Cloning and analysis of a Trichinella britovi gene encoding a cytoplasmic heat shock protein of 72 kDa.

A gene encoding a protein of 646 amino acid residues with a molecular mass of 71.3 kDa showing homology to the cytoplasmic form of the 70 kDa heat shock protein was cloned and sequenced from the nematode parasite Trichinella britovi (Tb). The gene was expressed in vitro as a protein of 71 kDa that was immunoprecipitated by a Trichinella-infected rabbit serum. Monospecific polyclonal antibodies raised against the recombinant Tb Hsp70 expressed in Escherichia coli, recognized a protein of 70 kDa by Western blot analysis of Tb soluble antigen (muscular stage). Tb Hsp70 was located in the nuclei of the muscle larvae as determined by the indirect immunofluorescent pattern on cross-sections of the worm. The expression of this protein was not detected in adult worm nuclei suggesting a differential expression of Hsp70 between the 2 stages of Trichinella.

Characterization of eleven antigenic groups in Trichinella genus and identification of stage and species markers.

Several monoclonal antibodies (Mabs) were raised against the L1 muscle stage (L1M) of Trichinella spiralis (Ts) and Trichinella pseudospiralis (Tp). Western blot analysis of various antigenic preparations established that Mabs described by different authors recognized 8 antigenic fractions (TSL1-TSL8) in crude extracts of infective larvae. The TSL1 fraction was immunodominant and present on the cuticle of different parasite stages. Mabs against Trichinella T5 (T5) and Ts were selected in order to extend the previous studies to another Trichinella phenotype. Only 35% of the selected Mabs recognized linear epitopes and 71% reacted with soluble or excretory-secretory antigens in a dot blot procedure and ELISA test. The targets of the Mabs were identified by immunoprecipitation with [35S]methionine-labelled L1M worm lysate. Mabs prepared from mice immunized with the whole parasite (T5) recognized a wider panel of antigens in different parasitic organs. Seven antigenic structures were distinguished on the cuticle and several epitopes were identified in the gut, haemolymph and stichocytes. Eleven antigenic groups were established according to their indirect immunofluorescence pattern on cross-sections of the worm. Monoclonal antibodies raised against Ts soluble antigen mainly recognized epitopes in stichocytes and on the cuticle surface. All the selected Mabs recognized T5 and Trichinella britovi (Tb) strengthening the link between these 2 species. Four Mabs were used to differentiate antigenic structures among 6 Trichinella phenotypes and to develop a new tool to follow gene flow within the Trichinella genus.