Ardura, A.

Garcia-Vazquez, E.

Pochon, X.

Samuiloviene, A.

Wood, S.

Zaiko, A.

OceanRep

Molecular approaches for non-indigenous species surveillance –from introduction pathways to established populations  Anastasija Zaiko,  Aurelija Samuiloviene, Alba Ardura, Eva Garcia-Vazquez,  Xavier Pochon, Susie Wood  Email: anastasija@corpi.ku.lt  Courtesy: JM Weslawski Challenges of surveying marine ecosystem Increasing human pressures, declining taxonomic expertise • Morphological identification methods are laborious • Require considerable taxonomic expertise • Often fail to identify cryptic species • Or species at the larval stage Photo credits: www.musselfree.org www.cefas.defra.gov.uk www.fao.org Crassostrea gigas Dreissena polymorpha Ostrea edule Risk to overlook or misidentify  non-indigenous, pathogen or indicator species Opportunities of the eDNA analysis with next-generation sequencing • High-throughput  • Parallel sequencing  • Millions of reads at once  • Low-cost sequencing  • Multiplex capability   • Multiple species detection    (DNA Metabarcoding) MiSeq IlluminaTM Technology The consecutive stages of NIS introduction Olenin et al. (2011) Mar Poll Bull 62: 2598-2604 1. NIS detection on a pathway: BW survey ALGAE METAZOA PROTOZOA # sequences # counts Peringia ulvae – an European hydrobiid gastropod seemed to withstand the harsh BW conditions Zaiko et al. (2015) Mar Poll Bull 92: 25-34 2. Early detection of NIS: biofilm study Syringe (SYR) Tape (TAP) Sponge (SPO) Swab (SWA) 0%20%40%60%80%100%SYR-Air SYR-Water SPO-Air SPO-Water SWA-Air SWA-WaterNumber of sequences, % UnassignedMetazoaChromalveolataRhizariaFungiArchaeplastidaAverage ratio of taxa per treatment (Super-Group Level) Pochon et al. (2015) Biofouling 31: 241-251  No statistical differences in biodiversity between devices nor sampling environments MiSeq IlluminaTM Technology => 450 Operational Taxonomic Units  Diversity  5 Supergroups 22 Phyla 42 Classes 83 Families 96 Genera  46 Species   NIS Ciona savignyi NZ Inventories 31 genera 28 species  Never reported in NZ 3. Early detection of NIS: plankton study   Plume area (PA) “Sea gates” (SG) Klaipeda Strait (KS) Species detected by metabarcoding Annelida Polychaeta Marenzelleria viridis Marenzelleria neglecta Crustacea Cladocera Bosmina coregoni Bosmina spp. Cercopagis pengoi Daphnia galeata Daphnia cucullata Evadne nordmanni Leptodora kindtii Pleopis polyphemoides Podon leukartii Copepoda Acartia tonsa Mesocyclops  leukarti Mollusca Bivalva Dreissena polymorpha Macoma balthica Mytilus sp. Gastropoda Hydrobia ulvae Rotifera Eurotatoria Keratella quadrata  ll ria viridis  >800 sequences found in all samples, aligned with high confi ence (>97% identity, >95% coverage) 4. Detection of a target NIS from plankton Rangia cuneata – a recent invasive species, abundant in the Vistula lagoon and with the potential to spread further Cerastoderma  Dreissena Macoma Mytilus Rangia Marker sensitivity ~ 40ng/uL DNA Location R. cuneata larvae density, ind/m3  Juodkrante Not detected Nida Not detected 1 6791 3 3058 7 1168 9 1946 4 20262 Ardura et al. (2015) Mar Environ Res, in press Summing up: • Molecular techniques can assist in NIS detection and surveillance  • They may help to overcome the issues with the lack of  taxonomic expertise • They allows identification of cryptic life stages (eggs or larvae), detection of rare and sparsely distributed organisms  • Suitable for general biodiversity assessment, detection of target organisms  • May help to obtain information necessary for development of environmental quality metrics or pressure indicators  BUT • Comprehensive reference databases needed • Quantification  should be elaborated • Marker validation / application of multiple markers Thank you! Collaborators: Yaisel J. Borell, Jose L. Martinez, Deni Ribicic, Julia Schmidt-Petersen, Anna Semenova, Jonathan Banks  

Molecular approaches for non-indigenous species surveillance – from introduction pathways to established populations

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