Yildiz, Fitnat, Bilecen, Kivanc, Kothary, Mahendra, McCardell, Barbara, Datta, Atin, Tall, Ben D., Sprando, Robert, Cinar, Hediye N.
[
International Worm Meeting,
2009]
Vibrio cholerae (VC), a natural dweller of aquatic ecosystem, causes acute gastroenteritis in humans, using virulence factors such as cholera toxin (CT) and toxin co-regulated pili (TCP). Vibrio cholerae O1 and O139 serogroups are associated with cholerae epidemics, while non-O1 non O-139 strains, which mostly lack CT and TCP, and vaccine strains with deleted CT and TCP loci are also capable of causing diseases such as diarrhea, soft tissue infections, sepsis, and inflammatory enterocolitis in humans in a sporadic fashion through mechanisms that are currently unclear. VC causes lethal infection in Caenorhabditis elegans via a CT / TCP independent process providing an excellent model to determine the roles of other virulence factors in pathogenesis (1, 2). We found that hemolysin (hlyA) deletion mutants showed attenuated killing in C. elegans which is an effect that can be complemented by the presence of an intact hlyA locus (3). Furthermore, O1 classical strains, which produce less hemolysin than O1 El tor strains, showed milder lethality and reduced developmental delay. It has been reported that VC hemolysin causes extensive vacuolization and lysis in cultured cells. We examined worms fed with either wild type or hlyA deletion mutants of VC for intestinal pathologies. Forty-six percent of the worms exposed to wild type VC exhibited intestinal vacuoles while none one of the worms fed an hlyA deletion mutant showed this defect after 48-hour exposure. Other intestinal pathologies such as distention of the intestinal lumen and intestinal cell shrinkage were observed in both hly+ and hly- conditions, although the defects were more severe in hly+ conditions. Intestinal lumen distention and cell shrinkage preceded vacuolization suggesting that intestinal colonization of the bacteria may be necessary for vacuole formation. Hemolysin A is a pore forming exotoxin whose role in VC pathogenesis is not fully understood. Further studies in C. elegans infection model will contribute to a greater understanding of the role of hemolysin virulence in the pathogenesis of cholera. (1) Vaitkevicius K. et al. PNAS, 103 (2006) 9280-9285 (2) Cinar HN. et al. 16 th International C. elegans Meeting, 2007 (3) Cinar HN. et al. Aging Stress and Pathogenesis Meeting, 2008.
Franco, Augusto A., Cinar, Hediye N., Tall, Ben D., Hahn, Justin, Kim, Sungji, Grim, Christopher, Kothary, Mahendra, Sahu, Surasri N., Datta, Atin
[
International Worm Meeting,
2013]
Vibrio parahaemolyticus (Vp), a Gram-negative marine bacterium, is an emerging pathogen causing gastroenteritis, wound infection, and septicemia. Gastroenteritis caused by Vp is associated with the production of thermostable direct (TDH) and TDH-related hemolysins (TRH), and the inflammatory responses are produced by effector proteins secreted by a type III secretion system (T3SS-2a, or b) contained within pathogenicity islands (1). In order to study innate immune responses to known virulence factors, and to discover new virulence factors, we used Caenorhabditis elegans as a host organism. Vp isolates, previously serotyped and characterized by PCR for the presence of T3SS-2, TRH, TDH, and urease genes, were tested using a C. elegans lethality assay for their virulence. Six groups of Vp strains, each representing the following genotypes, were tested in the C. elegans model: 1) urease+ trh+ tdh+ T3SS-2b+, 2) urease+ trh+ tdh- T3SS-2b + , 3) urease+ trh- tdh- T3SS-2b + , 4) urease+ trh- tdh+ T3SS-2a+ , 5) urease- trh- tdh- T3SS-2- , 6) urease- trh- tdh+ T3SS-2a + . Median survival rates of worms tested against Vp isolates ranged between five and sixteen days. Vp strains with functional urease gene loci resulted in shorter median survival rates of C. elegans. Our data suggest a strong correlation between the presence of Vp gene and C. elegans lethality. These results taken together with those previously reported by Honda et al.(2) that urease-positive, TDH- and TRH-negative strains were also capable of causing intestinal fluid accumulation in the rabbit ligated ileal loop assay, suggest a significant role for urease in disease development. Alternatively, urease might be needed to activate other virulence factor/s or a marker linked with a currently unknown virulence factor gene(s) in Vp. Further molecular and functional characterization of urease in C. elegans infection, is in progress. (1) Ham H, Orth K, 2012 (2) Honda S, et al. 1992.