Knowl. Manag. Aquat. Ecosyst. 2020, 421, 40 Knowledge & © M.O. Son et al., Published by EDP Sciences 2020 Management of Aquatic https://doi.org/10.1051/kmae/2020032 Ecosystems Journal fully supported by Office www.kmae-journal.org français de la biodiversité RESEARCH PAPER Occurrence of two exotic decapods, Macrobrachium nipponense (de Haan, 1849) and Procambarus virginalis Lyko, 2017, in Ukrainian waters 1 2 3 2 Mikhail O. Son , Halyna Morhun , Roman O. Novitskyi , Serhii Sidorovskyi , 2 2,* Maryna Kulyk and Serge Utevsky 1 Institute of Marine Biology of the National Academy of Sciences of Ukraine, Department of water quality, 37 Pushkinska str., 65148 Odesa, Ukraine 2 V.N. Karazin Kharkiv National University, Department of Zoology and Animal Ecology, 4 Maidan Svobody, 61022 Kharkiv, Ukraine 3 Dnipro State Agrarian and Economic University, Department of Water Bioresources and Aquaculture, 25 Serhii Yefremov str., 49600 Dnipro, Ukraine Received: 9 July 2020 / Accepted: 29 September 2020 Abstract – New information on the occurrence of two alien decapod crustaceans, Macrobrachium nipponense and Procambarus virginalis, was summarized for Ukraine. The invasive potential of both species was estimated in the context of local environmental and socio-economic issues. Their history as non- native species is marked by the early use of alien decapods as a natural resource by local residents. Both intentional and unintentional introductions occurred for M. nipponense, while the ornamental trade was an important source of non-native P. virginalis. New records of M. nipponense and P. virginalis from open freshwater habitats in the Odesa and Kharkiv provinces suggest continuing expansion of these decapods. We conducted a molecular phylogenetic analysis of COI sequences, which confirmed morphological identifications of invasive M. nipponense. This reinforces the presumed pathway of this species, including its pathway from China to south-western Ukraine. Procambarus fallax was found to be paraphyletic in relation to P. virginalis, which was probably caused by analyzing only one mitochondrial gene, which could be impacted by the recent emergence of the latter species. Keywords: Decapoda / Macrobrachium nipponense / Procambarus virginalis / Ukraine / invasive species Résumé – Présence de deux décapodes exotiques, Macrobrachium nipponense (de Haan, 1849) et Procambarus virginalis Lyko, 2017, dans les eaux ukrainiennes. De nouvelles informations sur la présence de deux crustacés décapodes exotiques, Macrobrachium nipponense et Procambarus virginalis, ont été compilées pour l’Ukraine. Le potentiel d’invasion des deux espèces a été estimé dans le contexte des problèmes environnementaux et socio-économiques locaux. Leur histoire en tant qu’espèces non indigènes est marquée par l’utilisation précoce des décapodes exotiques comme ressource naturelle par les résidents locaux. Des introductions intentionnelles et non intentionnelles ont eu lieu pour M. nipponense, tandis qu’un commerce ornemental a été une source importante de P. virginalis non indigène. De nouveaux signalements de M. nipponense et de P. virginalis dans des habitats d’eau douce ouverts dans les provinces d’Odesa et de Kharkiv suggèrent une expansion continue de ces décapodes. Nous avons effectué une analyse phylogénétique moléculaire des séquences COI, qui a confirmé les identifications morphologiques de M. nipponense envahissante. Cela renforce la voie présumée de cette espèce, y compris sa trajectoire de la Chine vers le sud-ouest de l’Ukraine. On a constaté que Procambarus fallax était paraphylétique par rapport à P. virginalis, ce qui est probablement dû à l’analyse d’un seul gène mitochondrial, qui pourrait être affecté par l’émergence récente de cette dernière espèce. Mots-clés : Décapode / Macrobrachium nipponense / Procambarus virginalis / Ukraine / espèce envahissante *Corresponding author: [email protected] This is an Open Access article distributed under the terms of the Creative Commons Attribution License CC-BY-ND (https://creativecommons.org/licenses/by-nd/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. If you remix, transform, or build upon the material, you may not distribute the modified material. M.O. Son et al.: Knowl. Manag. Aquat. Ecosyst. 