1a.Objectives (from AD-416):
1. Develop serological assays that differentiate swine chronically infected with zoonotic T. spiralis from swine only transiently exposed to T. murrelli.
2. Estimate how frequently pastured pigs and feral swine are exposed to infection with enzootic and zoonotic species of trichinella.
1b.Approach (from AD-416):
Screen expression libraries of T. spiralis and T. murrelli with hyper-immune sera from swine to identify clones encoding species-specific diagnostic antigens. Develop antibodies that discriminate among the two types of infection. Test in experimentally infected swine. Apply to panels of serum collected from feral swine and pastured pigs. Validate against genotyped parasite specimens obtained from tissue samples matched with serum.
This work seeks to develop serological assays to differentially diagnose swine exposure to Trichinella spiralis or Trichinella murrelli. The reason for doing so is that swine exposed to T. spiralis are likely to harbor abundant, life-long larval burdens posing significant risk to food safety, whereas swine exposed to T. murrelli generally mount successful immune responses. Current methods to screen serum, however, make no distinction between the two conditions. Experimental infections needed to procure larvae of each parasite type were completed; crude extracts and preparations of Excretory-Secretory products (containing immunodominant antigens) were obtained. Peptides from each were fluorescently labeled and run on 2D gels, transferred to membranes, and probed with hyperimmune serum from pigs harboring infections with either T. spiralis or T. murrelli. By refining conditions of electrophoresis, candidate diagnostic epitopes (differentially or exclusively expressed in one parasite or the other, as well as epitopes exclusively recognized by one type of immune serum or the other) are being evaluated. This year, assays were also performed on protein preparations from which sugar residues (glycans) were first removed, because some of these sugars, though highly immunogenic, are common to each parasite (and therefore unsuited for use in discriminating one type of infection from the other). Finally, a substantial portion of the T. spiralis genome was tested for its suitability as a scaffold for short sequence reads derived from the T. murrelli genome. Parameters derived from this 'proof of principle' will be used in ongoing efforts to assemble and compare the T. murrelli genome. Preliminary data derived from this project were also presented at the 13th International Congress on Trichinellosis.