Zoosyst. Evol. 99 (2) 2023, 545-556 | DO! 10.3897/zse.99.108032 yee BERLIN A new freshwater species of Gnorimosphaeroma (Crustacea, Isopoda, Sphaeromatidae) from Chichi-jima Island, Ogasawara Islands, Japan Ko Tomikawa', Junpei Yoshii', Akari Noda!, Chi-Woo Lee?, Tetsuro Sasaki?, Naoya Kimura*, Noboru Nunomura® Graduate School of Humanities and Social Sciences, Hiroshima University, Higashihiroshima, Hiroshima 739-8524, Japan Nakdonggang National Institute of Biological Resources, 137, Donam 2-gil, Sangju-si, Gyeongsangbuk-do 37242, Republic of Korea Institute of Boninology, Nishi-machi, Chichi-jima, Ogasawara, Tokyo 100-2101, Japan Tokiwazaka 1-7-18, Hirosaki, Aomori 036-8263, Japan Noto Marine Laboratory, Institute of Nature and Environmental Technology, Kanazawa University, Ogi, Noto-ché, Kanazawa, Ishikawa 927- 0553, Japan oF WN FP https://zoobank. org/55 EB7C8D-3 FA 4-4DDF-9340-A54B33999627 Corresponding author: Ko Tomikawa (tomikawa@hiroshima-u.ac.jp) Academic editor: Luiz F. Andrade # Received 15 June 2023 # Accepted 16 October 2023 Published 21 November 2023 Abstract This study describes Gnorimosphaeroma rivulare sp. nov. from a stream on Chichi-jima Island, Ogasawara Islands, Japan. This is the second freshwater species of Gnorimosphaeroma and the third Sphaeromatidae from oceanic islands. Gnorimosphaeroma rivulare sp. nov. is morphologically similar to G. boninense Nunomura, 2006, G. naktongense Kwon & Kim, 1987 and G. saijoense Nunomura, 2013. However, G. rivulare sp. nov. differs from these species in various morphological features, such as the shape of pleotelson and pereopod 2, relative length of antennule peduncular articles and pleopod 3 rami, number of setae on maxillula and maxilliped, and seta- tion on pereopod 3. Phylogenetic analyses revealed that G. akanense is sister to G. saijoense, and together they are sister to G. hokuri- kuense. This three taxa clade is sister to G. rivulare sp. nov. with G. iriei basal to them all. Our analysis concludes that G. boninense from Haha-jima Island, Ogasawara Islands is only distantly related to G. rivulare and may represent an independent colonization event. Key Words freshwater, inland water, isopod, molecular phylogeny, oceanic island, stream, taxonomy Introduction The Ogasawara Islands, oceanic islands, are a group of approximately 30 islands located in the Pacific Ocean Generally, oceanic islands do not occur on continental shelves. These are islands that have never been connected to a continental landmass. Inland water-dwelling organ- isms cannot reach oceanic islands without crossing the ocean or speciating from organisms that find themselves in a new habitat to which they have adapted, e.g., marine and then isolated in freshwater. Therefore, the occurrence of freshwater fish and invertebrates on oceanic islands 1s generally limited (Lévéque et al. 2008; Strong et al. 2008; Vainola et al. 2008; Wilson 2008). approximately 1000 km southeast of the Japanese archi- pelago. The Ogasawara Islands have small and well-de- veloped rivers inhabited by freshwater crustaceans, mol- luscs, and caddisflies. These taxa have limited diversity on oceanic islands (Satake and Cai 2005; Nunomura and Satake 2006; Miura et al. 2008; Tomikawa et al. 2022; Ito et al. 2023). More than 10,600 species of Isopoda have been de- scribed worldwide, occurring tn diverse aquatic and terres- trial environments (Boyko et al. 2023). Approximately 950 Copyright Tomikawa, K. et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 546 species of isopods have been recorded from inland waters; however, the number of species occurring 1n inland waters of oceanic islands is far fewer, with only a few species recorded from Pacific Ocean islands (Jaume and Quein- nec 2007; Wilson 2008). The aquatic sphaeromatid genus Gnorimosphaeroma Menzies, 1954 has 26 described spe- cies (Boyko et al. 2023). They are restricted to the Japa- nese Islands, Korean Peninsula, Mainland China, Alaska, and the eastern Pacific coast of North America (Tattersall 1921; Jang and Kwon 1993; Nunomura 2013; Wetzer et al. 2021). Most Sphaeromatidae are marine (Sket and Bruce 2004; Bruce 2005; Jaume