Retrospective Multi-Locus Sequence Analysis of African Swine Fever Viruses by "PACT" Confirms Co-Circulation of Multiple Outbreak Strains in Uganda.

African swine fever (ASF) is a haemorrhagic fever of swine that severely constrains pig production, globally. In Uganda, at least 388 outbreaks of ASF were documented from 2001 to 2012. We undertook a retrospective serological and molecular survey of ASF virus (ASFV) using banked samples collected from seven districts (Pallisa, Lira, Abim, Nebbi, Kabarole, Kibaale, and Mukono) of Uganda. Six assays (ELISA for antibody detection, diagnostic gene PCR and genomic amplification, and sequencing of four gene regions ( [P], [A], CVR of the -ORF [C], and [T]), hereinafter referred to as P-A-C-T (PACT)) were evaluated. Antibodies to ASFV were detected in the Abim district (6/25; 24.0%), and the remainder of the serum samples were negative (187/193; 96.9%). For the tissue samples, ASFV detection by assay was 8.47% for P, 6.78% for A, 8.47% for C, and 16.95% for T. The diagnostic PCR ( gene) detected seven positive animals from four districts, whereas the assay detected ten positives from all seven districts. In addition to the superior detection capability of TK, two virus variants were discernible, whereas CVR recovered three variants, and and sequencing each identified a single variant belonging to genotype IX. Our results indicate that dependence on serology alone underestimates ASF positivity in any infected region, that multi-locus sequence analysis provides better estimates of outbreak strain diversity, and that the assay is superior to the WOAH-prescribed conventional diagnostic PCR, and warrants further investigation.