Resistance-associated substitutions (RASs) have been identified in vitro for all of the DAAs approved for clinical use. NS3 and NS5A RASs may arise spontaneously due to the error-prone HCV RNA polymerase and therefore are present before DAA therapy. NS3 and NS5A RASs are selected during DAA therapy and enriched in people in whom treatment fails with NS3 and NS5A inhibitor-containing regimens, respectively. NS5B RASs have been reported but are very rare. For most regimens currently listed on the PBS, there is no clinical role for baseline HCV resistance testing in treatment-naive people or prior non-responders to either pegIFN-based therapy or protease inhibitor-based triple therapy, because such high SVR rates are achieved.
There is a theoretical role for NS5A RAS testing in people with Gt 1a HCV for whom treatment with the combination of elbasvir and grazoprevir for 12 weeks is planned (see Elbasvir plus grazoprevir).  In these people, the presence of an RAS at amino acid position 28, 30, 31 or 93 in the NS5A protein is associated with lower rates of SVR. In this population, SVR can be increased by prolonging treatment duration to 16 weeks and combining treatment with ribavirin. However, the frequency of these RASs is very low in the Australian population (< 5%–10% using population sequencing),  meaning that the clinical yield from testing is low. Furthermore, RAS testing is not widely available, nor is it currently reimbursed by the government. Given the low frequency of relevant NS5A RASs in the Australian population, we do not recommend routine resistance testing before treatment with elbasvir and grazoprevir.
Where available, resistance testing for NS3, NS5B and NS5A RASs should be considered after failure of combination DAA treatment. Resistance testing involves direct sequencing of the HCV genome and is available through specialised laboratories. As noted, this testing is not currently reimbursed in Australia. HCV sequencing may also be used as a research tool to differentiate relapse from reinfection, and to document transmission. Patients in whom combination DAA therapy fails should be managed in specialist centres.