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Biomedical subjects

L S Baron

Publications and source records attributed to L S Baron.

At least 37 records · Page 2Linked to original sources
PubMed1985

Development of a DNA probe to detect Salmonella typhi.

This study was undertaken to identify a DNA sequence that could be used to facilitate the diagnostic identification of Salmonella typhi, the causative agent of typhoid fever. All virulent S. typhi strains encode a relatively unique capsular antigen termed the virulence (Vi) antigen. Two distinct genetic loci, viaA and viaB, are involved in the synthesis of this antigen. The structural genes, located at viaB, were considered as a possible specific DNA probe. The viaB locus, contained in a recombinant cosmid, was subcloned to various plasmid vectors for this purpose. Selected viaB-region DNA fragments were then analyzed for specificity in DNA colony hybridization reactions with more than 170 strains representing a variety of enteric bacteria. An 8.6-kilobase EcoRI fragment was highly specific for the viaB gene region and was considered a good hybridization probe. This DNA probe should prove useful in rapid diagnostic assays set up to detect S. typhi in mixed bacterial samples (e.g., stools) within a few hours of specimen collection.

Antigens, Bacterial↗
PubMed1984

Unusual lipopolysaccharide antigens of a Salmonella typhi oral vaccine strain expressing the Shigella sonnei form I antigen.

Salmonella typhi 5076-1C, a potential live, oral vaccine for protection against typhoid fever and Shigella sonnei shigellosis, expresses the S. sonnei form I antigen and normal S. typhi somatic antigens. Polysaccharide antigens of this galactose epimeraseless genetic derivative strain were hot phenol-water extracted from cells grown with (+gal) and without (-gal) galactose. Ultracentrifugation of the aqueous layer from (+gal) cells resulted in a lipopolysaccharide (LPS) pellet having core-linked S. typhi O-antigen but no core-linked form I antigen; the LPS from (-gal) cells lacked O-antigen. The form I antigen, obtained from the supernatant, was purified by alcohol precipitation and ion exchange chromatography. Unlinked form I and S. typhi O-polysaccharide antigens, both present in the (+gal) supernatant, were further separated by gel filtration. Chemical analyses revealed the 5076-1C form I antigen to be a polymer (Mr = 14,000-20,000) having O-disaccharide repeating units comprised of 2-acetamido-4-amino-2, 4,6-trideoxy-D-galactose and 2-acetamido-2-deoxy-L-altruronic acid. Unlike parental S. sonnei form I LPS, the 5076-1C form I antigen lacked core lipid A, had low phosphorus content, and migrated in polyacrylamide gels with lower relative mobility. In contrast to current concepts of LPS assembly, these data indicate that 5076-1C form I antigen is transported to the cell surface without covalent linkage to core lipid A, and exists as a polymerized, antigenic surface entity.

Administration, Oral↗
PubMed1984

Oral vaccination of monkeys with an invasive Escherichia coli K-12 hybrid expressing Shigella flexneri 2a somatic antigen.

A living oral vaccine, designed to protect against Shigella flexneri 2a infections, was constructed by using Escherichia coli K-12 as a carrier strain. The hybrid strain, designated EC104, contained both chromosomal and plasmid genes from S. flexneri donor strains. In addition to expressing the S. flexneri 2a somatic antigen, it had inherited the property of epithelial-cell invasion. After the oral administration to rhesus monkeys, EC104 was isolated from the feces for up to 3 days, but by day 4 all stool cultures were negative. The serum antibody response against S. flexneri 2a somatic antigen was variable, but the vaccine conferred significant protection against an oral challenge with virulent S. flexneri 2a.

Administration, Oral↗
PubMed1984

Expression of lipopolysaccharide O antigen in Escherichia coli K-12 hybrids containing plasmid and chromosomal genes from Shigella dysenteriae 1.

The requirement for both plasmid and chromosomal genes in the biosynthesis of Shigella dysenteriae 1 lipopolysaccharide O antigen was demonstrated in Escherichia coli-Shigella hybrids. A 6-megadalton S. dysenteriae 1 plasmid, designated pWR23, was phenotypically tagged with the Tn3 ampicillin-resistance transposon. The tagged plasmid, designated pWR24, was transferred by transformation or conjugal mobilization to a rough E. coli K-12 recipient. Although the resultant hybrids were agglutinated in S. dysenteriae 1 antiserum, they did not remove all of the anti-Shiga agglutinins in absorption experiments. Modified lipid A core structure was detected in these hybrids, but Shiga O antigen was not expressed. When the his+ locus of the S. dysenteriae 1 chromosome was transferred by transduction to E. coli K-12 containing pWR24, complete Shiga O antigen was expressed. Lipopolysaccharide extracted from these hybrids was indistinguishable chemically, electrophoretically, and serologically from native S. dysenteriae 1 lipopolysaccharide.