2020, 421, 40 Table 1. Samples used in our study and main characteristics of localities. Locality Coordinates Date Salinity Substrate Collectors (PSU) Procambarus virginalis 1. Kharkiv, pond “Rodnyk” 50.030453, 36.348038 10.10.2019 No data Sand and clay S.A. Sidorovskyi, M.O. Kulyk 2. Dnipro, Kotlovan Lake 48.506056, 35.102197 13.11.2015 0.957 Sand R.O. Novitskyi, A. Artemenko Macrobrachium nipponense 3. Fish pond 46.3441966, 30.5963854 23.07.2018 2.920 Mud M.O. Son, H. Morhun, (Akkarzhanka River Basin) 20.08.2018 S.Utevsky 23.08.2018 4. Olexandrivka Reservoir 46.336806, 30.625972 02.09.2018 1.555 Mud M.O. Son (Akkarzhanka River Basin) 5. Reservoir in the Great 46.610917, 30.878972 13.03.2020 1.286 Mud with reed M.O. Son Adzhalyk Liman Basin fragments 1 Introduction resulted in instant reproductive isolation (Vogt et al., 2015). This reproductive isolation is the basis for the assignment of Over the past decades, expansions of five non-native the invasive marmorkrebs to its own species (Lyko, 2017). fi decapods, Rhithropanopeus harrisii (Gould, 1841), Eriocheir This cray sh is known as a non-native species with a high sinensis Milne-Edwards, 1853, Macrobrachium nipponense invasive potential, and it occurs throughout Europe (Martin (De Haan, 1849), Orconectes limosus (Rafinesque, 1815) and et al., 2010b; Bohman et al., 2013; Chucholl, 2014; Vojkovská ő Procambarus virginalis Lyko, 2017 have been recorded in et al., 2014; L kkös et al., 2016; Lipták et al., 2017; Ukraine’s inland waters (Son et al., 2013; Novitsky and Son, Pârvulescu et al., 2017; Ercoli et al., 2019). 2016; Kudryashov, 2020). While invasive crabs R. harrisii and This research was aimed at tracing the invasion history of E. sinensis are common along the Azov-Black Sea coast and M. nipponense and P. virginalis in Ukraine over the last few penetrate river basins through estuarine systems, O. limosus, years based on all currently available data. Both morphological fi M. nipponense and P. virginalis are distributed exclusively in and molecular characters were used to con rm the species inland waters (Son et al., 2013; Novitsky and Son, 2016; identity and to reveal phylogenetic relationships of their Kudryashov, 2020). populations in Ukraine. This is rather a large number of non-native species, considering that the native Ukrainian freshwater fauna includes only four decapod species, Astacus leptodactylus 2 Material and methods (Eschscholtz, 1823), Astacus astacus (Linnaeus, 1758), 2.1 Samples and sampling methods Astacus pachypus (Rathke, 1837) and Potamon ibericum (Bieberstein, 1808), of which the last three are not widespread In 2018–2019, decapod specimens were collected in and are considered to be vulnerable (Akimov, 2009; Policar different types of habitats in three of Ukraine’s provinces et al., 2018). representing a variety of climatic and ecological conditions of Macrobrachium nipponense is a widespread Indo-Pacific the country (Tab. 1, Figs. 1 and 2). species that occurs in estuarine and fresh waters (Yu and In a former quarry called Kotlovan Lake (Dnipropetrovsk Miyake, 1972; Cai and Ng, 2002). It has been introduced to Region), which was transparent enough to see objects of natural habitats in Bangladesh, Singapore, the Philippines, interest situated under water, samples were simply collected by Kazakhstan, Uzbekistan, Iraq, Iran, Moldova and Ukraine hand when diving. A transect video was used to estimate (Alekhnovich and Kulesh, 2001; Cai and Shokita, 2006; population density during this collection. In waterbodies in the De Grave and Ghane, 2006; Salman et al., 2006; Son et al., Akkarzhanka River Basin, a kick-net (ISO 7828) was dragged 2013). In addition, the species is cultivated in cooling along the shore over 5 m distances. A collector moved in reservoirs of thermal power stations in Russia, Belarus and zigzags to cover depths from the water edge to a depth of one Moldova (Alekhnovich and Kulesh, 2001). meter for 15 min, with 3–4 replicates. In the Great Adzhalyk The marmorkrebs (P. virginalis) is a triploid parthenoge- Liman Basin, one specimen was sampled by hand. Similarly, in netic species that was originally found in aquariums (Martin Kharkiv, crayfish samples were collected by hand in a man- et al., 2010a, 2016). This crayfish is viewed as a recent made pond. descendant of Procambarus fallax (Hagen, 1870), which is a Collected specimens were immediately preserved in 96% diploid sexually reproducing species known from Florida and ethanol for further morphological examination and molecular Georgia, USA (Taylor et al., 1996). analysis. The specimens are stored in the collections of The emergence of the marmorkrebs is explained by a very invertebrates at the Institute of Marine Biology, National recent, in terms of evolution, macromutation in P. fallax that Academy of Sciences of Ukraine, V. N. Karazin Kharkiv Page 2 of 11 M.O. Son et al.: Knowl. Manag. Aquat. Ecosyst. 2020, 421, 40 rostrum (González-Ortegón and Cuesta, 2006). Its easily noticeable distinguishing feature is its long second pereiopods (they exceed the length of the rest of the body in larger adults) that have claws bearing numerous setae. 2.3 DNA extraction and PCR The collected specimens of P. virginalis