Cloning, Expression And Antigenicity Analysis Of Thioredoxin Peroxidase Gene Of Taenia Multiceps

Cerebral coenurosis, which is a kind of serious fatal zoonoses, is caused by the parasitism of Coenurus cerebralis, the larvae of Taenia multiceps in central nerve tissue of host. Coenurosis is widly spread in China, especially in pasture fields of Northwestern China, Northern China and Northestern China with a rising morbidity. This disease not only causes enormous economic loss in husbandry production but also is a threat to human heath. Thioredoxin peroxidase (TPx), a member of peroxiredoxin (Prx) family, is a hydrosulfuryl specific antioxidant which could remove the excess reactive oxygen species produced by the normal cells of organism. TPx of parasite could protect against the damage of reactive oxygen species from host and parasite metabolism, functioning in the evasion of host immunity.The specific primers derived from T. solium Prx gene were used to amplify the T. multiceps thioredoxin peroxidase (TmTPx) cDNA fragment from Coenurus cerebralis isolated from naturally infected sheep in Jingtai county,Gansu province. Sequence analysis showed that TmTPx gene fragment had 99% identity to TaHc2-D11 mRNA of T. asiatica (EF420372) and 96% identity to T. solium Prx (AY728092). The recombinant expression plasmid pGEX-4T-TmTPx was constructed and transformed to E. coli BL21, then induced by IPTG to express the recombinant protein. The expressed product was analyzed by SDS-PAGE and a 43 kDa fusion protein was successfully expressed. The purified recombinant protein was used to immunize rabbit and the recombinant protein could detect the antibody response in the immunized rabbit and the antibody level increased with time of immunization. Furthermore, the specific rabbit antiserum against the purified TmTPx recombinant protein could specificly reacted to the coenurus protoscoleces antigen, suggesting that TmTPx recombinant protein had a good immunogenicity. Furthermore, specific primers were designed according to TaHc2-D11mRNA sequence and a 614 bp full length cDNA was amplyfied by RT-PCR, which contained a 591 bp full ORF encoding 196 aa with a molecular weight of 20.69 kDa and an isoelectric point of 7.61. Bioinformatics analysis indicated that TmTPx had a typical 2-Cys Prx conserved domain and the phylogenetic analysis of the known Prx of Taenia species revealed that T. multiceps had the closest relationship to T. asiatica, followed by T. solium and T. crassiceps, E. granulosus and E. multilocularis. These data will lay the foundation for the further study of the function of TmTPx.