Heterosporis saurida is a microsporidian that infects lizardfish, Saurida undosquamis (Richardson, 1848), in the Arabian Sea. Spores were isolated from infected lizardfish and used to infect derived fish cell lines: common carp brain (CCB), epithelioma papulosum cyprinid (EPC), fathead minnow epithelial (FHM), rainbow trout gonad (RTG), bluegill fry (BF-2) and chinook salmon embryo (CHSE). Non-fish cell lines were also tested that include: insect (SF-9), rabbit (RK-13) and African green monkey (Vero E6). No growth of H. saurida was observed in any fish cell line, SF-9 or Vero E6 cell lines. H. saurida spores grew only in RK-13 cell line and were detected by immunofluorescence. Developmental stages of H. saurida were seen in RK-13 cells by light and transmission electron microscopy, and species identification was confirmed by sequencing. This study demonstrated that H. saurida was able to proliferate in the mammalian RK-13 cell line, which thus represents an in vitro model for conducting molecular genetics and cell-pathogen interaction studies of Heterosporis.