Microsatellite Variation Research Articles

Analyses of Hybrid Viability across a Hybrid Zone between Two Alnus Species Using Microsatellites and cpDNA Markers.

Diploid Alnus glutinosa s. str. and autotetraploid A. rohlenae form a narrow hybrid zone in a study area in southern Serbia, which results in triploid hybrid formation. The vast majority of previous studies have been focused on studies of maternal plants, but the offspring resulting from their crossing have not been much studied. Here, we use the variability of microsatellites and chloroplast DNA between these species and their putative hybrids to create an overall picture of the development of the hybrid zone and its predicted type. To elucidate the gene transfer within both species, the origins of individual ploidies and especially the role of triploid hybrids, a germination experiment was carried out linked with a flow cytometry study of the resulting seedlings. The tension zone model seems to offer the most adequate explanation of our observations, with selection against triploid hybrids and the spatial positioning of the hybrid zone. Despite selection against them, the triploid hybrids play an important role in the exchange of genes between the two species and therefore serve as a bridge for introgression. The presence of fertile triploids is essential for enriching the haplotype diversity between these species and for the development of new genetic lineages.

Genomewide analysis of microsatellite markers based on sequenced database in two anuran species

Eukaryotic and prokaryotic cell genomes exhibit multiple microsatellites. In this study, we characterized microsatellites in genomes and genes of Nanorana parkeri and Xenopus laevis. This characterization was used for gene ontology (GO) analysis of coding sequences (CDS). Compared to the genome of N. parkeri, the genome of X. laevis is larger and contains more number of microsatellites, but the diversity of both species are similar. Trinucleotide repeats in the genome of N. parkeri and dinucleotide and tetranucleotide repeats inthe genome of X. laevis were the most diverse. In both the species, diversity of microsatellites was highest in intergenic regions, followed by intron and exon regions, and lowest in coding regions. Microsatellites in CDS are thus subject to higher selective pressure. Many microsatellites are concentrated upstream and downstream of genes in both species, suggesting suppression of repeats in the middle of protein-CDS. Repeats are enriched in regions near gene termini purely due to the biophysical constraints of protein structure. In GO analysis, two and five unique GO terms, only found in N. parkeri and X. laevis, respectively, indicate advantageous mutations during species evolution. Biological process, cellular component and molecular function ontology reflected in the GO analysis predicted that the microsatellites located in CDS can alter protein function and may provide a molecular basis for species adaptation to new and changing environments.

Genetic diversity analysis of specialty glutinous and low-amylose rice (Oryza sativa L.) landraces of Assam based on Wx locus and microsatellite diversity

The sticky rice of Assam is traditionally classified as bora (glutinous) and chokuwa (semi-glutinous) based on their stickiness after cooking. The Waxy (Wx) gene encodes for granule-bound starch synthase (GBSS) that controls the synthesis of amylose, which is a key determinant of rice end-use quality attributes. In this report, we analysed the level of variation in grain quality traits in a collection of bora and chokuwa cultivars, and examined the nucleotide diversity at the Wx locus of selected rice accessions to identify the possible cause of low-amylose in these rice cultivar groups. The Wx gene sequencing from 24 bora and chokuwa cultivars revealed several nucleotide variations that can explain the variation in the amylose phenotypes. The nucleotide polymorphisms in the downstream intron regions were similar to those reported in Bangladeshi Beruin cultivars. Among the Wx polymorphisms, the CTn microsatellite in exon 1 and G/T SNP in intron 1 (G/T-Int1) should be considered for marker assisted breeding involving bora cultivars. The Wx gene tree, classified the bora accessions possessing the G/T-Int1 SNP as japonicas. However, cluster analysis using microsatellite markers classified the bora and chokuwa cultivars as indica, and intermediate of indica-aus. The findings of this study supplemented our understanding on the evolution of the Wx gene under human selection. The results will assist plant breeders to effectively improve the bora and chokuwa landraces.

Temporal changes in gene expression and genotype frequency of the ornithine decarboxylase gene in native silverside Basilichthys microlepidotus: Impact of wastewater reduction due to implementation of public policies.