and Queinnec 2007). The ge- nus Gnorimosphaeroma is unusual because it includes not only marine species, but also has seven brackish and eight freshwater described species (Fig. 1, Table 1). Satake and Ueno (2013) reported an unidentified species of Gnorimosphaeroma from a stream on Chi- chi-jima Island, Ogasawara Islands. A recent collection of Gnorimosphaeroma was made on Chichi-jima Island during the author’s KT and NN research expedition to the Ogasawara Islands. A colleague provided an addi- tional collection of seven specimens. In this study, we describe this previously undescribed species of Gnorim- osphaeroma. To clarify the phylogenetic position of this Gnorimosphaeroma species, we performed a molecular phylogenetic analyses based on nuclear 18S rRNA and mitochondrial 16S rRNA genes. Materials and methods Samples Specimens of an unidentified species were collected from under the boulders in the upper stream of Nagatani Riv- er, Chichi-jima Island, Ogasawara Islands, Japan (Fig. 2). Our molecular phylogenetic analysis includes nine de- scribed species of Gnorimosphaeroma, one unidentified species from Chiba, Japan, one unidentified species from San Francisco Bay, and the new undescribed freshwater species described in this paper (Table 2). The specimens were collected using a fine-mesh hand-net and samples were subsequently fixed in 70% or 99% ethanol in-situ. Specimens fixed in 70% ethanol were transferred to 99% ethanol in the laboratory. Morphological observation All appendages of G. rivulare sp. nov. were dissected in 80% ethanol and mounted in gum-chloral medium. The slides were examined using a stereomicroscope (Olym- pus SZX7, Japan) and a light microscope (Nikon Eclipse Ni, Japan), and the body and appendages were illustrat- ed using a camera lucida. One male (paratype, NSMT-Cr 31507) was dehydrated through a graded ethanol series, and dried using hexamethyldisilazane (HMDS) (Nation zse.pensoft.net Tomikawa, K. et al.: New Gnorimosphaeroma from Ogasawara Islands 1983). They were then sputter-coated with gold and ob- served using scanning electron microscopy (SEM, JSM- 6510LV). Body length was measured as a straight-line distance from the rostral point to the posterior margin of pleotelson within the nearest 0.1 mm. Type specimens were deposited at the National Museum of Nature and Science, Tsukuba, Japan (NSMT). Molecular phylogenetic analyses Genomic DNA was extracted from the appendage mus- cles of the specimen following procedures detailed by Tomikawa et al. (2014). The primer set 16Sar and 16Sbr (Palumbi et al. 1991) was used to target the mitochondri- al 16S rRNA (16S), whereas the 18SF [5'-AAGATTA- AGCCATGCATGTC-3'] and 18SR [5'- GCTGGAAT- TACCGCGGCTGC-3'] primer pair were designed to target the nuclear 18S rRNA (18S). PCR and DNA se- quencing were performed using the method detailed by Tomikawa (2015). The newly obtained DNA sequences were deposited in the International Nucleotide Sequence Databases (INSD) through the DNA Data Bank of Japan (DDBJ) (Table 2). The phylogenetic analyses were conducted based on 16S and 18S sequence data generated for this project and also includes previously published sequences. Ancinus sp. (Ancinidae) and two Chitonosphaera species (Sphaero- matidae), C. /ata (Nishimura, 1968) and C. salebrosa (Nishimura, 1969) were used as the outgroup (Wetzer et al. 2013, 2018). The sequences were aligned using the Muscle algorithm implemented in MEGA XI (Tamura et al. 2021). The aligned lengths of the 16S and 18S were found to be 518 and 559 bp, respectively. Moreover, the concatenated sequences yielded 1077 bp of align- ment positions. Phylogenetic relationships were reconstructed via maximum likelihood (ML) and Bayesian inference (BI) and partitioned by 16S and 18S datasets. ML anal- yses were conducted using IQ-TREE web server (ver. 1.6.12, see http://www. igtree.org/; Trifinopoulos et al. 2016) with 1000 ultrafast bootstrap replicates (Hoang et al. 2018). The best evolutionary models were select- ed based on the corrected Bayesian information criteri- on (BIC) using ModelFinder (Kalyaanamoorthy et al. 2017): for 16S, TIM2+F+G4; for 18S, TNe+G4. BI