Genetic variability of minnows and loaches in rivers and high mountain lakes from the Pyrenees and Italian Alps

Abstract

[eng] The invasion of alien species is a major global threat to ecosystems, with significant impacts, especially on freshwater ecosystems. The species diversity in freshwater ecosystems has significantly declined worldwide, and in Europe, freshwater fish represent one of the animal groups with the highest number of invasive species. Due to human activities, restrictions on geographical distribution have disappeared, and fish are being introduced into new and distant areas for recreational and other economic benefits. The purpose of this thesis was to identify and assess the distribution of two invasive fish introduced by human activities, infer the spread process and introduction pathways. Through the use of various genetic analysis techniques, we aimed to characterize introduced individuals across a broad area and, beyond species identification, obtain information about inter-species hybridization and intra-species admixture. The first chapter aimed to accurately identify the species and assess the distribution of the genus Phoxinus in high-altitude lakes in the Pyrenees and the Italian Alps. The goal was to understand the introduction pathways of these species in order to shed light on their presence and distribution. Mitochondrial DNA (COI and Cytb; mtDNA) was sequenced in 201 individuals sampled from 53 sites including alpine lakes and adjacent water bodies. This data was utilized for species identification through phylogenetic tree analysis and for inferring introduction pathways using haplotype networks. P. dragarum has been discovered in its native habitat, including the Garonne basin, and in lakes to the east of the Pyrenees, and rivers in Catalonia. P. bigerri was primarily found in its native range, the Adour basin, and lakes to the west of the Pyrenees, suggesting it has been introduced. Lower species diversity of genus Phoxinus was observed in areas where fishing is prohibited, indicating that most introductions are associated with the use of live bait by anglers. The regulation of fishing has been confirmed as an effective measure to prevent the introduction of invasive species. The second chapter involved a more detailed using a total of 890 individuals from 62 locations, and nuclear DNA (microsatellite) sequencing was conducted. With the analyses of microsatellites (nuclear DNA; nDNA) we identified various lineages, including information on interspecies and intraspecies hybridization. 2 The nDNA results generally aligned with mtDNA, but some differences were noted. Individuals from Catalonia identified as P. septimaniae based on mtDNA were identified as P. dragarum or P. bigerri based on nDNA, suggesting that the individuals known as P. septimaniae in Catalonia are hybrids. Contrary to the assumption that there would be a high rate of intraspecies hybridization in isolated lake environments, a low rate was observed. An interesting observation was the recording of nuclear and mitochondrial three-way hybrids of P. septemaniae (mtDNA) and P. dragarum x P. bigerri (nDNA) in the River Tenes. The third chapter focused on the introduced species of loaches in the Catalonia, particularly in lowland rivers with greater human activity and accessibility. Individuals of Barbatula, Cobitis, and Misgurnus/Paramisgurnus were collected from 21 locations and were sequenced for COI and RAG-1. These data were utilized for species identification through phylogenetic tree analysis and introduction pathway inference using haplotype networks. We identified at least five non-native fish species (C. bilineata, M. anguillicaudatus, M, bipartitus, P. dabryanus, and an undescribed Barbatula species) in the Catalonia, and most of them were spreading in their distribution. The characteristics of invasive species were evolving over time after introduction, and the established invasive species in Catalonia shifted from being large and long-lived fish species used for culture or game fishing to small, benthic fish species used as live bait or for ornamental purposes.