Vol. 9, Issue 10, Part F (2025)

In small ruminants, Haemonchus contortus is a major blood-feeding parasite, resulting in symptoms from minor anemia to mortality. It has caused substantial economic losses to the global sheep industry, estimated between US $42 million and US $222 million annually. Current control methods rely on anthelmintic treatment resulting in growing anthelminthic resistance. The development of immunological and molecular strategies for controlling these parasites shows promise for future research, and in this area, it has already been observed that there is ongoing research focused on developing recombinant protein-based vaccines against H. contortus. This study aimed to clone, express, and characterize arginine kinase (AK), an excretory/secretory (ES) protein of H. contortus. Hc-AK, a member of the phosphagen kinase family, plays essential roles in cellular energy metabolism in both vertebrates and invertebrates and is recognized as an immune modulator influencing pro-inflammatory cytokine responses. The complete Hc-AK gene 1036 base pairs in length was amplified from H. contortus cDNA and inserted into the pET-32a(+) expression vector using Escherichia coli TOP10 cells. For protein production, the recombinant construct was introduced into E. coli BL21(DE3)pLysS cells and expression was induced with 1 mM IPTG at 37 °C with shaking at 180 rpm for 2, 4, 6, and 8 hours. SDS-PAGE followed by Coomassie Brilliant Blue staining displayed a prominent band of approximately 57.4 kDa, consistent with the predicted size of Hc-AK together with an 18 kDa fusion tag. Western blot analysis employing Ni-HRP conjugate and sera from H. contortus-infected sheep demonstrated specific recognition of the recombinant Hc-AK (rHc-AK), whereas no reactivity was observed with sera from uninfected controls. These findings suggest that the recombinant Hc-AK protein may serve as a promising vaccine candidate against H. contortus infection.