and Bayesian posterior probabilities (PPs) were estimat- ed using MrBayes v. 3.2.5 (Ronquist et al. 2012). The best-fit partition scheme and models for each partition were selected with the BIC using PartitionFinder with the ‘greedy’ algorithm, and for 16S, GTR+G; for 28S, JC+I. Two independent runs of four Markov chains were conducted for 10 million generations, and the tree was sampled every 100 generations. The parameter es- timates and convergence were checked using Tracer v. 1.7.1 (Rambaut et al. 2018), and the first 50,001 trees were considered burn-in and discarded. Zoosyst. Evol. 99 (2) 2023, 545-556 @ Marine ® Brackish @ Freshwater Sea of Okhotsk G. hoestlandti¥ G. anchialos / G. ovatum ae a i * G. naktongense | _ G. rebunense ’ _— G. kurilense / / f \ Se ~~ G, akanense j_ —— G. hokurikuense __--— G. pulchellum ~~ G. izuense “a ———— “yp ® ~ G. shikinense a : G. chejuense | oes “~ G. albicauda i ine Ly \ \ \ \ \ \ \ G. iriei \ \ \ G. boninense . iriei a \ \ G. tondaense \ G. trigonocaudum \ G. saijoense G. paradoxa G. tsushimaense Bering Sea North Pacific Ocean ay ae , 1 \ ~ G. hachijoense \ \ \ \ ™ G. rivulare sp. nov. Figure 1. Map showing the type localities of Gnorimosphaeroma species. Table 1. The type localities and habitats of Gnorimosphaeroma species. Species G. akanense Nunomura, 1998 G. albicauda Nunomura, 2005 G. anchialos Jang & Kwon, 1993 G. boninense Nunomura & Satake, 2006 G. chejuense Kim & Kwon, 1988 G. chinense (Tattersall, 1921) G. hachijoense Nunomura, 1999 G. hoestlandti Kim & Kwon, 1985 G. hokurikuense Nunomura, 1998 G. insulare (Van Name, 1940) G. iriei Nunomura, 1998 G. izuense Nunomura, 2007 G. kurilense Kussakin, 1974 G. naktongense Kwon & Kim, 1987 G. noblei Menzies, 1954 G. oregonense (Dana, 1853) G. ovatum (Gurjanova, 1933) G. paradoxa (Nunomura, 1988) G. pulchellum Nunomura, 1998 G. rayi Hoestlandt, 1969 G. rebunense Nunomura, 1998 G. rivulare sp. nov. G. saijoense Nunomura, 2013 G. shikinense Nunomura, 1999 G. tondaense Nunomura, 1999 G. trigonocaudum Nunomura, 2011 G. tsutshimaense Nunomura, 1998 Type locality Akan River, Hokkaido Hatsuneura, Chichi-jima Island, Ogasawara Islands Lake Songjiho Haha-jima Island, Ogasawara Islands Cheju Island Whangpoo River, Shanghai Hachijo Island, Tokyo Mukho Yomokuro-ike Pond, Toyama San Nicolas Island, California Lake Ezu, Kumamoto Izu, Shizuoka off Shikotan Island, Kuril Islands Nakdong River, Busan Tomales Bay, California Vancouver, British Columbia Sea of Japan (details unknown) Uwajima, Ehime Obitsu River, Chiba Tomales Bay, California Rebun Island, Hokkaido Nagatani River, Chichijima Island, Ogasawara Islands Shiotori River, Ehime Shikine Island, Tokyo Tonda River, Wakayama Hijikuro River, Nagasaki Tsushima Island me G. oregonense~ G. noble; —— G. rayi ; oa G. insulare ~ 1,000 km a a a es fe Habitat freshwater marine brackish freshwater marine brackish marine marine freshwater marine freshwater freshwater marine freshwater brackish marine marine marine brackish marine freshwater freshwater brackish marine brackish brackish freshwater zse.pensoft.net 548 Tomikawa, K. et al.: New Gnorimosphaeroma from Ogasawara Islands Table 2. Samples used for the phylogenetic analyses. Sequences marked with an asterisk (*) were newly obtained in the present study. ND, no sequence available. Species Voucher or isolate # Locality INSD # Note 16S 18S Sphaeromatidae Gnorimosphaeroma akanense G1913 Lake Akan, Hokkaido, Japan LC765314* LC765326* Topotype Gnorimosphaeroma boninense G1814 Chibusa Dam, Haha-jima |., Ogasawara Is., Japan LC765315* LC765327* Topotype Gnorimosphaeroma hokurikuense G1943 Ota, Toyama, Japan LC765316* LC765328* Topotype Gnorimosphaeroma iriei G1894 Lake Ezu, Kumamoto, Japan LC765317* LC765329* Topotype Gnorimosphaeroma noblei RWO2.021.1541 Tomales Bay, California, USA KU248168 JF699554 Topotype Gnorimosphaeroma oregonense RW10.003.3131 Vancouver, British Columbia, Canada MH427781 ND Topotype Gnorimosphaeroma rayi RW09.002.2567 Tomales Bay, California, USA MH427784 ND Topotype Gnorimosphaeroma Saijoense G1902 Kamo R., Ehime, Japan LC765319* LC765331* Topotype Gnorimosphaeroma rivulare sp. nov. G1820 Nagatani