Abstract
Tetmemena saprai n. sp. was isolated from fresh-water samples collected from Okhla Bird Sanctuary, Delhi, India and described based on its morphology, morphogenesis and molecular markers, namely the small-subunit (SSU) rRNA gene and internal transcribed spacers (ITS1-5.8S-ITS2). The morphological features of T. saprai n. sp. are as follows: colourless; rigid body with no cortical granules; water expelling vesicle about 15 mm in size, located below the adoral zone of membranelles near the left body margin; body size about 125-140 x 50-60 mu m in live and about 100 x 45 mu m in protargol preparations, with body length : width ratio of around 2:1; undulating membranes in a Stylonychia-pattern; adoral zone about 44 mu m in length with (on average) 45 adoral membranelles; 18 frontal-ventral-transverse cirri; one right and one left marginal row; six dorsal rows including two dorsomarginals, of which the fourth row is shortened anteriorly; three caudal cirri which are not equidistant; two macronuclear nodules; 2-4 micronuclei. A detailed morphogenetic study revealed the oral primordium to originate near the left most transverse cirrus, and the involvement of five parental cirri (three frontals and two ventrals) in the formation of streaks I-VI for the proter and opisthe. The genetic distance between T. saprai n. sp. and its congeners T. pustulata and T. vorax (S. vorax) varied from between 1% and 2% in the SSU rRNA gene and 1% and 6% in the ITS1-5.8S-ITS2 sequence. The secondary structures of the ITS1 and ITS2 RNA transcripts of four different species of genus Tetmemena (T. saprai n. sp., T. pustulata, T. vorax and T. bifaria) were also compared, revealing that the ITS1 region was more variable than ITS2. The morphological and morphogenetic characterisation, and phylogenetic analyses based on the molecular markers, confirm that the present species is a distinct species of the genus Tetmemena belonging to the subfamily Stylonychinae. This study also reveals that the macronuclear ITS region can be a suitable candidate for species identification. (C) 2020 The Authors. Published by Elsevier B.V. on behalf of King Saud University.