On some squat lobsters from India (Decapoda, Anomura, Munididae), with description of a new species of Paramunida Baba, 1988

Abstract Squat lobster specimens belonging to the family Munididae were recently collected along the southwestern coast of the mainland of India and in the Andaman Islands. The specimens belong to two known species, Agononida prolixa (Alcock, 1894) and Munida compacta Macpherson, 1997, and a new species, Paramunida bineeshisp. nov. We here redescribe A. prolixa and describe and figure the new species. Munida compacta is newly recorded from India, and we figure the live coloration. In addition, molecular and phylogenetic analyses of two mitochondrial markers (16S rRNA and COI) revealed the phylogenetic relationships of M. compacta and P. bineeshisp. nov. with their most closely related congeners. The genetic similarity among the individuals of M. compacta from different locations is also addressed.


Introduction
Squat lobsters are a very diverse and abundant group of anomuran decapods that are distributed throughout the world (Schnabel et al. 2011). A high level of squat lobster diversity is found in the western-central Pacific Ocean, particularly in the Solomon-Vanuatu-New Caledonia region, the Coral Sea, the Indo-Malay-Philippine archipelago, and French Polynesia Schnabel et al. 2011;Rodríguez-Flores et al. 2019a).
During the nineteenth century, several expeditions were carried out in the Indian Ocean by the Indian survey steamer 'Investigator'. The results of these expeditions -in which abundant material was gathered and many new species discovered -had been published by several workers, e.g., Wood-Mason and Alcock (1891), Alcock (1894Alcock ( , 1901, Alcock and Anderson (1894, 1899a, 1899b, Anderson (1896), McArdle (1901), MacGilchrist (1905), and Lloyd (1907). Of the many new species described by these authors, most were from the Bay of Bengal and the Arabian Sea. Deep-sea expeditions by the German vessel 'Valdivia' also gathered extensive material, including five new species (Doflein and Balss 1913). In addition to these expeditions, other relatively recent investigations in the Indian waters have revealed many new findings on squat lobsters, including descriptions of new species (e.g., George and Rao 1966;Rao 1974;Tirmizi and Javed 1993;Thirumilu 2011;Vaitheeswaran and Venkataramani 2012;Vaitheeswaran 2014Vaitheeswaran , 2015Vaitheeswaran , 2017Komai et al. 2019). Although the seas around India are considered to be a moderately rich biogeographic region (cf. Spalding et al. 2007;Schnabel et al. 2011), the current diversity of squat lobsters from the region is not very high, and more research is needed to improve our knowledge on the group in this interesting oceanic province.
Recently, we collected specimens from India that constitute three species belonging to the genera Agononida Baba & de Saint Laurent, 1996, Munida Leach, 1820, and Paramunida Baba, 1988, all from the family Munididae Ahyong, Baba, Macpherson & Poore, 2010. Among our material, the lone specimen of Paramunida from the Andaman Islands is described herein as a new species, Paramunida bineeshi sp. nov. Agononida prolixa (Alcock, 1894) is redescribed based on a male specimen from the Andaman Islands. Several specimens of Munida compacta Macpherson, 1997 were collected from Kerala (southwestern coast of the Indian mainland), and the species newly recorded in Indian waters. The live colouration of M. compacta is described and figured here. We also conducted molecular and phylogenetic analyses using two mitochondrial markers, cytochrome oxidase subunit I (COI) and 16S rRNA (16S), for M. compacta and P. bineeshi sp. nov. to determine the phylogenetic relationships with their most closely related congeners. Furthermore, we noticed the genetic similarity among the individuals of M. compacta from different localities.

Material and methods
The material, including the holotype of the new species, is located in the Department of Aquatic Biology and Fisheries, University of Kerala, Thiruvananthapuram, Kerala, India (DABFUK). The specimens from the Andaman Islands were collected by K.K. Bineesh of the Andaman and Nicobar Regional Centre, Zoological Survey of India. The terminology and measurements follow Baba et al. (2009Baba et al. ( , 2011. The size of the specimens is indicated by the postorbital carapace length (CL), measured along the midline from the base of the rostrum to the posterior margin of the carapace. The rostrum was measured from its base, situated at the level of the orbit, to the distal tip. Measurements of appendages were taken in dorsal (pereopod 1), lateral (antennule, pereopods 2-4) and ventral (antenna) midlines. Abbreviations used are: Mxp3, maxilliped 3; P1-4, pereopods 1-4.

