Marine Biology

, 166:82 | Cite as

The invasive sea slug Pleurobranchaea maculata is a vector of two potent neurotoxins in coasts of Argentina

  • Nahuel E. FariasEmail author
  • Alejandra B. Goya
  • Evangelina Schwindt
  • Sandra Obenat
  • Monika Dhanji-Rapkova
  • Andrew D. Turner
Original paper


Toxic exotic organisms can have profound effect as new vectors of keystone compounds in non-native areas. In recent years the invasive sea slug Pleurobranchaea maculata has been reported as thriving along the oriental coasts of South America. The same species had been previously found to contain high levels of tetrodotoxins (TTXs) in its native range. With the aim of determining toxin contents for the introduced individuals we performed mouse bioassays (MBA) and liquid chromatography tandem–mass spectrometry analyses (LC–MS/MS) in three distant populations (− 38°02′11″, − 57°31′28″; − 40°29′59″, − 60°14′09″; − 42°44′15″, − 65°01′40″) and followed the temporal variation in toxin contents in one of them, from June 2014 to January 2015. Relative low levels of TTXs were detected jointly with high levels of PSTs. This is the first identification of TTXs in the temperate coasts of the southwestern Atlantic and the first detection of PSTs in a pleurobranch, in both adults and eggs. Concentrations of PSTs and TTXs varied widely among individuals and populations, and through time. Our results provide new hints on the origin and acquisition mechanisms of these toxins in P. maculata and highlight the risk posed by the introduction of this new vector of potent neurotoxins for seafood safety and marine communities in the invaded area.



We want to acknowledge Carolina Kelly and Sonia Ortega from SENASA’s Mar del Plata Laboratory, for their help to perform mouse bioassays. We would like to thank Dr. Alejandro Bortolus (GEAC-IPEEC-CONICET) for encouraging us to undertake this study and for his contribution to the first discussions. We also thank two anonymous reviewers whose comments and criticism contributed significantly to improve this manuscript.

Authors contribution

NF has performed the field sampling, designed the study, analyzed the data and written a first draft. AG performed the mouse bioassays. ES provided the individuals from Puerto Madryn. MDR and AT performed the LC–MS/MS analyses. AG, ES, SO, MDR and AT contributed to the writing of the manuscript.


Field work was supported by ANPCyT-PICT 2016 # 1083, PIP CONICET 508 granted to ES. Toxin identification and quantifications were funded with Internal Cefas Seedcorn Funding (contract code DP345).

Compliance with ethical standards

Conflict of interest

All authors declare that they have no conflict of interest.

Ethical approval

All bioassays in this study have been conducted following the regulation 617/2002 for biological tests and animal facilities of the National Service of Agri-Food Health and Quality of Argentina (SENASA), and in fulfillment of the ISO/IEC 17,025 norm, that conforms to the Directive 2010/63/EU of the European Parliament, the Council of 22 September 2010 on the protection of animals used for scientific purposes and the Code of Ethics of the World Medical Association (Declaration of Helsinki) for animal experiments.

Supplementary material

227_2019_3529_MOESM1_ESM.pdf (716 kb)
Supplementary material 1 (PDF 716 kb)


