Applicability of DNA-based identifications for the WFD-guided monitoring using macroinvertebrates: a large-scale DNA metabarcoding study for implementing routine ecological status assessments in Iberian rivers

Over the last decade, remarkable improvements have been made in the field of metabarcoding-based tools for routine ecological status assessments. However, important issues are yet to be solved to fulfil the European Water Framework Directive (WFD) requirements and standards. These limitations, which include problems related to e.g. the lack of a complete COI macroinvertebrate barcode database available for the Iberian Peninsula Murria 2020, or the scarce recovery of specific taxa due to DNA extraction and/or PCR amplification bias, are especially difficult to overcome for routine freshwater macroinvertebrate monitoring. For that purpose, a large-scale study is on going to test how metabarcoding data can infer existing macroinvertebrate morphotaxonomy-based biotic indexes and ecological status of Iberian rivers. Freshwater macroinvertebrates were selected as a Biological Quality Element and identified by using both morphological and metabarcoding approaches. The mitochondrial gene for cytochrome c oxidase subunit I (COI) was used as a DNA Barcode. Taxonomic coverage, taxonomic composition metrics and ecological status obtained from both approaches were analysed. Physical and chemical variables obtained during the routine biomonitoring, as well as other ecological parameters including biodiversity indexes, were also assessed. Multivariate data analysis of these environmental and biotic data obtained from both approaches were compared. Results seem to support the hypothesis Kuntke 2019 that the DNA-metabarcoding approach might deliver similar quality assessments results to the morphological approach, though some refinement must be done at the different steps of the process prior to establish a reliable procedure allowing the alternative use of both methods giving similar results for the ecological status classes marked by the WFD.

Interpopulation Variability in Dietary Traits of Invasive Bleak Alburnus alburnus (Actinopterygii, Cyprinidae) Across the Iberian Peninsula

The bleak Alburnus alburnus is native to most of Europe. This cyprinid fish is a successful invader in the Iberian Peninsula. No studies exist on its foraging strategies on a large scale for this ecoregion. The aim of the present study was to compare dietary traits of invasive bleak among the main Iberian rivers and a ‘reference’ native bleak population from France. Bleak were sampled during May–June 2019 from the Iberian Rivers Ebro, Tagus, Guadiana, Segura and Guadalquivir and the River Saône (France). Diptera larvae and zooplankton were common food categories in the River Saône. Insect nymphs were more important in the River Ebro. The intake of plant material was higher in the River Tagus. Flying insects were more consumed in the River Guadiana. Nektonic insects were important in the River Guadalquivir. Detritus was a frequent food category for all populations, in terms of occurrence and mass. Dietary parameters followed a unimodal response in relation to the latitudinal gradient, with the maximum values for the Tagus and Guadiana populations. Overall, results suggest that this wide interpopulation variability will contribute to the species’ successful establishment throughout Mediterranean Europe, which poses a serious risk to its highly valuable native fish fauna.

Behavioral Interactions and Trophic Overlap between Invasive Signal Crayfish Pacifastacus leniusculus (Decapoda, Astacidae) and Native Fishes in Iberian Rivers

The signal crayfish Pacifastacus leniusculus Dana, 1852 is a successful invasive species in the Iberian Peninsula. This is of particular conservation concern, as fish fauna is highly endemic and also threatened within this ecoregion. The aim of this study was to analyze behavioral interactions and trophic overlap between signal crayfish and native fishes in Iberian rivers (northern Spain). Video cameras were used to record fish “dominance/evasion” after spatial encounters with crayfish. Diet composition and isotopic signatures (δ13C and δ15N) were compared to evaluate the niche overlap. Fish were dominant in 25% of the encounters with juvenile crayfish, whereas this percentage was only 4% with adult crayfish. Observations showed a high fish “evasion” response for Pyrenean stone loach Barbatula quignardi (Bǎcescu-Meşter, 1967) (>30%). Dietary results showed a high trophic overlap between signal crayfish with the pelagic Pyrenean minnow Phoxinus bigerri Kottelat, 2007 and the benthic loach. However, the isotopic niche overlap was low, with brown trout Salmo trutta L., 1758 showing the highest area (only 0.1 ‰2). Overall, our findings suggest that interferences may occur with native species for food (i.e., benthic invertebrates). Consequently, measures should be applied to control invasive crayfish in Iberian rivers.

Inter-population variability in growth and reproduction of invasive bleak Alburnus alburnus (Linnaeus, 1758) across the Iberian Peninsula

The native European freshwater cyprinid fish, common bleak Alburnus alburnus (Linnaeus, 1758), is an invasive non-native fish in the Iberian Peninsula, where it threatens the valuable endemic fish fauna. Despite the bleak’s invasive status, there is a dearth of studies on the biological traits of non-native populations in Europe’s Mediterranean region, and the present study aimed to compare bleak growth and reproductive traits across the principal rivers of Iberia with those from a native ‘reference’ bleak population in France. Non-native bleak from the River Tagus had the highest back-calculated total lengths (TLs), growth rate and body condition, but the lowest reproductive investment and smallest egg size of all studied populations. Whereas, these latter two traits were the highest in the River Ebro, where fecundity was the lowest for all Iberian rivers. The youngest age and the smallest TL at maturity were observed in the River Ebro. The River Segura had the lowest back-calculated TLs and growth rate, the highest fecundity and proportion of females, and the oldest age at maturity. Population traits of bleak in the River Saône (the native ‘reference’ population) were generally different from those in Iberian rivers, except for the River Ebro. This wide inter-population variability contributes to the species’ successful establishment in novel habitats.