Molecular analysis
Tissue, taken from one of the pereopods, was used to extract genomic DNA with the DNeasy (Qiagen) kit following manufacturer's protocol. A prior digestion of the sample was performed during 18-24 hours, and RNase was included before the extraction. Partial sequences of the mitochondrial COI and 16S partial genes were amplified by polymerase chain reaction (PCR) using combinations of the following primers: tenuiCOIFwint/ tenuiCOIRev1int/ tenuiCOIRev2int (Rodríguez-Flores et al. 2019a), LCO1490 (Folmer et al. 1994), COI-H (Machordom et al. 2003) for COI; and 16SAR and 16SBR (Palumbi et al. 1991) for 16S markers. The amplified fragments were purified using ExoSAP-IT (Affymetrix). Sequencing of both strands was performed using BigDye Terminator in an ABI 3730 genetic analyzer in the SECU-GEN service (Madrid, Spain). Forward and reverse DNA sequences were obtained for each specimen, which were checked and assembled using the program Sequencher 4.8 (Gene Code Corporation). Multiple alignment for the 16S marker was carried out using MAFFT (Katoh et al. 2002) with a posterior manual correction in AliView alignment editor (Larsson 2014).
Pleon: tergites 2-4 with seven or eight, five, and four spines, respectively, on anterior ridge; tergite 4 with median spine on posterior ridge.
Antennule: article 1 (distal spines excluded) about one-third CL, slightly overreaching cornea, with two distal spines, mesial spine clearly shorter than lateral spine; two spines on lateral margin, proximal one small, located at midlength of article, distal one long, not overreaching distolateral spine.
Antenna: article 1 with one distal spine on mesial margin, reaching end of article 2; article 2 with minute distomesial spine, distolateral angle unarmed; article 3 with long distomesial spine, slightly exceeding article 4.
Mxp3: ischium about twice length of merus measured along dorsal margin, distoventrally bearing long spine; merus with one strong median spine on flexor margin; extensor margin unarmed. P1: long, subequal, squamous, 4× CL, with few plumose setae and few scattered iridescent setae on mesial borders of articles. Merus 1.8× CL, 2.6× as long as carpus, with few dorsal and mesial spines. Carpus 0.6× length of palm, nearly 4× as long as broad, with few spines along mesial and dorsal sides. Palm about 7× longer than broad, with few minute dorsal spines. Fingers 0.7× length of palm, unarmed.
Remarks. Agononida prolixa was originally described as Munida squamosa var. prolixa by Alcock (1894) (Anonymous 1914). It was subsequently found in the Gulf of Mannar, the Andaman Sea, the Arabian Sea off the coast of Sri Lanka, and south-west of Nicobar Islands (Alcock and Anderson 1894; Alcock 1901; Doflein and Balss 1913). More recently, the species was recorded near Kollam (= Quilon), south-western India at depths of 220-360 m (Rao 1974). The variety described by Alcock (1894) was raised to the rank of species by Ahyong and Poore (2004), which was subsequently followed by Baba (2005) and Baba et al. (2008). Agononida prolixa was figured by Alcock and Anderson (1895), and its antennule, antenna, and male pleopod by Rao (1974). We here redescribe the species and illustrate its additional body parts for clarifying its taxonomy.
Agononida prolixa belongs to the group of species that have the carapace without protogastric spines, the lateral branchial margin with three spines, the pleonal tergite 4 armed with a spine on the posterior transverse ridge, and the antennal peduncle article 1 with a moderate-sized process that does not overreach article 4. The closest relatives of A. prolixa are A. isabelensis Cabezas, Macpherson & Machordom, 2009, from the Solomon Islands and Papua New Guinea (Cabezas et al. 2009;Macpherson et al. 2020), and A. nielbrucei Vereshchaka, 2005, from New Zealand and the Norfolk Ridge (Vereshchaka 2005;Ahyong 2007). Agononida prolixa is easily distinguished from A. isabelensis by the presence of a pair of median spines directly anterior to the posterior margin of the carapace, whereas these spines are absent A. isabelensis. Differences with A. prolixa and A. nielbrucei include the following: • In A. nielbrucei, the dorsal surface of the carapace has one branchiocardiac spine behind the postcervical spine on each side, and a second cardiac spine behind the median large spine. These spines are absent in A. prolixa.
