Cryptic species of the subgenus Morphnosoma Lutshnik (Coleoptera: Carabidae; genus Pterostichus) from Japan

Pterostichus thunbergi Morawitz and its related species of Japan are revised based on the structure of membranous parts of their genitalia for the first time, under the name of the thunbergi species group. This species group is monophyletic and is placed in the subgenus Morphnosoma Lutshnik. The monotypic subgenus Moritapterus Berlov, erected for P. thunbergi, is synonymized with Morphnosoma. Comparative study of the endophallus revealed that P. thunbergi michinoku Nakane is a junior synonym of P. habui and that four species should be recognized in this group. Three of them are sympatric with P. thunbergi and one of them, Pterostichus (Morphnosoma) robustistylis, is described as a new species. Molecular phylogeny based on the COI gene (mitochondrial [mt]DNA) supported its independence as a distinct species. The resultant trees of mtDNA and comparative study of morphology revealed that P. thunbergi is probably non‐monophyletic. In the present study we suggest the presence of further cryptic species of Morphnosoma in Japan.     

Utility of geometric morphometrics for inferring feeding habit from mouthpart morphology in insects: tests with larval Carabidae (Insecta: Coleoptera)

Feeding habits are important life‐history traits in animals; however, methods for their determination are not well established in many species. The larvae of the beetle family Carabidae are an example. The present study tested the utility of geometric morphometrics of mouthpart morphology to infer the feeding habits of carabid larvae. Using Pterostichus thunbergi as a model system, larval feeding habits were inferred using geometric morphometrics of mouthparts and the results were compared with those obtained from rearing experiments. The rearing experiments indicated that P. thunbergi larvae are carnivores that require snails as an essential part of the diet. Through geometric morphometrics, associations between mouthpart morphology and larval feeding habits were confirmed for species in which these two traits are known. A discriminant analysis using these associations classified P. thunbergi larvae as snail/slug feeders, which is a result compatible with the rearing experiments. Geometric morphometrics also revealed that morphological integration and ontogenetic shape change might play roles in the diversification of mouthpart morphology. Overall, these results demonstrate the utility of the geometric morphometrics of mouthparts to infer feeding habit and to clarify the mechanisms of mouthpart morphological diversification in the study group, and the results also serve as a basis for future studies of other insect groups.     

Patterns and processes in the evolutionary history of parrotfishes (Family Labridae)

Phylogenetic reconstruction of the evolutionary relationships among 61 of the 70 species of the parrotfish genera Chlorurus and Scarus (Family Labridae) based on mitochondrial and nuclear gene sequences retrieved 15 well‐supported clades with mid Pliocene/Pleistocene diversification. Twenty‐two reciprocally monophyletic sister‐species pairs were identified: 64% were allopatric, and the remainder were sympatric. Age of divergence was similar for allopatric and sympatric species pairs. Sympatric sister pairs displayed greater divergence in morphology, ecology, and sexually dimorphic colour patterns than did allopatric pairs, suggesting that both genetic drift in allopatric species pairs and ecologically adaptive divergence between members of sympatric pairs have played a role in diversification. Basal species typically have small geographical ranges and are restricted to geographically and ecologically peripheral reef habitats. We found little evidence that a single dominant process has driven diversification, nor did we detect a pattern of discrete, sequential stages of diversification in relation to habitat, ecology, and reproductive biology. The evolution of Chlorurus and Scarus has been complex, involving a number of speciation processes. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 107 , 529–557.     

Spatial and host related genomic variation in partially sympatric cactophagous moth species

Surveys of patterns of genetic variation in natural sympatric and allopatric populations of recently diverged species are necessary to understand the processes driving intra- and interspecific diversification. The South American moths Cactoblastis cactorum, Cactoblastis doddi and Cactoblastis bucyrus are specialized in the use of cacti as host plants. These species have partially different geographic ranges and differ in patterns of host plant use. However, there are areas that overlap, particularly, in northwestern Argentina, where they are sympatric. Using a combination of genome-wide SNPs and mitochondrial data we assessed intra and interspecific genetic variation and investigated the relative roles of geography and host plants on genetic divergence. We also searched for genetic footprints of hybridization between species. We identified three well delimited species and detected signs of hybridization in the area of sympatry. Our results supported a hypothetical scenario of allopatric speciation in the generalist C. cactorum and genetic interchange during secondary geographic contact with the pair of specialists C. bucyrus and C. doddi that probably speciated sympatrically. In both cases, adaptation to new host plants probably played an important role in speciation. The results also suggested the interplay of geography and host plant use as drivers of divergence and limiting gene flow at intra and interspecific levels.     

