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PubMed1992

Monoclonal antibodies directed to the O antigen of Salmonella serogroup E cross-react with lipopolysaccharides of Salmonella serogroups C, F and S.

Four murine hybridoma clones secreting monoclonal antibodies (mAb) directed against lipopolysaccharide (LPS) antigen of S. newington, serogroup E1, were obtained after a fusion of spleen cells of mice immunized with formaldehyde-killed bacteria and mouse myeloma cells of the X63-Ag8.653 line. Antigen binding properties and specificity of the mAb were studied in bacterial agglutination tests, passive hemolysis and its inhibition, passive hemagglutination tests, passive hemolysis and its inhibition, passive hemagglutination and immunoenzyme tests (ELISA and immunoblotting). Three of the mAb (24E6, 29E1 and 45F6) were agglutinating and were active in all tests used, while mAb 31H12 did not agglutinate bacteria but revealed a high reactivity in the immunoenzyme reactions. It was found that the mAb reacted with LPS and Salmonella strains from serogroup E (E1, E2, E3 and E4) as well as from serogroups C (C1 and C4), F and S thus showing that the O3 antigen possesses more than one epitope, one of which is represented on the LPS antigens of the serovars from the cross-reacting groups mentioned. According to the known chemical the most probable recognized epitope consists of mannose with beta-linkage to the next monosaccharide residue in the LPS chain.

Antibodies, Bacterial↗
PubMed2000

The antibiotic resistance patterns of Salmonella Typhi isolates in Italy, 1980-96. The Italian SALM-NET Working Group. Salmonella Network.

In this paper we report the distribution of Salmonella Typhi isolates in Italy and their resistance patterns to antibiotics. The data were collected by the Italian SALM-NET surveillance system in a pilot retrospective study of the period 1980-96. Data on drug-resistance were available for 82 isolates out of 176 S. Typhi isolated in Italy. Of these 82 isolates, 32 (39%) were resistant or intermediate to 1 or more antibiotics. Eight isolates were resistant and 7 intermediate to streptomycin; 4 isolates were resistant to ampicillin alone or in association with other antibiotics; only 2 strains (1 isolated in Lombardia in 1993 and the other 1 in Lazio in 1994) were resistant to chloramphenicol, and 2 (isolated in Sardegna and Piemonte in 1995 and 1996, respectively) showed intermediate resistance to chloramphenicol. The strains showing resistance to 3 or more antibiotics were very scarce: 1 (with 5 complete resistances) was isolated in Lazio in 1994, and another 1 (with complete resistance to 10 antibiotics and intermediate resistance to 2 antibiotics) was isolated in Molise in 1988. In conclusion, besides the routine activities to control typhoid fever, an accurate and continuous surveillance is necessary in order to quickly identify multidrug-resistant (MDR) S. Typhi strains and prevent their spread, even though their level, in our country, is still quite low.

Drug Resistance, Microbial↗
PubMedDate not supplied

[Isolation of a highly purified capsular polysaccharide from Salmonella enterica serovar Typhi (Salmonella typhi)--Vi-antigen and its use in serological diagnosis of typhoid fever].

For use in differential diagnostics of typhoid fever, samples of the capsular polysaccharide from Salmonella enterica serovar Typhi (usually named Vi-antigen) were isolated and characterized by physicochemical and serological methods. It was shown that only the sample of Vi-antigen with the minimal (0.57%) admixture of the corresponding lipopolysaccharide (LPS) from S. typhi retained a high serological activity in the tests with monoreceptor anti-Vi sera. However, it exhibited a substantially weaker reaction with sera from normal donors and patients with acute nontyphoid salmonelloses, than Vi-antigen preparations with a higher (0.8-1.2%) LPS content. The chromatographically pure Vi-antigen was purified by triple reprecipitation with hexadecyltrimethylammonium bromide. The content of the LPS admixture in the resulting Vi-antigen samples was quantitatively determined by GC. A high purification level of the Vi-antigen from the LPS admixture allows us to hope that this preparation could serve as a basic component of the test system for the diagnostics of typhoid fever. The English version of the paper.

