Biomonitoring of Ferreira and Sousa rivers using diatoms: comparison between morphological and molecular approaches

• Autor(a) principal: Teixeira, Marco José Neto
• Data de Publicação: 2021
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10773/30712
• topic: Diatoms; DNA metabarcoding; Biomonitoring; Ecological assessment of water; Anthropogenic pressures

Official monitoring programs implemented by the Portuguese Environmental Agency (APA), do not fully survey all streams and rivers due to their nationalwide coverage, budget, and practical limitations. Two rivers in the north of Portugal, the Sousa and Ferreira rivers, with a recent past of different types of pollution and that run over areas with a high degree of alteration and dense population, were analyzed ecologically based on biological (diatom communities), physicochemical, and hydromorphological data. Diatoms are excellent ecological indicators of water quality. They are broadly distributed, show high species diversity, and respond rapidly to anthropogenic pressures. The Water Framework Directive (WFD) gives the legal basis for the use of this ecological indicator for water quality assessment, through the calculation of autoecological indexes. Portugal officially adopted the use of the Specific Polluosensitivity Index (SPI), which after determined, is converted into Ecological Quality Ratio (EQR). To determine the SPI, a morphological identification and cunt of diatom species using a microscope is required. This approach requires high taxonomic skills and several hours of analysis. DNA metabarcoding combined with high-throughput sequencing techniques (HTS), offer a promising alternative to classic methodologies, reducing time and costs. The aim of this study was to verify what kind of pressures affect these rivers and streams in the municipality of Paredes, develop plans to improve the sites under study, compare the EQR values obtained for both methodologies, and verify the effectiveness and reliability of DNA metabarcoding on diatoms in the bioassessment of these watercourses. Diatoms were sampled following WFD standards in 2 rivers and streams (21 samples) from the municipality of Paredes, in autumn 2019. All samples were submitted in parallel to the molecular and morphological methodologies. Molecular methodology started with eDNA extraction, followed by DNA sequencing (Illumina MiSeq) using 312 bp rbcL DNA barcode. In the Mothur software, the sequences obtained were analyzed producing Operational Taxonomic Units (OTUs) taxonomically assigned to the R-Syst::diatom reference library. Inventories of diatom communities revealed only 24.2% of the species were common among methodologies, which is explained by the incompleteness of reference library. Regarding the EQR values, the results indicated a good correlation between morphological and molecular methods. PCO analysis revealed a gradient of quality classification in both methodologies, supporting the hypothesis that DNA metabarcoding may be a good option for ecological quality assessment. However, there is work to be done so that this new methodology can integrate in WFD standards. Biological, physical-chemical and hydromorphological data showed negative impacts on watercourses in urbanized and agricultural areas, requiring the implementation of measures and awareness of the population for the dangers involved.