Comparisons of Salmonella conjugation and virulence gene hyperexpression mediated by rumen protozoa from domestic and exotic ruminants.

Recent studies have identified a phenomenon in which ciliated protozoa engulf Salmonella and the intra-protozoal environment hyperactivates virulence gene expression and provides a venue for conjugal transfer of antibiotic resistance plasmids. The former observation is relegated to Salmonella bearing the SGI1 multiresistance integron while the latter phenomenon appears to be a more generalized event for recipient Salmonella. Our previous studies have assessed virulence gene hyperexpression only with protozoa from the bovine rumen while conjugal transfer has been demonstrated in rumen protozoa from cattle and goats. The present study examined virulence gene hyperexpression for Salmonella exposed to rumen protozoa obtained from cattle, sheep, goats, or two African ruminants (giraffe and bongo). Conjugal transfer was also assessed in these protozoa using Salmonella as the recipient. Virulence gene hyperexpression was only observed following exposure to the rumen protozoa from cattle and sheep while elevated virulence was also observed in these animals. Conjugal transfer events were, however, observed in all protozoa evaluated. It therefore appears that the protozoa-based hypervirulence is not universal to all ruminants while conjugal transfer is more ubiquitous.

Involvement of a Salmonella genomic island 1 gene in the rumen protozoan-mediated enhancement of invasion for multiple-antibiotic-resistant Salmonella enterica serovar Typhimurium.

Multiple-antibiotic-resistant Salmonella enterica serotype Typhimurium is a food-borne pathogen that may be more virulent than related strains lacking the multiresistance phenotype. Salmonella enterica serotype Typhimurium phage type DT104 is the most prevalent of these multiresistant/hypervirulent strains. Multiresistance in DT104 is conferred by an integron structure, designated Salmonella genomic island 1 (SGI1), while we recently demonstrated DT104 hyperinvasion mediated by rumen protozoa (RPz) that are normal flora of cattle. Hyperinvasion was also observed in other Salmonella strains, i.e., other S. enterica serovar Typhimurium phage types and other S. enterica serovars, like S. enterica serovar Infantis, possessing SGI1, while DT104 strains lacking SGI1 were not hyperinvasive. Herein we attempted to identify SGI1 genes involved in the RPz-mediated hyperinvasion of Salmonella strains bearing SGI1. Transposon mutagenesis, coupled with a novel reporter system, revealed the involvement of an SGI1 gene previously designated SO13. Disruption of SO13 expression led to an abrogation of hyperinvasion as assessed by tissue culture invasion assays and by bovine challenge experiments. However, hyperinvasion was not observed in non-SGI1-bearing strains of Salmonella engineered to express SO13. That is, SO13 and another SGI1 gene(s) may coordinately upregulate invasion in DT104 exposed to RPz.

Exposure to rumen protozoa leads to enhancement of pathogenicity of and invasion by multiple-antibiotic-resistant Salmonella enterica bearing SGI1.

Multiple-antibiotic-resistant Salmonella enterica serotype Typhimurium is a food-borne pathogen that has been purported to be more virulent than antibiotic-sensitive counterparts. The paradigm for this multiresistant/hyperpathogenic phenotype is Salmonella enterica serotype Typhimurium phage type DT104 (DT104). The basis for the multiresistance in DT104 is related to an integron structure designated SGI1, but factors underlying hyperpathogenicity have not been completely identified. Since protozoa have been implicated in the alteration of virulence in Legionella and Mycobacterium spp., we attempted to assess the possibility that protozoa may contribute to the putative hypervirulence of DT104. Our study reveals that DT104 can be more invasive, as determined by a tissue culture invasion assay, after surviving within protozoa originating from the bovine rumen. The enhancement of invasion was correlated with hypervirulence in a bovine infection model in which we observed a more rapid progression of disease and a greater recovery rate for the pathogen. Fewer DT104 cells were recovered from tissues of infected animals when protozoa were lysed by preinfection chemical defaunation of the bovine or ovine rumen. The protozoan-mediated hypervirulence phenotype was observed only in DT104 and other Salmonella strains, including serovars Agona and Infantis, possessing SGI1.