Takahiko Mukai

A review of the research on introduced freshwater fishes: New perspectives, the need for research, and management implications

Although freshwater fishes have a long history of human-induced introduction, recent globalization has accelerated worldwide introduction events even more, and those introduced fish species are now perceived to be a major threat to ecosystems. Over the last two decades, numerous studies have been published on introduced fish species; however, it has been challenging for researchers to understand the magnitude of the impact and the underlying mechanism of invasions. Recently, new perspectives in understanding invasive freshwater fish biology have been presented in a number of studies, which can be largely attributed to advances in analytical techniques and also to a growing need for proactive analysis in management strategies. The aim of this paper is to summarize new ecological perspectives, the need for research, and/or management implications with emphasis on technological advances in, for example, statistics, molecular analysis, modeling techniques, and landscape analysis addressed under the following five categories: introduction pathways, predicting spatial patterns, biotic homogenization, hybridization, and control and eradication. The conservation of native fish fauna and the management of introduced fish species will benefit from combining these new perspectives with fundamental studies such as those on life history and population biology.

GEDIMAP: a database of genetic diversity for Japanese freshwater fishes

Genetic diversity is a key component of biodiversity, and thus represents important information for evolutionary and conservation biology. DNA sequence data are primary sources of information on genetic diversity and are now accumulated and managed in international databases, such as the DNA Data Bank of Japan (DDBJ), the European Molecular Biology Laboratory (EMBL) Nucleotide Sequence Database, and GenBank of the US National Center for Biotechnology Information (NCBI). Such databases provide basic information that is used in most areas of contemporary bioscience (Brunak et al. 2002). MitoFish (Nishida 2009), a DNA database devoted to the mitochondrial DNA (mtDNA) genomes of fish, supports research into fish biology, and contains links to other international DNA databases and general fish databases (e.g., FishBase; Froese and Pauly 2009). Genetic diversity and divergence within and among local populations within a species are primary elements of biodiversity at the genetic level and are of essential conceptual and practical importance (Avise 2004). Genetic data from population genetic and phylogeographic studies based on various genetic markers, especially mtDNA sequences, are increasingly being reported. However, the DNA databases mentioned above do not provide effective functions for accumulating and searching for population genetic data, which are based on allele frequencies in each population with geographical data. Thus, when researchers want to browse or reuse data from previous studies, they typically need to reconstruct such a dataset by combining information described in original papers with that deposited in the DNA databases. Further, probably because of the lack of a platform for accumulating such information, a significant proportion of the papers do not provide fundamental information for reconstructing the population genetic data analyzed in those papers. Thus, it is difficult for anyone to use these valuable data secondarily. This situation impedes evolutionary and biogeographic research and applications in biodiversity conservation based on comprehensive population genetic data, and it leads to a waste of human, monetary, and wildlife resources. To improve this situation, we have constructed a comprehensive database for genetic diversity in Japanese freshwater fish. Freshwater fish are good targets for studying evolution and biogeography because of their restricted dispersal ability and conspicuous intraspecific divergence (Avise 2000; Watanabe et al. 2006). Research in this area will benefit from accumulated population genetic data. Here, we outline and describe the application of the database. ‘‘GEDIMAP’’ (genetic diversity and distribution map) is a public database primarily for mtDNA genetic diversity data within populations of freshwater fish in and around Japan (URL: http://gedimap.zool.kyoto-u.ac.jp/). The beta version was released in January 2008, and the current K. Watanabe (&) R. Kakioka K. Tominaga Department of Zoology, Division of Biological Science, Graduate School of Science, Kyoto University, KitashirakawaOiwakecho, Sakyo-ku, Kyoto 606-8502, Japan e-mail: watanak@terra.zool.kyoto-u.ac.jp

Genetic Relationships of the Genus Tridentiger (Pisces, Gobiidae) Based on Allozyme Polymorphism

Abstract The genetic relationships and taxonomic status of 7 taxa belonging to the genus Tridentiger (Pisces, Gobiidae) were investigated by means of analysis of allozymic variation at 14 loci. The results suggest that the two taxa “T. obscurus” and “T. brevispinis” which are sympatric and morphologically similar are reproductively isolated and are highly divergent from each other (the genetic distance values are 0.501-0,707). It is also suggested that “T. brevispinis” and “T. kuroiwae” are genetically different enough from each other to deserve subspecies at least. The other 4 taxa, “T. barbatus”, “T. nudicervicus”, “T. trigonocephalus” and “T. bifasciatus”, are genetically divergent each and are considered to be 4 biological species. A dendrogram showing the phylogenetic relationships of the 7 taxa was constructed from the genetic distances.

