Axenic Cultivation of the First Human-Derived Breviata anathema from a Patient with Fascioliasis in West Bengal, India

PURPOSE: To establish axenic cultivation of a human-derived Breviata anathema (strain T76) isolated from the stool of a patient with fascioliasis and to characterize its biological properties. METHODS: T76 was maintained in American Type Culture Collection (ATCC) medium 207, supplemented with sulfate-reducing Desulfovibrio desulfuricans and co-cultured with Pseudomonas aeruginosa, at 30 °C. Axenic cultivation was attempted by transferring T76 flagellates to various axenic culture media supplemented with antibiotics for parasitic intestinal protozoan species. RESULTS: The 18 S rRNA gene sequence of T76 exhibited significant homology with the free-living, bacterivorous B. anathema (ATCC 50338), a member of the rare clade Obazoa. Axenic cultivation of B. anathema (T76) was achieved in YIGADHA-S medium, in which gluconic acid and dihydroxyacetone replaced glucose as energy substrates. CONCLUSION: B. anathema (T76) was isolated from a patient with fascioliasis, exposed through consumption of aquatic plants from ponds, a known habitat of this organism. Axenic cultivation of T76 was established in YIGADHA-S medium, originally developed for Entamoeba dispar, and cyst-like spherical bodies harboring flagellates were observed and exhibited resistance to 0.1 N HCl.