Species-specific markers provide molecular genetic evidence for natural introgression of bullhead catfishes in Hungary

Abstract

Since three bullhead catfish species were introduced to Europe in the late 19th century, they have spread to most European countries. In Hungary, the brown bullhead (Ameiurus nebulosus) was more widespread in the 1970s–1980s, but the black bullhead (Ameiurus melas) has gradually supplanted since their second introduction in 1980. The introgressive hybridization of the two species has been presumed based on morphological examinations, but it has not previously been supported by genetic evidence. In this study, 11 different Hungarian habitats were screened with a new species-specific nuclear genetic, duplex PCR based, marker system to distinguish the introduced catfish species, Ameiurus nebulosus, Ameiurus melas, and Ameiurus natalis, as well as the hybrids of the first two. More than 460 specimens were analyzed using the above markers and additional mitochondrial sequence analyses were also conducted on >25% of the individuals from each habitat sampled. The results showed that only 7.9% of the specimens from two habitats belonged to Ameiurus nebulosus, and 92.1% were classified as Ameiurus melas of all habitats, whereas the presence of Ameiurus natalis was not detected. Two specimens (>0.4%) showed the presence of both nuclear genomes and they were identified as hybrids of Ameiurus melas and Ameiurus nebulosus. An additional two individuals showed contradicting results from the nuclear and mitochondrial assays as a sign of a possible footprint of introgressive hybridization that might have happened two or more generations before. Surprisingly, the level of hybridization was much smaller than expected based on the analyses of the North American continent’s indigenous stock from the hybrid zones. This phenomenon has been observed in several invasive fish species and it is regarded as an added level of complexity in the management of their rapid adaptation.

Morphometric and genetic evidence for cryptic diversity inGyrodactylus(Monogenea) infecting non-native European populations ofAmeiurus nebulosusandA. melas

“…Following Béres et al . (2017), Cytochrome b gene was amplified in the volume of 12 μ L reaction using PCR Multiplex kit (Qiagen) with primers CytB_L_14724F (5´-GACTTGAAAACCCACCGTTG-3´) and Cytbasa_R (5´-GAAGGCGGTCATCATAACTAG-3´). Cycling conditions for PCR were 95 °C at 15 min, then 35 cycles of 95 °C at 30 s, 56 °C at 20 s, 72 °C at 60 s. All PCR products were sequenced with specific primers (Eurofins, Germany), sequence data were analysed using Geneious ® 9.0.5. and compared with sequences from GenBank ( A. nebulosus -AY184271 and A. melas -AY184263).…”

“…Brown bullhead, A. nebulosus (LeSueur, 1819) is the most widespread; only a few decades after introduction, the species spread to most European countries(Rutkayová et al ., 2013). Black bullhead, A. melas (Rafinesque, 1820) has expanded its European range in recent decades (Béres et al ., 2017), while yellow bullhead, A. natalis (LeSueur, 1819) appears limited to Italy (Rutkayová et al ., 2013). The occurrence of A. nebulosus in the Czech Republic was first documented in 1890 (Hanel and Lusk, 2005) with its maximum population density occurring predominantly in the Elbe River basin during the mid-20th century.…”

“…Species identifications based on potentially ambiguous characters, for both A. melas and A. nebulosus , has resulted in uncertainty regarding the occurrence of these species in some European countries (Rutkayová et al ., 2013). Recently, molecular methods have contributed to the rapid identification of both species and their potential hybrids (Walter et al ., 2014; Béres et al ., 2017).…”