• The distomesial spine of the antennular article 1 is larger than the distolateral spine in A. nielbrucei while it is shorter in A. prolixa.  Diagnosis. Carapace with spiniform rostrum, five spines on branchial margin. Pleonal tergite 2 with more than six spines along anterior ridge, more than five transverse ridges. Thoracic sternum smooth, with few scales on sternite 4; sternite 4 narrowed, subtriangular, narrowly fitting to sternite 3. Eyes large, corneal width distinctly more than one-third distance between base of anterolateral spines. Antennular basal article with distomesial and distolateral spines subequal (or slightly different) in size. Antenna with distomesial spine of article 1 not exceeding end of article 2; distomesial spine of article 2 slightly overreaching end of article 3. Mxp3 merus unarmed on extensor distal margin. P1 fixed finger laterally with (one or two) subterminal spines only, occasionally with proximal spine; movable finger with proximal mesial spine only. P2-4 dactyli with movable spines along entire flexor margin.
Some specimens from Papua New Guinea have an orange carapace and whitish supraocular spines .
Genetic data. 16S and COI. Remarks. The species was originally described from the specimens collected near the Kei Islands of Indonesia at depths 246-694 m (Macpherson 1997). It was also recently reported from Papua New Guinea at depths 220-1012 m ). The specimens from India and Papua New Guinea are morphologically similar except for their colour differences (see notes on the colouration of M. compacta). Very low genetic distances were observed among the specimens from south-western India and Papua New Guinea (0.3-0.8% for COI and 0.0-0.6% for 16S), which support they are conspecific (Fig. 1A).
The most morphologically similar species to M. compacta is M. rhodonia from the south-western Pacific. Munida compacta and M. rhodonia can be readily distinguished from each other by the size of the second lateral marginal spine of the carapace, which is located immediately behind the anterolateral spine. The second lateral marginal spine of the carapace is well developed in M. compacta but very small in M. rhodonia. The distal part of the P2-4 propodi is distally broadened in M. compacta, while it is uniform in M. rhodonia. Furthermore, the ultimate flexor marginal spine (movable) of the P2-4 dactyli is nearly equidistant between the penultimate spine and the tip of the terminal claw in M. compacta, whereas it is much closer to the penultimate spine than to the tip of the terminal claw in M. rhodonia . A very low genetic divergence was observed between the two species (1.4% for COI and 0.3-0.5% for 16S) (Fig. 1A). In fact, the low genetic distance values may even be within the thresholds considered for intraspecific divergences. However, the morphological differences among the specimens of each species examined from Indonesia, Papua New Guinea, and India are constant (Macpherson 1997;Macpherson et al. 2020; this study), which suggests that M. compacta and M. rhodonia are separate species. A similar pattern has also been observed in other species comparisons. For instance, a low genetic distance (2.7% for COI, 0.3% for 16S), but with constant morphological differences, was observed between Munida iris A. Milne Edwards, 1880  In crustacean studies, a threshold to delimit species has been around 3% genetic divergence for COI (Lefébure et al. 2006;Rodríguez-Flores et al. 2019b). As such, M. compacta could be considered a junior subjective synonym of M. rhodonia. It is also possible that the sequence data is currently insufficient to differentiate these species. Nei (1987) pointed out that the speciation is based on the patterns of nucleotide substitutions, which occurs at certain rates. These two species seem to constitute lineages in an early stage of speciation. In these situations, further investigations are needed to clarify the taxonomy of M. compacta.
Munida andamanica Alcock, 1894, which occurs from India to the western Pacific Ocean, is also morphologically similar to M. compacta. These species are usually distinguished by the number of transverse ridges on the pleonal tergite 2: M. compacta has more than five transverse ridges while M. andamanica possesses only up to four transverse ridges (Baba 2005). The specimens of M. compacta examined by us, however, possess four to seven transverse ridges, which overlap with the count for M. andamanica. Therefore, this character state should be used with caution. Other characters are also variable. For instance, the P1 fixed finger has only one or two subterminal spines on the lateral margin in the specimens from Indonesia and Papua New Guinea, whereas the Indian specimens have both proximal and subterminal spines. Unfortunately, no genetic data is currently available for M. andamanica.
The interspecific and intraspecific variations in morphology and colour pattern seen among these three species (M. andamanica, M. compacta, and M. rhodonia) strongly demand a revision of the taxa involving more specimens from different localities, with both morphological and genetic data.
