A new species of Tortanus (Atortus) (Copepoda, Calanoida, Tortanidae) from Minicoy Island, southeastern Arabian Sea

* Correspondence: sanuvf@gmail.com


Introduction
The calanoid copepod genus Tortanus Giesbrecht, 1898 (Calanoida, Tortanidae) currently comprises 47 species (Walter and Boxshall, 2018; http://www.marinespecies. org); these are mainly distributed in coastal waters of the Indo-West Pacific and the northwestern Atlantic (Mulyadi et al., 2017). They are known to be typical carnivores, feeding on copepods and other zooplankters (Ambler and Frost, 1974;Uye et al., 1994;Hooff and Bollens, 2004;Nishida et al., 2015;Mulyadi et al., 2017). Five subgenera have so far been recognized in this genus: Tortanus (Tortanus) Giesbrecht, 1898;T. (Eutortanus) Smirnov, 1933;T. (Boreotortanus) Ohtsuka, 1992; T. (Acutanus) Ohtsuka, 1992;and T. (Atortus) Ohtsuka, 1992. The subgenus Atortus, accommodating 32 nominal species (Mulyadi et al., 2017), is regarded as the most advanced and is distributed exclusively in the Indo-West Pacific region. It tends to occur in more or less oligotrophic, clear, high-salinity (34-40 PSU) waters sometimes strongly influenced by warm currents, or in coral reefs and seagrass beds (Ohtsuka and Kimoto, 1989;Ohtsuka and Reid, 1998;Ohtsuka et al., 2000;Nishida and Cho, 2005;Nishida et al., 2015;Mulyadi et al., 2017). It must also be mentioned that cooccurrence of multiple closely related species of Tortanus (Atortus) makes it difficult to determine their femalemale correspondence solely on the basis of morphology (Nishida et al., 2015;Mulyadi et al., 2017). Morphological studies with the analysis of genetic markers are essential to establish the female-male correspondence . The present study describes a new species of Tortanus (Atortus) from a coral lagoon of Minicoy Island, southeastern Arabian Sea; molecular data of the new species are also provided to determine the female-male correspondence.

Materials and methods
The zooplankton samples were collected from two stations (S1 and S2) during the daytime on 24 January 2014, from the coastal lagoon waters of Minicoy Island (8°05′N, 73°37′E) in the southeastern Arabian Sea ( Figure 1) using a plankton net (mesh size 200 µm; mouth area 0.28 m 2 ). The net was towed horizontally just below the water surface at a speed of 1 knot for 10 min. For morphological analysis, the samples were fixed in 4% buffered formalin. Tortanus (Atortus) specimens were sorted from the original samples, and mouthparts and swimming legs were dissected with needles in a 50:50 solution of glycerin and distilled water. Line illustrations were made using a drawing tube attached to a Lynx bright-field compound microscope (LM 52-1704, Vision Engineering, Palmdale, CA, USA) with 40-1000× magnifications. Type specimens were deposited in the Zoological Survey of India (ZSI), Kolkata, and nontype specimens were deposited in the museum of the Department of Marine Biology, Microbiology, and Biochemistry, School of Marine Sciences, Cochin University of Science and Technology. The morphological terminology follows Huys and Boxshall (1991) and Ohtsuka and Reid (1998), and the total length of the specimen was measured from forehead margin to caudal rami.
Etymology: The specific name minicoyensis refers to the name of the type locality of this species. It therefore is a toponymic term, agreeing in gender with the masculine generic name.