Antigens, Bacterial↗
PubMed1983

Genetic studies of hybrids between coliphage phi 80 and Salmonella phage P22.

Hybrids between Escherichia coli phage phi 80 and Salmonella typhimurium phage P22 were isolated after superinfection by P22 of a smooth E. coli-S. typhimurium hybrid lysogenic for phi 80. These hybrid phages, designated phi 80immP22 and phi 80immP22dis, possessed the phi 80 protein coat and tail genes. The phi 80immP22 hybrids acquired the immunity (immC) region of P22 and some adjacent P22 genes, but E. coli-S. typhimurium strains lysogenic for phi 80immP22 hybrids remained sensitive to P22. The phi 80immP22dis hybrids, found ten times more frequently than the phi 80immP22 hybrids, contained a more extensive portion of the P22 genome which encompassed the immI as well as the immC region of P22. Therefore, the phi 80immP22dis hybrids conferred on their hosts immunity to P22 infection. Further analyses have revealed that the phi 80immP22dis hybrids carry the P22 attachment region and either P22 tail gene 9 or antigen conversion gene a1, but not both of these genes.

Coliphages↗
PubMed1982

Genetic and molecular studies of the regulation of atypical citrate utilization and variable Vi antigen expression in enteric bacteria.

1. The atypical citrate-utilizing ability to two strains of E. coli has been shown to be plasmid-encoded. Strain V414 carries a 130 Mdal conjugative Cit+ plasmid that also specifies Tcr and Cmr. Strain V517 carries 9 different plasmid species but only the 36 Mdal species is correlated with Cit+ ability. These plasmids are different from previously reported Cit+ plasmids of E. coli and Salmonella, which express thermosensitive conjugal transfer systems. 2. A 9 kb Pstl fragment, carrying the Cit+ genes of pWR60, has been cloned into the pBR325 plasmid. 3. Metabolic studies indicate that intact citrate is not incorporated directly into whole cells. Rather, atypical citrate utilization by these E. coli strains appears to involve partial metabolism of citrate at the cell surface before or during uptake. 4. The expression of atypical Cit+ ability by the parental pWR60 plasmid or by the recombinant pWR61 plasmid appears reversible and may involve an expression switch mechanism (i.e., insertion sequence element). 5. Two widely separated genetic loci, viaA and viaB, are necessary for Vi antigen synthesis in Salmonella and Citrobacter. In some strains of C. freundii, Vi antigen expression is reversible, a phenomenon which can be visualized by a colonial morphology transition between Vi-expressing and -nonexpressing forms. 6. The C. freundii viaB locus appears to encode the Vi antigen as well as the genetic "switch" mechanism controlling reversible Vi antigen expression. The viaA locus, which is found in several different bacterial species, may encode some common property (e.g., cell surface structure or enzymatic activity) that is needed for Vi antigen expression. 7. S. typhi and E. coli K12 hybrid strains which carry the C. freundii viaB locus have been constructed. These hybrid strains express reversible Vi antigen expression, even in the absence of general recombination (i.e., functional recA gene product). 8. The C. freundii viaB locus was transposed via Mu-mediated events to an F'lac plasmid in the E. coli K12 hybrid strain WR2376. F' plasmids carrying the viaB locus should serve as a highly enriched source of viaB DNA for physical examination of the switch mechanism. 9. Genetic manipulations such as those described herein can be used to study virtually any plasmid, viral, or chromosomally-encoded property. The resultant better understanding of biochemical pathways and of genetic regulatory control systems, and the isolation of desired gene sequences should provide ample information and materials for improving chemical processes and constructing vaccines against various organisms.

Antigens, Bacterial↗
PubMed1982

Intestinal immunoglobulin A responses in rabbits to a Salmonella typhi strain harboring a Shigella sonnei plasmid.

