Reduced proviral loads at seroconversion in sheep infected with attenuated mutants. A. At seroconversion day, DNA was extracted from blood isolated from wild type or mutant-infected sheep, as indicated. Proviral sequences were amplified by 25 cycles of PCR using tax specific primers. The amplification products were resolved on a 1% agarose gel and analyzed by Southern blotting with a BLV tax probe. Blood samples from uninfected sheep n° 4533 and 4534 were used as controls for PCR contamination. A PCR amplification of the gapdh gene was used as an internal control for DNA integrity. B. Mean proviral loads at seroconversion. For each category of mutant-infected sheep, the mean values of the proviral loads (in arbitrary units ± standard deviations as determined after Phosphorimaging scanning) were statistically compared using the Student t test to the mean proviral load of the wild type group (WT). The data result from three independent experiments using DNAs extracted around the seroconversion period (NS: p > 0.05 non-significance; * 0.01 < p < 0.05; *** p < 0.001, Student t test). Using real-time PCR, the proviral loads were estimated to be 270 and 245 copies per 1,000 cells in sheep infected, respectively, with the wild type virus and the GP30 recombinant whereas the levels yielded by the other mutants were significantly reduced (27, 40 and 24 copies / 1,000 cells for IG4, CRX3 and CRE, respectively).