Generation of chimeric chickens resistant to infection by avian leukemia virus subgroups A, B, D, E, and K through primordial germ cells.

Avian leukosis virus (ALVs), an oncogenic retrovirus, induces immunosuppression and various tumors in chickens, posing a severe global threat to poultry health and causing substantial economic losses. Natural resistance alleles, notably tva and tvb, confer genetic resistance to specific ALV subgroups (ALV-A, -K, and ALV-B, -D, -E, respectively). This study aimed to exploit this resistance by creating resistant chicken lines via primordial germ cells (PGCs)-mediated chimera. We first genotyped the tva and tvb loci in Qingyuan Partridge Chickens (QYPCs) and Zhanjiang Yellow Chickens (ZJYCs). Through selective mating, we generated homozygous tva and tvb QYPC embryos as PGC donors, and homozygous susceptible tva and tvb ZJYC embryos as recipients. Donor PGCs harvested from QYPC embryos were successfully cultured in vitro, as confirmed by the expression of germ cell-specific markers (DDX4, c-Kit, PRDM14) and scanning electron microscopy revealing characteristic microvilli. These PGCs were microinjected into recipient embryos, producing viable chimeric chickens. Genotyping confirmed the successful integration of the resistant donor-derived PGCs. The germline transmission efficiency reached 35.7%. Our findings demonstrate that PGC-mediated germline transmission provides an efficient strategy for rapidly establishing ALV-resistant poultry flocks, offering a potential strategy for establishing ALV-resistant poultry flocks by introgressing resistance alleles.