Development of an indirect ELISA and an immunochromatographic test method for early detection of Haemonchus contortus infection in sheep.

Haemonchosis, caused by the parasitic nematode Haemonchus contortus, poses a significant global threat to small ruminant health and productivity. Current diagnostic methods, such as fecal egg flotation, cannot detect early-stage or latent infections, hindering timely intervention. Building on prior identification of the secreted BPTI/Kunitz inhibitor domain-containing protein SPI-I8 as a key H. contortus antigen released during initial infection, we developed two diagnostic methods using recombinant Hc-SPI-I8B (rHc-SPI-I8B). First, an indirect ELISA with rHc-SPI-I8B as the coating antigen demonstrated detection of H. contortus infection in sheep as early as 10 days post-infection (dpi), with a sensitivity of 1:200. Concurrently, an immunochromatographic test (ICT) was established by conjugating rHc-SPI-I8B with 20 nm gold particles, achieving a sensitivity of 1:40, detecting antibodies at 11 dpi, and remaining effective beyond 100 dpi. Both methods exhibited high specificity, showing no cross-reactivity with positive sera of Echinococcus granulosus, Fasciola hepatica, or Fasciola gigantica. Additionally, they showed high stability and reproducibility (inter-/intra-assay CV <10 % for ELISA). Both approaches provide efficient tools for early diagnosis of haemonchosis.