Seven ascidian (Tunicata) species from the North Eastern Mediterranean


Published: Oct 4, 2023
Keywords:
Botryllid ascidians Didemnum Polyclinum Symplegma COI species delimitation
ARZU KARAHAN
BERIVAN TEMIZ
ESRA ÖZTÜRK
JACOB DOUEK
BARUCH RINKEVICH
Abstract

Members of the tunicates, a subphylum of marine filter-feeder chordates, inhabit all marine and oceanic habitats from the subtidal to the abyssal. Considered the closest relatives to the vertebrates, the tunicates are widely used as model organisms for evo-devo, allorecognition, senescence, and whole-body regeneration studies. However, species boundaries are poorly understood due to the high morphological and genetic plasticity that characterizes many tunicate taxa. Here, we present findings on seven tunicate species (Botrylloides israeliense, Botrylloides sp., Botryllus humilis, Botryllus schlosseri, Symplegma brakenhielmi, Polyclinum constellatum and Didemnum perlucidum) sampled from six Turkish sites at the North Eastern Mediterranean Sea and employed the mitochondrial barcoding marker (COI) for evaluating the relationships among geographically restricted and widely spread ascidian species. Species delimitation was conducted using sequences generated in the current study in addition to sequences obtained from GenBank. General morphological features and colors of colonies were recorded at sampling sites. Then, all Styelidae colonies were attached and cultured on slides in an aquaculture room, enabling the study of other features, such as zooid distributions and sizes, oral tentacle  numbers, and life cycles, using stereo and light microscopes. The spicules of formalin-fixed Didemnum perlucidum samples were examined under a light microscope. Then, scientific names were assigned to all species based on the results of the species delimitation and on comparisons of the obtained COI sequences with GenBank sequences. A putative new Botrylloides species (Botrylloides sp.) from the Antalya region was revealed, with a 99% match with the COI gene of another specimen from Saudi Arabia; further waiting for detailed traditional taxonomy.

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References
Aydın-Önen, S.A., 2018. Distribution of ascidians with a new record of the non-indigenous species Polyclinum constellatum savigny, 1816 from the Aegean coast of Turkey. Turkish Journal of Fisheries and Aquatic Sciences, 18 (9), 1077-1089.
Ballarin, L., Karahan, A., Salvetti, A., Rossi, L., Manni, L. et al., 2021. Stem Cells and Innate Immunity in Aquatic Invertebrates: Bridging Two Seemingly Disparate Disciplines for New Discoveries in Biology. Frontiers in Immunology, 12 (688106).
Ben-Hamo, O., Rosner, A., Rabinowitz, C., Oren, M., Rinkevich, B., 2018. Coupling astogenic aging in the colonial tunicate Botryllus schlosseri with the stress protein mortalin. Developmental Biology, 433 (1), 33-46.
Ben-Hamo, O., Rinkevich, B., 2021. Botryllus schlosseri – a model colonial species in basic and applied studies. p. 385- 402. In: Handbook of Marine Model Organisms in Experimental Biology.
Boutet, A., Schierwater, B. (eds.). Established and Emerging. CRC Press. Ben-Hamo, O., Izhaki, I., Ben-Shlomo, R., Rinkevich, B., 2023. The novel Orshina Rhythm in a colonial urochordate signifies the display of recurrent aging/rejuvenation sequels. Scientific Reports, 13 (1), 9788.
Berná, L., Alvarez-Valin, F., 2014. Evolutionary genomics of fast evolving tunicates. Genome Biology and Evolution, 6 (7), 1724-38.
Bitar, G., Kouli-Bitar, S., 2001. Nouvelles données sur la faune et la flore benthiques de la cote Libanaise. Migration Lessepsienne [New data on benthic fauna and flora on the coast Lebanese. Lessepsian migration]. Thalassia Salentina, 25, 71-74.
Bock, D.G., MacIsaac, H.J., Cristescu, M.E., 2012. Multilocus genetic analyses differentiate between widespread and spatially restricted cryptic species in a model ascidian. Proceedings of the Royal Society B: Biological Sciences, 279 (1737), 2377-2385.
Brunetti, R., 2009. Botryllid species (Tunicata, Ascidiacea) from the Mediterranean coast of Israel, with some considerations on the systematics of Botryllinae. Zootaxa, 2289 (1).