Salmonella typhi 5076-IC, which contains a plasmid that encodes the form I antigen of Shigella sonnei and which expresses S. typhi 9 and 12 and S. sonnei form I antigens, was used to immunize rabbits via chronically isolated ileal loops. Intestinal immunoglobulin A activity was detected against S. typhi, S. sonnei form I, and S. typhi strain 5076-IC. Thus S. typhi 5076-IC can effectively elicit mucosal immunoglobulin A to both S. typhi and S. sonnei.

Animals↗
PubMed1982

Evaluation of two Salmonella typhimurium hybrids as challenge organisms in a system for the assay of typhoid vaccines.

A mouse-virulent Salmonella typhimurium hybrid (H42), which expresses the Salmonella typhi Vi antigen in addition to S. typhi O antigens 9 and 12, and a mouse-virulent S. typhimurium hybrid (H1), which expresses only the 9 and 12 antigens of S. typhi, were compared in their behavior as challenge organisms in a system developed to assay the protective capabilities of typhoid vaccines. Swiss-Webster white mice, vaccinated intraperitoneally with live Escherichia coli hybrids expressing the S. typhi O antigens 9 and 12, were significantly protected against death from intraperitoneal challenge with each of the S. typhimurium hybrid strains. Vaccination with an E. coli hybrid expressing the S. typhi Vi antigen in addition to O antigens 9 and 12 was seen to confer no advantage in protection against either S. typhimurium hybrid challenge organism over that obtained by vaccination with an E. coli hybrid expressing only the O antigens of S. typhi. However, a notable difference in the behavior of the two S. typhimurium hybrids was seen in mice vaccinated with the parent of the E. coli hybrid vaccinating strains, E. coli F464, which expresses no surface antigens common to either of these S. typhimurium hybrid challenge organisms. A nonspecific (with respect to the vaccinating strain) protective effect, believed to be associated with Vi antigen expression by the challenge organism, was seen against the challenge with S. typhimurium hybrid H42 after F464 vaccination, whereas no protection was conferred by F464 vaccination against the challenge with Vi-nonexpressing S. typhimurium hybrid H1. Inasmuch as neither S. typhimurium hybrid discriminates between the expression or nonexpression of the Vi antigen in a vaccinating strain, it is concluded that the Vi-nonexpressing S. typhimurium hybrid H1, which more clearly indicates the vaccine-specific protective role of the S. typhi O antigens and does not exhibit the nonspecific protection response of hybrid H42, is the better choice as challenge organism for this vaccine assay system.

Animals↗
PubMed1982

Evaluation of two Salmonella typhi strains with reduced virulence for use in teaching and proficiency testing.

A total of 21 cases of laboratory-acquired typhoid fever associated with teaching and proficiency tests occurred in the United States during a 33-month period, prompting a search for less virulent strains of S. typhi which would be suitable for teaching purposes. Two strains were evaluated which are reported to have reduced virulence for mice. Strain Ty21a is a genetically constructed mutant that lacks the enzyme UDP-glucose-4-epimerase. This strain has reduced virulence for humans if grown under special laboratory conditions (in the presence of 0.1% d-galactose) and has been evaluated as a candidate for use as a live, oral vaccine. Strain H901 was originally isolated in Russia in 1918. It has not been tested in humans, but its nonmotile variant, O901, has been found to be somewhat less virulent for humans; however, it can cause infection with doses of 10(7) organisms. In teaching exercises, all strains should be treated as though they are fully virulent. Ty21a and H901 were satisfactory, but not ideal, for teaching purposes. Biochemically, they could be identified by conventional tests and by commercially available diagnostic systems, although Ty21a was H(2)S negative. Serologically, both strains posed problems. Both Ty21a and H901 were Vi antigen negative, and Ty21a was rough and grew poorly. Both strains were susceptible to antibiotics, including chloramphenicol, ampicillin, and trimethoprim-sulfameth-oxazole. When Ty21a and H901 were mixed with Escherichia coli and plated, Hektoen and salmonella-shigella agars were most useful for their recovery. The appearance of Ty21a and H901 on differential plating media was typical, although Ty21a had smaller colonies. The plating efficiency on MacConkey agar for Ty21a was 0.6 compared with 1 for H901. Neither strain can be recommended unequivocally for teaching purposes; instead, the advantages and disadvantages of each must be considered. Both strains have been deposited in the American Type Culture Collection (Ty21a = ATCC 33459 = CDC 2861-79; H901 = ATCC 33458 = CDC 2862-79).

Agglutination Tests↗
PubMed1981

Evidence that ribosomal protein S10 participates in control of transcription termination.

