Nobuaki Nagata

Diversification in a fluctuating island setting: rapid radiation of Ohomopterus ground beetles in the Japanese Islands

The Japanese Islands have been largely isolated from the East Asian mainland since the Early Pleistocene, allowing the diversification of endemic lineages. Here, we explore speciation rates and historical biogeography of the ground beetles of the subgenus Ohomopterus (genus Carabus) based on nuclear and mitochondrial gene sequences. Ohomopterus diverged into 15 species during the Pleistocene. The speciation rate was 1.92 Ma–1 and was particularly fast (2.37 Ma–1) in a group with highly divergent genitalia. Speciation occurred almost solely within Honshu, the largest island with complex geography. Species diversity is highest in central Honshu, where closely related species occur parapatrically and different-sized species co-occur. Range expansion of some species in the past has resulted in such species assemblages. Introgressive hybridization, at least for mitochondrial DNA, has occurred repeatedly between species in contact, but has not greatly disturbed species distinctness. Small-island populations of some species were separated from main-island populations only after the last glacial (or the last interglacial) period, indicating that island isolation had little role in speciation. Thus, the speciation and formation of the Ohomopterus assemblage occurred despite frequent opportunities for secondary contact and hybridization and the lack of persistent isolation. This radiation was achieved without substantial ecological differentiation, but with marked differentiation in mechanical agents of reproductive isolation (body size and genital morphology).

Application of RAD-based phylogenetics to complex relationships among variously related taxa in a species flock

Restriction site-associated DNA (RAD) sequences from entire genomes can be used to resolve complex phylogenetic problems. However, the processed data matrix varies depending on the strategies used to determine orthologous loci and to filter loci according to the number of taxa with sequence data for the loci, and often contains plenty of missing data. To explore the utility of RAD sequences for elucidating the phylogenetics of variously related species, we conducted RAD sequencing for the Ohomopterus ground beetles and attempted maximum-likelihood phylogenetic analyses using 42 data matrices ranging from 1.6×10(4) to 8.1×10(6) base pairs, with 11-72% missing data. We demonstrate that robust phylogenetic trees, in terms of bootstrap values, do not necessarily result from larger data matrices, as previously suggested. Robust trees for distantly related and closely related taxa resulted from different data matrices, and topologically different robust trees for distantly related taxa resulted from various data matrices. For closely related taxa, moderately large data matrices strongly supported a topology that is incompatible with morphological evidence, possibly due to the effect of introgressive hybridization. Practically, exploring variously prepared data matrices is an effective way to propose important alternative phylogenetic hypotheses for this study group.

Mechanical barriers to introgressive hybridization revealed by mitochondrial introgression patterns in Ohomopterus ground beetle assemblages

To reveal the role of diverged body size and genital morphology in reproductive isolation among closely related species, we examined patterns of, and factors limiting, introgressive hybridization between sympatric Ohomopterus ground beetles in central Japan using mitochondrial NADH dehydrogenase subunit 5 (ND5) gene sequences. We sampled 17 local assemblages that consisted of two to five species and estimated levels of interspecific gene flow using the genetic distance, DA, and maximum‐likelihood estimates of gene flow. Sharing of haplotypes or haplotype lineages was detected between six of seven species that occurred in the study areas, indicating mitochondrial introgression. The intensity and direction of mitochondrial gene flow were variable among species pairs. To determine the factors affecting introgression patterns, we tested the relationships between interspecific DA and five independent variables: difference in body size, difference in genital size, phylogenetic relatedness (nuclear gene sequence divergence), habitat difference, and species richness of the assemblage. Body and genital size differences contributed significantly to preventing gene flow. Thus, mechanical isolation mechanisms reduce the chance of introgressive hybridization between closely related species. Our results highlight the role of morphological divergence in speciation and assemblage formation processes through mechanical isolation.

Divergence of ovipositor length and egg shape in a brood parasitic bitterling fish through the use of different mussel hosts.