Brunetti, R., Manni, L., Mastrototaro, F., Gissi, C., Gasparini, F., 2017. Fixation, description and DNA barcode of a neotype for Botryllus schlosseri (Pallas, 1766) (Tunicata, Ascidiacea). Zootaxa, 4353 (1), 29-50.
Çınar, M.E., Bilecenoglu, M., Öztürk, B., Can, A., 2006. New records of alien species on the Levantine coast of Turkey (June 2006). Aquatic Invasions, 1 (2), 84-90.
Dias, G.M., Rocha, R.M., Lotufo, T.M.C., Kremer, L.P., 2013. Fifty years of ascidian biodiversity research in São Sebastião, Brazil. Journal of the Marine Biological Association of the United Kingdom, 93 (1), 273-282.
Dias, J., Rocha, R., Godwin, S., Tovar Hernández, M.A., Delahoz, M. et al., 2016. Investigating the cryptogenic status of the sea squirt Didemnum perlucidum (Tunicata, Ascidiacea) in Australia based on a molecular study of its global distribution. Aquatic Invasions, 11 (3), 239-245.
Ferrier, D.E., 2011. Tunicates push the limits of animal evo-devo. BMC Biology, 9 (3).
Galil, B.S., Marchini, A., Occhipinti-Ambrogi, A., 2018. East is east and West is west? Management of marine bioinvasions in the Mediterranean Sea. Estuarine, Coastal and Shelf Science, 201, 7-16.
Griggio, F., Voskoboynik, A., Iannelli, F., Justy, F., Tilak, M.K. et al., 2014. Ascidian Mitogenomics: Comparison of Evolutionary Rates in Closely Related Taxa Provides Evidence of Ongoing Speciation Events. Genome Biology and Evolution, 6 (3), 591-605.
Halim, Y., Messeih, A. M., 2016. Aliens in Egyptian waters. A checklist of ascidians of the Suez Canal and the adjacent Mediterranean waters (December 2016). Egyptian Journal of Aquatic Research, 42 (4), 449-457.
Hebert, P.D., Cywinska, A., Ball, S.L., deWaard, J.R., 2003. Biological identifications through DNA barcodes. Proceedings of Royal Society B: Biological Science, 270, 313-321.
Herdman, W.A., 1891. A revised classification of the Tunicata, with definition of the orders, sub-orders, families, subfamilies and genera, and analytical keys to the species. Zoological Journal of the Linnean Society, 23 (148), 558-652.
Holland, L.Z., 2016. Tunicates. Current Biology, 26 (4), R146-52.
Hyams, Y., Paz, G., Rabinowitz, C., Rinkevich B., 2017. Insights into the unique torpor of Botrylloides leachi, a colonial urochordate. Developmental Biology, 428 (1), 101-117.
Hyams, Y., Panov, Y., Rosner, A., Brodski, L., Rinkevich, Y. et al., 2022. Transcriptome landscapes that signify Botrylloides leachi (Ascidiacea) torpor states. Developmental Biology, 490, 22-36.
Jukes, T.H., Cantor, C.R., 1969. Evolution of protein molecules. Academic Press, New York and London, 412 pp.
Karahan, A., Öztürk, E., Temiz, B., Blanchoud, S., 2022. Studying tunicata WBR using Botrylloides anceps. p. 311- 332. In: Whole-Body Regeneration. Methods in Molecular Biology. Vol. 2450. Blanchoud, S., Galliot, B. (Eds). Humana Press, New York.
Katoh, K., Rozewicki, J., Yamada, K.D., 2018. MAFFT online service: multiple sequence alignment, interactive sequence choice and visualization. Briefings in Bioinformatics, 20, 1160-1166.
Kayış, Ş., 2011. Ascidian Tunicate, Botryllus schlosseri (Pallas, 1766) infestation on seahorse. Bulletin of the European Association of Fish Pathologists, 21 (2), 81-84.
Kimura, M.A., 1980. Simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution, 16, 111-120.
Lee, T., Shin, S., 2021. First Record of Colonial Ascidian, Botrylloides diegensis Ritter and Forsyth, 1917 (Ascidiacea, Stolidobranchia, Styelidae), in South Korea. Water, 13, 2164.
Lefort, V., Longueville J.E., Gascuel, O., 2017. SMS: Smart Model Selection in PhyML. Molecular Biology and Evolution, 34 (9), 2422-2424.