Chemical Precipitation↗
PubMed1981

Artificial Salmonella vaccines: Salmonella typhimurium O-antigen-specific oligosaccharide-protein conjugates elicit protective antibodies in rabbits and mice.

Several saccharides representative of the O-antigenic polysaccharide chain of Salmonella typhimurium (O antigens 4 and 12) were used as haptenic groups covalently linked to bovine serum albumin. The disaccharide abequose 1 --> 3 D-mannose was synthesized, and the [Formula: see text] tetra-, octa- and dodecasaccharides were isolated after cleavage of isolated S. typhimurium O-polysaccharide chains by using bacteriophage endo-glycosidases. Rabbits immunized with the saccharide-protein conjugates suspended in Freund complete adjuvant readily responded with O4 antibody titers as high, or almost as high, as those elicited by heat-killed bacteria. The octa- and dodecasaccharide conjugates also gave rise to O12 antibody titers. The antibody response in mice differed in two ways from that seen in rabbits: mice did not respond with measurable antibody production against the disaccharide haptens, and the highest S. typhimurium lipopolysaccharide antibody response elicited by the saccharide haptens was still approximately 50-fold lower than that elicited by heat-killed bacteria. The latter difference may be a consequence of the fact that the mouse preferentially produces antibodies against the galactose residue which is terminal in the hapten but not in the native O-antigenic polysaccharide chain. Antibodies elicited in rabbits against all saccharide-protein conjugates protected passively transferred mice against intraperitoneal challenge with 100 times the 50% lethal dose of S. typhimurium SH 2201 (O4, 12) but not against challenge with S. enteritidis SH 2204 (O9, 12). The antibodies elicited by the saccharide-protein conjugates protected as well as antibodies elicited by heat-killed bacteria.

Animals↗
PubMed2001

Flagellar phase variation of Salmonella enterica serovar Typhimurium contributes to virulence in the murine typhoid infection model but does not influence Salmonella-induced enteropathogenesis.

Although Salmonella enterica serovar Typhimurium can undergo phase variation to alternately express two different types of flagellin subunit proteins, FljB or FliC, no biological function for this phenomenon has been described. In this investigation, we constructed phase-locked derivatives of S. enterica serovar Typhimurium that expressed only FljB (termed locked-ON) or FliC (termed locked-OFF). The role of phase variation in models of enteric and systemic pathogenesis was then evaluated. There were no differences between the wild-type parent strain and the two phase-locked derivatives in adherence and invasion of mouse epithelial cells in vitro, survival in mouse peritoneal macrophages, or in a bovine model of gastroenteritis. By contrast, the locked-OFF mutant was virulent in mice following oral or intravenous (i.v.) inoculation but the locked-ON mutant was attenuated. When these phase-locked mutants were compared in studies of i.v. kinetics in mice, similar numbers of the two strains were isolated from the blood and spleens of infected animals at 6 and 24 h. However, the locked-OFF mutant was recovered from the blood and spleens in significantly greater numbers than the locked-ON strain by day 2 of infection. By 5 days postinfection, a majority of the mice infected with the locked-OFF mutant had died compared with none of the mice infected with the locked-ON mutant. These results suggest that phase variation is not involved in the intestinal stage of infection but that once S. enterica serovar Typhimurium reaches the spleens of susceptible mice those organisms in the FliC phase can grow and/or survive better than those in the FljB phase. Additional experiments with wild-type S. enterica serovar Typhimurium, fully capable of switching flagellin type, supported this hypothesis. We conclude that organisms that have switched to the FliC(+) phase have a selective advantage in the mouse model of typhoid fever but have no such advantage in invasion of epithelial cells or the induction of enteropathogenesis.

Animals↗
PubMed1969

Biosynthesis of cell wall lipopolysaccharide in mutants of Salmonella. V. A mutant of Salmonella typhimurium defective in the synthesis of cytidine diphosphoabequose.

