Mapping Full-Length Porcine Endogenous Retroviruses in a Large White Pig

Abstract

Xenotransplantation may bridge the widening gap between the shortage of donor organs and the increasing number of patients waiting for transplantation. However, a major safety issue is the potential cross-species transmission of porcine endogenous retroviruses (PERV). This problem could be resolved if it is possible to produce pigs that do not contain replication-competent copies of this virus. In order to determine the feasibility of this, we have determined the number of potentially replication-competent full-length PERV proviruses and obtained data on their integration sites within the porcine genome. We have screened genomic DNA libraries from a Large White pig for potentially intact proviruses. We identified six unique PERV B proviruses that were apparently intact in all three genes, while the majority of isolated proviruses were defective in one or more genes. No intact PERV A proviruses were found in this pig, despite the identification of multiple defective A proviruses. Genotyping of 30 unrelated pigs for these unique proviruses showed a heterogeneous distribution. Two proviruses were uncommon, present in 7 of 30 and 3 of 30 pigs, while three were each present in 24 of 30 pigs, and one was present in 30 of 30 animals examined. Our data indicate that few PERV proviruses in Large White pigs are capable of productive infection and suggest that many could be removed by selective breeding. Further studies are required to determine if all potentially functional proviruses could be removed by breeding or whether gene knockout techniques will be required to remove the residuum.

Characterization of Porcine Endogenous Retrovirus γ pro-pol Nucleotide Sequences

“…Differences in the proviral load have been observed in the genome of different animals, and the correlation of an increased proviral load in highly inbred pig breeds has been discussed previously (1,5,13,(15)(16)(17)23). Although most of the copies of PERV A and B are defective, several full-length functional PERV A and B from genomic pig loci have been sequenced (11,19). Recently, additional PERV ␥ clones, including defective sequences as well as mutant full-length copies (PERV E), have been described (17,22).…”

Developing Xenostandards for Microbiological Safety: New Zealand Experience

“…This is because the majority of PERV sequences appear to be either defective or deleted, i.e., they contain only one or two intact retro-viral open reading frames of gag, pol and env (Bosch, Arnauld, & Jestin, 2000;Czauderna, Fischer, Boller, Kurth, & Tonjes, 2000;Herring et al, 2001;Niebert, RogelGaillard, Chardon, & Tonjes, 2002). Consequently PERV proviral copy number is another important characteristic that should be investigated when assessing a donor herd for infectious risk.…”

Genotyping of Porcine Endogenous Retroviruses from a Family of Miniature Swine

“…Genomic mapping studies have shown that there are between approximately 10 and 100 proviral PERV loci in the genome of various pigs (1,3,7,9,10,20; Linda Scobie, unpublished data). However, most of these loci possess gross deletions or frameshift or point mutations rendering them replication defective.…”

“…However, most of these loci possess gross deletions or frameshift or point mutations rendering them replication defective. In our studies on miniature swine a number of full-length proviruses were found which upon transfection into human (293) and porcine (ST-IOWA) cells were unable to replicate (9). Furthermore, reporter gene analysis of the long terminal repeat (LTR) sequences from these proviruses revealed that their transcriptional activity was very low compared with tissue-culture adapted LTR sequences (24; Linda Scobie, personal communication).…”

“…Using a gene expression profiling strategy to analyze PERV, we identified transmission-specific fingerprints using restriction fragment length polymorphism (RFLP) analysis of single nucleotide polymorphisms (SNPs) in the gag region. This strategy was based on the PERV gag gene because (i) the number of gag-positive proviral loci present in Large White (9) and miniature swine (Linda Scobie, personal communication) genomes is less than that of env-positive loci; (ii) the gag sequence is highly conserved in PERV; and (iii) gag expression is essential for virus particle production from a cell.…”