Recombinant HcGAPDH Protein on the Surface of the Probiotic Bacillus subtilis Spore Protects Sheep against Haemonchus contortus Infection

Haemonchosis caused by Haemonchus contortus (H. contortus) has become increasingly prevalent in recent years. Effective prevention strategies are urgently needed to control this infectious disease. Anthelmintic drugs, as the most effective means of controlling parasitic infections, the safety of drug residues is worth of concern. Nowadays, there is increasing recognition that probiotics effectively prevent the infection of pathogens, which could be a replacement of anthelmintic drugs. In this study, by 16S ribosomal RNA (16S rRNA) sequencing, we found that the Bacillales in the abomasal microbiota of sheep with H. contortus infection was depleted. According to previous studies, we generated a recombinant Bacillus subtilis (B. subtilis) spores expressing Glyceraldehyde-3-phosphate dehydrogenase from H. contortus (HcGAPDH) on the surface. The expression of HcGAPDH did not affect the production and structure of spores from B. subtilis. Compared to that of control (Ctrl), the CotB-HcGAPDH fusion protein expressing recombinant B. subtilis spores (RSCotB-HcG) showed a favorable immunogenicity and induced significant immune responses in mice. Interestingly, the oral gavage of RSCotB-HcG triggered Th1-dominated immune responses. Consistently, RSCotB-HcG promoted the proliferation of peripheral blood lymphocytes of sheep, and increased anti-HcGAPDH IgG levels in the serum and anti-HcGAPDH sIgA levels in intestinal mucus. Moreover, the RSCotB-HcG promoted the relative abundance of probiotics in the abomasal microbiota in sheep. We found the average weight gain of RSCotB-HcG–received sheep with H. contortus infection (H.c+RSCotB-HcG) significantly increased 44.5% compared to the H. contortus-infected sheep (H.c). The numbers of H. contortus eggs per gram of faeces and worm burdens in the H.c+RSCotB-HcG sheep were reduced by 84.13% and 71.50%, respectively, compared to that of H.c sheep. Meanwhile, the damage caused by parasite infection were alleviated. Together, all these data provide insights into how to best use the probiotic-based strategy as a replacement of anthelmintic drugs.