Single Base Primer Extension Assay (SNaPshot) for Rapid Detection of Human Immunodeficiency Virus - 1 Drug Resistance Mutations

Abstract

The 3' primer extension assay can be employed to interrogate multiple known single nucleotide polymorphisms (SNPs) simultaneously using a multiplex approach. Over 200 Human Immunodeficiency Virus-1 (HIV-1) mutations are currently associated with drug resistance. Identifying these mutations assist in taking appropriate therapeutic decisions such that superior drug regimens can be designed for keeping the viral load below pathogenic level. Presently, reverse transcription of HIV-1 RNA followed by amplification of the pol gene and subsequent nucleotide sequencing is a popular method of HIV-1 drug resistance genotyping. However, it is still a time consuming and laborious protocol and require significant bioinformatics analysis in order to decipher the significance of mutations encountered during analysis. In this study we analyzed 75 HIV-1 positive clinical samples for 2 representative mutations, viz., M41L and M184V using a 3' primer extension protocol popularly known as SNaPshot assay. Twenty four and 36% of the patients were found to harbor the critical M41L and M184V mutations respectively while 20% were found to contain both the mutations together. The results were 100% concordant with DNA sequencing assay which was used as a gold standard. The results substantiate the cost, speed and economy-related advantages of this technology platform for interrogating known mutations of significance within the HIV-1 genome.