A mutant of Salmonella typhimurium LT2 was found to be unable to convert cytidine diphospho-4-keto-6-deoxy-d-glucose into cytidine diphosphoabequose. The mutation maps in the rfb gene cluster, which is known to be involved in the biosynthesis of the peripheral, "O side-chain" portion of cell wall lipopolysaccharide. In spite of the fact that, in the O side chains, abequose is not a part of the main chain but occurs as short branches, the mutant appears to be unable to polymerize oligosaccharide "repeat units" into long O side chains. The following evidence indicates that this failure is the result of the absence of cytidine diphosphoabequose rather than that of a superimposed second mutation in other genes of the rfb cluster. (i) The mutant does not behave like a multisite mutant in genetic crosses, and it gives rise, at a high frequency, to "revertants" where the ability to synthesize cytidine diphosphoabequose and the ability to synthesize normal lipopolysaccharide with O side chains are both restored. (ii) The mutant strain has normal levels of activity of all of the other enzymes known to be involved in O side-chain synthesis, except that the levels of several enzymes were lowered by about 30% owing to the polarity effect of the mutation. That the lowering of these enzymes is not responsible for the failure of the mutant to synthesize O side chains is clear from the fact that there were revertants which had regained some ability to synthesize abequose but still had lowered levels of these other enzymes, and that this type of revertant produced lipopolysaccharide with considerable amounts of O side chains.

Carbohydrates↗
PubMed1975

Isolation of a hybrid F' factor-carrying Escherichia coli lactose region and Salmonella typhimurium histidine region, F42-400 (F' ts114 lac+, his+): its partial characterization and behavior in Salmonella typhimurium.

Episome F' ts114 lac+ (F42-114) was transferred into Salmonella typhimurium carrying an F'his+ (FS400) episome, and fused episome F' ts114 lac+, his+ (F42-400) was obtained. Episome F42-400 could be transferred to S. typhimurium, Escherichia coli and Klebsiella pneumoniae. Identification of the episome was based on: (i) temperature sensitivity of the Lac+ and His+ phenotypes; (ii) the fact that F- segregants, obtained after temperature curing or acridine orange curing, were simultaneously Lac- and His-; and (iii) linkage of lac+ with his+ in episomal transfers to E. coli and S. typhimurium. The frequency of episome transfer was influenced by the genotype of the donor. Plasmid LT2, prevalent in S. typhimurium LT2 strains, was suggested to be responsible for the low fertility of S. typhimurium donors. Episome F42-400 was capable of chromosome mobilization, and the extent of chromosome mobilization was not influenced by the presence or absence of the histidine region on the donor chromosome. Growth in a defined medium with acridine orange was able to cure F42-400. The frequency of curing was increased (the frequency of His+ cells was 0.0001%) if the cells were grown at 40 C in the presence of acridine orange. Selection for temperature-resistant Lac+, His+ derivatives in a strain without histidine deletion yielded Hfr strains. However, similar and stronger selections in strains without the chromosomal histidine region failed to yield Hfr strains. Our inability to obtain Hfr's in strains without the chromosomal histidine region was explained by assuming that the episome F42-400 has lost the F sites involved in integration into the S. typhimurium chromosome.

Acridines↗
PubMed1979

Salmonella typhimurium newD and Escherichia coli leuC genes code for a functional isopropylmalate isomerase in Salmonella typhimurium-Escherichia coli hybrids.

The supQ newD gene substitution system in Salmonella typhimurium restores leucine prototrophy to leuD mutants by providing the newD gene product which is capable of replacing the missing leuD polypeptide in the isopropylmalate isomerase, a complex of the leuC and leuD gene product. Mutations in the supQ gene are required to make the newD protein available. An Escherichia coli F' factor was constructed which carried supQ- newD+ from S. typhimurium on a P22-specialized transducing genome. This F' pro lac (P22dsupQ394newD) episome was transferred into S. typhimurium strains containing th leuD798-ara deletion; the resulting merodiploid strains had a Leu+ phenotype, indicating that supQ- newD+ is dominant over supQ+ newD+, and eliminating the possibility that the supQ gene codes for a repressor of the newD gene. Furthermore, transfer of the F' pro lac (P22dsupQ39newD) into E. coli leuD deletion strains restored leucine prototrophy, showing that the S. typhimurium newD gene can complment the E. coli leuC gene. Growth rates of the S. typhimurium-E coli hybrid strains indicated that the mutant isopropylmalate isomerase in these strains does not induce a leucine limitation, as it does in S. typhimurium leuD supQ mutants. In vitro activity of the mutant isopropylmalate isomerase was demonstrated; the Km values for alpha-isopropylmalate of both the S. typhimurium leuC-newD isomerase and the S. typhimurium-E. coli hybrid isomerase were as much as 100 times higher than the Km values for alpha-isopropylmalate of the wild-type enzyme, which was 3 x 10(-4) M. Mutagenesis of E. coli leuD deletion strains failed to restore leucine prototrophy, indicating that E. coli does not have genes analogous to the S. typhimurium supQ newD genes, of that, if present, activation of a newD is a rare event or is lethal to the cell.