Bitterling fishes deposit their eggs on the gills of living mussels using a long ovipositor. We examined whether ovipositor length (OL) and egg shape correlated with differences in host mussel species in the family Unionidae among populations of the tabira bitterling (Acheilognathus tabira) in Japan. Bitterling populations that use mussels in the sub‐family Anodontinae possessed longer ovipositors and more elongated eggs than those using mussels of Unioninae, as expected from the difference in host size between the sub‐families (anodontine mussels are larger than unionine mussels). Based on a robust phylogeny of A. tabira populations, we demonstrated that the evolution of both OL and egg shape were correlated with host differences, but not with each other, suggesting that these traits have been selected for independently. Our study demonstrates how adaptive traits for brood parasitism may diverge with host shift due to different host availability and/or interspecific competition for hosts.

Phylogeography and Introgressive Hybridization of the Ground Beetle Carabus yamato in Japan Based on Mitochondrial Gene Sequences

Abstract To study the phylogeography of the ground beetle Carabus yamato in Japan, we compared 1,020-bp sequences of the mitochondrial NADH dehydrogenase subunit 5 (ND5) gene from 373 specimens from 37 localities with those of three parapatric species (C. albrechti, C. kimurai, and C. japonicus) that might share mitochondrial lineages with C. yamato through introgressive hybridization. We found 81 haplotypes from C. yamato. Of these, 17 haplotypes were considered to be of an introgressed lineage from C. albrechti, based on the phylogeny and geographic distribution. In addition, one haplotype of C. kimurai was likely an introgressant from C. yamato. Putative introgression events among the four species were restricted to these two directional cases. We analyzed the phylogeography of C. yamato using nested clade phylogeographical analysis and population genetic parameters. The mitochondrial lineages of C. yamato were estimated to have diverged no more than approximately 1.12 million years ago, implying that the estimated historical events occurred after the Early Pleistocene. Carabus yamato was inferred to have experienced a contraction of its distribution range, followed by recent range expansion. Populations in the western and eastern regions, segregated by Ise Bay and the Nobi and Okazaki Plains, diverged in the mitochondrial clades. The northern and most western populations possessed one clade only (except an introgressed lineage), whereas eastern and some southwestern populations possessed several diverged clades, which were considered to be ancestral; these populations may have been associated with refugia during glacial periods.

Factors determining the direction of ecological specialization in snail-feeding carabid beetles.

A stout–slender dimorphism in body shape is observed among carabid beetles of the subtribe Carabina, which feed on land snails. We hypothesized that this dimorphism has resulted from divergent ecological specialization for feeding on different‐sized land snails. Therefore, we examined whether the geographic variation in the body shape of Damaster blaptoides, a representative snail‐feeding species in Japan, is correlated with the size of Euhadra, a genus of land snails frequently consumed by D. blaptoides. An analysis of beetle specimens from the whole distribution area of D. blaptoides determined that more slender beetle populations occurred in localities harboring larger snails, whereas more stout beetles inhabited localities harboring smaller snails. This pattern could be adaptive because slender beetles exhibit high feeding performance for large snails by inserting their heads into the shells, whereas stout beetles do so for small snails by crushing the shells. The D. blaptoides populations showed a clear genetic isolation‐by‐distance pattern, which could be effective in promoting such local adaptation. Thus, food resources as well as geographic isolation may have promoted adaptive divergence of external morphology in the snail‐feeding carabid beetles.

Antagonistic selection factors induce a continuous population divergence in a polymorphism

Understanding the relative importance of selection and stochastic factors in population divergence of adaptive traits is a classical topic in evolutionary biology. However, it is difficult to separate these factors and detect the effects of selection when two or more contrasting selective factors are simultaneously acting on a single locus. In the damselfly Ischnura senegalensis, females exhibit color dimorphism and morph frequencies change geographically. We here evaluated the role of selection and stochastic factors in population divergence of morph frequencies by comparing the divergences in color locus and neutral loci. Comparisons between population pairwise FST for neutral loci and for the color locus did not detect any stochastic factors affecting color locus. Although comparison between population divergence in color and neutral loci using all populations detected only divergent selection, we detected two antagonistic selective factors acting on the color locus, that is, balancing and divergent selection, when considering geographical distance between populations. Our results suggest that a combination of two antagonistic selective factors, rather than stochastic factors, establishes the geographic cline in morph frequency in this system.

