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Molecular identification of poisonous pufferfishes and cross-atlantic genetic divergence patterns based on the mitochondrial COI gene

Published:September 30, 2019DOI:https://doi.org/10.1016/j.fsigss.2019.09.082

      Abstract

      The Tetraodontidae is the most speciose family within the order Tetraodontiformes, being characterized by beak-like jaws and the presence of powerful neurotoxins Tetrodotoxin/Saxitoxin associated with soft tissues, inflation behavior under stress, a condition shared with its accepted sister-family Diodontidae. Although several studies, including both morphological and molecular analyses have been conducted in the last decade the phylogeny and biogeography of Tetraodontidae and its species remain under debate. Several fatal intoxication cases had been observed in the last years related with the ingestion of Tetraodontidae species all around the world. Although recent technological advances have facilitated the sequencing of an entire mitogenome (∼16Kb), increasing the use of mtDNA as a phylogenetic marker, several studies primarily focusing on small mtDNA regions are continuously conducted. Therefore, the aim of the present study was to investigate the molecular identification and the cross-atlantic genetic divergence patterns observed from the mitochondrial COI gene of the poisonous genus Sphoeroides based on newly determined and previously published sequences from both North and South Atlantic Oceans.

      Keywords

      1. Introduction

      The Amazon River is one of the largest rivers in the world, with 6,996 km and over a thousand tributaries. Currently, the amazon river releases around 209 thousand cubic meters of water every second in the Atlantic Ocean. This huge quantity of water creates a current of great strength heading to the middle of the ocean, forming a barrier between the Atlantic seas south and north of the river, making it difficult for the passage of reef animals [
      • Coles V.J.
      • Brooks M.T.
      • Hopkins J.
      • et al.
      The pathways and properties of the Amazon River Plume in the tropical North Atlantic Ocean.
      ,
      • Floeter S.R.
      • Rocha L.A.
      • Robertson D.R.
      • et al.
      Atlantic reef fish biogeography and evolution.
      ]. The two species this study emphasizes (Sphoeroides greeleyi and Sphoeroides testudineus) have populations both north and south of were Amazon river meets the Atlantic, thus leading them to being used to discuss the permeability of the barrier. They are also important due to the fact that these species of puffer, just like other individuals of the Tetraodontidae family, are poisonous, and cause many incidents across the planet related with the ingestion of fish meat contamined with Tetrodotoxin. This toxin is produced in the gonads, liver and skin of the Tetraodontidae. Several cases of poisoning by this toxin have already been reported in the literature, including fatal cases. Tetradotoxin acts blocking cell’s sodium channel, causing great damage to the nervous system in humans [
      • Goto T.
      • Kishi Y.
      • Takahashi S.
      • et al.
      Tetrodotoxin.
      ]. In the present manuscript we discussed the permeability of the Amazon barrier using these two pufferfish species, and the mitochondrial COI gene.

      2. Methodology

      The material used in this study were from individuals of the species Sphoeroides greeleyi and Sphoeroides testudineus collected in Angra dos Reis, Rio de Janeiro, Brazil. The COI sequences were amplified using the standard PCR protocol using QIAGEN Mastermix and primers VF2 and Fish-R2. These sequences were then compared to existing ones in the literature (GenBank and BOLD). The DNA Barcoding methodology was used and the neighbor-joining tree was constructed based on K2P diastances. Alignment and sequence analysis was conducted using the the Geneious v4.82 and MEGA-X softwares.

      3. Results and discussion

      Results of the comparison of specimens north and south of the Amazon barrier showed almost no genetic differentiation between the populations of Sphoeroides testudineus (0,007 K2P distance) (Fig. 1).
      Fig. 1
      Fig. 1A. Sphoeroides greeleyi; B. Sphoeroides testudineus; C. COI K2P tree of Caribbean and North-South Atlantic Sphoeroides.
      As observed by [
      • Rocha L.A.
      Patterns of distribution and processes of speciation in Brazilian reef fishes.
      ], there are 248 reef fish species in common between the northern and southern oceans. These species represent 42% of the reef fish fauna of the sea north of the Amazon River plume and 74% of the fauna in the south. The effectiveness of the Amazon barrier may have been strongly influenced by change in the sea levels, not only in the current speed but also its salinity and the availability of marine reef habitats beneath the plume. In a high sea level the fresh water can pass above the Amazon shelf thanks to saltwater beneath it, creating a corridor below the current of river water [
      • Rocha L.A.
      Patterns of distribution and processes of speciation in Brazilian reef fishes.
      ]. This allows organisms to thrive in this corridor under the plume and connects the Brazilian reef fauna to the Caribbean, and vice-versa. The low COI K2P divergence observed in our results suggest that this could be the case for the Sphoeroides testudineus. In low-sea level (the present sea-level is considered low when compared to ones in the past) the freshwater flow passes directly on the Amazon shelf, combined with the high sedimentation, it’s unlikely that any coral reefs can develop on the north-eastern coast of South America, separating the Brazilian from the Caribbean coral and reef-associated faunas [
      • Rocha L.A.
      Patterns of distribution and processes of speciation in Brazilian reef fishes.
      ].

      4. Conclusions

      The observed COI K2P divergence (0,007) suggest that albeit the Amazon barrier, both populations seem to belong to a single species. We do also agree with [
      • Rocha L.A.
      Patterns of distribution and processes of speciation in Brazilian reef fishes.
      ] about the Amazon river restricting the dispersal of reef associated species, but, in the past, high sea levels have allowed the faunal interchange that resulted in the establishment of new species and populations in the both Caribbean and Brazilian coasts. These interchange created the situation we see today, with many species co-existing in both regions, despite the Amazon barrier.

      Role of funding

      This project was supported by the Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ) and the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior.

      Declaration of Competing Interest

      None.

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