Development of a novel molecular tool for the rapid assessment of the biodiversity changes of benthic nematodes assemblages. Mares Conference Marine Ecosystems Helath and conservation

The molecular profiling system was developed using directed terminal-restriction fragment length polymorphism (dT-RFLP) to characterize soil nematode assemblages by relative abundance of feeding guilds and validation by comparison to traditional morphological method. The good performance of these molecular tools applied to soil nematodes assemblages create an opportunity to develop a novel approach for rapid assessment of the biodiversity changes of benthic nematodes assemblages of marine and estuarine sediments. The main aim of this research is to combine morphological and molecular analysis of estuarine nematodes assemblages, to establish a tool for fast assessment of the biodiversity changes within habitat recovery of Zostera noltii seagrass beds; and validate the dT-RFLP as a high-throughput tool to assess the system recovery. It was also proposed to develop a database of sequences related to individuals identified at species level to develop a new taxonomic reference system. A molecular phylogenetic analysis of the estuarine nematodes has being performed. After morphological identification, barcoding of 18S rDNA are being determined for each nematode species and the results have shown a good degree of concordance between traditional morphology-based identification and DNA sequences. The digest strategy developed for soil nematodes is not suitable for marine nematodes. Then five samples were cloned and sequenced and the sequence data was used to design a new dT-RFLP strategy to adapt this tool to marine assemblages. Several solutions were presented by DRAT and tested empirically to select the solution that cuts most efficiently, separating the different clusters. The results of quantitative PCR showed differences in nematode density between two sampling stations according the abundance of the nematode density obtained by the traditional methods. These results suggest that qPCR could be a robust tool for enumeration of nematode abundance, saving time.

Fixing, storage time and DNA extraction applied to dT-RFLP for ecological and molecular studies of marine nematodes assemblages

The application of molecular methods offers an alternative faster than traditional methods based on morphology It is nearly impossible to process all the samples in short period using traditional methods, and the deterioration of marine sediments rapidly occurs The dT-RFLP (directed Terminal-Restriction Fragment Length Polymorphism) allows a rapid assessment of biodiversity changes of nematodes assemblages The use of a not suitable fixing, storage time and DNA extraction could be a limitation in molecular analysis like dT-RFLP and real time PCR.Objetives: the best fixative •the level of DNA degradation over the time •the best DNA extraction method for marine nematodes and suitable for dT-RFLP analysis

Changes in fish assemblage structure after implementation of Marine Protected Areas in the south western coast of Portugal.

Marine Protected Areas (MPAs) are increasingly being recommended as management tools for biodiversity conservation and fisheries. With the purpose of protecting the region's biodiversity and prevent the over exploitation of marine resources, in February 2011 the no-take MPAs of Ilha do Pessegueiro and Cabo Sard~ao were implemented within the “Parque Natural do Sudoeste Alentejano e Costa Vicentina “(PNSACV) Marine Park, south western coast of Portugal. As such, commercial and recreational fishing became prohibited in these areas. In order to evaluate the effects of these no-take MPAs, the structure of their fish assemblages and of adjacent control areas without fishing restrictions were studied between 2011 (immediately after implementation) and 2013 (two years after implementation). A total of 4 sampling campaigns were conducted (summer 2011, winter 2012, summer 2013 and winter 2013) using trammel nets and bottom trawl. Ichthyofaunal assemblages from the no-take MPAs (treatment) were compared with adjacent areas (controls) and changes evaluated as a function of time since protection. Results revealed significant increase in fish abundance after the implementation of the no-take MPAs. Furthermore, significant differences in the structure of fish assemblages (abundance and fish size) between protected and neighbouring areas were rapidly observed upon the implementation of the no-take MPAs. In addition, specimens of larger size occurred more frequently within Ilha do Pessegueiro no-take MPA in the last year of the study. Overall, despite the young age of these no-take MPAs, changes on the structure of their fish assemblages were already evident after only two years of protection, indicating that management measures such as MPA designation may play an important role to promote fisheries sustainable exploitation as well as to protect species with conservation interest.