Comparative intra‐ and interspecific sexual organ reciprocity in four distylous Primula species in the Himalaya‐Hengduan Mountains

• Distyly is a mechanism promoting cross‐pollination within a balanced polymorphism. Numerous studies show that the degree of inter‐morph sexual organ reciprocity (SOR) within species relates to its pollen‐mediated gene flow. Similarly, a lower interspecific SOR should promote interspecific isolation when congeners are sympatric, co‐blooming and share pollinators. In this comparative study, we address the significance of SOR at both intra‐ and interspecific levels.
• Seventeen allopatric and eight sympatric populations representing four Primula species (P. anisodora, P. beesiana, P. bulleyana and P. poissonii) native to the Himalaya‐Hengduan Mountains were measured for eight floral traits in both long‐ and short‐styled morphs. GLMM and spatial overlap methods were used to compare intra‐ and interspecific SOR.
• While floral morphology differed among four Primula species, SOR within species was generally higher than between species, but in species pairs P. poissonii/P. anisodora and P. beesiana/P. bulleyana, the SOR was high at both intra‐ and interspecific levels. We did not detect a significant variation in intraspecific SOR or interspecific SOR when comparing allopatric versus sympatric populations for all species studied.
• As intraspecific SOR increased, disassortative mating may be promoted. As interspecific SOR decreased, interspecific isolation between co‐flowering species pairs also may increase. Hybridisation between congeners occurred when interspecific SOR increased in sympatric populations, as confirmed in two species pairs, P. poissonii/P. anisodora and P. beesiana/P. bulleyana.

     

Mitochondrial capture and incomplete lineage sorting in the diversification of balitorine loaches (Cypriniformes,Balitoridae) revealed by mitochondrial and nuclear genes

Tang, Q.‐Y., Liu, S.‐Q., Yu, D., Liu, H.‐Z. & Danley, P.D. (2012) Mitochondrial capture and incomplete lineage sorting in the diversification of balitorine loaches (Cypriniformes, Balitoridae) revealed by mitochondrial and nuclear genes. —Zoologica Scripta, 41, 233–247. Understanding the diversification of species is a central goal of evolutionary biological studies. One powerful tool to investigate the speciation process is molecular systematics. Here, we use molecular methods to investigate the evolution of balitorine loaches belonging to two genera, Lepturichthys and Jinshaia. Both genera contain only two species (Lepturichthys fimbriata, Lepturichthys dolichopterus and Jinshaia sinensis and Jinshaia abbreviata), all of which are endemic to China. These species share many morphological and ecological characters and exhibit overlapping distributions in the Upper Yangtze River. In this study, we used two mitochondrial genes (Cytb and COI) and one nuclear gene (RAG1) to investigate the phylogenetic relationships within and between these two genera. Phylogenetic analyses and network construction based on mitochondrial and nuclear genes consistently supported the monophyly of Jinshaia. In contrast, the mitochondrial and nuclear genes yielded conflicting results in Lepturichthys. The phylogenetic analyses of mitochondrial sequences identify two distinct Lepturichthys lineages, Lepturichthys A and Lepturichthys B. Lepturichthys A includes most of L. fimbriata individuals from the Upper Yangtze River and is the sister group to all Jinshaia species. Lepturichthys B consists of the remaining L. fimbriata individuals from the Upper and Middle Yangtze River, and all L. dolichopterus individuals from the Minjiang River in Southeastern China. However, the analysis of the nuclear sequence indicates that the genus Lepturichthys is monophyletic and is only distantly related to Jinshaia. This incongruence suggests that introgressive hybridization might have occurred between L. fimbriata (Lepturichthys A) and Jinshaia species. As a result of this hybridization event, L. fimbriata captured the mitochondrial genome of the sympatric Jinshaia species. This capture event appears to have occurred at least 1.74 million years ago. Additionally, L. fimbriata appears to be paraphyletic; the nuclear data indicated that L. dolichopterus forms a monophyletic clade nested within L. fimbriata. Because L. dolichopterus and L. fimbriata are allopatric and hybridization may not be possible, we suggest that the observed paraphyly of L. fimbriata is a product of incomplete lineage sorting. In addition, the reciprocal monophyly of J. sinensis and J. abbreviata could not be resolved. This may be the result of interspecific hybridization as these species occur sympatrically. However, incomplete lineage sorting may have caused the observed topology of the Jinshaia species. The data presented here illustrate the complex evolutionary history of the balitorine loach species: intergeneric hybridization and interspecific hybridization have likely occurred in this lineage. In addition, possible incomplete lineage sorting may further obscure the evolutionary history of this group. The complex relationships of the balitorine loaches provide a rich evolutionary system to study the creation of sympatric and sister species.     

