Molecular genetic characterization using the ribosomal RNA (rDNA) gene accrues a wealth of knowledge regarding the true nature of species diversity of Kudoa Meglitsch, 1947 (Myxozoa: Myxosporea: Multivalvulida) and the biogeographical relationships of isolates from different host fish and sea areas. In the present study, we characterized morphologically and genetically three Kudoa spp. with four shell valves and polar capsules (SV/PC), forming pseudocysts in the myofiber of trunk muscles of Cheilodactylus zonatus or Acanthogobius hasta in the natural seawater around Japan. Myxospores from C. zonatus fished in the western Pacific Ocean off Kochi, Japan, were unequal quadrangular pyramids with one large and three smaller SV/PC, morphologically closest to Kudoa whippsi recorded in various pomacentrid and apogonid fish from the Australian Coral Sea. The 18S and 28S rDNA nucleotide sequences of the Japanese isolate were highly similar to some Australian K. whippsi isolates, but also displayed less similarity to other K. whippsi isolates from the same sea mainly due to instability of nucleotides at certain base positions and/or segments of different isolates. All the K. whippsi isolates including the present Japanese isolate, however, were distinct from Kudoa gunterae, K. whippsi's closest kudoid species in morphology, molecular phylogeny, and biogeography. Our detection of K. whippsi from C. zonatus in the natural seawater around Japan is a new host and geographical record. Kudoid myxospores from A. hasta from the Sea of Ariake, a deep bay of the western part of Japan, exhibited two morphotypes, one resembling K. whippsi and the other Kudoa quadricornis with distinct posteriolateral SV projections. However, rDNA nucleotide sequencing revealed that these two Kudoa spp. were distinct from any known congeners; thus, Kudoa akihitoi n. sp. and Kudoa empressmichikoae n. sp. were erected. The morphological differentiation of K. akihitoi n. sp. from multiple Kudoa spp. with scalene stellate myxospores containing one large and three smaller SV/PC was difficult, whereas K. empressmichikoae n. sp. with spherical spore bodies extending small posteriolateral SV projections was distinct from known congeners with similar but elongated spore bodies and PC, i.e., K. quadricornis and Kudoa paraquadricornis, found in the trunk muscle of carangid fish from the Australian Coral Sea.
Keywords: Acanthogobius hasta; Cheilodactylus zonatus; Genetic diversity; Japan; Kudoa akihitoi n. sp.; Kudoa empressmichikoae n. sp.; Kudoa whippsi; Multivalvulida.