Paradoxapseudes shimojiensis sp. nov. (Crustacea: Tanaidacea: Apseudidae) from a submarine limestone cave in Japan, Title with notes on its chelipedal morphology and sexual system Author(s) Kakui, Keiichi; Fujita, Yoshihisa Marine Biology Research, 16(3), 195-207 Citation https://doi.org/10.1080/17451000.2020.1720249 Issue Date 2020-03-20 Doc URL http://hdl.handle.net/2115/80409 This is an Accepted Manuscript of an article published by Taylor & Francis in Marine Biology Research on 12/02/2020, Rights available online: https://www.tandfonline.com/doi/full/10.1080/17451000.2020.1720249. Type article (author version) File Information MAR BIOL RES16_195-207.pdf Instructions for use Hokkaido University Collection of Scholarly and Academic Papers : HUSCAP This is an Accepted Manuscript of an article published by Taylor & Francis in Marine Biology Research on 12 Feb. 2020, available online: http://www.tandfonline.com/10.1080/17451000.2020.1720249. 1 1 Paradoxapseudes shimojiensis sp. nov. (Crustacea: Tanaidacea: 2 Apseudidae) from a submarine limestone cave in Japan, with notes on 3 its chelipedal morphology and sexual system 4 5 Keiichi Kakuia and Yoshihisa Fujitab 6 7 aFaculty of Science, Hokkaido University, Sapporo 060-0810, Japan; bOkinawa 8 Prefectural University of Arts, Naha 903-8602, Okinawa, Japan 9 10 Corresponding author. Keiichi Kakui: [email protected]; Faculty of Science, 11 Hokkaido University, N10 W8, Kita-ku, Sapporo 060-0810, Japan 12 Email address for 2nd author: [email protected] 13 14 Acknowledgments 15 We thank Masaru Mizuyama, Katrine Worsaae, Peter Rask Møller, Yuji Ise, Hiroki Ichi, 16 Hirotaka Ichi, and Go Tomitani for assistance in scuba sampling in the submarine cave; 17 and Matthew H. Dick for reviewing the manuscript and editing our English. 18 19 Funding 20 This study was supported by the Japan Society for the Promotion of Science (JSPS) 21 under KAKENHI grants JP16K18597 and JP19K06800 to KK, and JP16K07490 and 22 JP16H06309 to YF. 23 24 25 26 2 27 ABSTRACT 28 We describe Paradoxapseudes shimojiensis sp. nov. from a submarine limestone cave at 29 Shimoji-jima Island, Ryukyu Islands, southwestern Japan. This species resembles five 30 species (P. basibidens, P. bassoprofundo, P. bermudeus, P. edgari, and P. heroae, 31 among 17 congeners) that bear a long pleotelson, but differs from them in having i) a 32 naked antennal article 1, ii) the maxillipedal basis with one inner distal plumose seta, 33 iii) the chelipedal basis with one dorsodistal and one ventro-subproximal simple setae, 34 iv) the pereopod-1 basis with four dorsal simple setae longer than the width of the 35 pereopod-1 basis, but without ventrodistal spiniform setae, v) the pereopod-1 merus 36 with three mid-inner ventral simple setae, and vi) the pleopodal protopod with two inner 37 plumose setae. We determined partial nucleotide sequences for the cytochrome c 38 oxidase subunit I (COI) and 18S rRNA (18S) genes in P. shimojiensis for future use in 39 DNA barcoding and phylogeny. Paradoxapseudes shimojiensis has serial ridges on the 40 inner surfaces of the left and right chelipedal bases that quite resemble the stridulatory 41 organs in harvestmen (Opiliones); by analogy, we speculate that these ridges may be 42 stridulatory sound-producing organs. Two specimens had both a fully-developed 43 marsupium and genital cone, suggesting that P. shimojiensis is simultaneously 44 hermaphroditic. 45 46 KEYWORDS: Malacostraca; Peracarida; stygophilic; hermaphrodite; acoustic; benthos 47 48 49 50 51 3 52 Introduction 53 Since the pioneering work by Băcescu (1980), 17 species of tanaidaceans in 15 genera 54 have been reported from submarine caves (Kakui and Fujita 2018; Kakui et al. 2019), 55 although most of these species are not restricted to caves (García-Herrero et al. 2019). 56 For species that have been reported only from caves, it remains unclear whether they are 57 obligate cave dwellers, both because they lack obvious external adaptations to cave life 58 and because there are insufficient data from the benthic areas outside the caves (Kakui 59 and Fujita 2018). As submarine caves are difficult to access and investigate, and minute 60 animals like tanaidaceans can easily be overlooked, research on submarine-cave 61 tanaidaceans, including their taxonomy, is still in its infancy. 62 The apseudid genus Paradoxapseudes Guţu, 1991, consisting of 17 species 63 (Tzeng and Hsueh 2014; Bird 2015), is the only tanaidacean genus with multiple 64 submarine-cave dwellers, including P. bermudeus (Băcescu, 1980), P. intermedius 65 (Hansen, 1895), and P. mortoni (Bamber, 1997) (Guţu and Iliffe 1985; Bamber 2000; 66 Gerovasileiou et al. 2016). Only one congener, P. littoralis (Shiino, 1952), has been 67 reported from Japanese waters, but not as a cave dweller (Shiino 1952). In 2018, our 68 faunal survey of a submarine limestone cave at Shimoji-jima Island, Miyako Island 69 Group, southwestern Japan, yielded one Paradoxapseudes species, which turned out to 70 be undescribed. Here we describe this species as new, provide partial sequences for the 71 cytochrome c oxidase subunit I (COI) and 18S rRNA (18S) genes, and discuss the 72 possible function of unusual serial ridges on the chelipeds, as well as indications that the 73 species is simultaneously hermaphroditic. 