Genomic footprints of an old affair: Single nucleotide polymorphism data reveal historical hybridization and the subsequent evolution of reproductive barriers in two recently diverged grasshoppers with partly overlapping distributions

Secondary contact in close relatives can result in hybridization and the admixture of previously isolated gene pools. However, after an initial period of hybridization, reproductive isolation can evolve through different processes and lead to the interruption of gene flow and the completion of the speciation process. Omocestus minutissimus and O. uhagonii are two closely related grasshoppers with partially overlapping distributions in the Central System mountains of the Iberian Peninsula. To analyse spatial patterns of historical and/or contemporary hybridization between these two taxa and understand how species boundaries are maintained in the region of secondary contact, we sampled sympatric and allopatric populations of the two species and obtained genome‐wide single nucleotide polymorphism data using a restriction site‐associated DNA sequencing approach. We used Bayesian clustering analyses to test the hypothesis of contemporary hybridization in sympatric populations and employed a suite of phylogenomic approaches and a coalescent‐based simulation framework to evaluate alternative hypothetical scenarios of interspecific gene flow. Our analyses rejected the hypothesis of contemporary hybridization but revealed past introgression in the area where the distributions of the two species overlap. Overall, these results point to a scenario of historical gene flow after secondary contact followed by the evolution of reproductive isolation that currently prevents hybridization among sympatric populations.     

Reproductive isolation via divergent genital morphology due to cascade reinforcement in Ohomopterus ground beetles

Secondary contact between incipient species and selection against maladaptive hybridization can drive reinforcement between populations in contact and result in reproductive character displacement (RCD). Resultant divergence in mating traits within a species may generate downstream reproductive isolation between populations with displaced and non-displaced traits, referred to as the cascade reinforcement hypothesis. We examined this hypothesis using three allopatric populations of the ground beetle Carabus maiyasanus with a genital lock-and-key system. This species shows RCD in male and female genital morphologies in populations in contact with the sister species C. iwawakianus. In a reciprocal mating experiment using three allopatric populations with differences in male and female genital sizes, insemination failure increased as the difference in genital size increased. Based on the reproductive isolation index, insemination failure was the major postmating-prezygotic isolation barrier, at least in one population pair with comparable total isolation to those of other species pairs. By contrast, there was only incomplete premating isolation among populations. These results suggest that RCD in genital morphologies drives incipient allopatric speciation, supporting the cascade reinforcement hypothesis. These findings provide insight into the roles of interspecific interactions and subsequent trait diversification in speciation processes.     

Evolution of the elaborate male intromittent organ of Xiphophorus fishes

Internally fertilizing animals show a remarkable diversity in male genital morphology that is associated with sexual selection, and these traits are thought to be evolving particularly rapidly. Male fish in some internally fertilizing species have “gonopodia,” highly modified anal fins that are putatively important for sexual selection. However, our understanding of the evolution of genital diversity remains incomplete. Contrary to the prediction that male genital traits evolve more rapidly than other traits, here we show that gonopodial traits and other nongonopodial traits exhibit similar evolutionary rates of trait change and also follow similar evolutionary models in an iconic genus of poeciliid fish (Xiphophorus spp.). Furthermore, we find that both mating and nonmating natural selection mechanisms are unlikely to be driving the diverse Xiphophorus gonopodial morphology. Putative holdfast features of the male genital organ do not appear to be influenced by water flow, a candidate selective force in aquatic habitats. Additionally, interspecific divergence in gonopodial morphology is not significantly higher between sympatric species, than between allopatric species, suggesting that male genitals have not undergone reproductive character displacement. Slower rates of evolution in gonopodial traits compared with a subset of putatively sexually selected nongenital traits suggest that different selection mechanisms may be acting on the different trait types. Further investigations of this elaborate trait are imperative to determine whether it is ultimately an important driver of speciation.     

