An HIV-1 integrase genotype assay for the detection of drug resistance mutations.
BACKGROUND
The integrase inhibitors (e.g. Raltegravir) are a new class of antiretroviral drugs that have recently become available for the treatment of patients with multi-drug resistant HIV-1 within Australia. The emergence of mutations that confer resistance to the integrase inhibitors has been observed in vivo; however, no commercial genotyping assay is currently available to screen for resistance to these drugs.


METHODS
The HIV-1 integrase gene was amplified from plasma-derived HIV-1 viral RNA via reverse transcription-polymerase chain reaction and genotype determined via population DNA sequencing. Drug resistance mutations and polymorphisms were detected using the Stanford University online HIV database. Assay sensitivity and reproducibility were determined using clinical and laboratory-derived samples.


RESULTS
Our in-house assay was capable of genotyping the integrase gene from all samples tested (n = 30) of HIV-1 subtypes B, C, D, F, CFR01_AE and CRF02_AG and can amplify the integrase region from plasma samples containing as few as 50 HIV RNA copies/mL. The assay is highly reproducible (average nucleotide concordance = 99.6%, n = 4) and is capable of detecting resistance-associated mutations.


CONCLUSIONS
This assay is suitable for routine drug resistance screening of plasma samples from HIV-infected patients receiving integrase inhibitor antiretroviral drugs and also serves as a useful research tool.
Hearps
Anna C
AC
Clinical Research Laboratory, Centre for Virology, Burnet Institute for Medical Research and Public Health, Melbourne, Vic. 3004, Australia. annah@burnet.edu.au
Greengrass
Vicki
V
Hoy
Jennifer
J
Crowe
Suzanne M
SM
D001315:Australia; D024921:Drug Resistance, Multiple, Viral / Q000235:genetics*; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D019427:HIV Integrase / Q000235:genetics*; D019428:HIV Integrase Inhibitors / Q000494:pharmacology; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D012367:RNA, Viral / Q000032:analysis* / Q000302:isolation & purification
D019428:HIV Integrase Inhibitors; D012367:RNA, Viral; D019427:HIV Integrase