Evaluating the potential for invasion by alien freshwater fishes in northern Kyushu Island, Japan, using the Fish Invasiveness Scoring Kit

The potential invasiveness of 28 freshwater fishes in northern Kyushu Island, Japan, was evaluated using the Fish Invasiveness Scoring Kit (FISK). The five co-authors scored the level of invasiveness for each species and calculated the total FISK scores; the maximum and minimum scores were then eliminated, and the mean of the remaining three scores was used as the final score for each species. The mean scores ranged from 11.0 (Hypomesus nipponensis) to 31.0 (Cyprinus carpio). The receiver operating characteristic curve indicated that the threshold value between fishes that present a high risk of invasion and the other species were 19.8.

Genetic Differentiation of the Gobies Gymnogobius castaneus and G. taranetzi in the Region Surrounding the Sea of Japan as Inferred from a Mitochondrial Gene Genealogy

Abstract The phylogenetic relationships between gobies of the genus Gymnogobius were analyzed using mitochondrial cytochrome b gene sequences, focusing on the species currently classified as G. taranetzi and G. castaneus that occur in Japan, South Korea, and Russia. Gobies of the two species collected at 12 localities in Japan, South Korea, and Russia formed a monophyletic clade (called the “castaneus species complex” here) with G. breunigii as the sister clade. Within the species complex, six lineages were recognized: (L1) G. castaneus from the Akigawa River, Tokyo, Japan; (L2) G. castaneus from Yuza, Yamagata, Japan; (L3) G. taranetzi from Russia and South Korea; (L4) G. castaneus from the Tonegawa River, Chiba, Japan; (L5a) G. taranetzi from Shimane, Japan; and (L5b) G. castaneus + G. taranetzi from the Japan Sea coast of northern Japan. The two local lineages of G. castaneus (L1 and L2) are highly divergent from the others. The Japanese populations of G. taranetzi have diverged from the continental G. taranetzi populations, while one mitochondrial lineage (L5b) is shared with G. castaneus of northeast Japan. Therefore, the current species G. taranetzi and G. castaneus as defined morphologically are polyphyletic, necessitating a taxonomic revision. The genetic differentiation of isolated local lineages and the evolution of taranetzi- and castaneus-type gobies have likely occurred repeatedly in brackish/freshwater habitats around the Sea of Japan. We discussed the time of divergence for these gobies based on a tree with the molecular clock assumption.

Genetic and geographical differentiation of Pandaka gobies in Japan

The mitochondrial DNA (mtDNA) phylogeny of Japanese Pandaka species (Perciformes: Gobiidae) was inferred from partial nucleotide sequences of the mitochondrial 12S and 16S rRNA genes (1083 bp). The resultant mtDNA tree showed two major clades (clade I and clade II), which were inconsistent with the present taxonomic classification. One of the major clades was further divided into two geographical groups, distributed on the Japanese Major Islands (clade I-A) and from Amami-oshima Island to Iriomote Island (clade I-B). The mtDNA haplotypes in clade II were found only on Iriomote Island. The mtDNA divergences in clade I indicated that the Japanese Major Island (clade I-A) and Ryukyu (clade I-B) groups have been geographically isolated from each other for millions of years, based on the putative molecular divergence rate. The geographical distributions of mtDNA haplotypes in clade I-A and clade I-B also suggested that Pandaka gobies had not dispersed to distant offshore islands, indicating that their geographical differentiation may be closely associated with the geological history of the Japanese and Ryukyu Archipelagos.

Phylogeny, hybridization, and life history evolution of Rhinogobius gobies in Japan, inferred from multiple nuclear gene sequences.

Rhinogobius fishes (Gobiidae) are distributed widely in East and Southeast Asia, and represent the most species-rich group of freshwater gobies with diversified life histories (i.e., amphidromous, fluvial, and lentic). To reveal their phylogenetic relationships and life history evolution patterns, we sequenced six nuclear and three mitochondrial DNA (mtDNA) loci from 18 species, mainly from the mainland of Japan and the Ryukyu Archipelago. Our phylogenetic tree based on nuclear genes resolved three major clades, including several distinct subclades. The mtDNA and nuclear DNA phylogenies showed large discordance, which strongly suggested mitochondrial introgression through large-scale interspecific hybridization in these regions. On the basis of the molecular dating using geological data as calibration points, the hybridization occurred in the early to middle Pleistocene. Reconstruction of the ancestral states of life history traits based on nuclear DNA phylogeny suggests that the evolutionary change from amphidromous to freshwater life, accompanied by egg size change, occurred independently in at least three lineages. One of these lineages showed two life history alterations, i.e., from amphidromous (small egg) to fluvial (large egg) to lentic (small egg). Although more inclusive analysis using species outside Japan should be further conducted, the present results suggest the importance of the life history evolution associated with high adaptability to freshwater environments in the remarkable species diversification in this group. Such life history divergences may have contributed to the development of reproductive isolation.