Human activity has caused a deterioration in the health and population size of riverine species; thus, public policies have been implemented to mitigate the anthropogenic impacts of water use, watercourse transformation, and pollution. We studied the Maipo River Basin, one of the most polluted with untreated wastewater in Chile, for a period of 12 years (2007‐2019). Since the implementation of new public policies, including the operation of a wastewater collector (2012), the Maipo River Basin is currently much less polluted by untreated water than before. To analyze the impact of wastewater reduction in this river basin, we studied the native silverside (Basilichthys microlepidotus), which inhabits both polluted and unpolluted areas of the river basin. Previous studies reported the overexpression of the ornithine decarboxylase (odc) gene, heterozygote deficit, and high frequency of a homozygote odc genotype in silverside populations that inhabit wastewater‐polluted sites, suggesting a phenotypic change and genotypic selection in response to pollution. Here, a population affected and another population unaffected by wastewater were studied before and after implementing the wastewater collector. The physicochemical data of water samples, changes in odc expression and microsatellite variability, and odc genotype frequencies were analyzed. The results showed physicochemical changes in the affected site before and after the operation of the wastewater collector. The microsatellite loci showed no changes in either population. The odc expression in the affected site was higher before the operation of the wastewater collector. Significant changes in the genotype frequencies of the odc gene before and after the wastewater collector operation were detected only at the affected site, wherein the homozygous dominant genotype decreased from >59% to <25%. Our results suggest that public policies aimed at mitigating aquatic pollution can indirectly affect both gene expression and genotype frequencies of important functional genes.

Microhaplotype genotyping-by-sequencing of 98 highly polymorphic markers in three chestnut tree species

Chestnut species have large ecological, cultural and economic importance. Developing genetic markers for these species is of interest for conservation, breeding or evolutionary studies. We designed 192 primer pairs targeting microsatellites detected in the Castanea mollissima reference genome and tested them on C. sativa and C. crenata. We PCR amplified 3 × 50 microsatellites in 106 chestnut trees. Microhaplotype calling accounting for all polymorphisms resulted in a total of 98 high confidence polymorphic markers. Mean number of haplotypes per marker was 9.05 with respectively 71%, 12% and 16% of the variation corresponding to microsatellite variation in repeats number, SNP within the repeat motif and SNP or INDEL in the flanking sequence. Overall, the simple protocol described here generated a powerful multilocus genetic dataset for chestnut genetic investigations.

Genetic Evaluation of Population Structure in Tautog

AbstractTautog Tautoga onitis supports both recreational and growing commercial fisheries from Virginia to Massachusetts. Tautogs, which aggregate around structured habitats (wrecks, rocks, bridge pilings, etc.), are slow to mature and have a low reproductive rate relative to other teleosts. Slow growth and easy catchability result in Tautogs being extremely vulnerable to overexploitation, and their populations are slow to rebuild. At present, Tautogs are managed by the Atlantic States Marine Fisheries Commission as four separate regions: Massachusetts–Rhode Island, Long Island Sound, New Jersey–New York Bight, and Delaware–Maryland–Virginia, despite a lack of genetic information to support this management scenario. The objective of the current study was to investigate the genetic connectivity of Tautogs throughout their management zone and provide stock structure information in support of fisheries management. We developed and used 18 variable microsatellite loci to evaluate the genetic structure of 789 Tautogs sampled across their effective U.S. East Coast range from Massachusetts to North Carolina. We also amplified and sequenced a 1,267 base pair portion of the mitochondrial DNA control region from Tautogs sampled from Massachusetts and Virginia. Comparison of pairwise FST values among samples collected from locations south of Cape Cod, Massachusetts, were low and nonsignificant, suggesting that gene flow is sufficient to prevent the accumulation of genetic differences. The collection from Cape Cod Bay, Massachusetts, was genetically distinct from all other samples (FST = 0.021 to 0.046), indicating that Cape Cod is a significant barrier to dispersal for Tautogs and suggesting that Tautogs north and south of Cape Cod should be managed as separate stocks.

