Characterisation of potential vaccine targets in Haemonchus contortus

In a recent study, bioinformatics and RNA interference (RNAi) were used to look for the genes responsible for the acute lethal or developmental arrest in Haemonchus contortus . Out of the fifty-six H. contortus genes selected as potential targets, 41 provided strong to moderate phenotype. Seven genes were then selected for expression, purification and characterisation in the present study. A 981 bp full-length cDNA encoding H. contortus fatty acid synthase thioesterase domain ( Hc FAS-TE) was cloned, expressed in Escherichia coli, and the recombinant protein purified and biochemically characterised. The protein sequence of Hc FAS-TE showed the greatest similarity with FAS-TE of Teladorsagia circumcincta and Ancylostoma caninum . The optimum substrate concentration for recombinant Hc FAS-TE was 125 µM, whereas the addition of 150 µM Orlistat reduced the Hc FAS-TE activity by 85%. A 1563 bp full-length aspartyl tRNA synthetase ( Hc DRS-1), a 3048 bp full-length glutamyl tRNA synthetase ( Hc ERS-2), a 1482 bp vacuolar ATPase subunit A ( Hc VHA-12), a 1362 bp transcriptional co-activator histone acetyltransferase domain ( Hc CBP-1) and a 1362 bp full-length serum glucocorticoid-inducible kinase ( Hc SGK-1) and a 1152 bp full-length NADH-dependent oxidoreductase ( Hc F36A2.3) were also cloned, expressed in E. coli and purified. Despite varying assay conditions, there were no detectable activities for Hc DRS-1, Hc ERS-2 or Hc F36A2.3. Due to the assay complexity, no attempts were made to assay Hc SGK-1 or Hc VHA-12. Antibodies in serum and saliva from field-immune, but not parasite-naïve, animals recognised recombinant Hc FAS-TE, Hc VHA-12, Hc CBP-1, Hc F36A2.3, Hc DRS-1, and Hc ERS-2, but not Hc SGK-1, showing that an immune host has developed a response against the proteins. • Full-length cDNAs of 8 selected targets were cloned, expressed in E. coli , and purified. • Recombinant HcFAS-TE was biochemically characterised, its enzymatic activity was significantly inhibited by Orlistat • Seven out of eight recombinant proteins were recognised by antibodies in serum and saliva from field-immune sheep, confirming their immunogenicity.