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A single specimen of Arothron reticularis (398 mm in total length), belonging to the family Tetraodontidae, was collected in the coastal waters off Hansan island using a set net in May 2019. The morphological characteristics of the specimen are as follows: large eyes and mouth, white spots on the body, and a white line surrounding the eyes in an annular shape. A result of maximum likelihood tree showed that A. reticularis is related to A. hispidus (93.6%). We proposed the Korean name “Geu-murl-mu-nui-kkeo-kkeurl-bok,” in accordance with the characteristics of the specimen. Keywords: Tetraodontidae, New species, Arothron, Arothron reticularis, Reticulated toadfish Background southern Red Sea, as Tetraodon reticularis (Randall The family Tetraodontidae (Order Tetraodontiformes) et al. 2012). consists of 2 subfamilies, 26 genera, and 196 species The genus Arothron is represented along the Korean worldwide (Nelson et al. 2016), of which 7 genera coast by four species (Chyung 1977; Lee 1993): A. firma- and 52 species occur in Japan (Nakabo and Yagishita mentum, Arothron stellatus (Anonymous 1798), Aro- 2013) and 5 genera and 31 species occur in Korea thron hispidus (Linnaeus 1758), and Arothron (MABIK 2020). Of the 196 species, the genus Aro- nigropunctatus (Bloch and Schneider, 1801). For the first thron comprises 10 species that are present in Japan time, A. reticularis, including morphological traits and and 4 species in Korea (MABIK 2020). mitochondrial DNA cytochrome c oxidase subunit I (mt The genus Arothron is distributed in the tropical COI) sequences, was recorded based on single specimen regions of the Indo-Pacific, except for Arothron fir- collected from Korea. mamentum (Temminck and Schlegel 1850), which is antitropical (Matsuura 2016). The reticulated toad- Materials and methods fish, Arothron reticularis (Bloch and Schneider 1801), On May 9, 2019, a single reticulated pufferfish, A. reticu- is distributed in the tropical waters of Okinawa, laris, was captured from the coastal waters off Hansan Taiwan, and the Indo-West Pacific (Masuda et al. Island using a set net (Fig. 1). The methods used for the 1984). Thefirst report from theRed Seawas ob- fin ray counts and morphometric characters follow those tained in 1956, when Roux-Estève identified a speci- of Han (1995). All morphometric measurements were men (197 mm in total length), which was from the measured to the nearest 0.1 cm using Vernier calipers. island of Abulat (Abu Latt) in the Farasan Islands, Soft x-ray microscopy (LISTEM, REX-525R, Korea) was used to count the number of vertebrae and fin rays. * Correspondence: email@example.com For genetic analysis, the total genomic DNA was ex- Department of Fisheries Science, Chonnam National University, Yeosu tracted from the prepared part of the tissue using the 59626, South Korea Full list of author information is available at the end of the article QIAamp DNA Micro Kit (Qiagen, Hilden, Germany), © The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. Kang et al. Fisheries and Aquatic Sciences (2020) 23:31 Page 2 of 5 Fig. 1 Map showing the capture site of Arothron reticularis in the coastal waters off Hansan Island following the manufacturer’s instructions. The polymer- maximum likelihood (ML) method based on the GTR+ ase chain reaction (PCR) amplification was performed G+I model and 10,000 bootstrap replications in the TM using the 2X TOPsimple PreMIX (aliquot) (Enzy- raxmlGUI 2.0 program (Edler et al. 2020). nomics, Daejeon, Korea) with the universal mitochon- drial COI primer set, LCO1490 forward primer (5′-GGT Results and discussion CAA ATC ATA AAG ATA TTG G-3′) and HCO2198 Family Tetraodontidae Bonaparte, 1831 reverse