Lack of a role of cytotoxic necrotizing factor 1 toxin from Escherichia coli in bacterial pathogenicity and host cytokine response in infected germfree piglets

Some Escherichia coli strains isolated from intestinal or extraintestinal i nfections in pigs produce cytotoxic necrotizing factor 1 (CNF1), In order t o analyze the role of CNF1 in the pathogenesis of porcine colibacillosis, n ewborn colostrum-deprived germfree piglets were orally inoculated with a wi ld-type CNF1-producing strain (M623) or with an isogenic cnf1 mutant (M623 Delta CNF1). The two isogenic strains induced a high mortality with similar lung and serosal inflammatory lesions, indicating that both strains were p athogenic in these piglets. Bacterial counts in various organs of inoculate d piglets revealed an intestinal predisposition of M623 and M623 Delta CNF1 strains for the cecum and colon. Extraintestinal organs (lungs, liver, spl een, and kidney) were also colonized by both strains. Similar colonization of intestinal and extraintestinal tissues in animals inoculated with either strain was observed, except in the ileum, where M623 showed a higher colon ization than M623 Delta CNF1. Intestinal (ileum and colon), extraintestinal (lung and kidney), and immune (mesenteric lymph nodes and spleen) tissues were sampled at 1 day postinoculation and analyzed for cytokine expression by a reverse transcriptase PCR technique. Inoculation with E. coli M623 ind uced an enhanced expression of inflammatory cytokines (interleukin-1 alpha [IL-1 alpha], tumor necrosis factor alpha, and IL-12p40) in the intestinal organs compared to uninoculated piglets or piglets inoculated with nonpatho genic intestinal E. coli 862B, which is also able to colonize the intestina l tract. There was little difference in cytokine transcript levels in the i ntestinal and extraintestinal organs in piglets inoculated with E. coli str ains M623 or M623 Delta CNF1, except in the ileum, where IL-1 alpha and IL- 8 mRNA levels correlated with bacterial colonization. Expression of regulat ory cytokines (gamma interferon and IL-4) was weak in immune tissues from p iglets inoculated with M623 or M623 Delta CNF1. Taken together, our data in dicate that the CNF1-producing strain, M623, is pathogenic and induces infl ammatory cytokine expression in germfree, colostrum-deprived piglets. Never theless, in this model, the CNF1 toxin does not appear to be a major factor for pathogenicity or cytokine response, as demonstrated by the use of an i sogenic cnf1 mutant.