Analytical and clinical performance of a Chikungunya qRT-PCR for Central and South America()()

Chikungunya was introduced into the Americas in 2015 causing a pandemic across the continent. Testing during the acute phase of infection relies on qRT-PCR, but available assays have a number of limitations. A qRT-PCR assay specific to the chikungunya E1 gene was designed using sequence data from contemporary strains. A probit analysis established the 95% limit of detection as 19.6 copies per reaction. We compared the assay with a US Centers for Disease Control (CDC) chikungunya qRT-PCR as the reference standard. The assay had a sensitivity and specificity of 98.4% and 100% in 90 samples retrospectively collected in Guatemala. In a further 74 febrile samples prospectively collected in Ecuador and Guatemala the test had a sensitivity and specificity of 100% and 98.4%, respectively. Sequencing the nsp4 gene of the discordant positive sample indicated the presence of chikungunya RNA, and mismatches to the primer binding sites of the CDC assay.