Severe Perkinsea infection (SPI) in frog tadpoles of the USA

Since 1999 severe systemic infections of tadpoles with a previously unreported protist have been described causing mass mortalities in the United States (US). Based on the phylogenetic analysis of the 18S ribosomal DNA, the... [ view full abstract ]

Since 1999 severe systemic infections of tadpoles with a previously unreported protist have been described causing mass mortalities in the United States (US). Based on the phylogenetic analysis of the 18S ribosomal DNA, the causal agent was classified within the phylum Perkinsea clustering with other non-pathogenic Perkinsea organisms in a monophyletic group (NAG01). In this study we describe severe Perkinsea infection (SPI) mortality events investigated by the USGS–National Wildlife Health Center (NWHC) from an epidemiological and pathological perspective and characterize genetically the implicated organism. We have reviewed wild anuran mortality events and prospective studies investigated by USGS-NWHC since 1999. Diagnosis of SPI was based on the presence of characteristic gross and microscopic lesions and intralesional presence of the organism. Liver from 19 SPI frogs and 79 grossly and microscopically normal immature frogs were processed for standard PCR targeting NAG01. The products were sequenced and phylogenetically analyzed. Severe Perkinsea infection was diagnosed in 225 frogs (196 larvae and 29 metamorphs) from 20 mortality events and eight prospective studies. Twelve species were affected over a broad geographic area from Alaska to Florida. Severe Perkinsea infection was pathologically characterized by greatly enlarged and pale-discolored organs with massive numbers of protozoa replacing preexistent tissues but with minimal inflammation. NAG01 sequences were amplified from all tested SPI frogs (19/19, 100%) and from 2 of 79 (2.5%) apparently normal immature frogs (subclinical infections). All the sequences from PCR-positive frogs (19 SPI and 2 subclinically infected) shared 99.5% identity with previously published SPI-associated Perkinsea sequences clustering in a distinct clade within NAG01. These findings suggest that: 1) SPI is the second most important infectious disease of tadpoles in the US after ranavirus infections; 2) SPI mortalities are broadly distributed with numerous host species; and 3) SPI is caused by a genetically distinct Perkinsea lineage within NAG01.