Diagnostic polymerase chain reaction assay to detect Kudoa neurophila (Myxozoa: Multivalvulida) in a marine finfish hatchery

A single-round polymerase chain reaction (PCR) diagnostic assay was developed from a small subunit ribosomal DNA (SSU rDNA) gene sequence to detect the myxozoan parasite Kudoa neurophila, the causative agent of myxozoan disease in the hatchery reared marine finfish, striped trumpeter Latris lineata (Forster). The assay was developed for use as a disease control management tool in a hatchery system specifically designed to research and produce marine finfish such as striped trumpeter juveniles for aquaculture. The assay is sufficiently species specific and sensitive enough to detect a small fragment of the parasite's SSU rDNA. At the lower limits of detection, the test is con- sistently positive to an estimated 0.1 spore or 60 fg of parasite DNA per 25 µl PCR reaction in serial dilution and positive to an estimated 0.1 spore in 25 mg of infected fish CNS tissue (4 spores g -1 ). Specifically, the test is capable of detecting early stages of the life cycle within the fish host and con- sequently diagnosing an infection not normally detected using traditional histological techniques. The test is also effective for screening water supplies and prey species cultures throughout the hatch- ery system to determine bio-security efficacy, to assist in identification of an alternate or other primary fish host, to indicate the location of potential disease reservoirs, and to enable a targeted approach to disease prevention.