Hybridization between two bitterling fish species in their sympatric range and a river where one species is native and the other is introduced

Abstract

The distributions of two bitterling fish (subfamily: Acheilognathinae), Tanakia lanceolata and T. limbata, overlap in western Japan. Acheilognathinae fish lay their eggs in the gills of freshwater bivalves, and the early juvenile stage develops in the gills. Populations of freshwater bivalves are declining worldwide, which has limited the number of spawning substrate for bitterlings. T. limbata has been artificially introduced to some rivers in Ehime, Japan, where it coexists with native T. lanceolata, and some hybrids have been observed. We collected both species from several sites in western Japan, and from the Kunichi River system in Ehime, and analyzed genetic population structure based on six microsatellite loci and sequences of the mitochondrial cytochrome b gene. Structure analysis identified three genetically distinct populations: T. lanceolata, T. limbata “West Kyushu”, and T. limbata “Setouchi”. Two clades of T. limbata were also supported by molecular phylogenetic analyses based on cytochrome b. Hybrids in Ehime originated mostly from interbreeding between male T. lanceolata and female T. limbata “West Kyushu”, and made up 10.2% of all collected fish, suggesting that hybrids occurred frequently between females of colonizing species and males of native species. On the other hand, interspecific hybrids were detected at rates of 40.0%, 20.0%, and 17.6% in the Ima River (Fukuoka), Midori River (Kumamoto), and Kase River (Saga), respectively, which are naturally sympatric regions. We found a few T. limbata “Setouchi” in the Midori and Kase Rivers, which were supposed to be introduced from other regions, coexisting with native T. limbata “West Kyushu”, and this cryptic invasion may have triggered the interspecific hybridization. These results suggest that artificial introduction of a fish species, a decline in the unionid population, and degradation of habitat have caused broad hybridization of bitterlings in western Japan.

Hybridization between an endangered freshwater fish and an introduced congeneric species and consequent genetic introgression

“…lanceolata and T . limbata on the Matsuyama plain [15,18]. We collected Tanakia individuals from a 50 m section along the river/streams at each site to limit negative effects on the endangered T .…”

“…lanceolata and T . limbata in different locations [15]. Hha I produced fragments of 91, 218, 252 and 369 bps for T .…”

“…In western Japan, Tanakia limbata has been artificially transplanted to some rivers outside its native range, where it generates invasive hybrids with the native congeneric species T . lanceolata [15]. Tanakia limbata was introduced to Ehime Prefecture in the 1970s [16] and now competes with two other bitterlings there, the native T .…”

“…lanceolata and the exotic R . ocellatus ocellatus , for the use of the same unionid species as breeding substrata [15]. Populations of the unionid mussels, Pronodularia japanensis and Nodularia douglasiae nipponensis , which serve as spawning sites for bitterlings on the Matsuyama Plain, have decreased rapidly over the past 30 years because of habitat degradation [17].…”

Novel genotyping system for distinguishing among native, non‐native and admixed individuals of rosy bitterling Rhodeus ocellatus subspecies

“…The first studies using molecular data were based on allozyme ( e.g ., Avise & Saunders, 1984; Konishi et al ., 2003), random amplified polymorphic DNA (RAPD) analysis ( e.g ., Elo et al ., 1997; Yamazaki et al ., 2005) or restriction fragment length polymorphism (RFLP; e.g ., Padhi & Mandal, 1997; Hashimoto et al ., 2010). More recent studies have used genotyping by microsatellite DNA ( e.g ., Uemura et al ., 2018; Bradbeer et al ., 2019), sequencing of mitochondrial DNA (mtDNA) and nuclear DNA (nDNA; Jang‐Liaw et al ., 2019) or single‐nucleotide polymorphisms (SNPs) ( e.g ., Amish et al ., 2012; Lamer et al ., 2015). These methods, however, are expensive and demand a large amount of effort.…”