Application of the ViroKey? SQ FLEX assay for detection of cytomegalovirus antiviral resistance

Background: Cytomegalovirus (CMV) is a viral infection which establishes lifelong latency, often reactivating and causing disease in immunosuppressed individuals, including haematopoietic stem cell transplant (HSCT) re-cipients. Treatment can be problematic due to antiviral resistance which substantially increases the risk of pa-tient mortality. Diagnostic testing capabilities for CMV antiviral resistance in Australia and elsewhere have traditionally relied on gene-specific Sanger sequencing approaches, however, are now being superseded by next generation sequencing protocols.Objective: Provide a snapshot of local mutations and explore the feasibility of the ViroKey?(R) SQ FLEX Genotyping Assay (Vela Diagnostics Pty Ltd) by examining sequencing success.Method: Performed sequencing on adult (n = 38) and paediatric (n = 81) plasma samples, over a large range of viral loads (above and below the assay recommended threshold of >1,000 International Units (IU)/mL; noting most of our paediatric samples have loads <1,000 IU/mL).Results: Eleven test runs (including three repeat runs; 14 to 15 samples per run) were conducted, and four runs were deemed valid. The overall individual sample success rate for the four evaluable test runs was 71.2% (42/59 samples); 80.4% (37/46) samples >1,000 IU/mL were valid. Ten clinically important antiviral resistance mu-tations were detected, the most common being A594V in the UL97 gene, found in 6 (5%) samples.Conclusions: A range of technical issues were experienced, however with improvement this platform could be a useful addition to routine pathology workflows, providing timely antiviral resistance results for patients un-dergoing HSCT.