Ultradeep pyrosequencing technologies permit an assessment of the genetic diversity and the presence and frequency of minority variants in a viral population.
Rilpivirine, a second-generation NNRTI, will be used in first-line regimen therapy, but the prevalence of minority variants that are resistant to rilpivirine is unknown.
The current widely applied standard method to screen for HIV-1 genotypic resistance is based on Sanger population sequencing (Sseq), which does not allow for the identification of minority variants (MVs) below the limit of detection for the Sseq-method in patients receiving integrase strand-transfer inhibitors (INSTI).
We believe that the observed mutant reads reflect minority variants undetectable by bulk sequencing methodology.
Varghese et al. also detected additional minority variants with major etravirine mutations only in patients failing an NNRTI-containing regimen [14].
The impact of context could potentially be mitigated by the inclusion of additional oligonucleotides that would specifically interrogate known minority variants.
These results highlight the complex nature of drug resistance and the ability of the MDAP platform to identify minority variants within a viral population.
This finding suggests that these V3 loops are not generated and lost with each replication cycle, but in fact may be maintained as minority variants during chronic infection.
For example, our platform was less sensitive for minority variants in which two mutations are separated by less than six bases, and we were unable to detect variants with neighboring mutations (Figure S9 and data not shown).
Our results show that T215A and T215I not only evolve following treatment interruption in patients with failing therapy, but also can exist as minority variants prior to any therapy.
This diversity is characteristic of RNA viruses, and the degree to which minority variants influence the adaptability and phenotype of the population has remained a much debated, but largely unanswered, question.