We report the isolation of an Escherichia coli K-12 strain with a mutation, nusE71, that results in a change in ribosomal protein S10. Phage lambda fails to grow in hosts carrying the nusE71 mutation because the lambda N gene product is not active. The N product regulates phage gene expression by altering transcription complexes so that they can overcome termination barriers. This suggests that a ribosomal protein is involved in antitermination of transcription.

Bacteriophage lambda↗
PubMed1981

Construction of a potential bivalent vaccine strain: introduction of Shigella sonnei form I antigen genes into the galE Salmonella typhi Ty21a typhoid vaccine strain.

Shigella sonnei, an intestinal pathogen, produces a characteristic form I cell surface antigen now known to be plasmid encoded. We considered that the GalE Salmonella typhi Ty21a oral vaccine strain, highly effective against typhoid, might be modified so as to be protective also against shigellosis due to S. sonnei. The plasmid responsible for form I antigen synthesis was therefore conjugally transferred to the galE S. typhi strain. Serological studies revealed that the derivative strain produces the form I antigen in addition to the normal S. typhi somatic antigens. Testing in mice demonstrated that the derivative form I galE S. typhi strain is protective against both S. sonnei and S. typhi challenges. These data suggest that the galE S. Ty21a oral vaccine strain, which presumably stimulates the local immune system in the intestine, may also serve as a useful carrier for other antigenic determinants to protect against different intestinal infections.

Animals↗
PubMed1981

Genetic regulation of variable Vi antigen expression in a strain of Citrobacter freundii.

Certain strains of the genus Citrobacter exhibit a variable expression of the Vi surface antigen that appears to involve a special mechanism for regulation of gene expression. Two nonlinked chromosomal loci, viaA and viaB, are known to determine nonvariable Vi antigen expression in strains of Salmonella. To confirm the presence of analogous loci in Citrobacter and to ascertain whether either of them is involved in variable Vi antigen expression in this organism, donor strains were constructed from Citrobacter freundii WR7004 and used to transfer their Vi antigen-determining genes to ViaA- and ViaB- Salmonella typhi recipient strains. Vi antigen expression in C. freundii was found to be controlled by loci analogous to the Salmonella via genes. S. typhi recipients of the C. freundii viaA+ genes were restored to the full, continuous expression of the Vi antigen normally seen in S. typhi. Thus, the C. freundii viaA genes appeared to play no role in the variable expression of the Vi antigen. In contrast, S. typhi recipients of the C. freundii viaB+ genes exhibited the rapid, reversible alternation between full Vi antigen expression and markedly reduced Vi antigen expression that was seen to occur in the C. freundii parent. The C. freundii viaB locus was thus identified as the one whose genes are regulated so as to produce variable Vi antigen expression. Genes determining another C. freundii surface antigen, the synthesis of which is not affected by the mechanism regulating Vi expression, were coinherited with the C. freundii viaB+ genes. An invertible, insertion sequence element located within the C. freundii viaB locus is proposed to account for the regulation of variable Vi antigen expression.

Antigens, Bacterial↗
PubMed1980

L factor that is required for beta-galactosidase synthesis is the nusA gene product involved in transcription termination.

The DNA-dependent in vitro synthesis of Escherichia coli beta-galactosidase requires the presence of a soluble protein referred to as L factor [Kung, H., Spears, C. & Weissbach, H. (1975) J. Biol. Chem. 250, 1556-1562]. In the present study, comparison of physical, immunological, and biological properties shows that L factor is the product of the E. coli nusA gene. The nusA gene product is known to interact with bacteriophage lambda N gene protein and to prevent premature termination of transcription from the early lambda promoters. Our results suggest that premature transcription termination in the lac operon of E. coli may also be overcome by the nusA protein.

Bacterial Proteins↗
PubMed1979

Mouse protective capabilities of Escherichia coli hybrids expressing Salmonella typhi antigens.