River, Chichijima |., Ogasawara Is., Japan LC765320* LC765332* Paratype Gnorimosphaeroma rivulare sp. nov. NSMT-Cr 31490; G1826 Nagatani River, Chichijima |., Ogasawara Is., Japan LC765321* LC765333* Paratype Gnorimosphaeroma rivulare sp. nov. NSMT-Cr 31486; G1892 Nagatani River, Chichijima |., Ogasawara Is., Jagoan LC765322* LC765334* Paratype Gnorimosphaeroma sp. G1954 Obitsu R., Chiba, Japan LC765318* LC765330* Gnorimosphaeroma sp. RW02.060.2550 San Francisco Bay, California, USA MH427743 ND Gnorimosphaeroma tondaense G1972 Takase River, Wakayama, Japan LC765323* LC765335* Topotype Outgroup Ancinidae Ancinus sp. RW05.010.1475 Naos Island, Republic of Panama KU248307 JF699514 Sphaeromatidae Chitonosphaera lata G1935 Takase R., Wakayama, Japan LC765324* LC765336* Chitonosphaera salebrosa G1937 Edura,Wakayama, Japan NC765375%- IC 765337 Results 2 February 2021. Paratypes: 3 males, 5.3 mm (NSMT-Cr 31486; G1892), 4.7 mm (NSMT-Cr 31487; G1893), Taxonomy 5.2 mm (NSMT-Cr 31507) 1 female 3.2 mm (NSMT-Cr Family Sphaeromatidae Latreille, 1825 Genus Gnorimosphaeroma Menzies, 1954 Gnorimosphaeroma rivulare Tomikawa, Yoshii & Nunomura, sp. nov. https://zoobank.org/E97CCBDD-FS5FE-473F-BD78-B98DB4230BEB Figs 2-7 New Japanese name: Chichijima-kotsubumushi Type materials. Holotype: male 4.9 mm (NSMIT-Cr 31485), upper stream of Nagatani River (27°04.051'N, 142°12.938'E), alt. 160 m, Chichi-jima Island, Ogasawara Islands, Tokyo, Japan, collected by Tetsuro Sasaki on 31488), data same as for holotype; 2 males, 4.1 mm (NS- MT-Cr 31489), 4.0 mm (NSMT-Cr 31490; G1826), local- ity same as for holotype, collected by Noboru Nunomura on 16 December 2019. Type locality. Japan, Tokyo: Ogasawara Islands, Chichi-jima Island, upper stream of Nagatani River (Figs 1, 2A). Diagnosis. Pleonites incompletely fused; anterior su- ture line longer than posterior one. Pleotelson posterior margin rounded. Maxillula medial lobe with 4 plumose setae and a short single seta; lateral lobe with 10 robust setae. Maxilla medial lobe with 17 setae; middle lobe with 12 setae; lateral lobe with 13 setae. Pereopod 1 ba- sis with a single seta on posterodistal corner; merus with + cy " tn RE EY Figure 2. Habitat and live specimen of Gnorimosphaeroma rivulare sp. nov. A. Habitat; B. Living state of the holotype, male 4.9 mm (NSMT-Cr 31485). zse.pensoft.net Zoosyst. Evol. 99 (2) 2023, 545-556 549 FERC A Oars SS [| : y 0.1 mm: K, L « 1.0mm:B, C-E, J,M : aC eis oe Figure 3. Gnorimosphaeroma rivulare sp. nov. A-F, H, J-M. Holotype, male 4.9 mm (NSMT-Cr 31485); G, I. Paratype, male 5.3 mm (NSMT-Cr 31486). A. Habitus, dorsal view; B. Antennula, medial view; C. Antenna, medial view; D. Labrum, dorsal view; E. Right mandible, medial view; F. Incisor and setal row of right mandible, medial view; G. Incisor of right mandible, lateral view; H. Incisor, lacinia mobilis and setal row of left mandible, medial view; I. Incisor and lacinia mobilis of left mandible, medial view; J. Paragnaths, posterior view; K. Maxillula, anterior view; L. Maxilla, anterior view; M. Maxilliped, anterior view. zse.pensoft.net 550 4 setae on anterodistal corner. Pereopod 2 propodus sub- rectangular, not swollen. Pereopod 3 merus, carpus, and propodus sparsely setulose. Uropod exopod length 0.7 times as long as endopod. Description. Male [NSMIT-Cr 31485, holotype, 4.9 mm]. Body (Figs 2B, 3A) ovate, vaulted, ca. 1.5 times longer than wide, dorsal surface unornamented, without marginal setae; eyes ovate, simple. Coxal plates (Figs 3A, 7A) without visible distinct articulation to pere- onites, partly overlapped, distal coxal margins rounded without setae; coxa 3 tapered anteriorly and posteriorly such that the animal can conglobulate. Pleon (Figs 3A, 7B) incompletely fused; pleonite 1 almost invisible, hid- den by pereonite 7; anterior suture line between pleonites 2 and 3 longer than posterior one between pleonites 3 and 4: pleonite 5 fused with pleonite 4, barely recognizable. Pleotelson (Figs 3A, 7B) slightly narrower than pleon, length 0.5 times longer than wide, posterior margin en- tire, rounded, with minute setae. Antennule (Fig. 3B) exceeding posterior margin of head; length