  1. Alcaraz A, Whiplle RE, Gregg HR, Anderson BD, Grant PM (1999) Analysis of Tetrodotoxin. Forensic Sci Int 99:35–45CrossRefGoogle Scholar
  2. Anon (2005) AOAC official method 959.08. Paralytic shellfish poison. Biological method. Final action. In: Truckses MW (ed) AOAC Official methods for analysis Chapter 49: natural toxins, 18th edn. AOAC International, Gaithersburg, pp 79–80Google Scholar
  3. Asakawa M, Ito K, Kajihara H (2013) Highly toxic ribbon worm Cephalothrix simula containing tetrodotoxin in Hiroshima bay, Hiroshima prefecture, Japan. Toxins (Basel) 5:376–395. CrossRefGoogle Scholar
  4. Bane V, Lehane M, Dikshit M, O’Riordan A, Furey A (2014) Tetrodotoxin: chemistry, toxicity, source, distribution and detection. Toxins (Basel) 6:693–755. CrossRefGoogle Scholar
  5. Battini N, Farías NE, Giachetti CB, Schwindt E, Bortolus A (2019) Staying ahead of invaders: can we cope with niche shifts? Mar Ecol Prog Ser 612:127–140. CrossRefGoogle Scholar
  6. Blanco J, Reyero MI, Franco J (2003) Kinetics of accumulation and transformation of paralytic shellfish toxins in the blue mussel Mytilus galloprovincialis. Toxicon 42:777–784. CrossRefPubMedGoogle Scholar
  7. Bökenhans V, Fernández Alfaya JE, Bigatti G, Averbuj A (2018) Diet of the invasive sea slug Pleurobranchaea maculata in Patagonian coastal waters. New Zeal J Zool. CrossRefGoogle Scholar
  8. Boundy MJ, Selwood AI, Harwood DT, McNabb PS, Turner AD (2015) Development of a sensitive and selective liquid chromatography-mass spectrometry method for high throughput analysis of paralytic shellfish toxins using graphitised carbon solid phase extraction. J Chromatogr A 1387:1–12. CrossRefPubMedGoogle Scholar
  9. Bricelj VM, Lee HJ, Cembella AD, Anderson DM (1990) Uptake kinetics of paralytic shellfish toxins from the dinoflagellate Alexandrium fundyense in the mussel Mytilus edulis. Mar Ecol Prog Ser 63:177–188. CrossRefGoogle Scholar
  10. Bricelj VM, Haubois AG, Sengco MR, Pierce RH, Culter JK, Anderson DM (2012) Trophic transfer of brevetoxins to the benthic macrofaunal community during a bloom of the harmful dinoflagellate Karenia brevis in Sarasota Bay, Florida. Harmful Algae 16:27–34. CrossRefGoogle Scholar
  11. Carreto JI, ElBusto C, Sancho H, Carignan M, Yasumoto T, Oshima Y (1996) Comparative studies on paralytic shellfish toxin profiles of marine snails, mussels and an Alexandrium tamarense isolate from the Mar del Plata coast (Argentina). Rev lnvest Des Pesq 10:101–107Google Scholar
  12. Ciocco NF, Lasta ML, Narvarte M, Bremec C, Bogazzi E, Valero J, Orensanz JM (2006) Chapter 26 Argentina. In: Shumway SE (ed) Parsons GJBT-D in A and FS scallops: biology, ecology and aquaculture. Elsevier, New York, pp 1251–1292CrossRefGoogle Scholar
  13. Cusick KD, Sayler GS (2013) An overview on the marine neurotoxin, saxitoxin: genetics, molecular targets, methods of detection and ecological functions. Mar Drugs 11:991–1018. CrossRefPubMedPubMedCentralGoogle Scholar
  14. Dao HV, Takata Y, Sato S, Fukuyo Y, Kodama M (2009) Frequent occurrence of the tetrodotoxin-bearing horseshoe crab Carcinoscorpius rotundicauda in Vietnam. Fish Sci 75:435–438. CrossRefGoogle Scholar
  15. Derby CD, Aggio JF (2011) The neuroecology of chemical defenses. Integr Comp Biol 51:771–780. CrossRefPubMedGoogle Scholar