Bighorn Sheep Genetic Structure in Wyoming Reflects Geography and Management

ABSTRACTAligning wildlife management boundaries with accurate biological units promotes effective conservation and management practices that reflect ecological and evolutionary processes. Neutral genetic markers allow for quantitative delineation of population structure without a priori assumptions or biases. In the United States, bighorn sheep (Ovis canadensis) are a charismatic component of Wyoming's biodiversity and a species that provides important viewing and hunting opportunities. Bighorn sheep abundances are relatively stable throughout Wyoming, and the species is managed by administrative units identified using expert knowledge, distribution and movement data, and geographic and administrative boundaries. We used a panel of 38 variable microsatellite loci and 512 base pairs of mitochondrial DNA sequence to identify the genetic structure throughout the state and in translocation source herds, quantify the extent of genetic diversity within each genetic cluster, and estimate the degree of gene flow among herds using blood and tissue samples collected 1989–2017. We identified genetic structure of Rocky Mountain bighorn sheep in the major mountain ranges of Wyoming, with strong support for ≥5 genetic clusters using microsatellite loci. These genetic clusters generally aligned with current management units, whereas mitochondrial data showed a more complex mosaic that was not geographically patterned. Genetic variation estimated from both markers was high within each herd and comparable among herds. The assignment of individuals reflected a combination of geographic isolation and translocation, which has been extensive. Our results provide a state‐wide assessment of genetic diversity and structure that will enhance management by understanding the outcomes of translocation, identifying the source of unknown individuals, and parameterizing disease ecology models. © 2020 The Wildlife Society.

A first attempt of citizen science in the genetic monitoring of a Posidonia oceanica meadow in the Italian Southern Adriatic Sea

The endemic Mediterranean seagrass Posidonia oceanica (L.) Delile forms meadows of great ecological importance that are presently under elevated anthropogenic pressures. In this study, a model of citizens’ observatory has been translated into practice to observe and monitor the health status of the Site of Community Importance, the Posidonia oceanica meadow named San Vito-Barletta extending along the coast of Apulia region in the Italian Southern Adriatic Sea. This meadow has been lately classified as “disturbed”. With the goal of gathering relevant data for the management and conservation of this meadow, we conducted underwater visual census of the epiphytes on P. oceanica leaves. Complementarily, population genetics analysis using highly variable microsatellites markers was carried out. Two campaigns of surveys in three different stations (Molfetta, Giovinazzo and Mola di Bari) revealed that the distribution of the epiphytic communities was not uniform among the meadow. In Molfetta station, the leaf-epiphytes biomass was visibly more abundant with increased filamentous epiphytes. The genetic analysis revealed that the San Vito-Barletta meadow shows an excess of heterozygosity and demonstrated the existence of at least two subpopulations. Since the sampling areas are situated along the coasts of different Apulian cities, this situation may reflect the diverse anthropogenic factors that vary from city to city. Therefore, as local citizens’ observatory, we now have to undertake the necessary actions to raise general public awareness of the need of this species protection at Apulian level and to promote good conservation practices before the meadow regresses irreversibly.

Genetic population structure of invasive raccoons (Procyon lotor) in Hokkaido, Japan: Unique phenomenon caused by pet escape or abandonment

Phylogeographic studies can resolve relationships between genetic population structure of organisms and geographical distributions. Raccoons have become feral in Japan, and in Hokkaido island, they have been rapidly increasing in number and spreading since the 1970s. We analyzed mitochondrial (mtDNA) and microsatellite DNA to understand the current phylogenetic distribution and invasive founder events. Overall, Hokkaido raccoons maintained high genetic diversity (i.e., the level of heterozygosity was comparable to the original habitat, North America). Based on mtDNA distribution and microsatellite diversity, Hokkaido raccoons were divided into six management units. However, mtDNA haplotype distributions and genetic structures based on microsatellites did not always correspond to each other (e.g., two geographically and genetically separated populations showed similar mtDNA distributions). In addition, a high degree of genetic admixture was observed in every unit, and the degree of genetic differentiation was low even between regions separated by long distances. Compared with other countries in Europe where genetic distribution of introduced raccoons is more clearly structured, the current results represent a unique and complex phenomenon of pet escape/abandonment in Hokkaido: i.e., genetically related colonies were introduced into multiple regions as founder events, resulting in the current state in which raccoons are not clearly genetically differentiated even 40 years after introduction.

First steps towards the identification of evolutionarily significant units in Mexican native trout: An assessment of microsatellite variation

