Toxigenicity testing of clinical isolates of non-typhoidal salmonellae in Vero cell culture & Caenorhabditis elegans model.

ABSTRACT

The non-typhoidal salmonellae (NTS) are recognized agents of gastroenteritis worldwide. Some of the NTS do not produces cytotoxic changes in tissue culture and not much is known about the endotoxicity of the clinical isolates of NTS (mostly Salmonella enterica serotype Typhimurium and Salmonella enterica serotype Enteritidis). We examined the exotoxic (cytotoxin) and endotoxic activity of clinical isolates of NTS in two assay models namely Vero cell culture and the nematode, Caenorhabditis elegans. Bacteria-free culture supernatants of 40 isolates NTS were tested in 96 well microtitre plate containing confluent monolayers of Vero cells. For the effects on C. elegans, the worms were exposed to bacteria free culture supernatants in 24 well microtitre plate for 24 h and then transferred to OP50 Escherichia coli lawn culture. The endotoxic activity of the live bacterium was studied by feeding the worms in the lawn culture of NTS separately. No cytopathic effect was observed with NTS tested in Vero cell culture assay. Likewise, the worms exposed to the bacteria-free culture supernatants were found active up to 7 days. In the co-culture killing assay, worms were found dead with characteristic stiff and straight appearance by 16(th) day. The worms were alive up to 21 days in OP50 E. coli. Bacteria-free culture supernatants did not have any deleterious effect on worms or in Vero cell culture, suggesting that there is no soluble toxic factor (diffusible toxin) in the culture supernatants. However, live NTS were found to be lethal to the worms; indicating that direct interaction between viable NTS and C. elegans is necessary for killing.