Environmental DNA Assay for the Detection of the American Bullfrog (Lithobates catesbeianus) in the Early Stages of the Invasion in the Ebre Delta.

The American bullfrog (Lithobates catesbeianus) is considered to be one of the most harmful invasive species. In the Iberian Peninsula, this species had been cited occasionally until the year 2018, when L. catesbeianus appeared in the Ebre Delta, and, for the first time, it started breeding in a territory of the Peninsula. Using environmental DNA (eDNA) analysis and visual surveys, the American bullfrog invasion in the Ebre Delta was monitored across two consecutive years (2019-2020). No specimens were observed in 2019, and results for the eDNA survey also failed to detect this species in the Delta. In 2020, two individuals were captured and, under the most conservative criteria to constrain the number of positive detections, eDNA analyses detected the presence of the American bullfrog in at least five locations. Performing an eDNA assay yielded a higher sensitivity with a lower sampling effort than traditional methods. Although the American bullfrog does not appear to still be well-established in the Ebre Delta, only a few bullfrog individuals could be enough for their establishment in suitable habitats. In this context, eDNA assays are essential tools to facilitate the detection, control, and eradication of this species in the first stage of the invasion process.

Effects of water pollution and river fragmentation on population genetic structure of invasive mosquitofish.

We analyzed variation at the GPI-2 locus and eleven microsatellite loci of eastern mosquitofish Gambusia holbrooki populations introduced to the Ebro River (Spain), sampling above and below a dam (Flix Reservoir) where severe chronic pollution has been well documented. Allele frequency changes at the GPI-2 locus in the sites nearest to the polluted sediments agree with previous results from studies in mercury-exposed populations of this highly invasive fish. Genetic distinction of the mosquitofish collected close to the polluted sediments was detected at the GPI locus but also at the presumptive neutral microsatellite loci. Recent migration rates estimated from microsatellites indicated that around 30% of fish collected in a specific location were immigrants from upstream and downstream sources. Such high migration rates probably contribute to the mosquitofish's invasive success and suggest that the consequences on the mosquitofish regional genetic structured of high levels of water toxicants could be mediated by immigration from other sites, but the effect of pollutants on local diversity might be higher than observed here.

Gene flow and maintenance of genetic diversity in invasive mosquitofish (Gambusia holbrooki).

Genetic analyses contribute to studies of biological invasions by mapping the origin and dispersal patterns of invasive species occupying new territories. Using microsatellite loci, we assessed the genetic diversity and spatial population structure of mosquitofish (Gambusia holbrooki) that had invaded Spanish watersheds, along with the American locations close to the suspected potential source populations. Mosquitofish populations from the Spanish streams that were studied had similar levels of genetic diversity to the American samples; therefore, these populations did not appear to have undergone substantial losses of genetic diversity during the invasion process. Population structure analyses indicated that the Spanish populations fell into four main clusters, which were primarily associated with hydrography. Dispersal patterns indicated that local populations were highly connected upstream and downstream through active dispersal, with an average of 21.5% fish from other locations in each population. After initially introducing fish to one location in a given basin, such dispersal potential might contribute to the spread and colonization of suitable habitats throughout the entire river basin. The two-dimension isolation-by-distance pattern here obtained, indicated that the human-mediated translocation of mosquitofish among the three study basins is a regular occurrence. Overall, both phenomena, high natural dispersal and human translocation, favor gene flow among river basins and the retention of high genetic diversity, which might help retain the invasive potential of mosquitofish populations.