In the remote high-elevation streams and rivers in the northwest of Mexico, there is an endemic and highly diverse group of native trout related to the rainbow trout lineage (Oncorhynchus mykiss). Currently, only two species have been formally described from this trout complex, the Mexican golden trout, O. chrysogaster, and the San Pedro Martir rainbow trout, O. m. nelsoni, but only the former occurs in the Sierra Madre Occidental (SMO). South and north of the O. chrysogaster range, there are other undescribed trout that have been recently discovered. Given the various human impacts on aquatic systems occupied by these Mexican native trout, the lack of taxonomic descriptions and the imminent risk of losing populations without their even being documented, we sought to infer a hierarchical population structure of the Mexican native trout, determine the geographic patterns of genetic diversity and gene flow among them, and identify evolutionarily significant units for a prompt conservation strategy. The allelic diversity across eleven microsatellite loci was used to assess genetic divergences of wild trout collected throughout its range in Mexico. Our results reject the old hypothesis that many of the trout in Mexico are introduced or translocated hatchery-reared rainbow trout and strongly support the new hypothesis of distinct genetic populations and lineages in the northwest of Mexico. According to the different statistical analysis of the genetic data, we distinguish between species and subspecies (O. chrysogaster, O. mykiss, and O. m. nelsoni) of the other regional lineages of undescribed trout (northern and southern) inhabiting the SMO. We also found a strong genetic substructure within these large clusters. We propose the presence of seven Evolutionarily Significant Units in the drainage basins of the SMO (two occur in the northern region, four in the southern, and one for the area of distribution of O. chrysogaster) and one in north-western Baja California. Based in microsatellite data a minimum gene flow among populations is evidenced, resulting the presence the highly divergent populations.

Genetic consequences of being a dwarf: do evolutionary changes in life-history traits influence gene flow patterns in populations of the world's smallest goldenrod?

Contrasting life-history traits can evolve through generations of dwarf plant ecotypes, yet such phenotypic changes often involve decreased plant size and reproductive allocation, which can configure seed dispersal patterns and, subsequently, population demography. Therefore, evolutionary transitions to dwarfism can represent good study systems to test the roles of life-history traits in population demography by comparing genetic structure between related but phenotypically divergent ecotypes. In this study, we examined an ecotypic taxon pair of the world's smallest goldenrod (stem height 2.6 cm) in alpine habitats and its closely related lowland taxon (30-40 cm) found on Yakushima Island, Japan. Genetic variation in chloroplast DNA sequences, nuclear microsatellites and genome-wide single-nucleotide polymorphisms were used to investigate 197 samples from 16 populations, to infer the population genetic demography and compare local genetic structure of the ecotypes. We found a pronounced level of genetic differentiation among alpine dwarf populations, which were much less geographically isolated than their lowland counterparts. In particular, several neighbouring dwarf populations (located ~500 m apart) harboured completely different sets of chloroplast haplotypes and nuclear genetic clusters. Demographic modelling revealed that the dwarf populations have not exchanged genes at significant levels after population divergence. These lines of evidence suggest that substantial effects of genetic drift have operated on these dwarf populations. The low-growing stature and reduced fecundity (only 3.1 heads per plant) of the dwarf plants may have reduced gene flow and rare long-distance seed dispersal among habitat patches, although the effects of life-history traits require further evaluation using ecological approaches.

Population structure of the New Zealand whelk, Cominella glandiformis (Gastropoda: Buccinidae), suggests sporadic dispersal of a direct developer

AbstractWe examined phylogeographic structure in the direct-developing New Zealand endemic intertidal mud whelk, Cominella glandiformis. Two hundred and ninety-six whelks from 12 sites were collected from sheltered shores around New Zealand’s four largest islands (North Island, South Island, Stewart Island and Chatham Island), encompassing the geographical range of this species. Despite being direct developers, gene flow among C. glandiformis populations may occur over short distances by adult floating, and over larger distances by rafting of egg masses. Primers were developed to amplify variable microsatellite regions at six loci. All loci were variable, with 8–34 alleles/loci. Observed and expected heterozygosities were high across all alleles, with minimal evidence of null alleles. The average number of alleles varied from 3.5 (Chatham Island) to 7.5 (Waitemata Harbour). Strong genetic structure was evident, with distinct ‘eastern’ and ‘western’ groups. Each group extended over a large geographic area, including regions of unsuitable habitat, but were linked by oceanic currents. We suggest that the intraspecific geographic genetic structure in C. glandiformis has arisen due a combination of ocean currents (promoting gene flow between geographically distant regions) and upwelling areas (limiting gene flow between certain regions).