primer (5′-TAA ACT TCA GGG TGA CCA Genus Arothron Müller, 1841 AAA AAT CA-3′) (Folmer et al. 1994). The amplified Tetraodon Linnaeus, 1758: 333 (type species: Tetrao- PCR product was sequenced using the Applied Biosys- don lineatus Linnaeus) tems 3730xl DNA Analyzer (ThermoFisher, Foster City, Arothron reticularis (Bloch and Schneider 1801) CA, USA). The obtained mt COI sequence (658 bp) was (New Korean name: Geu-murl-mu-nui-kkeo-kkeurl- compared with 16 Tetraodontiformes mt COI sequences bok) (Fig. 2) (14 Tetraodontidae species and 2 Ostraciidae species [as Tetrodon reticularis Bloch & in Schneider, 1801: 506 an outgroup]) retrieved from GenBank (Appendix 1). In (type locality: Malabar coast, India) total, 17 sequences were aligned using MAFFT v.7 Arothron reticularis: Matsuura et al. 1984: 364 (Japan); (Katoh and Standley 2013) in Geneious v.9 (Biomatters Nakabo 2000: 1429 (Japan); Allen and Adrim, 2003: 65 Ltd.). The phylogenetic tree was reconstructed using the Fig. 2 Arothron reticularis (MABIK PI00049568). a Lateral view. b X-ray radiograph (scale bars indicate 10 mm) Kang et al. Fisheries and Aquatic Sciences (2020) 23:31 Page 3 of 5 (Indonesia); Veeruraj et al., 2011: 4 (India); Sonoyama Table 1 Comparison between the morphological traits of the genus Arothron in this study and previous studies et al., 2020: 130 (Japan). Morphological Present Sujatha and Han (1995) characteristics study Padmavathi (2015) Materials examined A. reticularis A. A. firmamentum stellatus Arothron reticularis (MABIK PI00049568) 330 mm standard length (SL) (398 mm total length [TL]) was col- Standard length 330 264–478 269–350 391–410 (SL, mm) lected in the coastal waters off Hansan Island, Total length as 121 118–125 –– Gyeongsangnam-do, Republic of Korea, using a set net %SL on May 9, 2019 (Fig. 3). % in body 32 34–41 36.0 ± 4.6 48.7 ± depth 2.3 Description % in head 39 33–36 29.5 ± 0.9 30.8 ± length 1.0 The morphological trait measurements for the present % in predorsal 80 73–81 69.9 ± 1.2 66.7 ± specimen are shown in Table 1. The measurements length 1.3 expressed as a percentage of the SL include the follow- %in 44 34––40 –– ing: body depth, 26.1; head length, 39.4; predorsal length, prepectoral 79.7; prepectoral length, 44.1; preanal length, 85.8; length length of dorsal fin, 17.0; length of pectoral fin, 12.7; % in preanal 86 72–82 75.0 ± 2.3 70.5 ± length 1.3 length of anal fin, 13.6; caudal peduncle depth, 14.5; pre- anus length, 80.2; dorso-caudal peduncle length, 18.2; % in length of 17 16–19 20.3 ± 1.9 15.4 ± dorsal fin 1.1 and ventro-caudal length, 17.3. The measurements % in length of 13 13–16 14.6 ± 1.1 11.5 ± expressed as a percentage of the head length include the pectoral fin 0.7 following: nasal length, 35; snout length, 38; eye diam- % in length of 14 15–17 –– eter, 27; and interorbital width, 54. The fin ray counts anal fin are as follows: dorsal fin rays, 10; pectoral fin rays, 10; % in caudal 15 21–27 12.2 ± 1.8 14.1 ± anal fin rays, 10; caudal fin rays, 11; and vertebrae, 18 peduncle 0.7 depth (10 + 8). The nostril has two fleshy flaps, formed by the bifur- % in preanus 80 –– – length cation of a single base (Fig. 4). The body is elongated % in dorso– 18 – 16.2 ± 0.7 20.5 ± and covered with prickles, except for the regions around caudal 0.1 the mouth and caudal peduncle. The eyes are notably peduncle large, broad, and flat. The mouth is large with four teeth, length % in ventro– 17 – 18.9 ± 1.2 21.5 ± caudal 1.1 peduncle length % in head length % in nasal 35 –– – length % in snout 38 38–44 –– length % in eye 27 30–36 –– diameter %in 54 51–62 –– interorbital width Counts Dorsal fin rays 10 10–11 14 11–12 Pectoral fin rays 18 17–18 13–14 11 Anal fin rays 10 10–11 15–16 18–19 Caudal