Abstract: Recent studies have revealed a high diversity of pleurostomatid ciliates in brackish habitats. Here, a novel species, Pseudolitonotus spirelis gen. et sp. n., isolated from a mangrove wetland of southern China, was investigated based on living observation, protargol staining, and molecular analyses. The new genus Pseudolitonotus gen. n. is characterized by the last left somatic kinety (LKn) being shortened and none of the right somatic kineties extending to the anterior end of the cell, thus distinguishing it from all known pleurostomatid genera. The type species, Pseudolitonotus spirelis sp. n., is characterized by the possession of two macronuclear nodules, 11–15 right and 7–9 left kineties, a single contractile vacuole subterminally located, extrusomes evenly spaced along the entire ventral margin and some forming an "apical group", two types of cortical granules, and the bottom of the oral slit invariably being twisted. Litonotus gracilis (Pan et al. Eur J Protistol 51:494–506, 2015) is believed to be another member of this new genus as its LKn and right somatic kineties are all shortened. Hence, a new combination, Pseudolitonotus gracilis (Pan et al., 2015) comb. n., is suggested and its diagnosis is improved. Molecular phylogenetic analyses based on SSU rDNA sequence data reveal that Pseudolitonotus gen. n. is monophyletic and groups with Apolitonotus (Pan et al. J Eukaryot Microbiol 67:252–262, 2020) of the family Protolitonotidae (Wu et al. Zool Scr 46:245–253, 2017). However, the familial assignment of this new genus is uncertain based on current data.