever, cold, and generalized convulsion. On examination, the child was febrile and was having seizures. The anterior fontanelle was not bulging. Lumbar puncture was done and Salmonella typhimurium was isolated from cerebrospinal fluid. Initially the infant improved clinically with appropriate management, but had a fatal outcome due to nosocomial pneumonia.     

Pneumococcal Capsular Polysaccharide Preparations May Contain Non-C-Polysaccharide Contaminants That Are Immunogenic

We measured the capacity to opsonize Streptococcus pneumoniae serotype 6B and estimated the concentration of immunoglobulin G anti-6B capsular polysaccharide (PS) antibodies in 25 pre- and postimmune sera from adults immunized with a pneumococcal PS vaccine. We first studied two postvaccination serum samples displaying less opsonophagocytic capacity than expected. The majority of anti-6B antibodies in the two samples reacted with the capsular PSs of several unrelated serotypes (2, 4, 9V, 19F, and 23F) and with the lysate of noncapsulated S. pneumoniae bacteria but not with C-PS. The non-type-specific antibodies accounted for at least one-half of anti-6B antibodies in 40% of prevaccination sera and 10% of postvaccination sera from adults. The non-type-specific antibodies could be demonstrated in the enzyme-linked immunosorbent assays (ELISAs) for pneumococcal antibodies to other serotypes (4, 9V, 18C, 19F, and 23F). The nonspecific antibodies appear to bind a contaminant(s) in the current preparations of capsular PS. ELISA for antibodies to pneumococcal capsules may not be serotype specific for some samples.     

Plasmid-associated virulence of Salmonella typhimurium.

We investigated the role of the 100-kilobase (kb) plasmid of Salmonella typhimurium in the virulence of this organism for mice. Three strains, LT2-Z, SR-11, and SL1344, which possessed 100-kb plasmids with identical restriction enzyme digestion profiles, were cured of their respective 100-kb plasmids after Tnmini-tet was used to label plasmids. Curing wild-type virulent strains SR-11 and SL1344 raised peroral 50% lethal doses from 3 x 10(5) and 6 x 10(4) CFU, respectively, to greater than 10(8) CFU. Both wild-type strains had intraperitoneal 50% lethal doses of less than 50 CFU, whereas the intraperitoneal 50% lethal doses for cured SR-11 and SL1344 were less than 50 and 400 CFU, respectively. Reintroduction of the Tnmini-tet-labeled, 100-kb plasmid restored wild-type virulence. Invasion from Peyer's patches to mesenteric lymph nodes and spleens after peroral inoculation was the stage of pathogenesis most affected by curing S. typhimurium of the 100-kb plasmid. Wild-type S. typhimurium replicated in spleens of mice inoculated intravenously to a greater extent than did plasmid-cured derivatives. Wild-type and cured strains equally adhered to and invaded Henle-407, HEp-2, and CHO cells; furthermore, the presence of the 100-kb plasmid was not necessary for replication of S. typhimurium within CHO cells. The 100-kb plasmid had no effect on phagocytosis and killing of S. typhimurium by murine peritoneal macrophages in vitro and in vivo. Similarly, wild-type and plasmid-cured strains were resistant to killing by 90% normal human, rabbit, and guinea pig sera. All wild-type and plasmid-cured S. typhimurium strains possessed complete lipopolysaccharide, as determined by silver staining solubilized cells in sodium dodecyl sulfate-polyacrylamide gels. We have confirmed the role of the 100-kb plasmid of S. typhimurium in virulence, primarily in invasion to mesenteric lymph nodes and spleens after peroral inoculation of mice. Involvement of the 100-kb plasmid in infection of mesenteric lymph nodes and spleens suggests a role for the plasmid in the complex interaction of S. typhimurium with cells of the reticuloendothelial system.     

Biotypes of Salmonella typhimurium in Iraq

207 Salmonella typhimurium strains isolated from childrens below three years (190 strains), adult patients (8 strains), health carriers (7 strains) or animals (2 strains) were studied with the biotyping scheme of Cordano, Richard and Vieu (1971). Five biovars were found and 76.8% of the strains were TTR+, Ino+, Tre+, d. Tar- (biovar d). In Irak the epidemiology of the Salmonella typhimurium human infections is associated with an high frequency of strains of Salmonella typhimurium 0:5- (var. Copenhagen), biovar d.     

Salmonella typhimurium meningitis in an adult patient with AIDS

Salmonella meningitis is an unusual complication of Salmonella sepsis and occurs mainly in children. A rare case of Salmonella typhimurium meningitis occurring in an adult HIV positive man who presented with a history of fever and diarrhoea is reported. On examination he was dehydrated, and had oral thrush, weakness of lower limbs and neck stiffness. A septic diagnostic screen was performed and he was commenced on empiric intravenous cefotaxime therapy for meningitis. S typhimurium was cultured from cerebrospinal fluid and blood culture specimens. It was non-lactose fermenting, oxidase negative, H(2)S positive and motile. Cefotaxime was continued for 14 days and the patient responded without neurological sequelae.     

Immune response to infection with Salmonella typhimurium in mice

Infection of mice with Salmonella typhimurium results in systemic infection and a disease similar to that seen in humans after infection with S. typhi. The innate immune system can restrict replication of S. typhimurium to a certain degree, but for effective control and eradication of bacteria, acquired immunity is essential. Salmonella infection induces the generation of specific CD4+ and CD8+ T cells, and both T cell populations are important for protection during primary and secondary responses, although the mechanisms underlying T cell-mediated protection are not yet completely understood. Infection with S. typhimurium also results in a strong antibody response to Salmonella antigens and, in contrast to most other intracellular bacteria, this antibody response participates in protection. In summary, the response to S. typhimurium involves both T and B cell-mediated immunity, and mechanisms mediated by both lymphocyte populations are important for control of primary infection and protection against secondary infection.