fin rays 11 10–11 –– Vertebrae 18 – 20 18 Fig. 3 Frontal view of the fresh Arothron reticularis specimen Kang et al. Fisheries and Aquatic Sciences (2020) 23:31 Page 4 of 5 and a white line surrounds the eyes in an annular shape (Fig. 3) Distribution The distribution occurred in the Southern Sea of Korea (present study) and Indo-West Pacific Ocean, including the South China Sea and Ryukyu Islands (Veeruraj et al. 2011). Comparison with type species The difference between A. reticularis and the type spe- cies (Tetraodon lineatus) are as follows: T. lineatus has 2~3 dorsal spines, 9~10 dorsal soft rays, 2 anal spines, and 7~8 anal soft rays (Fishbase 2020). There was no Fig. 4 Olfactory organ of Arothron reticularis difference in the distribution of prickles; however, the white spots and white lines around the eyes were absent and the dentary is protruded further than the premaxil- in T. lineatus. lary. The lateral line is indistinct. The gill slit is large. Remarks Although the measurements were taken in a dry state, Coloration of the specimen the morphological traits of the present specimen are When fresh, there were white lines on the body that similar to those of Sujatha and Padmavathi (2015). There curved from around the eyes and then across the ab- were differences in morphometric traits between A. stel- domen. There were no ventral extensions of the dark latus and A. firmamentum (Table 1). body color into the pale ventral part of the head or The morphological classification keys of the genus body. White spots were present from the dorsal of Arothron fish are the spots and lines around the eyes, the head to the caudal fin. After preservation, the ventral part, and caudal fin. A. reticularis morphologic- ventral surface turned beige, and the body turned ally resembles the genus Arothron fish, but differs in the greenish brown. There are white spots on the body distribution of prickles (prickles absent on the snout and Fig. 5 Maximum likelihood analysis based on the mitochondrial cytochrome c oxidase subunit I (mt CO1) sequences of Tetraodontiformes species. Bootstrap replicates (BP) are indicated on the nodes (except for BP < 70) Kang et al. Fisheries and Aquatic Sciences (2020) 23:31 Page 5 of 5 at least the posterior half of the caudal peduncle in A. 59626, South Korea. Department of Animal Welfare, National Institute of Ecology, Seochen 33657, South Korea. hispidus), white line around the head (white spots around the head in A. hispidus) (Randall et al. 2012), Received: 22 July 2020 Accepted: 27 November 2020 white spots scattered around the caudal fin (caudal fin has no white spots of A. carduus and A. manilensis) References (Nakabo 2000), and discontinuous stripes on the body Chyung MK. The fishes of Korea. Seoul: Ilji Publishing Co; 1977. (continuous stripes on the body of A. multilineatus) Edler D, Klein J, Antonelli A, Silverstro D. raxmlGUI 2.0: a graphical interface and toolkit for phylogenetic analyses using RAxML. bioRxiv. 2020. https://doi.org/ (Matsuura 2016). This species has a reticulate pattern on 10.1111/2041-210X.13512. the body. In accordance with the characteristics, a new Fishbase. Sweden, Stockholm 2020. http://fishbase.org. Accessed 29 May 2020. Korean name was proposed for this species “Geu-murl- Folmer O, Black M, Goeh W, Lutz R, Vrijenhoek R. DNA primers for amplification of mitochondrial cytochromec oxidase subunit I from divers metazoan mu-nui-kkeo-kkeurl-bok.” invertebrates. Mol Mar Biol Biotechnol. 