Biological and genetic characterization of TnphoA mutants of Salmonella typhimurium TML in the context of gastroenteritis

Abstract

TnphoA transposon insertion mutants of phoN-negative derivatives of Salmonella typhimurium TML (of human gastroenteritic origin) were selected by growing mutagenized recipient bacteria under a variety of growth conditions. Ninety-seven individual mutants, which expressed alkaline phosphatase, were collected and tested for their ability to invade HEp-2 cells. Seven smooth mutants had a reduced ability to invade HEp-2 cells, and three smooth mutants were consistently more invasive than their corresponding parental strains. One rough mutant was of similar invasiveness and two were of reduced invasiveness when compared with that of parental strains. The seven smooth hypoinvasive mutants, the three smooth hyperinvasive mutants, and the three rough mutant strains were tested for their abilities to invade ileal enterocytes by the rabbit ileal invasion assay described previously (3). All smooth mutants exhibited parental levels of invasiveness. The rough mutants were hypoinvasive in the rabbit ileal invasion assay. The HEp-2 system is therefore not a good predictor of behavior in gut tissue in this model. DNA sequences flanking the transposon were determined for five mutants which were hypoinvasive in the HEp-2 cell assay. The mutations were found to be insertions in two previously identified invasion genes, invG and invH, and in a gene not normally associated with invasion, pagC. These observations lead one to be cautious in the interpretation of the biological significance of data obtained from invasion of tissue culture monolayers when extrapolated to gut tissue.

Cell-Contact-Stimulated Formation of Filamentous Appendages by Salmonella typhimurium Does Not Depend on the Type III Secretion System Encoded by Salmonella Pathogenicity Island 1

“…Immunocytochemical staining of S. typhimurium allowed the quantification of invading bacteria (Table 2). As previously described for MDCK or other cell lines, the S. typhimurium mutant strains examined, i.e., invA (20), invG (40), sspC (28), PhoP c (5), and prgH (5) strains, were all severely deficient for invasion of both MDCK and Caco-2 cells compared to their parent strains (Table 2).…”

“…These data demonstrate that expression of the short filamentous surface appendages does not correlate either with S. typhimurium invasiveness or SPI1-encoded protein secretion. S. typhimurium 83, which exhibits dramatically reduced invasiveness in vitro (10; this study) due to a mutation in invG encoding a type III secretion system component (40), produces appendages like those of wild-type strains when in contact with epithelial cells both in vivo and in vitro. Thus, InvG is not essential to form cell contact-induced short filamentous appendages, and, since InvG appears to be an integral part of the SPI1-encoded type III secretion apparatus (18,34,46), our data indicate that the entire system is not required for the formation of these appendages.…”

The Salmonella typhimurium InvH protein is an outer membrane lipoprotein required for the proper localization of InvG

“…Contrary to previous predictions by Altmeyer et al (1993), our own analysis also predicted invH to encode an outer membrane lipoprotein. The invH gene is essential for the pathogenicity of S. typhimurium (Altmeyer et al, 1993;Lodge et al, 1995;Watson et al, 1995). Its gene product has been previously described as a membrane protein when analysed ᮊ 1998 Blackwell Science Ltd, Molecular Microbiology, 28, 1367-1380 Fig.…”

Salmonella, the host and disease: a brief review

“…43 In vivo, this combination of effectors are essential for early inflammatory pathogenesis in mice and cows, explaining in part the overlap between intestinal invasiveness and inflammatory pathogenicity in some models of infection. [44][45][46][47][48][49][50][51][52][53] In vivo, SPI-1-mediated behaviors seem to be critical for early intestinal inflammation, [54][55][56] yet their absence does not influence systemic inflammation following intraperitoneal challenge, 31 and delayed intestinal inflammation occurs in their absence. 57,58 Interestingly, although incapable of inducing PEEC-mediated transmigration of neutrophils across model epithelia, comparison of the inflammatory gene expression profiles of cultured intestinal epithelial monolayers infected with wild-type serovar Typhimurium or those with a complete deletion of the SPI-1 pathogenicity island demonstrated little difference in the proinflammatory potential of these strains.…”