López-Legentil, S., Legentil, M.L., Erwin, P.M., Turon, X., 2015. Harbor networks as introduction gateways: contrasting distribution patterns of native and introduced ascidians. Biological Invasions, 17 (6), 1623-1638.
Mastrototaro, F., Montesanto, F., Salonna, M., Grieco, F., Trainito, E. et al., 2019. Hitch-hikers of the sea: Concurrent morphological and molecular identification of Symplegma brakenhielmi (Tunicata: Ascidiacea) in the western Mediterranean Sea. Mediterranean Marine Science, 20 (1), 197-207.
Michaelsen, W., 1904. Revision der compositen Styeliden oder Polyzoinen. p.1-124. In: Mitteilungen aus dem Naturhistorischen Museum. Vol. 21. Kommissionsverlag von Lucas Gräfe & Sillem, Hamburg.
Monniot, C., 1988. Ascidies de Nouvelle-Calédonie. IV: Styelidae. Bulletin du Muséum national d’histoire naturelle. Section A, Zoologie, biologie et écologie animales, 10 (2), 163-196.
Monniot, F., 1983. Ascidies littorales de Guadeloupe I. Didemnidae. Bulletin du Museum National d’Histoire Naturelle, 4a (5), 5-49.
Montesanto, F., Chimienti, G., Gissi, C., Mastrototaro, F., 2022. Polyclinum constellatum (Tunicata, Ascidiacea), an emerging non-indigenous species of the Mediterranean Sea: integrated taxonomy and the importance of reliable DNA barcode data. Mediterranean Marine Science, 23 (1), 69-83.
Mueller, W.A., Rinkevich, B., 2020. Cell communication-mediated nonself-recognition and intolerance in representative species of the animal kingdom. Journal of Molecular Evolution, 88 (6), 482-500.
Nydam, M.L., Lemmon, A.R., Cherry, J.R., Kortyna, M.L., Clancy, D.L. et al., 2021. Phylogenomic and morphological relationships among the botryllid ascidians (Subphylum Tunicata, Class Ascidiacea, Family Styelidae). Scientific Reports, 11 (1), 8351.
Pallas, P.S., 1766. Elenchus zoophytorum sistens generum adumbrationes generaliores et specierum cognitarum succinctas descriptiones cum selectis auctorum synonymis. Apud Petrum van Cleef, The Hague, 451 pp.
Palomino-Alvarez, L.A., Nydam, M.L., Rocha, R.M., Simões, N., 2022. New Botrylloides, Botryllus, and Symplegma (Ascidiacea: Styelidae) in Coral Reefs of the Southern Gulf of Mexico and Mexican Caribbean Sea. Diversity, 14 (11), 977.
Puillandre, N., Brouillet, S., Achaz, G., 2021. ASAP: assemble species by automatic partitioning. Molecular Ecology Resources, 21 (2), 609-620.
Rambaut, A., 2018. FigTree: tree figure drawing tool version 1.4.4. Institute of Evolutionary Biology, University of Edinburg.
Ratnasingham, S., Hebert, P.D.N., 2007. BOLD: The barcode of life data system. Molecular Ecology Notes, 7 (3), 355-364.
Reem, E., Douek, J., Rinkevich, B., 2017a. Ambiguities in the taxonomic assignment and species delineation of botryllid ascidians from the Israeli Mediterranean and other coastlines. Mitochondrial DNA Part A: DNA Mapping, Sequencing, and Analysis, 29 (7), 1073-1080.
Reem, E., Douek, J., Paz, G., Katzir, G., Rinkevich, B., 2017b. Phylogenetics, biogeography and population genetics of the ascidian Botryllus schlosseri in the Mediterranean Sea and beyond. Molecular Phylogenetics and Evolution, 107, 221-231.
Reem, E., Douek, J., Rinkevich, B., 2022. A critical deliberation of the “species complex” status of the globally-spread colonial ascidian Botryllus schlosseri. Journal of the Marine Biological Association of the United Kingdom, 101 (7), 1047-1060.
Rinkevich, B., Shlemberg, Z., Lilker-levav, T., Goren, M., Fishelson, L., 1993. Life history characteristics of Botrylloides (Tunicata) populations in Akko Bay, Mediterranean Coast of Israel. Israel Journal of Zoology, 39 (3), 197-212.