Molecular analysis
The mtCOI sequences were successfully generated using the primer pair, reaction mix, and thermal regime described above. The developed sequences of female and male T. minicoyensis sp. nov. were submitted to the NCBI database and assigned the following accession numbers: KP749951 to KP749953 for males, and KP749954 for females. ML analysis was performed, and pairwise sequence distances were generated and analyzed using the developed sequences as well as the mtCOI sequences of their 9 congeneric species acquired from the NCBI database (Table 2). Nassodonta insignis H. Adams, 1867 (KP739843) was selected as the outgroup. The ML tree clearly exhibited the differential assemblage of congeneric species of the genus Tortanus ( Figure 6). Female and male T. minicoyensis sp. nov. sequences were arrayed within a single clade with a high bootstrap value (100%), which is distinct from all other sequences of subgenus Atortus based on 1000 bootstrap pseudo-replicas. The outgroup N. insignis exhibited maximum divergence array. The level of inter-and intraspecific divergence persisting within the genus Tortanus was evident from the distance matrix data. Specifically, T. minicoyensis sp. nov. possessed an intraspecific sequence divergence ranging from 0%-0.4% (Table 3).

Species Abbreviation Accession No. Remarks
Tortanus ( (1) serrated ridge of the right antennule produced proximally over onethird of segment XIX and the ridge slightly raised from the surface plane of the segment, (2) beak-shaped medial process on the right fifth leg coxa, (3) triangular medial process with a depression on the distomedial margin on the basis, and (4) strongly curved subdistal seta on the left fifth leg. The species of the subgenus Atortus hitherto known from the Indian Ocean are Tortanus (Atortus) andamanensis and T. sigmoides Nishida, Anandavelu & Padmavati, 2015;T. magnonyx Ohtsuka & Conway, 2005; T. insularis Ohtsuka & Conway, 2003;T. nishidai Ohtsuka, El Sherbiny & Ueda, 2000;T. tropicus Sewell, 1932;and T. recticauda Giesbrecht, 1889 ( http:// copepodes.obs-banyuls.fr/en). Among them, T. tropicus is the largest (female 2.71 mm; male 2.29 mm) (Sewell, 1932). However, the new species herein described has the largest length range known in the genus (female 2.66-2.94 and male 2.25-2.74 mm). The differential diagnosis of T. minicoyensis sp. nov. with all other species of the subgenus Atortus from the Indian Ocean is represented in Tables 4 and 5. T. minicoyensis sp. nov. was found swarming along with Acartia bispinosa Carl, 1907, Labidocera madurae A. Scott, 1909, Undinula vulgaris Dana, 1849, and Centropages orsinii Giesbrecht, 1889 in the present collection. The swarming behavior of members of the subgenus Atortus have been observed in oligotrophic clear waters mainly in subtropical and tropical areas (Kimoto et al., 1988;Ohtsuka and Kimoto, 1989;Ohtsuka and Reid, 1998;Ohtsuka et al., 2000) as a response to the presence of prey copepods, and also to avoid visual predators (Ohtsuka et al., 2000). The mtCOI sequences were developed for T. minicoyensis sp. nov. to bring out the molecular variance from the congeners and establish the female-male correspondence of the species. The speciation of T. minicoyensis sp. nov. is exhibited in the genetic distance matrix, which showed an intraspecific divergence of 0%-0.4%, which is under the threshold value of 4% for calanoid copepods (Bucklin et al., 2010).

Key to the species of subgenus Atortus from the Indian Ocean:
Adult  Asymmetrical, right margin with a bulge at midpoint with 2 ventrolateral spines ( Figure 2D) Asymmetrical with 2 rounded medial processes, left ramus slightly larger than the right ( Figures  2C, 2D)

T. recticauda
Symmetrical (Bowman, 1987, fig. 1A) Not described Asymmetrical, expanded anterolaterally on both sides ending in small papilla with apical spinule; right expansion strongly convex to straight laterally (Bowman, 1987, figs. 1A-1C) Asymmetrical, left ramus slightly wider and longer than right (Bowman, 1987, figs. 1A, 1C) Produced proximally over one-third of segment XIX and the ridge slightly raised from the surface plane of the segment ( Figures 5B, 5C) Semitrapezoid with a beak-shaped medial process ( Figure 5E) Bearing triangular shaped medial process with depression on distomedial margin and fused at the distal side of its base ( Figure 5E) Strongly curved along hemispherical tip with the granular surface ( Figures  5F, 5G)