The Shiga-like toxins have DNA sequence homology to the toxins accountable for the dysentery brought about by the Shigella species. <i>Escherichia coli</i> which encode and produce shiga-like toxins are referred to as shiga toxin-producing E. coli (STEC). Upon infection with STEC, humans may develop a variety of clinical symptoms ranging in severity from bloody diarrhea to life threatening hemolytic uremic syndrome (HUS). Hemolytic uremic syndrome is the most fatal disease manifestation upon STEC infection for humans and has been documented to occur in up to 20% of patients upon STEC infection [29]. The Shiga toxins (Shiga toxin 1 and 2) are regarded as the principal virulence factor of STEC and are responsible for the clinical manifestations during HUS in humans [49].<p>
Cattle are the primary non-human reservoir for STEC and therefore represent an attractive target for pre-slaughter intervention as a means to reduce human infections. To date, vaccination with secreted proteins including Shiga toxin 2 (Stx2), has reduced the numbers of bacteria shed in feces [3]. Even though published data exists supporting vaccination in cattle as a means to reduce STEC, commercially available vaccines are not being used by farms and STEC remain a significant zoonotic pathogen of humans causing disease and death. To further our knowledge about STEC pathogenesis in cattle, we examined the effect of Shiga toxin 2 on bovine immune responses. Bovine lymphocyte function was determined in the presence of Shiga toxin 2 and the magnitude of bovine immunological responses was measure after immunization with Shiga toxin 2. In general, results suggest that Shiga toxin 2 downregulates bovine immune responses suggesting vaccination with effector molecules that exclude Shiga toxin 2 may induce a better immunological response and improve vaccine efficacy.<p>
To examine the possibility that Stx2 modulates bovine immune responses, we investigated lymphocyte function in the presence of Stx2. Menge et al [70] have reported that bovine lymphocytes express the Stx receptor and that Shiga toxin 1 inhibits lymphocyte proliferation in vitro. We isolated two populations of lymphocytes, peripheral blood mononuclear cells (PBMCs) and ileal Peyers patch lymphocytes (IPPL) and compared lymphocyte function in the presence and absence of Stx2. We found that Stx2 did not affect IPPL viability in vitro but did inhibit IPPL proliferation after 12 hours of incubation <i>in vitro</i>. In
contrast, no altered PBMC function could be observed in the presence of Stx2. These results suggest that receptor-bound Stx2 may inhibit IPPL proliferation and that the two populations of lymphocytes isolated are unique and distinct from each other in their response to Stx2.<p>
To determine the effect of Stx2 on bovine immune responses during STEC infection, a bovine ileal ligated loop model was employed. Ligated loops were inoculated with either a Stx2+ STEC strain or an isogenic Stx2- STEC strain. After 24 hours, IPPL populations were isolated from each ligated loop and immunophenotyped. The results indicated a significantly reduced CD4+ T cell population in the presence of Stx2. No differences in the levels of IFNá, TNFá, IL12 or IFNã could be detected between groups. These results suggest that Stx2 modulates bovine immune responses but not as a result of increased production of these cytokines. To extend this finding, we determined the effect of Stx2 on bovine immune responses during active immunization by using ELISA to measure serological responses in the presence and absence of Stx2. Serological responses to secreted proteins, as well as a co-administered antigen (hen egg lysozyme), were significantly reduced in the groups of cattle that were immunized with either purified Stx2 or secreted protein preparations isolated from STEC compared to groups vaccinated with antigens which did not contain the toxin. Bovine proliferative responses were also measured and the results indicated significantly reduced proliferation in the groups vaccinated with the formulations containing Stx2. Therefore, based on these results, we conclude that Stx2 downregulates bovine immune responses and thus may contribute to the colonization and persistence of cattle by STEC.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:SSU.etd-08252010-163545 |
| Date | 05 October 2010 |
| Creators | Chu, Audrey |
| Contributors | Deneer, Harry, Gannon, Vic, Griebel, Philip, Napper, Scott, Potter, Andrew |
| Publisher | University of Saskatchewan |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://library.usask.ca/theses/available/etd-08252010-163545/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Saskatchewan or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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