Hybridization patterns between two marine snails,Littorina fabalis and L. obtusata

Characterizing the patterns of hybridization between closely related species is crucial to understand the role of gene flow in speciation. In particular, systems comprising multiple contacts between sister species offer an outstanding opportunity to investigate how reproductive isolation varies with environmental conditions, demography and geographic contexts of divergence. The flat periwinkles, Littorina obtusata and L. fabalis (Gastropoda), are two intertidal sister species with marked ecological differences compatible with late stages of speciation. Although hybridization between the two was previously suggested, its extent across the Atlantic shores of Europe remained largely unknown. Here, we combined genetic (microsatellites and mtDNA) and morphological data (shell and male genital morphology) from multiple populations of flat periwinkles in north‐western Iberia to assess the extent of current and past hybridization between L. obtusata and L. fabalis under two contrasting geographic settings of divergence (sympatry and allopatry). Hybridization signatures based on both mtDNA and microsatellites were stronger in sympatric sites, although evidence for recent extensive admixture was found in a single location. Misidentification of individuals into species based on shell morphology was higher in sympatric than in allopatric sites. However, despite hybridization, species distinctiveness based on this phenotypic trait together with male genital morphology remained relatively high. The observed variation in the extent of hybridization among locations provides a rare opportunity for future studies on the consequences of different levels of gene flow for reinforcement, thus informing about the mechanisms underlying the completion of speciation.     

Niche partitioning and the effect of interspecific competition on microhabitat use by two sympatric galaxiid stream fishes

1. Numerous interacting abiotic and biotic factors influence niche use and assemblage structure of freshwater fishes, but the strength of each factor changes with spatial scale. Few studies have examined the role of interspecific competition in structuring stream fish assemblages across spatial scales. We used field and laboratory approaches to examine microhabitat partitioning and the effect of interspecific competition on microhabitat use in two sympatric stream fishes (Galaxias‘southern’ and Galaxias gollumoides) at large (among streams and among sites within streams) and small (within artificial stream channels) spatial scales. 2. Diurnal microhabitat partitioning and interspecific competition at large spatial scales were analysed among three sympatry streams (streams with allotopic and syntopic sites; three separate catchments) and four allopatry streams (streams with only allotopic sites; two separate catchments). Electro‐fishing was used to sample habitat use of fishes at 30 random points within each site by quantifying four variables for each individual: water velocity, depth, distance to nearest cover and substratum size. Habitat availability was then quantified for each site by measuring those variables at each of 50 random points. Diet and stable isotope partitioning was analysed from syntopic sites only. Diel cycles of microhabitat use and interspecific competition at small spatial scales were examined by monitoring water velocity use over 48 h in artificial stream channels for three treatments: (i) allopatric G. ‘southern’ (10 G. ‘southern’); (ii) allopatric G. gollumoides (10 G. gollumoides) and (iii) sympatry (five individuals of each species). 3. One hundred and ninety‐four G. ‘southern’ and 239 G. gollumoides were sampled across all seven streams, and habitat availability between the two species was similar among all sites. Galaxias‘southern’ utilised faster water velocities than G. gollumoides in both the field and in channel experiments. Both species utilised faster water velocities in channels at night than during the day. Diet differences were observed and were supported by isotopic differences (two of three sites). No interspecific differences were observed for the other three microhabitat variables in the field, and multivariate habitat selection did not differ between species. Interspecific competition had no effect on microhabitat use of either species against any variable either in the field (large scale) or in channels (small scale). 4. The results suggest that niche partitioning occurs along a subset of microhabitat variables (water velocity use and diet). Interspecific competition does not appear to be a major biotic factor controlling microhabitat use by these sympatric taxa at any spatial scale. The results further suggest that stream fish assemblages are not primarily structured by biotic factors, reinforcing other studies de‐emphasising interspecific competition.     