Intraspecific variation in the mitochondrial genome among local populations of Medaka Oryzias latipes

The draft genome data of Medaka Oryzias latipes shows that it has distinct intraspecific genetic variation. To survey the genetic variations contributing to environmental adaptation, we focused on the mitochondrial DNA (mtDNA). The complete mtDNA sequences of Medaka were compared among 8 local population stocks and 4 inbred strains established from genetically divergent groups. Inbred strain HSOK, derived from the Eastern Korean group of Medaka, has a mitochondrial gene order that was distinct from other Medaka groups. Phylogenetic trees based on the mitochondrial genome sequences indicated that the mitogenome from the Shanghai stock (China) and HSOK strain were highly diverged from Japanese Medaka, and that the Japanese Medaka mitogenome was diverged into two groups; this result was fully consistent with those of the previous study using mtDNA-encode gene sequences. Among tRNA genes, the most divergent was the tRNA(Thr) gene as reported in humans previously. The number of tandemly repeated 11 nucleotide units in the Medaka mtDNA control region (CR) varied greatly among local populations. The number of repeats was more variable in the Northern Japanese group (10-34) than in the Southern group (7-12), while two other Oryzias species, inhabiting tropical regions, had no repeats. A comprehensive comparison between the number of repeat units and meteorological data indicated that the number of repeats correlated to the index data of a cold environment and seasonal climatic change. In cold (5 degrees C) acclimated fish, the mRNA levels varied among mitochondria coding genes. mRNA of the cytochrome oxidase subunit I gene in some local stocks was induced by cold temperature and seemed to be correlated with the number of repeated sequences in the CR. This study revealed that the repeated sequences in the mtDNA CR might function for mtDNA gene expression and that the number of tandem repeats in Medaka mtDNA is likely related to adaptation to a harsh habitat.

Natural hybridization and gene flow between two Tridentiger gobies in Lake Hinuma (Ibaraki Prefecture, Japan)

Of 851 specimens ofTridentiger obscurus andt. brevispinis collected from Lake Hinuma (Ibaraki Prefecture, Japan) from July 1996 to February 1998, 49 (5.8%) comprised F1 hybrids and backcross progeny of the two species. Since the mitochondrial DNA haplotypes of the F1 hybrids reflected those ofT. brevispinis, most instances of hybridization are thought to have occurred between maleT. obscurus and femaleT. brevispinis. Although both allozyme and mitochondrial DNA analyses indicated backcrossing and introgression of mitochondrial DNA, the frequency of backcross progeny was relatively low, suggesting the existence of a natural selection to backrossing.

Geographical distribution and genetic diversity of Gymnogobius sp. “Chokai-endemic species” (Perciformes: Gobiidae)

The geographical distribution and intraspecific genetic diversity of an endangered freshwater goby, Gymnogobius sp. “Chokai-endemic species”, was surveyed based on mitochondrial cytochrome b gene sequences in Yamagata Prefecture and adjacent areas on the island of Honshu, Japan. An extensive field survey showed that the species is distributed in 11 locations from Gosen, Niigata Prefecture to Yuza, Yamagata Prefecture. The species was not collected together with Gymnogobius sp. “widely distributed species” at each location, although the distribution of these two species overlaps. A maximum likelihood phylogenetic analysis showed that Gymnogobius sp. “Chokai-endemic species” is genetically divergent from other Gymnogobius castaneus species complex and the species consists of Niigata and Yamagata sub-clades. The result suggests that the Niigata and Yamagata regional groups can be distinguished from each other by their sequence divergence and the two groups should be treated as provisionally distinct “evolutionarily significant units”. The indices of intra-population genetic diversity were noticeably low, suggesting that each population has probably experienced bottleneck events. The geographical distance between the two populations in the Yamagata group, where the haplotypes are completely different from each other, was within 3 km. This suggests that habitat loss, fragmentation and isolation occurred in the Yamagata area due to some artificial cause. Conservation measures may be needed not only to protect individual habitats but also to promote genetic exchange among habitats in the Yamagata area.