Rinkevich, B., Lilker-Levav, T., Goren, M., 1994. Allorecognition/ xenorecognition responses in Botrylloides (Ascidiacea) subpopulations from the Mediterranean coast of Israel. Journal of Experimental Zoology, 270 (3), 302-313.
Rinkevich, B., Ballarin, L., Martinez, P., Somorjai, I., Ben-Hamo, O. et al., 2022. A pan-metazoan concept for adult stem cells: the wobbling Penrose landscape. Biological Reviews, Cambridge Philosophical Society, 97 (1), 299-325.
Rocha, R.M., da Zanata, T.B., Moreno, T.R., 2012. Keys for the identification of families and genera of Atlantic shallow water ascidians. Biota Neotropica, 12 (1).
Rocha, R.M., Salonna, M., Griggio, F., Ekins, M., Lambert, G. et al., 2019. The power of combined molecular and morphological analyses for the genus Botrylloides: identification of a potentially global invasive ascidian and description of a new species. Systematics and Biodiversity, 17 (5), 509-526.
Ronquist, F., Teslenko, M., van der Mark, P., Ayres, D.L., Darling, A. et al., 2012. Mrbayes 3.2: Efficient bayesian phylogenetic inference and model choice across a large model space. Systematic Biology, 61 (3), 539-542.
Rosner, A., Kravchenko, O., Rinkevich, B., 2019. IAP genes partake weighty roles in the astogeny and whole-body regeneration in the colonial urochordate Botryllus schlosseri. Developmental Biology, 448 (2), 320-341.
Rubinstein, N.D., Feldstein, T., Shenkar, N., Botero-Castro, F., Griggio, F. et al., 2013. Deep sequencing of mixed total DNA without barcodes allows efficient assembly of highly plastic ascidian mitochondrial genomes. Genome Biology and Evolution, 5 (6), 1185-1199.
Saito, Y., Okuyama, M., 2003. Studies on Japanese botryllid ascidians. IV. A new species of the genus Botryllus with a unique colony shape, from the vicinity of Shimoda. Zoological Science, 20 (9), 1153-1161.
Savigny, J.C., 1816. Mémoires Sur Les Animaux Sans Vertèbres. Paris, 240 pp.
Shenkar, N., Gittenberger, A., Lambert, G., Rius, M., da Rocha, M.R. et al., 2023. Ascidiacea World Database. https://www. marinespecies.org/ascidiacea (Accessed 17 June 2023).
Tamura, K., Stecher, G., Kumar, S., 2021. MEGA11: Molecular evolutionary genetics analysis version 11. Molecular Biology and Evolution, 38 (7), 3022-3027.
Temiz, B., Öztürk, E., Blanchoud, S., Karahan, A., 2023. Phylogeographic and morphological analysis of Botrylloides niger Herdman, 1886 from the northeastern Mediterranean Sea. Diversity, 15 (3), 367.
Ulman, A., Ferrario, J., Occhpinti-Ambrogi, A., Arvanitidis, C., Bandi, A. et al., 2017. A massive update of non-indigenous species records in Mediterranean marinas. PeerJ, 5, e3954.
Viard, F., Roby, C., Turon, X., Bouchemousse, S., Bishop, J., 2019. Cryptic diversity and database errors challenge non-indigenous species surveys: An illustration with Botrylloides spp. in the English Channel and Mediterranean Sea. Frontiers in Marine Science, 6, 615.
Waterhouse, A.M., Procter, J.B., Martin, D.M.A., Clamp, M., Barton, G.J., 2009. Jalview Version 2-A multiple sequence alignment editor and analysis workbench. Bioinformatics, 25 (9), 1189-1191.
Williams, G.C., van Syoc, R., 2007. Methods of Preservation and Anesthetization of Marine Invertebrates. p. 37-42. In: The Light and Smith Manual: Intertidal Invertebrates from Central California to Oregon, 4th Ed. Carlton, J.T. (Ed). University of California Press, Berkeley and Los Angeles, CA.
Zhang, J., Kapli, P., Pavlidis, P., Stamatakis, A., 2013. A general species delimitation method with applications to phylogenetic placements. Bioinformatics, 29 (22), 2869-2876.
Zenetos, A., Çinar, M.E., Crocetta, F., Golani, D., Rosso, A. et al., 2017. Uncertainties and validation of alien species catalogues: The Mediterranean as an example. Estuarine, Coastal and Shelf Science, 191, 171-187.
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