Diversity of Eimeria tenella apical membrane antigen-1 from chickens in Mpumalanga province and its in silico epitope prediction as a vaccine candidate.

Abstract

Coccidiosis has been a significant challenge in the poultry industry. There is a high request for the modification of a cost-effective immunizing agent to curtail this disease. Apical membrane antigen 1 (AMA1) has been reported as a protective antigen in sub-unit vaccine development against several apicomplexan parasites such as Plasmodium falciparum, Eimeria tenella and Eimeria maxima. However, knowledge of genetic diversity in this vaccine candidate is imperative. Also, to minimize the cost and time involved in producing a vaccine, computational vaccine design has received much attention through the immunoinformatics method. Therefore, screening for the potential vaccine epitopes in AMA1 that can induce cellular and humoral immune response through the immunoinformatics technique looks promising. This investigation aimed to detect the level of genetic diversity amid Eimeria tenella Apical Membrane Antigen 1(EtAMA1) in selected farms in Mpumalanga province and predict vaccine epitopes from this antigen. Four hundred fresh faecal samples were collected from 10 selected broiler chicken farms in Mpumalanga. The samples were screened for Eimeria oocyst using a compound microscope, and samples containing oocyst were further screened for E. tenella using molecular methods. AMA1 (n=103) was amplified from positive samples for E. tenella, and resulted amplicons were sent to Inqaba Biotec for sequencing and analyzed using MEGA6.06 and DnaSP programs. The results revealed low levels of genetic diversity among Mpumalanga EtAMA1 sequences which were measured by nucleotide diversity (0.0007) diversity, haplotype diversity (0.113) and haplotype number (3). Correspondingly, the haplotype network revealed 4 haplotypes, 3 of which consist of samples from Mpumalanga. Identification of immunogenic B- and T-cell epitopes from EtAMA1 sequences was further carried out and were used to construct a multiepitope vaccine (MEV) using immunoinformatics approaches. The constructed MEV is 311 amino acids long. It was constructed by linking 6 B-cell, 3 CD8+ epitopes and 6 CD4+ epitopes with appropriate adjuvant and linkers. Both adjuvant and linkers were used to increase the immunogenicity of the MEV. The designed MEV was highly antigenic and non-allergenic. The results showed a strong binding affinity of MEV with TLR4. These results suggest that the predicted vaccine could be a significant vaccine candidate against chicken coccidiosis through further experimental validation is still necessary.