ratio of peduncular articles 1-3 as 1.0: 0.4: 0.5; peduncular article 1 ovate, length 1.6 times as long as wide, anterior margin with a few setae, posterior margin lined with setae; peduncular article 2 subquadrate, length 1.2 times as long as wide, anterior and posterior margins lined with setae; peduncular article 3 rectangular, slender, length 3.0 times as long as wide, with setae on anterior and posterior margins; flagellum 6-articulate, articles 2 and 3 each with 2 aesthetaescs and articles 4 and 5 each with an aesthetasc. Antenna (Fig. 3C) reaching distal margin of pereonite 2; length ratio of peduncular articles 1-5 as 1.0: 0.9: 0.9: 1.5: 1.5; peduncular articles 1-3 sub- quadrate with fine setae on anterior margin; peduncular articles 4 and 5 subrectangular, each length 1.9 times lon- ger than wide, anterior margin of article 4 with fine setae, article 5 with a few setae; flagellum 12-articulate (only the first five articles drawn in Fig. 3C). Labrum (Fig. 3D) anterior margin rounded with fine setae. Mandibles (Fig. 3E—I) left and right incisors with 4 and 3 cusps, respectively; lacinia mobilis of left man- dible with 3 cusps; setal row of left and right mandibles comprised 7 and 9 serrate setae; palp 3-articulate, length ratio of articles 1-3 as 1.0: 1.2: 0.8, article 1 with fine setae, article 2 with 10 setae on lateral margin and fine setae on medial margin, article 3 with 11 setae on later- al margin. Paragnaths (Fig. 3J) with rounded shoulders, bearing setae. Maxillula (Fig. 3K) medial lobe narrow, slightly shorter than lateral lobe, with 4 plumose setae and a short simple seta; lateral lobe with 10 robust setae, 4 of which serrate. Maxilla (Fig. 3L) medial lobe with 17 setae, some of which plumose; middle lobe with 12 setae; lateral lobe with 13 setae. Maxilliped (Fig. 3M) endite reaching distal margin of palp article 2, distal margin with 8 plumose setae and 5 simple setae, medial margin with a coupling hook and 4 plumose setae; palp 5-articulate, articles 2-4 with 3, 3 and 6 setae on lateral margin, re- spectively, produced mediodistally with setae, article 5 narrow with marginal setae. zse.pensoft.net Tomikawa, K. et al.: New Gnorimosphaeroma from Ogasawara Islands Pereopod | (Fig. 4A, B) basis with a ventrodistal simple seta, ventral margin with fine setae; merus lobate dorsodis- tally, dorsodistal corner with 4 setae, ventral margin with robust setae and fine setae; carpus short with 2 robust setae and fine setae on ventral margin; propodus oval, swollen, length 2.5 times as long as wide, with 2 robust setae on ventral margin and 5 setulate setae close to ventral margin; dactylus length 0.6 times as long as propodus. Pereopod 2 (Fig. 4C, D) basis with a ventrodistal simple seta; mer- us weakly lobate dorsodistally, with 5 setae on dorsodistal corner, ventral margin with fine setae; carpus with 6 robust and 3 slender setae on distal surface, ventral margin with fine setae; propodus subrectangular, not swollen, length 2.8 times as long as wide, with 3 robust setae on ventral mar- gin; dactylus length 0.5 times as long as propodus. Pereo- pod 3 (Fig. 4E, F) basis with a ventrodistal simple seta; merus lobate dorsodistally, dorsodistal corner with 6 setae, ventral margin with sparse fine setae; carpus with 5 robust and some slender setae on distal margin, ventral margin with sparse fine setae; propodus length 2.8 times as long as wide, with 2 robust setae and a slender single seta on ven- tral margin; dactylus length 0.5 times as long as propodus. Pereopod 4 (Fig. 4G) basis with a simple seta on ventrodis- tal corner and some broom setae on dorsal margin; ischium with fine setae on ventral margin; merus lobate dorsodis- tally with robust setae, dorsal and ventral margins with fine setae; carpus with sparse fine setae on ventral margin; propodus with 3 robust setae on ventral margin, dorsal and ventral margins with sparse fine setae. Pereopod 5 (Fig. 4H) basis with a simple seta on dorsodistal corner; merus and carpus with robust setae on distal margins; propodus with 3 robust setae on dorsal margin. Pereopods 6 and 7 (Fig. 5A, B) basis with a simple seta on dorsodistal