Phylogenetic analysis of African horse sickness virus segment 10: sequence variation, virulence characteristics and cell exit.

African horse sickness virus (AHSV) genome segment 10 encodes the non-structural proteins NS3/NS3a, which is involved in release of virus from cells. Full length segment 10 cDNAs were amplified by reverse transcription-polymerase chain reaction, from isolates of AHSV serotypes 2, 3, 4, 5, 7, 8 and 9. These cDNAs were cloned, sequenced and their phylogenetic relationships analysed. High levels of sequence homology were detected in segment 10 from some isolates of different serotypes, confirming that they could be grouped on this basis (serotypes 4, 5, 6 and 9 (group alpha); serotypes 3 and 7 (group beta); serotypes 1, 2, and 8 (group gamma). However, data from bluetongue virus (the prototype orbivirus) indicate that the AHSV serotype is determined exclusively by the structural outer coat proteins VP2 and VP5, encoded by genome segments 2 and 5 respectively. Therefore, as a direct consequence of genome segment reassortment between AHSV strains from different serotypes, the differences observed in segment 10 do not give a reliable indication of virus serotype. Segment 10 of AHSV 3 (virulent) and AHSV 3att (attenuated) were also analysed. These strains, together with AHSV 8, have been used to study of the genetic basis of virulence using reassortment (O'Hara et al., this publication). Virus release studies, using Culicoides cell cultures, indicate that differences in segment 10 of AHSV 3att and 8 can influence the timing of virus release from the infected cell.