The morphology and phylogeny of Pleuronema binucleatum n. sp., P. parawiackowskii n. sp., and P. marinum Dujardin 1841, collected from Hangzhou Bay estuary, China, were investigated using standard methods. Pleuronema binucleatum n. sp. can be identified by possessing about 90-120 × 35-50 μm cell size in vivo, reniform body outline, two macronuclei, six to eight preoral kineties, 32-41 somatic kineties, and posterior end of the anterior fragment of membranelle 2 (M2a) hook-like. Pleuronema parawiackowskii n. sp. is characterized by the combination of the following characters: body size about 60-90 × 20-40 μm in vivo, elliptical in outline, four to eight preoral kineties, 20-29 somatic kineties, posterior portion of the M2a slightly curved but nonhooked, and single macronucleus sausage-like. After comparison with other populations of P. marinum, it is suggested that many misidentifications exist in previous studies. And an improved diagnosis of P. marinum was supplied: cell about 95-180 μm long, elliptical in outline; 2-4 preoral kineties and 53-70 somatic kineties; both membranelle 1 and membranelle 3 three-rowed; posterior end of the M2a straight; single contractile vacuole characteristically positioned near mid-body. The small subunit rRNA genes of three forms were sequenced. Phylogenetic analyses indicate that the monophyly of the genus Pleuronema is still not supported.

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