  16. Esteves JL, Santinelli N, Sastre V, Díaz R, Rivas O (1992) A toxic dinoflagellate bloom and PSP production associated with upwelling in Golfo Nuevo, Patagonia, Argentina. Hydrobiologia 242:115–122. CrossRefGoogle Scholar
  17. Farias NE, Obenat S, Goya ABA (2015) Outbreak of a neurotoxic side-gilled sea slug (Pleurobranchaea sp.) in Argentinian coasts. New Zeal J Zool 42:51–56. CrossRefGoogle Scholar
  18. Farias N, Wood S, Obenat S, Schwindt E (2016) Genetic barcoding confirms the presence of the neurotoxic sea slug Pleurobranchaea maculata in southwestern Atlantic. New Zeal J Zool 43:292–298. CrossRefGoogle Scholar
  19. Ferrer RP, Zimmer RK (2013) Molecules of keystone significance: crucial agents in ecology and resource management. Bioscience 63:428–438. CrossRefGoogle Scholar
  20. Freitas JC, Ogata T, Veit CH, Kodama M (1996) Occurrence of tetrodotoxin and paralytic shellfish toxins in Phallusia nigra (Tunicata, Ascidiacea) from the Brazilian coast. J Venom Anim Toxins 2:28–38CrossRefGoogle Scholar
  21. Games PA, Howell JF (1976) Pairwise multiple comparison procedures with unequal N’s and/or variances: a Monte Carlo study. J Educ Stat 1:113–125. CrossRefGoogle Scholar
  22. Gayoso AM (2001) Observations on Alexandrium tamarense (Lebour) Balech and other dinoflagellate populations in Golfo Nuevo, Patagonia (Argentina). J Plankton Res 23:463. CrossRefGoogle Scholar
  23. Hall S, Strichartz G, Moczydlowski M, Ravindran A, Reichardt PB (1990) Marine toxins: origin, structure, and molecular pharmacology. American Chemical Society, Washington DCCrossRefGoogle Scholar
  24. Ikeda K, Emoto Y, Tatsuno R, Wang JJ, Ngy L, Taniyama S, Takatani T, Arakawa O (2010) Maturation-associated changes in toxicity of the pufferfish Takifugu poecilonotus. Toxicon 55:289–297. CrossRefPubMedGoogle Scholar
  25. Ito K, Asakawa M, Sida Y, Miyazawa K (2003) Occurrence of paralytic shellfish poison (PSP) in the starfish Asterina pectinifera collected from the Kure Bay, Hiroshima Prefecture, Japan. Toxicon 41:291–295. CrossRefPubMedGoogle Scholar
  26. Itoi S, Yoshikawa S, Asahina K, Suzuki M, Ishizuka K, Takimoto N, Mitsuoka R, Yokoyama N, Detake A, Takayanagi C, Eguchi M, Tatsuno R, Kawane M, Kokubo S, Takanashi S, Miura A, Suitoh K, Takatani T, Arakawa O, Sakakura Y, Sugita H (2014) Larval pufferfish protected by maternal tetrodotoxin. Toxicon 78:35–40. CrossRefPubMedGoogle Scholar
  27. Katikou P, Georgantelis D, Sinouris N, Petsi A, Fotaras T (2009) First report on toxicity assessment of the Lessepsian migrant pufferfish Lagocephalus sceleratus (Gmelin, 1789) from European waters (Aegean Sea, Greece). Toxicon 54:50–55. CrossRefPubMedGoogle Scholar
  28. Khor S, Wood SA, Salvitti LR, Taylor DI, Adamson J, McNabb P, Cary SC (2014) Investigating diet as the source of tetrodotoxin in Pleurobranchaea maculata. Mar Drugs 12:1–16. CrossRefGoogle Scholar
  29. Kwong RWM, Wang WX, Lam PKS, Yu PKN (2006) The uptake, distribution and elimination of paralytic shellfish toxins in mussels and fish exposed to toxic dinoflagellates. Aquat Toxicol 80:82–91. CrossRefPubMedGoogle Scholar
  30. Lago J, Rodriguez LP, Blanco L, Vieites JM, Cabado AG (2015) Tetrodotoxin, an extremely potent marine neurotoxin: distribution, toxicity, origin and therapeutical uses. Mar Drugs 13:6384–6406. CrossRefPubMedPubMedCentralGoogle Scholar
