Browsing by Author "Martins, Filipa M.S."
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- Have the cake and eat it: optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoringPublication . Martins, Filipa M.S.; Galhardo, Mafalda; Filipe, Ana Filipa; Teixeira, Amílcar; Pinheiro, Paulo; Paupério, Joana; Alves, Paulo C.; Beja, PedroDNA metabarcoding can contribute to improving cost‐effectiveness and accuracy of biological assessments of aquatic ecosystems, but significant optimization and standardization efforts are still required to mainstream its application into biomonitoring programmes. In assessments based on freshwater macroinvertebrates, a key challenge is that DNA is often extracted from cleaned, sorted and homogenized bulk samples, which is time‐consuming and may be incompatible with sample preservation requirements of regulatory agencies. Here, we optimize and evaluate metabarcoding procedures based on DNA recovered from 96% ethanol used to preserve field samples and thus including potential PCR inhibitors and nontarget organisms. We sampled macroinvertebrates at five sites and subsampled the preservative ethanol at 1 to 14 days thereafter. DNA was extracted using column‐based enzymatic (TISSUE) or mechanic (SOIL) protocols, or with a new magnetic‐based enzymatic protocol (BEAD), and a 313‐bp COI fragment was amplified. Metabarcoding detected at least 200 macroinvertebrate taxa, including most taxa detected through morphology and for which there was a reference barcode. Better results were obtained with BEAD than SOIL or TISSUE, and with subsamples taken 7–14 than 1–7 days after sampling, in terms of DNA concentration and integrity, taxa diversity and matching between metabarcoding and morphology. Most variation in community composition was explained by differences among sites, with small but significant contributions of subsampling day and extraction method, and negligible contributions of extraction and PCR replication. Our methods enhance reliability of preservative ethanol as a potential source of DNA for macroinvertebrate metabarcoding, with a strong potential application in freshwater biomonitoring.
- Origin and history of Phoxinus (Cyprinidae) introductions in the Douro Basin (Iberian Peninsula): an update inferred from genetic dataPublication . Garcia-Raventós, Aina; Martins, Filipa M.S.; Teixeira, Amílcar; Sousa, Ronaldo; Froufe, Elsa; Varandas, Simone; Lopes-Lima, Manuel; Beja, Pedro; Filipe, Ana FilipaThe number of non-native freshwater fishes in the Iberian Peninsula has been greatly increasing. In this study, individuals of the genus Phoxinus were detected in 18 out of 138 stream sites sampled across the Douro Basin in 2017 and 2018. A total of 26 individuals were barcoded using partial cytochrome c oxidase subunit I (COI) and cytochrome b (cytb) genes for species identification and determination of geographical origin. Molecular data provided the first record of a second Phoxinus species in western Douro (Portugal, Iberian Peninsula), with haplotypes closely matching those found in the Charente River (southern France). This species is suspected to be a recent introduction associated with the use of minnows as live bait by freshwater anglers, which was facilitated by human movements between France and Portugal. Individuals from watercourses in eastern Douro (Spain) were genetically assigned to Phoxinus bigerri, an introduced species previously known for that region, which confirms reports of introduction events from Ebro to Douro Basin probably also related to freshwater angling and facilitated by geographic proximity. The potential ecological impacts of this genus in the region are unknown and need further investigation.