cor- ner; merus and carpus with robust setae on distal margins; propodus with 2 robust setae on dorsal margin. Penial process (Fig. 5C) simple, length 3.54.2 times as long as basal width, close set. Pleopod 1 (Fig. 5D) peduncle length 0.5 times width with 4 setae on mediodistal corner; exopod oval, length 1.9 times width, 1.2 times length of endopod, with plumose setae marginally; endopod subtriangular, length 1.4 times width, with distal plumose setae. Pleopod 2 (Fig. SE) peduncle length 0.4 times width, mediodistal and laterodistal corners with 2 and 1 setae, respective- ly; exopod oval, length 1.8 times width, with marginal plumose setae; endopod subtriangular, length 1.6 times width, with plumose setae on distal margin; appendix masculina slender, length 6.0 times width, 1.2 times as long as endopod. Pleopod 3 (Fig. 5F) peduncle length 0.4 times width, mediodistal and laterodistal corners with 2 and 1 setae, respectively; exopod oval, length 1.9 times width, with transverse suture, bearing plumose setae marginally; endopod slightly shorter than exopod with plumose setae on distal margin. Pleopod 4 (Fig. 5G) pe- duncle small, length 0.5 times width, laterodistal corner with a single seta; exopod length 1.8 times width with transverse suture, bearing 4 plumose setae and simple setae marginally; endopod oval, length 1.4 times width, Zoosyst. Evol. 99 (2) 2023, 545-556 551 A 0.2mm:A, C, E, G, H Figure 4. Gnorimosphaeroma rivulare sp. nov., holotype, male 4.9 mm (NSMT-Cr 31485). A. Right pereopod 1, medial view; B. Merus, carpus, propodus and dactylus of right pereopod 1, medial view; C. Right pereopod 2, medial view; D. Carpus, propodus and dactylus of right pereopod 2, medial view; E. Right pereopod 3, medial view; F. Merus, carpus, propodus and dactylus of right pereopod 3, medial view; G. Right pereopod 4, medial view; H. Right pereopod 5, medial view. zse.pensoft.net 552 0.05 mm: | 0.3 mm: D-H ‘s JEL ee a fA! y xy Tomikawa, K. et al.: New Gnorimosphaeroma from Ogasawara Islands ~ tent ta x nee oes a eT. SS EKO CEE Na: STeTe SC G. sp. Chiba, Japan 9 54/— G. rayi @ G. tondaense & 39/- G. iriei © g5/1.9 | G: Mvulare sp. nov. r 69/0.99 G. rivulare sp. nov. @ G. rivulare sp. nov. @ 63/0.91 . G. hokurikuense @ 94/10 |r G- akanense @ 8910.84! G saijoense Ancinus sp. Figure 8. Maximum likelihood tree for 1077 bp of 16S rRNA and 18S rRNA markers. Numbers on nodes represent ultra-bootstrap val- ues for maximum likelihood and Bayesian posterior probabilities. Symbols beside the species name indicate the habitats shown in Fig. 1. zse.pensoft.net Zoosyst. Evol. 99 (2) 2023, 545-556 lotic waters. A stagnant bottom environment may not be favourable for G. rivulare sp. nov., and could severely limit its future persistence. For this newly described spe- cies survive, conservation of this restricted habitat is an urgent issue that needs to be addressed. Author contributions KT, JY, AN, TS, NK, and NN collected the samples used in this study. KT, JY, AN, and CL performed molecular phylogenetic analyses. KT, JY, and NN were responsible for describing and naming the new species. This manu- script was compiled by KT and carefully reviewed and finalized by all authors. Acknowledgements We thank Drs Regina Wetzer of Natural History Mu- seum of Los Angeles County and Brenda L. Doti of Universidad de Buenos Aires for their critical reading and valuable comments on our manuscript. This work was partly supported by the Japan Society for the Pro- motion of Science KAKENHI grants JP21H00910, JP22HO1011, JP22K06373, and JP22K06371 to KT. This work was also supported by grants from the Na- kdonggang National Institute of Biological Resourc- es (NNIBR) funded by the Ministry of Environment (MOE) of the Republic of Korea (NNIBR202302102) to CL. We would like to thank Editage (www.editage. com) for English language editing. References Boyko CB, Bruce NL, Hadfield KA, Merrin KL, Ota Y, Poore GCB, Taiti S (2023) World Marine, Freshwater and Terrestrial Isopod Crustaceans database. https://doi.org/10.14284/365 Bruce NL (2005) New sphaeromatids (Crustacea: Isopoda: Sphaeroma- tidea) from coastal and freshwater habitats in New