  31. Llewellyn LE, Endean R (1989) Toxicity and paralytic shellfish toxin profiles of the xanthid crabs, Lophozozymus pictor and Zosimus aeneus, collected from some Australian coral reefs. Toxicon 27:596–600. CrossRefPubMedGoogle Scholar
  32. Llewellyn L, Negri A, Robertson A (2006) Paralytic shellfish toxins in tropical oceans. Toxin Rev 25:159–196. CrossRefGoogle Scholar
  33. Lopes VM, Lopes AR, Costa P, Rosa R (2013) Cephalopods as vectors of harmful algal bloom toxins in marine food webs. Mar Drugs 11:3381–3409. CrossRefPubMedPubMedCentralGoogle Scholar
  34. MacKenzie AL (2014) The risk to New Zealand shellfish aquaculture from paralytic shellfish poisoning (PSP) toxins. New Zeal J Mar Freshw Res 48:430–465. CrossRefGoogle Scholar
  35. MacQuarrie SP, Bricelj VM (2008) Behavioral and physiological responses to PSP toxins in Mya arenaria populations in relation to previous exposure to red tides. Mar Ecol Prog Ser 366:59–74. CrossRefGoogle Scholar
  36. Magarlamov TY, Melnikova DI, Chernyshev AV (2017) Tetrodotoxin-producing bacteria: detection, distribution and migration of the toxin in aquatic systems. Toxins (Basel) 9:166. CrossRefGoogle Scholar
  37. Mazio CA, Dragani WC, Caviglia FJ, Pousa JL (2004) Tidal hydrodynamics in Golfo Nuevo, Argentina, and the adjacent continental shelf. J Coast Res 204:1000–1011. CrossRefGoogle Scholar
  38. McNabb P, Selwood AI, Munday R, Wood SA, Taylor DI, Mackenzie LA, van Ginkel R, Rhodes LL, Cornelisen C, Heasman K, Holland PT, King C (2010) Detection of tetrodotoxin from the grey side-gilled sea slug—Pleurobranchaea maculata, and associated dog neurotoxicosis on beaches adjacent to the Hauraki Gulf, Auckland, New Zealand. Toxicon Off J Int Soc Toxinology 56:466–473. CrossRefGoogle Scholar
  39. Miyazawa K, Noguchi T, Maruyama J, Jeon JK, Otsuka M, Hashimoto K (1985) Occurrence of tetrodotoxin in the starfishes Astropecten polyacanthus and A. scoparius in the Seto Inland Sea. Mar Biol 90:61–64. CrossRefGoogle Scholar
  40. Montoya NG, Carignan MO, Carreto JI (2018) Alexandrium tamarense/catenella blooms in the Southwestern Atlantic: paralytic shellfish toxin production and its trophic transference. In: Hoffmeyer M, Sabatini M, Brandini F, Calliari D, Santinelli N (eds) Plankton ecology of the southwestern Atlantic. Springer, Cham, pp 453–476CrossRefGoogle Scholar
  41. Noguchi T, Arakawa O (2008) Tetrodotoxin—distribution and accumulation in aquatic organisms, and cases of human intoxication. Mar Drugs 6:220–242. CrossRefPubMedPubMedCentralGoogle Scholar
  42. Noguchi T, Jeon JK, Arakawa O, Sugita H, Deguchi Y, Shida Y, Hashimoto K (1986) Occurrence of tetrodotoxin and anhydrotetrodotoxin in Vibrio sp. isolated from the intestines of a xanthid crab, Atergatis floridus. J Biochem 99:311–314. CrossRefPubMedGoogle Scholar
  43. Noguchi T, Miyazawa K, Daigo K, Arakawa O (2011) Paralytic shellfish poisoning (PSP) toxin-and/or tetrodotoxin-contaminated crabs and food poisoning by them. Toxin Rev 30:91–102. CrossRefGoogle Scholar
  44. Ottaway J (1977) Pleurobranchaea novaezelandiae preying on Actinia tenebrosa. New Zeal J Mar Freshw Res 11:125–130CrossRefGoogle Scholar
  45. Pratheepa V, Vasconcelos V (2013) Microbial diversity associated with tetrodotoxin production in marine organisms. Environ Toxicol Pharmacol 36:1046–1054. CrossRefPubMedGoogle Scholar