Escherichia coli↗
PubMed1993

New method for gene disruption in Salmonella typhimurium: construction and characterization of an ada-deletion derivative of Salmonella typhimurium TA1535.

A new method for gene disruption in Salmonella typhimurium was developed. The key steps of this method are to produce restriction fragments with compatible ends, preligate to produce concatemers, and then transform by electrotransformation. We developed and used this method to construct a mutant of S. typhimurium TA1535 in which the resident ada-like (adaST) gene was replaced with a kanamycin resistance gene to produce an adaST-deletion mutant derivative. The S. typhimurium adaST-deletion strain did not exhibit a higher level of mutability upon treatment with N-methyl-N'-nitro-N-nitrosoguanidine than did its wild-type parent strain. However, it did exhibit a higher sensitivity with respect to killing by N-methyl-N'-nitro-N-nitrosoguanidine. The ability of AdaST to function as a transcriptional activator is discussed.

Bacterial Proteins↗
PubMed1968

Interactions between macrophages of guinea pigs and Salmonellae. I. Fate of Salmonella typhimurium within macrophages of normal guinea pigs.

A suspended cell culture procedure was described for the cultivation of guinea pig macrophages infected with Salmonella typhimurium. The fate of the intracellular bacteria was assessed by quantitative recovery of viable bacteria with 0.5% solution of sodium desoxycholate. Two strains of S. typhimurium with different degrees of virulence for mice were compared. There was an initial destruction of intracellular bacteria of both strains; however, the extent of this destruction differed. Approximately 1% of the avirulent bacteria initially phagocytized survived at the end of 4 hr, whereas approximately 8% of the virulent bacteria survived at the end of 3 hr. After this initial killing, the intracellular bacteria began to multiply at a logarithmic rate between 3 and 21 hr after phagocytosis, and then a stationary phase was attained. The rate of this multiplication was comparable for both strains.

Animals↗
PubMed2000

Multiple clones within multidrug-resistant Salmonella enterica serotype Typhimurium phage type DT104. The Greek Nontyphoidal Salmonella Study Group.

Six distinct clones were present among Greek multidrug-resistant Salmonella enterica serotype Typhimurium phage type DT104, since isolates belonging to resistance phenotypes including the ACSSuT (ampicillin, chloramphenicol, streptomycin, sulfonamides, and tetracycline) core could be distinguished with respect to their pulsed-field gel electrophoresis patterns, int1 integron structures, and presence or absence of antibiotic resistance genes ant(3'')-Ia, pse-1, and tem-1.

Animals↗
PubMed1988

Physical and genetic mapping of the Salmonella dublin virulence plasmid pSDL2. Relationship to plasmids from other Salmonella strains.

