Introduction: Yucatan SLAD/D (SLA, swine leukocyte antigen) haplotype inbred miniature swine are candidates as organ donors for xenotransplantation. Porcine endogenous retroviruses (PERV) inherit infectious potential if pig cells, tissues, or organs were transplanted to immunosuppressed human recipients. Particularly, ecotropic PERV-C that could recombine with PERV-A to highly replication-competent human-tropic PERV-A/C should be excluded from pig breeds designed for xenotransplantation. SLAD/D pigs do not bear replication-competent PERV-A and –B, even if they carry PERV-C. Therefore, when using SLAD/D pigs, absence and/or elimination of genomic PERV-C is required to fully guarantee patient infectious safety in pig-to-human xenotransplantation.

Methods: Based upon the hypothesis that SLAD/D pigs inherit a proviral PERV-C, a bacteriophage lambda library of a female pig was generated and screened. PERV-C clone number 561 was isolated. The provirus, truncated in the envelope (env) and missing the 3’-LTR due to cloning in lambda, was complemented by PCR, and the recombinants were functionally characterized. Recombinant clone PERV-C(561) was chromosomally mapped by its 5’-proviral flanking sequences.

Results: Transfection of recombinant clone PERV-C(561) promoted retroviral expression including reverse transcriptase (RT) in ST-IOWA cells. Over time, stable transfectants of recombinant PERV-C(561) confirmed an increased infectivity in vitro compared to other PERV-C. Full-length PCR using 5’-and 3’-flanking primers specific to the genomic PERV-C(561) locus verified that this specific SLAD/D haplotype pig harbors at least one full-length PERV-C provirus. The chromosomal location is different from that of the previously described PERV-C(1312) provirus, which was derived from the porcine cell-line MAX-T.

Conclusion: The sequence data presented here provide further knowledge about PERV-C infectivity and contribute to targeted knockout in order to generate PERV-C-free founder animals.