Sequencing and Phylogenetic Analysis

By the end of 2012, more than 6.1 million people were infected with HIV-1 in South Africa. Subtype C was responsible for the majority of these infections and more than 300 near full-length genomes (NFLGs) have been published. Currently very few non-subtype C isolates have been identified and characterized within the country, particularly full genome non-C isolates. Seven patients from the Tygerberg Virology (TV) cohort were previously identified as possible non-C subtypes and were selected for further analyses. RNA was isolated from five individuals (TV047, TV096, TV101, TV218, and TV546) and DNA from TV016 and TV1057. The NFLGs of these samples were amplified in overlapping fragments and sequenced. Online subtyping tools REGA version 3 and jpHMM were used to screen for subtypes and recombinants. Maximum likelihood (ML) phylogenetic analysis (phyML) was used to infer subtypes and SimPlot was used to confirm possible intersubtype recombinants. We identified three subtype B (TV016, TV047, and TV1057) isolates, one subtype A1 (TV096), one subtype G (TV546), one unique AD (TV101), and one unique AC (TV218) recombinant form. This is the first NFLG of subtype G that has been described in South Africa. The subtype B sequences described also increased the NFLG subtype B sequences in Africa from three to six. There is a need for more NFLG sequences, as partial HIV-1 sequences may underrepresent viral recombinant forms. It is also necessary to continue monitoring the evolution and spread of HIV-1 in South Africa, because understanding viral diversity may play an important role in HIV-1 prevention strategies