Cochliopodium megatetrastylus n. sp. is described based on light microscopy, fine structure and molecular genetic evidence. Amoebae are broadly oval to somewhat triangular during locomotion with average length of 37 μm and breadth of 50 μm, and surrounded by a hyaloplasm margin, somewhat narrow when at rest but more expanded during locomotion (~ 5–10 μm at the anterior). Sparsely occurring subpseudopodia, barely emergent from the hyaloplasm, are blunt and finger-like, occasionally becoming adhesive laterally or at the posterior. Cysts develop after 2–3 weeks in culture and are round with a distinct margin, decreasing in size from 20 to 5 μm during maturation. The granuloplasm contains refractile crystals. A vesicular nucleus (~ 6 μm), containing a nucleolus (2–3 μm), is variable in shape from somewhat lenticular in section to irregularly rounded with undulating or lobed margins. Surface scales (~ 0.3 μm in height) have an apical deeply concave funnel-like collar (~ 0.15 μm deep), without a spine, composed of radial fine rays and concentric filaments forming a finemesh, supported on four non-cross-linked styles (~ 0.2 μm apart) attached to a round to broadly angular base plate (0.6–1 μm) with a fine gridtexture. Cysts are rounded and enclosed by an organic wall bearing remnants of the scales on its outer surface. Both concatenated analysis of SSU-rDNA and COI genes and comparative morphologies support the designation of Cochliopodium megatetrastylus n. sp. as a new species.