Comparison of mitochondrial genome and development of specific PCR primers for identifying two scuticociliates, Pseudocohnilembus persalinus and Uronema marinum.

Background: Pseudocohnilembus persalinus and Uronema marinum (Ciliophora, Scuticociliatia), as parasitic scuticociliatid ciliates, were isolated from Scophthalmus maximus and Takifugu rubripes, respectively, in our previous studies. These ciliates are morphologically very similar; hence, it is difficult to identify specific scuticociliate species using traditional classification methods for performing taxonomic research and disease control studies.

Methods: We annotated the mitochondrial genomes of these two scuticociliates on the basis of previous sequencing, including analyses of nucleotide composition, codon usage, Ka/Ks, and p-distance. We also compared the nucleotide and amino acid similarity of the mitochondrial genomes of P. persalinus, U. marinum, and other 12 related ciliates, and a phylogenetic tree was constructed using 16 common genes. We chose the nad4 and nad7 genes to design specific PCR primers for identification.

Results: P. persalinus and U. marinum were found to have a close evolutionary relationship. Although codon preferences were similar, differences were observed in the usage of codons such as CGA, CGC, and GTC. Both Ka/Ks and p-distance were less than 1. Except for yejR, ymf57, ymf67, and ymf75, the amino acid sequence similarity between P. persalinus and U. marinum was greater than 50%.

Conclusions: The mitochondrial genomes of P. persalinus and U. marinum were thoroughly compared to provide a reference for disease prevention and control. The specific PCR primers enabled us to identify P. persalinus and U. marinum rapidly and accurately at the molecular level, thus providing a basis for classification and identification.