Zealand. Zootaxa 1002(1): 1-20. https://doi.org/10.11646/zootaxa.1002.1.1 Hoang DT, Chernomor O, von Haeseler A, Minh BQ, Vinh LS (2018) UFBoot2: Improving the Ultrafast Bootstrap Approximation. Molec- ular Biology and Evolution 35(2): 518—522. https://doi.org/10.1093/ molbev/msx28 1 Hoestlandt H (1969) Sur un sphérome nouveau de la céte pacifique américaine, Gnorimosphaeroma rayi n. sp. (isopode flabellifére). Comptes rendus hebdomadaires des séances de |’ Académie des sci- ences, série D 268: 325-327. Ito T, Sasaki T, Takahashi C, Sugawara H, Hayashi F (2023) The fam- ily Hydroptilidae Curtis (Trichoptera) in the Ogasawara Islands, northwestern Pacific, with particular reference to adaptive radia- tion in the oceanic islands. Zootaxa 5231(2): 141-164. https://doi. org/10.11646/zootaxa.5231.2.2 JangIK, Kwon DH(1993)Anewspecies of the genus Gnorimosphaeroma (Crustacea, Isopoda, Sphaeromatidae) from a brackish-water lake in Korea. Tongmul Hakhoe Chi 36: 402-407. 595 Jaume D, Queinnec E (2007) A new species of freshwater isopod (Sphaeromatidea: Sphaeromatidae) from an inland karstic stream on Espiritu Santo Island, Vanuatu, southwestern Pacific. Zootaxa 1653(1): 41-55. https://doi.org/10.11646/zootaxa. 1653.1.3 Kalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS (2017) ModelFinder: Fast model selection for accurate phylogenetic estimates. Nature Methods 14(6): 587-589. https://doi.org/10.1038/ nmeth.4285 Kwon DH, Kim HS (1987) A new species of the genus Gnorimo- sphaeroma (Crustacea, Isopoda, Sphaeromatidae) from the Naktong River, with a key to the Korean species of the genus. Korean Journal of Systematic Zoology 3: 51-56. Latreille PA (1825) Familles naturelles du réegne animal, exposées succinctement et dans un ordre analytique, avec l’indication de leurs genres. Jean-Baptiste Baillicre, Paris, 570 pp. https://doi. org/10.5962/bh1 title. 16094 Lévéque C, Oberdorff T, Paugy D, Stiassny MLJ, Tedesco PA (2008) Global diversity of fish (Pisces) in freshwater. Hydrobiologia 595(1): 545-567. https://doi.org/10.1007/s10750-007-9034-0 Menzies RJ (1954) A review of the systematics and ecology of the genus “Exosphaeroma” with the description of a new genus, a new species, and a new sub-species (Crustacea, Isopoda, Sphaeromidae). Ameri- can Museum Novitates 1683: 1-24. Miura M, Mori H, Nakai S, Satake K, Sasaki T, Chiba S (2008) Mole- cular evidence of the evolutionary origin of a Bonin Islands endemic Stenomelania boninensis. The Journal of Molluscan Studies 74(2): 199-202. https://doi.org/10.1093/mollus/eyn003 Nation JL (1983) A new method using hexamethyldisilazane for preparation of soft insect tissues for scanning electron mi- croscopy. Stain Technology 58(6): 347-351. _ https://doi. org/10.3109/105202983090668 11 Nishimura S (1968) Gnorimosphaeroma lata n. sp., a new marine iso- pod from Kii, Japan. Publications of the Seto Marine Biological Laboratory 16(4): 273-280. https://doi.org/10.5134/175549 Nishimura S (1969) Gnorimosphaeroma salebrosa sp. nov. from the coast of Kii, Japan (Isopoda: Sphaeromatidae). Publications of the Seto Marine Biological Laboratory 16(6): 385-393. https://doi. org/10.5134/175563 Nunomura N (1998) On the genus Gnorimosphaeroma (Crustacea, Iso- poda, Sphaeromatidae) in Japan with descriptions of six new spe- cies. Bulletin of the Toyama Science Museum 21: 23-54. Nunomura N (1999) A new Species of the Genus Gnorimosphaeroma (Iso- poda, Sphaeromatidae) from the Mouth of Tonda River Kii Pemnsula, Southern Japan. Bulletin of the Toyama Science Museum 22: 1-5. Nunomura N (2013) Isopod crustaceans from Shikoku, western Ja- pan-1, specimens from Ehime Prefecture. Bulletin of the Toyama Science Museum 446: 19-78. Nunomura N, Satake K (2006) A new species of the genus Gnorimo- sphaeroma (Crustacea: Isopoda) from Hahajima, Bonin islands, southern Japan. Bulletin of the Toyama Science Museum 29: 1-6. Palumbi S, Martin A, Romano S, McMillan WO, Stice L, Grabowski G (1991) The simple fool’s guide to PCR. Version2.0. Honolulu, HL: Department of Zoology and Kewalo Marine Laboratory, University of Hawaii. Rambaut A, Drummond AJ, Xie D, Baele G, Suchard MA (2018) Posterior