Table 1.

Methods to detect minority variants of drug-resistant HIV-1

Assays	Attributes	Limitations	Methods	Max Sensitivity*	References
Point mutation assays (with or without additional ligation step and/or PCR)	High sensitivity/specificity Moderately labor intensive Easy interpretation Additional ligation step improves specificity Additional PCR-amplification improves sensitivity	False positive results at lower limits Polymorphisms can reduce sensitivity Single-allele analyses No genetic linkage	LiPA	2%–10%	[17, 18]
			AS-PCR	0.01%–<1%	[19–27]
			MultiCode-RTx		[30, 31]
			OLA	<1%–12%	[32–37]
			LigAmp		[38, 39]
			RCA		[40, 41]
Sequencing assays	Extended sequence analyzed Genetic linkage if clonal	Labor intensive Cost Recombination	SGS	<1%–10%	[42, 43]
			CSA		[12, 15, 44, 45]
			HTA		[47, 48]
			SSA		[49]
			UDPS	< 1%	[50–54]
			DNA microarray	0.01%–<1%	[13, 56]
			PASS	0.01%–0.1%	[57]
Phenotypic assays	High sensitivity for co-receptor tropism Additional phenotypic information Possibility of identify novel DRM Can discern reversal of resistance	Low sensitivity for drugs with viral targets Cost Turnaround time	ES trofile assay TyHRT	0.3%–<1%	[58, 61–64] [39, 60]

PCR: polymerase chain reaction, DRM: drug resistance mutation, LiPA: line probe assay, AS-PCR: allele-specific PCR, OLA: oligonucleotide ligation assay, RCA: rolling circle amplification, UDPS: ultra-deep pyrosequencing, SGS: single genome sequencing, CSA: clonal sequence analyses, HTA: heteroduplex tracking assay, SSA: successive specific amplification, PASS: parallel allele-specific sequencing, TyHRT: phenotypic assays based on Saccharomyces cerevisiae Ty1, ES: enhanced-sensitivity (trofile assay).

^*Sensitivity depends upon the assay as well as the number of copies being interrogated.