Promises and pitfalls of Illumina sequencing for HIV resistance genotyping

•The Illumina MiSeq currently has the most favorable parameters for HIV resistance genotype testing by next-generation sequencing.•The extensive genetic variability of HIV within hosts is a key challenge for next-generation sequence analysis.•The MiSeq system has exhibited both significant longitudinal (between run) and within-run cross-contamination issues.•The high throughput of the MiSeq provides an opportunity for cost-saving by processing hundreds of samples on the same run.

Genetic sequencing (“genotyping”) plays a critical role in the modern clinical management of HIV infection. This virus evolves rapidly within patients because of its error-prone reverse transcriptase and short generation time. Consequently, HIV variants with mutations that confer resistance to one or more antiretroviral drugs can emerge during sub-optimal treatment. There are now multiple HIV drug resistance interpretation algorithms that take the region of the HIV genome encoding the major drug targets as inputs; expert use of these algorithms can significantly improve to clinical outcomes in HIV treatment. Next-generation sequencing has the potential to revolutionize HIV resistance genotyping by lowering the threshold that rare but clinically significant HIV variants can be detected reproducibly, and by conferring improved cost-effectiveness in high-throughput scenarios. In this review, we discuss the relative merits and challenges of deploying the Illumina MiSeq instrument for clinical HIV genotyping.