Abstract
Malacosporeans are a group of endoparasitic cnidarians (Myxozoa) that use freshwater bryozoans and fish as final and intermediate hosts, respectively. The malacosporean Tetracapsuloides bryosalmonae causes proliferative kidney disease (PKD), an emerging disease in aquaculture and wild fish populations, including threatened salmonids in Europe and the USA. Mixed infections of malacosporeans are often encountered, and a monitoring tool for screening of multiple malacosporean species in either their fish or bryozoan hosts is therefore desirable. We describe an inexpensive method that combines PCR amplification of the partial 18S rRNA gene (~260 bp) and a single-step restriction fragment length polymorphism (RFLP) method for identification of 10 malacosporean lineages and species. We demonstrate and test this methodology on a set of DNA extracted from malacosporeans infecting fish kidney and tissues sampled from bryozoan colonies and compare the results with Sanger sequencing of the same parasite DNA isolates. The PCR-RFLP and Sanger sequencing methods agreed in 100% of cases. The PCR-RFLP method offers a number of opportunities, including screening large panels of host tissue samples to gain insights into infection patterns, characterizing mixed infections, and confirming highly pathogenic T. bryosalmonae infections. The method can also be further refined as new sequence data become available for malacosporeans.
Highlights
Myxozoans comprise a diverse but poorly sampled clade of endoparasitic cnidarians
The PCR-restriction fragment length polymorphism (RFLP) method offers a number of opportunities, including screening large panels of host tissue samples to gain insights into infection patterns, characterizing mixed infections, and confirming highly pathogenic T. bryosalmonae infections
All evidence suggests that freshwater bryozoans (Phylum Bryozoa; Class Phylactolaemata) are the only invertebrate hosts used by malacosporeans, with sporogonic stages developing in the body cavity (Canning et al 2000, Hartikainen et al 2014, Patra et al 2017)
Summary
Myxozoans comprise a diverse but poorly sampled clade of endoparasitic cnidarians. With some 2600 described species, they currently represent 19% of total cnidarian species diversity (Okamura et al 2018). All have complex life cycles, exploiting invertebrate and vertebrate hosts (Lom & Dyková 2006, Okamura et al 2015). Myxozoans are divided into 2 major groups, the highly derived and species-rich Myxosporea and the species-poor Malacosporea characterized by the retention of primitive features (Gruhl & Okamura 2015). Salmonids were the only fish hosts known to be exploited by malacosporeans, with the unplaced parasite (referred to as ‘PKX organism’) being detected in kidney of fish suffering from proliferative kidney disease (PKD) (Hedrick et al 1993).
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