DOUBLE-NESTED POLYMERASE CHAIN-REACTION FOR DETECTION OF CAPRINE ARTHRITIS-ENCEPHALITIS VIRUS PROVIRAL DNA IN BLOOD, MILK, AND TISSUES OF INFECTED GOATS

A nested polymerase chain reaction (PCR) for detecting proviral DNA of caprine arthritis-encephalitis virus (CAEV) in biological samples was developed. Primers for both gag and pol sequences of the CAEV genome were included in a single tube for simultaneous amplification ('double ' PCR), and the resulting bands were resolved visually in ethidium bro mide-stained agarose gels. Internal gag and pol probes were used to ve rify the identity of the amplified products by non-radioactive Souther n hybridization. Final confirmation of the identity of representative PCR bands was provided by DNA sequence analysis. A comparison between the PCR and an antibody ELISA (with recombinant CAEV p28 as target) us ing 141 caprine blood samples indicated very strong agreement between the two assays (kappa = 0.912). Four of 7 goats with indeterminate ELI SA results were PCR-positive as were 5 of 40 (12.5%) seronegative goat s, most probably indicating delayed seroconversion. Eleven of 27 goats (41%) PCR-positive on blood had detectable CAEV proviral DNA in milk. Proviral DNA was also detected in lung, mesenteric lymph node, bone m arrow, synovial membrane, and mammary gland of a seropositive, clinica lly affected goat, but not in equivalent tissues of a healthy seronega tive goat.