  46. Qiu J, Meng F, Ding L, Che Y, McCarron P, Beach DG, Li A (2018) Dynamics of paralytic shellfish toxins and their metabolites during timecourse exposure of scallops Chlamys farreri and mussels Mytilus galloprovincialis to Alexandrium pacificum. Aquat Toxicol 200:233–240. CrossRefPubMedGoogle Scholar
  47. Sabrah MM, El-Ganainy AA, Zaky MA (2006) Biology and toxicity of the pufferfish Lagocephalus sceleratus (Gmelin, 1789) from the Gulf of Suez. Egypt J Aquat Res 32:283–297Google Scholar
  48. Salvitti LR (2015) Elucidating the Origin of Tetrodotoxin in Pleurobranchaea maculata and Stylochoplana sp. University of Waikato, HamiltonGoogle Scholar
  49. Salvitti LR, Wood SA, Taylor DI, McNabb P, Cary SC (2015a) First identification of tetrodotoxin (TTX) in the flatworm Stylochoplana sp.; A source of TTX for the sea slug Pleurobranchaea maculata. Toxicon 95:23–29. CrossRefPubMedGoogle Scholar
  50. Salvitti LR, Wood SA, Winsor L, Cary SC (2015b) Intracellular immunohistochemical detection of tetrodotoxin in Pleurobranchaea maculata (Gastropoda) and Stylochoplana sp. (Turbellaria). Mar Drugs 13:756–769. CrossRefPubMedPubMedCentralGoogle Scholar
  51. Salvitti LR, Wood SA, McNabb P, Craig S (2015c) No evidence for a culturable bacterial tetrodotoxin producer in Pleurobranchaea maculata (Gastropoda: Pleurobranchidae) and Stylochoplana sp. (platyhelminthes: Polycladida). Toxins (Basel) 7:255–273. CrossRefGoogle Scholar
  52. Salvitti LR, Wood SA, Fairweather R, Culliford D, McNabb P, Cary SC (2016) In situ accumulation of tetrodotoxin in non-toxic Pleurobranchaea maculata (Opisthobranchia). Aquat Sci 79(2):335–344. CrossRefGoogle Scholar
  53. Soong TW, Venkatesh B (2006) Adaptive evolution of tetrodotoxin resistance in animals. Trends Genet 22:621–626. CrossRefPubMedGoogle Scholar
  54. Tatsuno R, Shikina M, Soyano K, Ikeda K, Takatani T, Arakawa O (2013) Maturation-associated changes in the internal distribution of tetrodotoxin in the female goby Yongeichthys criniger. Toxicon 63:64–69. CrossRefPubMedGoogle Scholar
  55. Taylor DI, Wood SA, McNabb PS (2011) Population surveys of Pleurobranchaea maculata and tetrodotoxin in Waitemata harbour. Nelson, New ZealandGoogle Scholar
  56. Taylor DI, Wood SA, McNabb P, Ogilvie S, Cornelisen C, Walker J, Khor S, Cary SC (2015) Facilitation effects of invasive and farmed bivalves on native populations of the sea slug Pleurobranchaea maculata. Mar Ecol Prog Ser 537:39–48. CrossRefGoogle Scholar
  57. R Core Team (2016) R: a language and environment for statistical computing. Version 3.2.2. R foundation for statistical computing, Vienna, Austria. Accessed 1 Dec 2016
  58. Turner AD, Tarnovius S, Goya AB (2014) Paralytic shellfish toxins in the marine gastropods Zidona dufresnei and Adelomelon beckii from Argentina: toxicity and Toxin Profiles. J Shellfish Res 33:519–530. CrossRefGoogle Scholar
  59. Turner AD, Powell A, Schofield A, Lees DN, Baker-Austin C (2015a) Detection of the pufferfish toxin tetrodotoxin in European bivalves, England, 2013 to 2014. Euro Surveill. CrossRefPubMedGoogle Scholar
  60. Turner AD, McNabb PS, Harwood DT, Selwood AI, Boundy MJ (2015b) Single-laboratory validation of a multitoxin ultra-performance LC-hydrophilic interaction LC–MS/MS method for quantitation of paralytic shellfish toxins in bivalve shellfish. J AOAC Int 98:609–621. CrossRefPubMedGoogle Scholar