Delimitation of sympatric Palaemon (Decapoda,Palaemonidae) species of the Laguna Madre,Mexico

Grass shrimps Palaemon mundusnovus, P. pugio and P. vulgaris sympatrically inhabit in the Laguna Madre, Mexico. They exhibit very close morphological similarity and overlap in their diagnostic characteristics, which has hindered certainty in their identification and has raised doubt regarding their taxonomic validity. In this work, we analyse intra‐ and interspecific morphometric and meristic variability through a multivariate analysis, and we determine the molecular variation using mitochondrial sequences of the 12S, 16S and COX1 genes to confirm the validity of the three taxa as having distinct lineages or to recognize a smaller number of species with phenotypic plasticity. Our results corroborate the taxonomic validity of the three species; however, there is intraspecific plasticity, interspecific overlap of characteristics and greater morphological and molecular similarity between the species P. mundusnovus and P. pugio, whereas P. vulgaris was better delimited. These species form a monophyletic group but the phylogenetic relationship obtained is discussed. Telson length was the primary variable in the principal components analysis, whereas the length of the propodus of the second pereiopod was the best discriminant. The range of variation reported in the characteristics linked to the rostrum is extended, and its influence on the separation of these species is recognized. The joint application of multivariate analysis from morphological variables and molecular tools is recommended to clarify the taxonomic status of species featuring close morphological similarity and sympatric distribution.     

Genital divergence in sympatric sister snails

A pattern of greater divergence in mating traits between sister‐species pairs with overlapping ranges than between allopatric species pairs is expected if reinforcement commonly contributes to speciation. Few large‐scale comparative analyses have addressed this prediction, especially for genital form. Here, we show that penial morphology follows the predicted pattern in 40 robustly identified sister‐species pairs in the marine gastropod subfamily Littorininae. Further work is needed to exclude other processes that may contribute to genital divergence between sympatric species, but the clear pattern we observe strongly suggests a role for genital form in reproductive isolation in this large clade.     

Conservation genetics of an endemic mountaintop salamander with an extremely limited range

Montane regions can promote allopatric speciation and harbor unique species with small ranges. The southern Appalachians are a biodiversity hotspot for salamanders, and several montane endemics occur in the region. Here, we present the first DNA sequence data for Plethodon sherando, a terrestrial salamander recently discovered in the Blue Ridge Mountains of Virginia. We sequenced two mitochondrial regions (cyt-b and CO1) from salamanders at reference sites near the center of P. sherando’s range and from two contact zones where P. sherando populations are replaced by Plethodon cinereus, the Northern Red-Backed salamander. We then used these sequence data to examine divergence and hybridization between the two taxa. We found P. sherando and P. cinereus morphotypes from contact zones to be reciprocally monophyletic and highly divergent (~17%). P. sherando exhibited very low sequence diversity (π = 0.0010) as compared to P. cinereus from the same locations (π = 0.0096). Salamander morphology in the contact zone was as distinct as morphology at reference sites, and discriminant function analysis based on morphology successfully classified 98% of salamanders to their mitochondrial lineage. Phylogenetic analysis of cyt-b sequences showed P. sherando to be sister to Plethodon serratus (the Southern Red-Backed salamander) rather than P. cinereus or any nearby mountaintop endemics. Our results suggest that P. sherando is a distinct lineage that is not subject to substantial introgression from P. cinereus and that may have a history of geographic isolation. Given its limited range (<80 km²), we believe P. sherando should merit a conservation status similar to that of other mountaintop salamanders in the region.     

REPRODUCTIVE CHARACTER DISPLACEMENT IN PHLOX

Plants of the red- and pink-flowered P. drummondii were introduced into a natural population of P. cuspidata. Their seeds were progeny-tested, and the percentage hybrid seed determined for each variant. Thirty-eight percent of the seed from the pink variant were hybrid compared to 13% of the red. The difference in hybrid production is best explained by corolla color. The self- and cross-compatibility of nine sympatric and nine allopatric populations were studied in the greenhouse. On the average, sympatric populations are more self-compatible than allopatric populations. Pollen germination is 13.7% vs. 7.3%, the difference being statistically significant. The self-compatibility differential was accompanied by a self seedset differential. On the average, sympatric populations are more cross-compatible with P. cuspidata than are allopatric populations. The germination of P. cuspidata pollen on sympatric P. drummondii stigmas was 13.3% vs. 9.8% on allopatric stigmas. However, the difference is not statistically significant. The presence of P. cuspidata has promoted reproductive character divergence in P. drummondii. The shift in corolla color and the increase in self-compatibility reduce the potential for gametic wastage and interspecific hybridization.     

