Abstract

Ranavirus has caused disease epidemics and mass mortality events globally in free-ranging amphibian, turtle, and tortoise populations. Viral isolation and conventional PCR are the most common methods for diagnosis. In this study, quantitative real-time PCR (qPCR) assays were developed using a TaqMan probe-based and two distinct SYBR Green primers from a highly conserved region of the major capsid protein of frog virus 3 (family Iridoviridae, genera Ranavirus). Standard curves were generated using each primer set, as well as the conventional PCR primer, from a viral DNA segment cloned within a plasmid. TaqMan qPCR primer sets detected virus at a level ten times lower than SYBR Green primers and 1000 times lower than conventional PCR. Thirty-one clinical samples (whole blood and oral swabs) from box turtles were tested using these assays and prevalences compared. The advancement of qPCR allows rapid, sensitive, and quantitative for nucleic acid detection and is advantageous for early detection and disease monitoring.