Historical divergence of mechanical isolation agents in the ground beetle Carabus arrowianus as revealed by phylogeographical analyses

In the carabid genus Carabus subgenus Ohomopterus, diverged body size and genital morphology serve as mechanical reproductive barriers. To elucidate the diverging process of body and genital sizes in Carabus arrowianus, which exhibits marked morphological diversity among geographical populations and may represent an early stage of speciation, we analysed a mitochondrial gene sequence for 1051 individuals from 63 populations and male morphology for 359 individuals from 47 populations. Two discrete morphological groups segregated by geographical barriers were distinguished, one of which possessed smaller bodies and shorter genitalia (S group) than the other (L group), which exhibited larger bodies and exaggerated genitalia. Genetic divergence between the two groups was significant but not large. Phylogeographical and population genetic analyses indicated that the L group was derived from the S group, and a coalescent simulation revealed that the two groups diverged during the latest middle Pleistocene (0.13 million years ago), with a much larger effective population size in the L group than the S group. Because the body size divergence could not be explained by adaptation to climatic conditions and genital morphology is considered to be subject to sexual selection, we postulated that a population division and colonization in favourable habitats caused by the Pleistocene climatic and geographical change might facilitate natural and sexual selection for enlarged body and genital sizes in the L group.

Relative importance of habitat use, range expansion, and speciation in local species diversity of Anolis lizards in Cuba

Variations in species richness of local assemblages may be explained by local ecological processes or large-scale evolutionary and biogeographical processes. In Anolis lizards, species with different ecomorphs can coexist by occupying different niches. In addition, several species with the same ecomorph (e.g., trunk-ground) can coexist, and the number of trunk-ground anole species varies among local species assemblages. In this study, we assessed the importance of ecological interactions, number of speciation events, and range expansion for local and regional species diversity of these lizards. We examined the species richness and thermal microhabitat partitioning (considered to be a measure of ecological interaction) of 12 trunk-ground anole species in 11 local assemblages in Cuba. The results indicated that the phylogenetic structure of trunk-ground anole lizard assemblages was random. However, there was an overdispersion of preferences for thermal microhabitat use, which indicates that differences in microhabitat use are likely to occur within assemblages. We suggest that the number of speciation events within regions and the number of sympatrically coexisting species increases species richness at the local level. Migration appeared to be limited, leading to the range expansion of only three species with different thermal requirements. The thermal niches of species were conserved within Anolis allogus clade, whereas species within the Anolis homolechis and Anolis sagrei clades tended to change their thermal niches. Our results suggest that the species composition and richness in local assemblages could be explained by evolutionary history (the number of speciation events and limits to range expansion) and ecological processes (habitat partitioning). Of the ecological factors, the number of thermal (microhabitat use) and structural niches (e.g., vegetation) could limit the potential number of coexisting species within a local assemblage n

Factors restricting the range expansion of the invasive green anole Anolis carolinensis on Okinawa Island, Japan

Abstract The green anole Anolis carolinensis invaded the Ogasawara Islands in Japan, drove various native species to extinction, and its distribution expanded 14 years after initial establishment. A. carolinensis invaded Okinawa Island, but it has not expanded its distribution in more than 25 years, although its density is extremely high in the southern region. To determine whether A. carolinensis has the potential to expand its distribution on Okinawa Island, we performed phylogenetic analysis of mitochondrial ND2 DNA sequences to study the origin of A. carolinensis that invaded Okinawa Island. We further used a species distribution model (MaxEnt) based on the distribution of native populations in North America to identify ecologically suitable areas on Okinawa Island. Nucleotide sequence analysis shows that the invader A. carolinensis originated in the western part of the Gulf Coast and inland areas of the United States and that a portion of the anoles on Okinawa was not introduced via the Ogasawara Islands. The MaxEnt predictions indicate that most areas in Okinawa Island are suitable for A. carolinensis. Therefore, A. carolinensis may have the potential to expand its distribution in Okinawa Island. The predictions indicate that habitat suitability is high in areas of high annual mean temperature and urbanized areas. The values of precipitation in summer in the northern region of Okinawa Island were higher compared with those of North America, which reduced the habitat suitability in Okinawa Island. Adaptation to low temperatures, an increase in the mean temperature through global warming, and an increase in open environments through land development will likely expand the distribution of A. carolinensis in Okinawa Island. Therefore, we must continue to monitor the introduced populations and be alert to the possibility that city planning that increases open environments may cause their range to expand.