Bacterial diversity and tetrodotoxin analysis in the viscera of the gastropods from Portuguese coast

To trace the pathway of tetrodotoxin (TTX) producing microorganism in the Atlantic coast of Portugal, culture-dependent evaluation of the bacterial isolates from the viscera of the gastropods Monodonta lineata, Gibbula umbilicalis, Nucella lapillus and Patella intermedia, and from the environmental samples (biofilm and surrounding sea water) was carried out. Samples were collected from eight different coastal locations of Northern Portugal. A total of 311 isolates were identified. The observed bacterial diversity was distributed over five different classes (Gammaproteobacteria, Alphaproteobacteria, Flavobacteria, Bacilli and Actinobacteria) with the greatest number of 16S rRNA gene sequence derived from the Gammaproteobacteria (75%). Phylogenetic analysis based on the 16S rRNA gene showed that bacterial isolates were highly diverse and most of which were found in other marine environment. Among the different species isolated, Vibrio was found abundant. Eventhough TTX was not detected (UPLC-MS/MS) in the isolates from this study, PCR screening identified some natural product biosynthesis genes (PKS and NRPS) involved in its assembly. Further PCR screening of the TTX producing two ATCC Vibrio sp. reveals that NRPS might be involved in the biosynthesis of TTX through the incorporation of arginine. © 2016 Elsevier.

This research was partially supported by the European Regional Development Fund (ERDF) through the COMPETE Operational Competitiveness Programme and national funds through FCT Foundation for Science and Technology under the project UID/Multi/04423/2013 and by the Structured Program of R&D&I INNOVMAR - Innovation and Sustainability in the Management and Exploitation of Marine Resources (reference NORTE-01-0145-FEDER-000035, Research Line NOVELMAR), funded by the Northern Regional Operational Programme (NORTE2020) through the European Regional Development Fund (ERDF). PV also acknowledges FCT for postdoctoral grant (SFRH/BPD/78269/2011), Joana Azevedo and Micaela Vale for the timely support. Also acknowledges CIFGA, Spain for the UPLC-MS/MS analysis.