Plasmids of approximately 80 kb in size are found in nearly all clinical isolates of Salmonella dublin and are believed to be essential for virulence. We have shown previously that the 80-kb plasmid pSDL2 is required for the S. dublin Lane strain to establish a lethal systemic infection in BALB/c mice after oral or intraperitoneal inoculation. We now present a physical and genetic characterization of pSDL2. We have established a complete restriction endonuclease cleavage map of pSDL2 for five enzymes: Xba I, Bam HI, Xho I, Sal I, and Hind III. The region specifying autonomous replication has been localized to a 10.5-kb region of the Sal I A fragment by subcloning on the vector pBR322. Using transposon insertion mutagenesis with Tn5-oriT, a region encoding the virulence phenotype has been mapped within a 6.4-kb portion of the Sal I B fragment. Deletions generated by partial Eco RI restriction digestion demonstrate that at least 50 kb of the plasmid DNA are not required for replication or virulence functions, confirming the map location of these phenotypes. Plasmids of different sizes and restriction patterns were found in mouse virulent strains of S. dublin Vi+, S. enteritidis, and S. choleraesuis. By Southern hybridization, these putative virulence plasmids share a common 4-kb Eco RI fragment with the virulence region of pSDL2, and the plasmids from S. dublin Vi+ and S. enteritidis were shown to express mouse virulence comparable to pSDL2.

Cloning, Molecular↗
PubMedDate not supplied

Salmonella in foods--a new enrichment procedure for use with the TECRA Salmonella visual immunoassay: collaborative study.

A collaborative study was conducted to compare a new enrichment procedure for the TECRA Salmonella Visual Immunoassay with the reference method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM 7th Ed.). Three food types (milk powder, black pepper, and soy flour) were analyzed in Australia, and 3 food types (milk chocolate, dried egg, and raw turkey) were analyzed in the United States. Thirty-eight collaborators participated in the study. No significant differences (p > 0.05) were observed for the pairwise comparison of the proportion of positive samples for the TECRA method with that for the reference method. The new enrichment procedure for the TECRA method has been adopted First Action by AOAC INTERNATIONAL.

Animals↗
PubMed1975

[Bacteriocinogenesis in cultures of salmonella isolated in Bulgaria. I. Type bacteriocins in several salmonellae].

A total of 1645 strains of Salmonella organisms isolated in Bulgaria were studied for the production of bactericins in the course of three years (1971-1973). The strains were isolated from farm and wild animals and birds, food products, environment, forage, human beings, poikilothermic animals, and insects. Seventy-two of them (4.37 per cent) proved bacteriocinogenic. The following types of bacteriocins were established: I+V, B+F+V, F+I+V, B+Y+V, E+F+Y+V, E+F+V, B+V, B+YmF.

Bacteriocins↗
PubMedDate not supplied

Salmonella in foods: new enrichment procedure for TECRA Salmonella Visual Immunoassay using a single rv(R10) only, TT only, or dual rv(R10) and TT selective enrichment broths (AOAC official method 998.09): collaborative study.

A collaborative study was conducted to compare a new enrichment procedure for the TECRA Salmonella Visual Immunoassay (TSVIA) with the reference method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (7th Ed.). Three food types (milk powder, pepper, and soy flour) were analyzed in Australia and 3 food types (milk chocolate, dried egg, and raw turkey) were analyzed in the United States. Thirty-eight collaborators participated in the study. The TECRA method was evaluated using both Rappaport-Vassiliadis R10 (RV(R10)) and tetrathionate (TT) broths for selective enrichment. M broth cultures arising from each of the 2 selective enrichment broths were tested in the TSVIA using 2 individual wells, one for each selective broth, and a single well to test the pooled selective enrichment broths. The results for the pooled enrichment broths were reported elsewhere. This study presents the results for the use of single enrichment broths, i.e., RV(R10) only or TT only, with the TSVIA. No significant differences (p > 0.05) were observed for the pairwise comparison of the proportion of positive samples for either RV(R10) or TT used as a single enrichment broth for the TSVIA with that for the reference method.

Animals↗
PubMed1979

[The antigenic relationships between Candida albicans, Salmonella cholerae suis, and Salmonella cholerae suis var. Kunzendorf O antigens].

In this study a common antigenic factor (s) between C. albicans CBS-562 and S. cholerae suis 211 (0:6(2),7) was shown with agglutination and absorption tests. This common antigenic factor (s) is different than previously shown Salmonella 0:7 antigen. This new factor (s) is not present in S. cholerae suis var. kunzendorf 1350 (0:6(1), 7) and S. cholerae suis var. kunzendorf 5210 (0:6(2),7) serotypes.

Antigens, Bacterial↗