Molecular Characterization and Expression Analysis of the Gene Encoding 3-Hydroxyacyl-CoA Dehydrogenase (EGR-03347) from Echinococcus Granulosus and the Evaluation of the Immune Protection of the Definitive Hosts (dogs)

Abstract

Background: Cystic echinococcosis, a serious parasitic zoonosis, is caused by tapeworm (Echinococcus granulosus) larvae. The development of an effective vaccine is a promising strategy to control echinococcosis. E. granulosus has a complete tricarboxylic acid cycle pathway, in which 3-hydroxyacyl-CoA dehydrogenase (EGR-03347) is a key enzyme.Methods: In the present study, the cDNA encoding EGR-03347 in Echinococcus granulosus (rEGR-03347) was successfully cloned and the molecular and biochemical characterizations carried out. The immunoreactivity of recombinant EGR-03347 (rEGR-03347) was investigated using western blotting. The immunolocalization of EGR-03347 in different life stages of E. granulosus was determined using specific polyclonal antibody, quantitative real-time reverse transcription polymerase chain reaction was used to analyze their transcript levels in PSCs and 28-day strobilated worms stages. In addition, recombinant protein rEGR-03347 was mixed with the adjuvant Quil A for vaccinating dogs, after three vaccine injections, all the dogs were orally challenge-infected with 100000 protoscoleces of E. granulosus. After 28 days of infection, all the dogs were euthanized and necropsied for collecting and counting E. granulosus worms, post-infection the antibody and cytokine were measured for the immunogenicity analysis of this protein.Results: rEGR-03347 is a highly conserved protein, consisting of 308 amino acids. Recombinant EGR-03347 could be identifed in the sera of patients with CE and in mouse anti-rEGR-03347 sera. Immunofluorescence analysis showed that EGR-03347 mainly localized in the tegument of protoscoleces and adults, and their transcript levels were high in the 28-day strobilated worms. Furthermore, enzyme-linked immunosorbent assays post‑infection showed that IgG gradually increased after the first immunization with rEGR-03347 compared with the control group, rEGR-03347 changed the interferon gamma and interleukin 4 levels. We observed an 87.2 % reduction in E. granulosus numbers and 66.7 % inhibition of the segmental development of E. granulosus in the rEGR‑03347‑vaccinated dogs compared with the nonvaccinated controls.Conclusions: This is the first report characterizing a 3-hydroxyacyl-CoA dehydrogenase from the tapeworm E. granulosus. We have characterized the sequence, structure and location of EGR‑03347 and investigated the immunoreactivity, immunogenicity and serodiagnostic potential of rEGR‑03347 . The results demonstrated rEGR-03347 as a potential vaccine against E. granulosus infection in dogs.