summarization in bayesian phylogenetics using Tracer 1.7. Systematic Biology 67(5): 901-904. https://doi.org/10.1093/ sysbio/syy032 zse.pensoft.net 556 Ronquist F, Teslenko M, van der Mark P, Ayres DL, Darling A, Hohna S, Larget B, Liu L, Suchard MA, Huelsenbeck JP (2012) MRBAYES 3.2: Efficient Bayesian phylogenetic inference and model selection across a large model space. Systematic Biology 61(3): 539-542. https://doi.org/10.1093/sysbio/sys029 Satake K, Cai Y (2005) Paratya boninensis, a new species of freshwa- ter shrimp (Crustacea: Decapoda: Atyidae) from Ogasawara, Ja- pan. Proceedings of the Biological Society of Washington 118(2): 306-311. https://doi.org/10.2988/0006-324X(2005)118[306:PBAN- SO]2.0.CO;2 Satake K, Ueno R (2013) Distribution of freshwater macroinverte- brates in streams with dams and associated reservoirs on a subtrop- ical oceanic island off southern Japan. Limnology 14(2): 211-221. https://doi.org/10.1007/s10201-012-0393-5 Satake K, Ueno R, Sasaki T (2019) Freshwater and brackish-water mac- roinvertebrates in the Ogasawara Islands. Global Environmental Re- search 23: 73-75. Sket B, Bruce NL (2004) Sphaeromatids (Isopoda, Sphaeromatidae) from New Zealand fresh and hypogean waters, with description of Bilistra n. gen. and three new species. Crustaceana 76: 1347-1370. https://doi.org/10.1163/156854003323009858 Strong EE, Gargominy O, Ponder WF, Bouchet P (2008) Global di- versity of gastropods (Gastropoda; Mollusca) in freshwater. Hyd- robiologia 595(1): 149-166. https://do1.org/10.1007/s10750-007- 9012-6 Tamura K, Stecher G, Kumar S (2021) MEGA11: Molecular Evolution- ary Genetics Analysis version 11. Molecular Biology and Evolution 38(7): 3022-3027. https://doi.org/10.1093/molbev/msab120 Tattersall WM (1921) Mysidacea, Tanaidacea, and Isopoda. Pt. 7. In: Zoological results of a tour in the Far East. Memoires of the Asiatic Society of Bengal 6: 403-443. Tomikawa K (2015) A new species of Jesogammarus from the Iki Island, Japan (Crustacea, Amphipoda, Anisogammaridae). ZooKeys 530: 15-36. https://doi.org/10.3897/zookeys.530.6063 zse.pensoft.net Tomikawa, K. et al.: New Gnorimosphaeroma from Ogasawara Islands Tomikawa K, Kobayashi N, Kyono M, Ishimaru S-i, Grygier MJ (2014) Description of a new species of Sternomoera (Crustacea: Amphipoda: Pontogeneiidae) from Japan, with an analysis of the phylogenetic relationships among the Japanese species based on the 28S rRNA gene. Zoological Science 31(7): 475-490. https://doi. org/10.2108/zs140026 Tomikawa K, Sasaki T, Aoyagi M, Nakano T (2022) Taxonomy and phy- logeny of the genus Melita (Crustacea: Amphipoda: Melitidae) from the West Pacific Islands, with descriptions of four new species. Zoologischer Anzeiger 296: 141-160. https://doi.org/10.1016/j.jcz.2021.12.005 Trifinopoulos J, Nguyen LT, von Haeseler A, Minh BQ (2016) W-IQ- TREE: A fast online phylogenetic tool for maximum likelihood analysis. Nucleic Acids Research 44(W 1): W232—W235. https://doi. org/10.1093/nar/gkw256 Vainola R, Witt JDS, Grabowski M, Bradbury JH, Jazdzewski K, Sket B (2008) Global diversity of amphipods (Amphipoda; Crustacea) in freshwater. Hydrobiologia 595(1): 241-255. https://doi.org/10.1007/ $10750-007-9020-6 Wetzer R, Pérez-Losada M, Bruce NL (2013) Phylogenetic relationships of the family Sphaeromatidae Latreille, 1825 (Crustacea: Peracarida: Isopoda) within Sphaeromatidea based on 18S-rDNA molecular data. Zootaxa 3599(2): 161-177. https://do1.org/10.11646/zootaxa.3599.2.3 Wetzer R, Bruce NL, Pérez-Losada M (2018) Relationships of the Sphaeromatidae genera (Peracarida: Isopoda) inferred from 18S rDNA and 16S rDNA genes. Arthropod Systematics & Phylogeny 76(1): 1-30. https://doi.org/10.3897/asp.76.e3 1934 Wetzer R, Wall A, Bruce NL (2021) Redescription of Gnorimosphaeroma oregonense (Dana, 1853) (Crustacea, Isopoda, Sphaeromatidae), designation of neotype, and 16S-rDNA molecular phylogeny of the north-eastern Pacific species. ZooKeys 1037: 23-56. https://doi. org/10.3897/zookeys. 1037.63017 Wilson GDF (2008) Global diversity of Isopod crustaceans (Crustacea; Isopoda) in freshwater. Hydrobiologia 595(1): 231—240. https://doi. org/10.1007/s10750-007-9019-z