Genetic analysis of sable (<Emphasis Type="Italic">Martes zibellina</Emphasis>) and pine marten (<Emphasis Type="Italic">M. martes</Emphasis>) populations in sympatric part of distribution area in the northern Urals

Analysis of nucleotide sequences of the mitochondrial DNA (mtDNA) control region (495 bp) of sables (Martes zibellina) and pine martens (M. martes) from allopatric parts of the species ranges has shown a considerable interspecific genetic distance (>3%). In sympatric populations of these species in the northern Urals, differences between two species-specific mtDNA lineages are still large; however, classification of each individual nucleotide sequence with one of the two lineages is not correlated with whether the given animal is phenotypically a sable, a pine marten, or a potential hybrid (the so-called “kidas”). This indicates a high degree of reciprocal introgression of the sable and pine marten mtDNA in the northern Urals and suggests that their interspecific hybridization is common in the sympatric zone.     

Character displacement in Hydrobia

Summary Fenchel's study of size variation in Hydrobiid snails in the Limfjord, Denmark, provides one of the most convincing cases of ecological character displacement available. In order to assess the generality of the phenomena within the Hydrobia genus, allopatric and sympatric Hydrobia ventrosa, H. neglecta and H. ulvae were collected from 24 coastal sites around Eastern England in July and October, 1982. Shell heights of 5,850 snails from 55 samples were recorded. These data were analysed for intraspecific allopatric-sympatric, and interspecific height differences. Such differences were not significant for H. ventrosa/H. neglecta, but the species exhibited parallel variation at sympatric sites. H. ventrosa/H. ulvae showed only significant interspecific differences, but in both allopatry and sympatry. The pattern of mean size variation for these species resembles that in the Limfjord. However the statistical analyses fail to provide support for character displacement. Differences in character states attributed to the process of character displacement may result from a number of other causes. Environmental conditions at sympatric and allopatric sites may act differentially on the heights of H. ulvae and H. ventrosa. The conditions that prevail at sympatric H. ulvae sites appear to lead to increased size in this species irrespective of the presence of H. ventrosa.     

Reproductive character displacement and the genetics of gamete recognition in tropical sea urchins

Reproductive character displacement occurs when sympatric and allopatric populations of a species differ in traits crucial to reproduction, and it is commonly thought of as a signal of selection acting to limit hybridization. Most documented cases of reproductive character displacement involve characters that are poorly understood at the genetic level, and rejecting alternative hypotheses for biogeographic shifts in reproductive traits is often very difficult. In sea urchins, the gamete recognition protein bindin evolves under positive selection when species are broadly sympatric, suggesting character displacement may be operating in this system. We sampled sympatric and allopatric populations of two species in the sea urchin genus Echinometra for variation in bindin and for the mitochondrial cytochrome oxidase I to examine patterns of population differentiation and molecular evolution at a reproductive gene. We found a major shift in bindin alleles between central Pacific (allopatric) and western Pacific (sympatric) populations of E. oblonga. Allopatric populations of E. oblonga are polyphyletic with E. sp. C at bindin, whereas sympatric populations of the two species are reciprocally monophyletic. There is a strong signal of positive selection (P(N)/P(S) = 4.5) in the variable region of the first exon of bindin, which is associated with alleles found in sympatric populations of E. oblonga. These results indicate that there is a strong pattern of reproductive character displacement between E. oblonga and E. sp. C and that the divergence is driven by selection. There is much higher population structure in sympatric populations at the bindin locus than at the neutral mitochondrial locus, but this difference is not seen in allopatric populations. These data suggest a pattern of speciation driven by selection for local gamete coevolution as a result of interactions between sympatric species. Although this pattern is highly suggestive of speciation by reinforcement, further research into hybrid fitness and egg-sperm interactions is required to address this potential mechanism for character displacement.