THE DEVELOPMENT OF PCR BASED ASSAYS FOR THE DETECTION AND DIFFERENTIATION OF SIMIAN IMMUNODEFICIENCY VIRUS IN-VIVO

Polymerase chain reaction based assays, which amplify a region of the gag gene, have been developed for the direct detection of simian immun odeficiency virus (SIV) DNA sequences in the blood of experimentally i nfected cynomolgus macaques. In macaques infected with a characterised virus pool (11/88 pool SIVmac 32H), an assay employing a single round of amplification was found to be highly sensitive and specific. Howev er, in animals infected with the SIV molecular clones J5 and C8 (Rud e t al., J. Gen. Virol. 75, 529-543), it was necessary to use two rounds of amplification and nested primer pairs in order to achieve sensitiv ity > 90%. In order to differentiate macaques infected with either of the two genetically distinct SN clones, J5 or Cs, a third PCR based as say has been developed, which amplifies a 492 bp region of the nef gen e. Sequence differences between the nef genes of the two molecular clo nes enabled the PCR product amplified from each virus to be distinguis hed by restriction analysis. These sensitive and specific assays compl ement virological detection of SIV and enable superinfection studies t o be evaluated; a prerequisite for the testing of live attenuated immu nodeficiency virus vaccines.