1994;3:294–9. In this study, a partial gene sequence of the mt COI Han KH. Morphology, osteology and phylogeny of the fishes of the family (658 bp) of A. reticularis was obtained for the first time. Tetraodontidae (Teleostei: Tetraodontiformes). Busan: PhD thesis: Pukyong National University, Department of Marine Biology; 1995. Using this sequence, we created an mt COI dataset, Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: including 16 Tetraodontiformes mt COI sequences from improvements in performance and usability. Mol Biol Evol. 2013;30:772–80. GenBank, and analyzed the genetic distance of A. reticu- Lee WO. New records of six species of the Tetraodontidae (Pisces: Tetraodontiforems) from Korea. Korean J Ichthyol. 1993;5:165–76. laris using Kimura-2-Parameter method. Neighbor- MABIK. National list of marine species. Seocheon: Namu Press; 2020. joining tree showed that A. reticularis belongs to the Masuda H, Amaoka K, Araga C, Uyeno T, Yoshino T. The fishes of the Japanese genus Arothron (family Tetraodontidae) and is related to archipelago 2nd ed. Tokyo: Tokai University Press; 1984. Matsuura K. A new pufferfish, Arothron multilineatus (Actinopterygii: A. hispidus (similarity 93.6%) (Fig. 5). Tetraodontiformes: Tetraodontidae), from the Indo-West Pacific. Ichthyol Res. 2016. https://doi.org/10.1007/s10228-016-0517-8. Nakabo T. Fishes of Japan with pictorial keys to the species. 2nd ed. Tokyo: Tokai Conclusions University Press; 2000. One specimen of reticulated toadfish was collected in Nakabo T, Yagishita N. Tetraodontidae: fishes of Japan with pictorial keys to the the coastal waters off Hansan Island on May 9, 2019. species. 3rd ed. Tokyo: Tokai University Press; 2013. Nelson JS, Grande TC, Wilson MVH. Fishes of the world. 5th ed. New York: John The specimen was identified as Arothron reticularis, Wiley & Sons; 2016. based on morphological traits and the first record of Randall JE, Bogorodsky SV, Rose JM. Color variation of the puffer Arothron species in Korea. hispidus (Linnaeus) and comparison with A. reticularis (Bloch & Schneider). Aqua International Journal of Ichthyology. 2012;18:41–54. Sujatha K, Padmavathi P. Taxonomy of puffer fish (Pisces: Tetraodontidae) Supplementary Information represented in the catches of Visakhapatnam, central eastern coast of India. The online version contains supplementary material available at https://doi. J Mar Biol Ass India. 2015. https://doi.org/10.6024/jmbai.2015.57.2.1798-14. org/10.1186/s41240-020-00176-5. Veeruraj A, Arumugam M, Ajithkumar T, Balasubramanian T. Distribution of Tetraodontiformes (Family: Tetraodontidae) along the Parangipettai Coast, Southeast coast of India. Zootaxa. 2011. https://doi.org/10.11646/zootaxa. Additional file 1: Appendix 1. List of 17 species used in the 3015.1.1. phylogenetic analysis of this study. Publisher’sNote Acknowledgements Springer Nature remains neutral with regard to jurisdictional claims in This work was supported by the Efficient Securement of Marine Bioresources published maps and institutional affiliations. and Taxonomic Research (2020 M00100), funded by the National Marine Biodiversity Institute of Korea (MABIK). We would like to thank the head of Bongam Village, Hansan Island, for providing the sample. Authors’ contributions CBK and TSY wrote the manuscript. SHL and HRL carried out the molecular genetic experiments. KHH designed the study and finalized the manuscript. All authors read and approved the final manuscript. Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare that they have no competing interests. Author details Department of Exhibition and Education, Marine Biodiversity Institute of Korea, Seochen 33662, South Korea. Department of Taxonomy and Systematics, Marine Biodiversity Institute of Korea, Seochen 33662, South Korea. Department of Fisheries Science, Chonnam National University, Yeosu
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Published: Dec 21, 2020
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