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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.

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