  61. Turner AD, Boundy MJ, Rapkova MD (2017) Development and single-laboratory validation of a liquid chromatography tandem mass spectrometry method for quantitation of tetrodotoxin in mussels and oysters. J AOAC Int 100:1469–1482. CrossRefPubMedGoogle Scholar
  62. Wägele H, Klussmann-Kolb A (2005) Opisthobranchia (Mollusca, Gastropoda)—more than just slimy slugs Shell reduction and its implications on defence and foraging. Front Zool 2:3. CrossRefPubMedPubMedCentralGoogle Scholar
  63. Wiese M, D’Agostino PM, Mihali TK, Moffitt MC, Neilan BA (2010) Neurotoxic alkaloids: saxitoxin and its analogs. Mar Drugs 8:2185–2211. CrossRefPubMedPubMedCentralGoogle Scholar
  64. Willan RC (1983) New Zealand side-gilled sea slugs (Opisthobranchia: Notaspidea: Pleurobranchidae). Malacologia 23:221–270Google Scholar
  65. Wood SA, Casas M, Taylor DI, McNabb P, Salvitti LR, Ogilvie S, Cary SC (2012a) Depuration of Tetrodotoxin and changes in bacterial communities in Pleurobranchea maculata adults and egg masses maintained in captivity. J Chem Ecol 38:1342–1350. CrossRefPubMedGoogle Scholar
  66. Wood SA, Taylor DI, McNabb P, Walker J, Adamson J, Cary SC (2012b) Tetrodotoxin concentrations in Pleurobranchaea maculata: temporal, spatial and individual variability from New Zealand populations. Mar Drugs 10:163–176. CrossRefPubMedPubMedCentralGoogle Scholar
  67. Wu Z, Xie L, Xia G, Zhang J, Nie Y, Hu J, Wang S, Zhang R (2005) A new tetrodotoxin-producing actinomycete, Nocardiopsis dassonvillei, isolated from the ovaries of puffer fish Fugu rubripes. Toxicon 45:851–859. CrossRefPubMedGoogle Scholar
  68. Yamada R, Tsunashima T, Takei M, Sato T, Wajima Y, Kawase M, Oshikiri S, Kajitani Y, Kosoba K, Ueda H, Abe K, Itoi S, Sugita H (2017) Seasonal changes in the tetrodotoxin content of the flatworm Planocera multitentaculata. Mar Drugs 15:56. CrossRefPubMedCentralGoogle Scholar
  69. Yasumoto T, Yasumura D, Yotsu M, Michishita T, Endo A, Kotak Y (1986) Bacterial production of tetrodotoxin and anhydrotetrodotoxin. Agric Biol Chem 50:793–795. CrossRefGoogle Scholar
  70. Yıldırım Y, Anderson MJ, Hansson B, Patel S, Millar CD, Rainey PB (2018) Genetic structure of the grey side-gilled sea slug (Pleurobranchaea maculata) in coastal waters of New Zealand. PLoS One 13:e0202197CrossRefGoogle Scholar
  71. Zimmer RK, Ferrer RP (2007) Neuroecology, chemical defense, and the keystone species concept. Biol Bull 213:208–225CrossRefGoogle Scholar
  72. Zimmer RK, Schar DW, Ferrer RP, Krug PJ, Kats LB, Michel WC (2006) The scent of danger: tetrodotoxin (TTX) as an olfactory cue of predation risk. Ecol Monogr 76:585–600.;2 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Laboratorio de InvertebradosFCEyN, Universidad Nacional de Mar del PlataMar del PlataArgentina
  2. 2.Instituto de Investigaciones Marinas y Costeras (IIMyC), CONICETMar del PlataArgentina
  3. 3.Departamento de Toxinas Marinas, Laboratorio Regional Mar del PlataCentro Regional Buenos Aires Sur SENASA (Servicio Nacional de Sanidad y Calidad Agroalimentaria)Mar del PlataArgentina
  4. 4.Grupo de Ecología en Ambientes Costeros (GEAC-IBIOMAR-CONICET)Puerto MadrynArgentina
  5. 5.Centre for Environment, Fisheries and Aquaculture Science (CEFAS)WeymouthUK

Personalised recommendations