Adhesion-Regulating Molecule from Haemonchus contortus: Potential Antigen for Diagnosis of Early Infection in Goats.

Kalibixiati Aimulajiang,Xiaowei Tian,Muhammad Ali-Ul-Husnain Naqvi,Xiangrui Li,Muhammad Ali Memon,Mingmin Lu,Ruofeng Yan,Wen Chu,Lixin Xu,Xiaokai Song,Yongqian Bu

doi:10.3390/pathogens9010034

Abstract

Haemonchus contortus, a blood-sucking nematode of ruminants, causes large economic losses worldwide. Diagnosis of infection mainly depends on the evaluation of clinical signs and fecal examination. However, this has limitations for the diagnosis of early or light infections, where serological diagnosis seems to be more accurate and reliable. In this study, the recombinant H. contortus adhesion-regulating molecule protein (rHCADRM) was expressed and purified, and its diagnostic potential was evaluated. Serum samples from goats experimentally infected with H. contortus (n = 5) were collected at 0 (before infection, negative control), 7, 14, 21, 35, 49, 63, 85, and 103 days post-infection (DPI). The reactions between rHcADRM and goat serum were tested using Western blot (WB) analysis. The results show that rHcADRM can be recognized in the serum as early as 14 DPI, and the antibody against rHcADRM in infected goat could be maintained for over 89 days. No reaction was found between rHcADRM and antibodies against Trichinella spiralis, Fasciola hepatica, or Toxoplasma gondii. An indirect enzyme-linked immune sorbent assay (ELISA) was developed based on rHcADRM. The optimal coating antigen (279 ng of rHcADRM/well) and serum dilutions (1:50) were determined by checkerboard titration. A total of 64 serum samples, including 32 from H. contortus infection goats and 32 from helminth-free goats, were used to determine the positive (0.362) and negative (0.306) cut-off values for the ELISA. The results show this serological diagnosis method is highly sensitive (90.6%) and specific (93.75%). The coefficient of variation within run and between runs was less than 11%. To apply this indirect ELISA during field examination, 51 serum samples were randomly collected from goat farms and tested using this method. The result showed that 19.6% (10/51) of goats were infected with H. contortus, which was 100% consistent with the necropsy result, higher than that of fecal examination (15.7%, 8/51). These results indicate that rHcADRM could be a potential antigen for diagnosis of H. contortus infection in goats.

Highlights

Haemonchus contortus is one of the most important gastrointestinal nematodes of small ruminants.This parasitic nematode feeds on the blood from capillaries in the abomasum of sheep, goat, cattle, and other ruminants [1,2]
We developed an indirect enzyme-linked immune sorbent assay (ELISA) based on rHcADRM to evaluate its diagnostic potential and to complement the results obtained through Western blotting for further supporting the diagnosis of H. contortus infection
These results proved that indirect ELISA has better ability to detect H. contortus infection compared to McMaster

Summary

Introduction

Haemonchus contortus is one of the most important gastrointestinal nematodes of small ruminants. This parasitic nematode feeds on the blood from capillaries in the abomasum of sheep, goat, cattle, and other ruminants [1,2]. Infection with this parasite often results in anemia, diarrhea, weight loss, Pathogens 2020, 9, 34; doi:10.3390/pathogens9010034 www.mdpi.com/journal/pathogens. Ruminants are infected by ingestion of the third-stage larva (L3). The worms develop into adults at about 21 days post-infection (DPI) and begin to shed eggs through feces [6]

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