Description of ovigerous female holotype. Carapace: as long as broad. Dorsal surface covered with numerous spines and spinules, each usually on very short arcuate striae, with few uniramous setae. Epigastric region with two spines, each behind supraocular spine; with median row of spines behind rostral spine. Mesogastric region with three well-developed spines in midline, anterior two spines thicker than anterolateral spine. Cervical groove distinct. Cardiac and anterior branchial regions circumscribed. Cardiac region with median row of three well-developed spines, first thicker than others. Each branchial region with row of moderate-sized spines near cardiac region. Posterior transverse ridge with one well-developed median spine. Frontal margin slightly concave. Lateral margins convex, with small spines. Anterolateral spine reaching sinus between rostral and supraocular spines. Rostrum short, triangular, with thin dorsal longitudinal carina; supraocular spines shorter than rostrum; margin between rostral and supraocular spines straight.
Pleon: tergites 2 and 3 each with four moderate-sized spines on anterior ridge, posterior ridge with two moderate-sized median spines. Tergite 4 with four to six spines on anterior ridge; posterior ridge with distinct, single median spine.
Eyes: maximum corneal diameter more than one-third distance between bases of anterolateral spines.
Antennule: article 1 exceeding cornea, with distomesial spine slightly shorter than distolateral; about twice longer than wide, with fringe of long setae along lateral margin; lateral margin with distal slender portion about half as long as proximal inflated portion.
Antenna: anterior prolongation of article 1 clearly overreaching antennular article 1 by about one-third of its length; article 2 (excluding spines) less than twice length of article 3, 1.5× as long as wide, ventral surface with small scales; distomesial spine long, slightly mucronated, slightly exceeding antennal peduncle, nearly reaching midlength of anterior prolongation of article 1, distolateral spine small, not reaching end of article 3; article 3 slightly longer than wide, unarmed.
Mxp3: ischium about twice length of merus measured along dorsal margin, distoventrally bearing 1 spine; merus with 1 strong median spine on flexor margin; extensor margin unarmed. P1: long, subequal, squamous, 3.0× CL, with dense plumose setae and scattered iridescent setae on mesial borders of articles. Merus 1.3× CL, 1.7× as long as carpus, with dorsal and mesial spines; distal spines strong, distomesial spine not reaching proximal quarter of carpus. Carpus slightly shorter than palm, 5.5× as long as broad, with spines along mesial and dorsal sides. Palm 6× longer than broad, with spines along mesial margin. Fingers 0.8× length of palm; movable finger with one small proximal mesial spine; fixed finger unarmed.
P2-4: long and slender, with scales on lateral sides of meri, carpi, and propodi; each scale with short setae; with dense plumose setae and scattered iridescent setae on extensor borders of articles. P2 2.8× CL, merus 1.2× longer than CL, 10.5× as long as high, 1.5× as long as propodus; propodus 10× as long as high, 1.6× length of dactylus. Merus with well-developed extensor marginal spines, increasing in size distally, flexor margin with few spines and one well-developed distal spine; row of small spines along flexolateral margin. Carpus with small extensor spines, distal spine on extensor and flexor margin. Propodus with small movable flexor spines. Dactylus gently curved, with longitudinal carinae along mesial and lateral sides, ventral border unarmed. P3 with similar spination and segment proportions as in P2; merus as long as P2 merus; propodus and dactylus as long as those of P2. P4 slightly shorter than P2; merus 1.2× CL; propodus and dactylus slightly longer than those of P3; merocarpal articulation clearly exceeding end of anterior prolongation of first segment of antennal peduncle.
Etymology. The new species is named after Kinattum Kara Bineesh who collected this species and kindly passed it to us for study. The species epithet is a noun in the genitive singular.
Remarks. Paramunida bineeshi sp nov. is closely related to P. mozambica from the south-western Indian Ocean. The two species can be easily distinguished by the following characters: • The two anterior spines of the cardiac region are larger than the supraocular spines in the new species, whereas they are smaller in P. mozambica.
• The antennal article 3 is slightly longer than wide in the new species, whereas it is nearly twice as long as wide in P. mozambica.
• The spines along the flexor and extensor margins of P2-4 are larger in the new species than in P. mozambica.
The new species is also related to P. marionis from the southwestern Indian Ocean. The two species can be differentiated on the basis of the following characters: • The spines of the median row on the gastric and cardiac regions are clearly stronger in the new species than in P. marionis.
• The posterior transverse ridge of the carapace has a well-developed median spine in the new species, whereas the spine is absent in P. marionis.
• The anterolateral spine reaches the sinus between the rostral and supraocular spines in the new species, whereas it clearly extends beyond the sinus in P. marionis.
• The distomesial spine of the antennal article 2 is slightly mucronated or blunt in the new species but spiniform in P. marionis.
Distribution. The new species is only known from the type locality in the Andaman Sea. Depth: 320 m.