title: Development, validation and clinical evaluation of a low cost in-house HIV-1 drug resistance genotyping assay for Indian patients.
abstract: Human Immunodeficiency Virus-1 (HIV-1) drug resistance genotyping assay is a part of clinical management of HIV-1 positive individuals under treatment with highly active antiretroviral therapy (HAART). Routine monitoring of drug resistance mutations in resource limited settings like India is not possible due to high cost of commercial drug resistance assays. In this study we developed an in-house, cost effective HIV-1 drug resistance genotyping assay for Indian patients and validated it against the US-FDA-approved ViroSeq HIV-1 drug resistance testing system. A reference panel of 20 clinical samples was used to develop and validate the assay against ViroSeq HIV-1 drug resistance testing system which was subsequently used to genotype a clinical panel of 225 samples. The Stanford HIV database was used to identify drug resistant mutations. The analytical sensitivity of the assay was 1000 HIV-1 RNA copies/ml of plasma sample while precision and reproducibility was 99.68 ± 0.16% and 99.76 ± 0.18% respectively. One hundred and one drug resistant mutations were detected by the in-house assay compared to 104 by ViroSeq system in the reference panel. The assay had 91.55% success rate in genotyping the clinical panel samples and was able to detect drug resistant mutations related to nucleoside reverse transcriptase inhibitor (NRTI), non-nucleoside reverse-transcriptase inhibitor (NNRTI) as well as protease inhibitor (PI) classes of antiretroviral drugs. It was found to be around 71.9% more cost effective compared to ViroSeq genotyping system. This evaluation of the assay on the clinical panel demonstrates its potential for monitoring clinical HIV-1 drug resistance mutations and population-based surveillance in resource limited settings like India.
authors.affiliation: Department of Molecular Biology, Dr. D. Y. Patil Biotechnology & Bioinformatics Institute, Dr. D. Y. Patil Vidyapeeth, Pune, Maharashtra, India.
authors.forename: Arpan
authors.initials: A
authors.lastname: Acharya
authors.affiliation: Department of Molecular Diagnostics, SN Genelab, Surat, Gujarat, India.
authors.forename: Salil
authors.initials: S
authors.lastname: Vaniawala
authors.affiliation: Department of Molecular Diagnostics, Supratech Micropath Laboratory & Research Institute, Ahmedabad, Gujarat, India.
authors.forename: Parth
authors.initials: P
authors.lastname: Shah
authors.affiliation: Department of Microbiology, Padmashree Dr. D. Y. Patil Medical College & Research Centre, Dr. D. Y. Patil Vidyapeeth, Pune, Maharashtra, India.
authors.forename: Rabindra Nath
authors.initials: RN
authors.lastname: Misra
authors.affiliation: Department of Molecular Biology, Dr. D. Y. Patil Biotechnology & Bioinformatics Institute, Dr. D. Y. Patil Vidyapeeth, Pune, Maharashtra, India.
authors.forename: Minal
authors.initials: M
authors.lastname: Wani
authors.affiliation: Department of Microbiology, Interdisciplinary Science, Technology and Research Academy, AISC, Pune, Maharashtra, India.
authors.forename: Pratap N
authors.initials: PN
authors.lastname: Mukhopadhyaya
mesh_terms: D000328:Adult; D019380:Anti-HIV Agents / Q000494:pharmacology; D003362:Cost-Benefit Analysis; D004252:DNA Mutational Analysis; D024882:Drug Resistance, Viral / Q000235:genetics*; D005260:Female; D060005:Genotyping Techniques / Q000191:economics*; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology*; D015497:HIV-1 / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D007194:India; D008297:Male; D010802:Phylogeny; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D019380:Anti-HIV Agents; D012367:RNA, Viral