A genetic analysis of HIV-1 from Punjab, India reveals the presence of multiple variants

Abstract

Objective: To determine the extent of HIV-1 genetic variation in Indian patients.

Design: To avoid any bias in selecting viral variants, HIV-1 DNA was amplified directly from the peripheral blood mononuclear cells of patients and sequenced. Genetic similarity between Indian sequences and other geographic isolates was analysed by phylogenetic analysis algorithms.

Methods: A fragment encompassing the C2/V3-V5 regions of HIV-1 gp120 was amplified from the lymphocyte DNA of 12 Indian patients. Multiple clones from each patient were sequenced. Nucleotide sequences encompassing about 650 base pairs were aligned for the Indian and other geographically distinct isolates. Inter-isolate relationships were analysed by means of distance, parsimony and neighbour-joining algorithms.

Results: Nucleotide sequence comparisons showed low interpatient variation. Amino-acid comparisons revealed a high degree of homology between Indian sequences in this study and those studied earlier. On distance and parsimony trees, most of the Indian sequences clustered together as subtype C. However, sequences from three patients also showed significant homologies and phylogenetic clustering outside of subtype C.

Conclusions: The predominant strain of HIV-1 in India belongs to subtype C and little interpatient nucleotide sequence divergence in the majority of cases suggests recent spread of HIV-1 in this region. This study also presents the first evidence for non-C subtypes in the Indian population with two epidemiologically linked samples remaining unclassified for any existing env subtype. The presence of variant subtypes in Indian patients sheds light on the transmission routes of HIV-1 to India and emphasizes the need to include these sequences in vaccine development strategies.

PIP: Health workers collected blood samples from 12 persons infected with HIV living in the Punjab in India to obtain peripheral blood mononuclear cells so researchers could determine the extent of HIV-1 genetic variation. They prepared multiple clones of the C2/V3-V5 regions of HIV-1 gp120 from the lymphocyte DNA of each patient. They used distance, parsimony, and neighbor-joining algorithms to analyze the inter-isolate relationships. They aligned nucleotide sequences of about 650 base pairs for the Indian and other geographically distinct isolates. All but two cases were males. The two females acquired HIV from their husbands. Based on the nucleotide sequence comparisons, there was low interpatient variation. Amino acid comparisons found a high degree of homology between Indian sequences in this study and those studied previously. Most Indian sequences clustered together as subtype C on the distant and parsimony trees. Three patients had significant homologies and phylogenetic clustering outside of subtype C. One patient had env gene homology to subtype B sequences prevalent in Europe and the Americas. The two others had env gene sequences that clustered away from all presently known subtypes of HIV-1. These three cases were the first sequences divergent from subtype C in India. All of these patients and one that clustered marginally with subtype C had possible contacts outside India. Variant subtypes in Indian patients provide clues on the transmission routes of HIV-1 to India. They also underscore the need for researchers to include these sequences as they develop an HIV/AIDS vaccine. Since the leading HIV-1 strain in India conforms to subtype C and there was limited nucleotide sequence variation in most cases, these findings indicate recent spread of HIV-1 in the Punjab.