74 75 Materials and methods 76 All tanaidaceans were collected by SCUBA diving on 26 October 2018 in the 4 77 submarine limestone cave Akuma-no-Yakata (“Devil’s Palace” in Japanese), located on 78 a reef slope at Shimoji-jima Island, Miyako Island Group, southwestern Japan 79 (24°49'22.51"N, 125°08'07.84"E), with the entrance at a depth of about 35 m; see 80 Osawa and Fujita (2019) for detailed information on the cave. Mud deposited in holes 81 or fissures in the cave wall was collected with a commercially made aquatic suction 82 sampler (yabby pump), from the “middle zone” of the cave (50–80 m from the entrance, 83 ca. 20 m depth, no light, ca. 28‰ salinity; rocky or rubble-covered substrate) (Osawa 84 and Fujita, 2019). Tanaidaceans were sorted from the mud sample and fixed and 85 preserved in 99% ethanol. The specimens studied were deposited in the University 86 Museum Fujukan, University of the Ryukyus (RUMF), Okinawa, Japan, and in the 87 Invertebrate Collection of the Hokkaido University Museum (ICHUM), Sapporo. The 88 methods used for dissection, preparation of slides, light microscopy, scanning electron 89 microscopy (SEM), and drawing were as described by Kakui and Angsupanich (2012). 90 Orientation and terminology here follow Larsen (2003), except that the term 91 “plumose sensory seta” (PSS; Bird 2011) is used instead of “broom seta” and the term 92 “protopod” is used instead of “basal article” for pleopods and uropods. Body length 93 (BL) was measured from the base of the antennules to the tip of the pleotelson, and 94 body width at the widest portion of the cephalothorax (CW, cephalothorax width). 95 Pleotelson length (PL) was measured between the anterior and posterior edges of the 96 pleotelson, and pleotelson width (PW) at the widest portion of the segment (in many 97 specimens, the width across the lateral processes on the pleotelson). Appendages were 98 measured in the holotype. Measurements were made axially from digital images by 99 using ImageJ (Rasband 2019): dorsally on the body, antennules, antennae, and uropods; 100 laterally on the chelipeds, pereopods, and pleopods. The length and width of congeners 5 101 were measured from original illustrations if these measurements were not provided in 102 descriptions. 103 Total DNA was extracted from the left cheliped of the holotype by using a 104 NucleoSpin Tissue XS Kit (TaKaRa Bio, Japan). Table I lists the primers used for DNA 105 amplification and sequencing. PCR amplification conditions for COI with TaKaRa Ex 106 Taq DNA polymerase (TaKaRa Bio) were 94°C for 1 min; 35 cycles of 98°C for 10 s, 107 62°C for 30 s, and 72°C for 50 s; and 72°C for 2 min. Conditions for 18S amplification 108 with KOD FX Neo polymerase (Toyobo, Japan) were 94°C for 2 min; 45 cycles of 109 98°C for 10 s, 65°C for 30 s, and 68°C for 90 s; and 68°C for 3 min. Nucleotide 110 sequences were determined by direct sequencing with a BigDye Terminator Kit ver. 3.1 111 and a 3730 Genetic Analyzer (Life Technologies, California, USA), and segments were 112 concatenated by using MEGA7 (Kumar et al. 2016). All sequences we determined were 113 deposited in the International Nucleotide Sequence Database (INSD) through the DNA 114 Data Bank of Japan (DDBJ). 115 The determined COI sequence (658 nt, encoding 219 amino acids) was aligned 116 with that of P. intermedius obtained from the INSD (accession number HM016216; 117 Drumm 2010; originally identified as Apseudes cf. bermudeus) by using CLUSTAL W 118 (Thompson et al. 1994); the aligned sequences were trimmed in MEGA7 to the shortest 119 length between them (402 nt). Our 18S sequence (2176 nt) was aligned with those of P. 120 bermudeus and P. littoralis obtained from INSD (accession numbers GQ175865 and 121 AB618175; Wilson 2009; Kakui et al. 2011) by using MAFFT ver. 7 (Katoh and 122 Standley 2013) with the Q-INS-i strategy (Katoh and Toh 2008); the aligned sequences 123 were trimmed to the shortest length among them (2036 nt). Kimura (1980) 2-parameter 124 (K2P) distances among the aligned sequences in each dataset were calculated with 125 MEGA7. 6 126 127 Systematics 128 Family Apseudidae Leach, 1814 129 Genus Paradoxapseudes Guţu, 1991 130 Paradoxapseudes Guţu, 1991: 349. Type species: Paradoxapseudes cubensis Guţu, 131 1991. 132 133 Amended diagnosis modified after Guţu (2008) 134 Apseudinae with small sized, dorsoventrally flattened body.