An Escherichia coli hybrid, F1061, expressing Salmonella typhi somatic antigens 9 and 12, and a derivative of this hybrid, E. coli hybrid WR3078, expressing the S. typhi Vi antigen in addition to somatic antigens 9 and 12, were compared with S. typhi Ty2 in experiments to test their ability, as live vaccines, to protect Swiss white mice against death from challenge with a mouse-virulent Salmonella typhimurium hybrid expressing the S. typhi antigens 9, 12, Vi, and d. When the live, vaccinating organisms were administered intraperitoneally, 87.5% of the mice immunized with S. typhi Ty2 survived challenge, as compared with 62.5% of those immunized with E. coli hybrid F1061 and 55% of those inoculated with E. coli hybrid WR3078. When live organisms were administered orally at a dose of 10(9), 67.5% of the mice immunized with S. typhi Ty2 survived challenge as compared with 47.5% of those immunized with E. coli hybrid F1061 and 40% of those administered E. coli hybrid WR3078. Thus, the protection conferred by E. coli hybrid F1061 expressing only the S. typhi somatic antigens, although significant in this system, was inferior to that conferred by S. typhi Ty2 and the addition of the S. typhi Vi antigen to this hybrid (creating E. coli hybrid WR3078) did not enhance that protection.

Administration, Oral↗
PubMed1978

lambdaimm P22dis: a hybrid of coliphage lambda with both immunity regions of Salmonella phage P22.

Genetically marked lambda and P22 phages were recombined in Escherichia coli-Salmonella typhimurium hybrid WR4028, a host sensitive to infection by both of these phages. Hybrid phages that acquired the immC region of P22, but retained the genes for the lambda protein coat were selected on WR4027 (lambda), a lambda-immune, P22-resistant derivative of WR4028. In these lambdaimmP22 hybrids, at least the c through P genes of lambda were replaced with functionally related P22 genes. Phage recombinants with more extensive regions of the P22 genome were selected on the double lysogen WR4027 (lambda, lambdaimmP22). One such hybrid, lambdaimmP22dis, was determined by heteroduplex analysis to contain approximately 40% of the P22 genome. Genetic studies established that lambdaimmP22dis possesses the two widely separated immunity control regions of P22 (immC and immI) and that these loci are expressed in E. coli K-12 lysogenic for lambdaimmP22dis. In addition, lambdaimmP22dis contains the P22 a 1 locus responsible for somatic 0--1 antigen conversion in Salmonella. Although the lambdaimmP22dis phage particle has the lambda head and tail, the phage genome also carried P22 tail gene 9 as evidenced by the production of free P22 tails. It also has the P22 att site as indicated by the integration of the lambdaimmP22dis prophage near the proA locus on the bacterial chromosome.

Antigens, Bacterial↗
PubMed1977

Genetic studies of hybrids between coliphage lambda and salmonella phage P22: genetic analysis of the P22-lambda hybrid class.

P22-lambda hybrids which retain the protein coat of P22 have been isolated and characterized into two types. Type 1 hybrids which have the c through O-P genes of lambda are unable to grow lytically on Salmonella typhimurium. On the other hand, type 2 hybrids which contain only the c region of lambda, plated on S. typhimurium. Both hybrid types retained the generalized transducing and antigenic conversion capabilities of P22.

Antigens, Viral↗
PubMed1977

Genetic basis of Vi antigen expression in Salmonella paratyphi C.

Analysis of hybrids formed in a cross between a Salmonella paratyphi C Hfr and an S. typhimurium recipient indicated that the structural genetic determinants of the S. paratyphi C Vi antigen are located closely adjacent to the mel determinant, between this marker and purA. A similar location was indicated for the structural genetic determinants of the S. typhi Vi antigen (the viaB locus) by the results of a mating in which a hybrid S. typhimurium Hfr bearing the S. typhi viaB determinants was used to transfer these genes to an S. typhimurium recipient. Mating experiments with a Vi-antigen-expressing S. typhi Hfr and an S. typhimurium hybrid recipient expressing the Vi antigen of S. paratyphi C yielded no recombinants in which loss of Vi antigen expression occurred, indicating that the chromosomal locus occupied by the genetic determinants of the S. paratyphi C Vi antigen is the same one at which, in S. typhi, the viaB genes reside. Introduction of a mutant S. typhi viaA gene into an S. typhimurium hybrid expressing the Vi antigen, as the consequence of prior receipt of the S. paratyphi C viaB determinants, resulted in that hybrid's loss of Vi antigen expression, demonstrating that the viaA determinant plays a role in Vi antigen expression in S. paratyphi C, as well as in S. typhi. Although the percentages of coinheritance of the viaB and mel determinants in the mating experiments suggested that their linkage is sufficiently close to allow cotransduction by P22, attempts to accomplish this with lysates prepared on S. typhimurium hybrids expressing either S. typhi or S. paratyphi C viaB determinants were not successful.

Antigens, Bacterial↗