The evolution of spring fen ecotypes in Rhinanthus: genetic evidence for parallel origins in Scandinavia after the last ice age

Locally adapted ecotypes can constitute an important part of the biodiversity, especially in young floras with few endemic species. However, the origins, distinctness and conservation value of many ecotypes remain uncertain because genetic data are lacking or no common-garden study has been carried out. In the present study, we evaluated the distinctness and genetic structure of a phenotypically deviating morph of Rhinanthus angustifolius, growing in calcareous spring fens on the Baltic island of Gotland. Using data from a common-garden experiment and analyses of nuclear microsatellite variation, we compared fen populations on Gotland with conspecific populations from habitats more typical of the study species. We also included the fen specialist R. osiliensis from the Baltic island of Saaremaa in the molecular analyses to make further inferences about the origin of the Gotlandic fen morph. Our data indicate that the Gotlandic fen populations constitute a phenotypically and genetically distinct ecotype that most likely has evolved at least two times on Gotland after the last ice age. In congruence with previous studies, we also infer that fen ecotypes have evolved independently on Gotland and Saaremaa. We propose a varietal status for the Gotlandic fen ecotype and give recommendations for the conservation of this taxon.

A comparison of neutral genetic differentiation and genetic diversity among migratory and resident populations of Golden-crowned-Kinglets (Regulus satrapa)

Many animals migrate seasonally between breeding and non-breeding territories and these annual movements can have a profound effect on population genetic structure. We genotyped 281 individuals from 11 populations at seven variable microsatellite loci and compared patterns of neutral genetic differentiation and neutral genetic diversity among migratory and resident breeding populations of the Golden-crowned Kinglet (Regulus satrapa), a widespread North American songbird. We predicted that resident populations would exhibit greater genetic differentiation and lower genetic diversity than migratory populations because migratory behaviour is thought to enhance gene flow. The magnitude of genetic differentiation and genetic diversity among migratory and resident populations was comparable, and the greatest levels of differentiation were observed for pairwise comparisons between Ontario and all western populations. Distance-based redundancy models and redundancy models revealed that patterns of neutral genetic differentiation and neutral genetic diversity follow an isolation-by-distance model and are not correlated with migratory behaviour. Overall it appears that genetic patterns are more closely associated with Pleistocene glacial history as proposed in a previous study.

Population Structure and Genetic Connectivity of Squat Lobsters (Munida Leach, 1820) Associated With Vulnerable Marine Ecosystems in the Southwest Pacific Ocean

Studies of genetic diversity and population genetic structure in deep-sea fauna mainly focus on vulnerable marine ecosystem (VME) indicator taxa, whilst relatively few studies have focussed on VME-associated taxa whose distributions are not exclusively limited to VMEs. Knowledge of genetic connectivity (gene flow) amongst populations of VME-associated taxa, such as squat lobsters, will contribute to ongoing management decision-making related to the protection of VMEs. To better understand the genetic diversity and genetic structure of squat lobster populations (Munida isos, M. endeavourae and M. gracilis) at different spatial scales (biogeographic provinces, regions, and geomorphic features) in the southwest Pacific Ocean, mitochondrial COI region and nuclear microsatellite markers were employed. Overall, the levels of genetic diversity were high for the COI region and moderate for the microsatellite loci across the three Munida species. AMOVA of COI variation revealed no significant genetic differentiation, whereas AMOVA of microsatellite variation revealed significant genetic differentiation amongst the three species, but at different spatial scales. Based on microsatellite variation, a range of analyses (Structure, PCoA, DAPC) provided some evidence of limited genetic differentiation at different spatial scales across the three species. Low to moderate levels of assignment success (~40–60%) based on microsatellite variation were achieved for the three Munida species, suggesting high levels of gene flow and possible panmixia. Nonetheless, for Munida isos, populations from the Tasmanian slope were genetically differentiated from all other populations and may act as source populations, whereas populations from the Kermadec Ridge region may be sink populations for all three Munida species. Our results highlight the need to consider gene flow at trans-national scales when managing anthropogenic impacts on VMEs. The results are discussed in the context of existing marine protected areas, which can contribute new information useful to the management of VMEs within the southwest Pacific Ocean.

Genetic structure features of cattle populations of Ukrainian selection by polymorphism of loci, associated with milk productivity traits

Aim. To study the genetic structure of cattle populations of Ukrainian selection by polymorphism of functional genes (PRL, PL) and microsatellites (BM027, RM185). Methods. The study was conducted using the method of polymerase chain reaction (PCR) and restriction analysis in case of loci PRL and PL, and using classic PCR with subsequent electrophoresis in polyacrylamide gel to analyze microsatellite variability. Results. The results of the studies demonstrated that the locus of prolactin by RsaI-polymorphism in the fourth exon was polymorphic in both experimental populations (Ukrainian Black-and-White and Ukrainian Red-and-White dairy breeds of cattle). The mutation (Indel) was fi rst determined in the fourth exon of prolactin gene, the variants of which correlated with some alleles of the locus by RsaI-polymorphism. The locus of placental lactogen by RsaI-polymorphism in the fi fth exon was monomorphic in both experimental populations. Microsatellite locus RM185 was polymorphic in both groups of animals, whereas BM027 – only in the Black-and-White dairy breed. Conclusions. The specifi cities of the genetic structure of the Ukrainian Black-and-White and Red-andWhite dairy breed populations by polymorphism of functional genes and microsatellite loci were determined. The locus of placental lactogen by RsaI-polymorphism in the fi fth exon cannot be used in further studies due to the absence of alternative variants of the gene in both studied populations of animals. The analysis of the distribution of haplotype frequencies demonstrated the absence of deviation from the equilibrium state by linkage for each of the investigated markers which makes their use impossible in the breeding programs as a separate functional unit.

Multi-year genetic sampling indicates maternal gene flow via colony emigrations in the army ant Eciton burchellii parvispinum

Sex-biased dispersal occurs when one sex disperses more frequently or farther than the opposite sex. In ants, dispersal is often male biased and males typically contribute more strongly to gene flow within and among ant populations. However, army ants may offer an exceptional case of female-biased dispersal, because army ant colonies (particularly species in the Neotropical genus Eciton) routinely emigrate among nest sites. We hypothesized that maternal mobility via successive colony emigrations would reduce the relative male bias in dispersal, and that emigrations would contribute to gene flow within an Eciton population. We genotyped workers (highly variable DNA microsatellite loci) from Eciton burchellii parvispinum colonies collected over a 10 × 10 km area in Monteverde, Costa Rica. Samples were spaced approximately 3 years apart (in 2006 and 2009), the typical time between colony reproductive bouts. We used worker genotypes to reconstruct the genotypes of their mother queens and of a subset of the males that sired the workers. We then analyzed the population genetic structure of the queen and male genotypes by comparing individual-based relatedness estimates for each sex across geographic distance and over time. There was no significant difference in spatial genetic structure between the sexes, either within or between the 2006 and 2009 samples. There was some evidence against genetic philopatry: 2009 queens were significantly unrelated to 2006 queens collected nearby (within 0.5 km). The patterns suggest maternal dispersal via emigrations contributes to gene flow, reducing or eliminating male biases in dispersal. From two populations, we estimated average queen dispersal distance to be 0.63–1.04 km, a greater distance than many winged ant queens disperse. Because emigrations over the lifetime of army ant queens may contribute to gene flow across the landscape, habitat connectivity should be maintained to permit colony emigrations and support genetic diversity in populations of this keystone species.

Incipient sympatric speciation via host race formation in Phengaris arion (Lepidoptera: Lycaenidae)

The plausibility of sympatric speciation is still debated despite increasing evidence, such as host races in insects. This speciation process may be occurring in the case of the two phenological forms of the obligatorily myrmecophilous Phengaris arion. The main goal of our research was to study the nature and causes of difference between these forms focusing primarily on the incipient speciation via host races. Molecular analyses based on highly variable microsatellites together with Wolbachia screening, male genitalia morphometrics and host ant studies were carried out on four syntopic sample pairs. Our results show that the two phenological forms of P. arion may meet the criteria for host plant races. They coexist in sympatry in certain parts of the species range which is allowed by the adaptation to the distinct phenology of the host plants. Negative selection acts against the intermediate individuals which are on the wing in the inappropriate time frame. Thus, disruptive selection affects and produces bimodal distributions of phenotypes. However, the phenology of food plants is not entirely distinct and fluctuates year by year. Therefore, the two forms can exchange genes occasionally depending on the length of the time slot when they can meet with each other. Consequently, the reproductive isolation could not be completed and the existence of the two arion forms may represent only an incipient stage of sympatric speciation. It is also clear that Wolbachia is likely not a driver of sympatric speciation in this case.

A genetic and morphological survey to trace the origin of Melipona beecheii (Apidae: Meliponini) from Cuba

The stingless bee Melipona beecheii is extensively distributed across Mexico and Central America and the only Melipona found in the Greater Antilles. The aim of this work was to establish possible continental affinities of M. beecheii from the Antilles. We compared populations from the main island (Cuba) with two clearly separated genetic lineages of M. beecheii on mainland, namely the Yucatan peninsula (Mexico) and Costa Rica. We used morphometrics, the degree of cephalic maculation, and microsatellite variation. Results indicate a greater morphological and genetic similarity between populations from Cuba and the Yucatan peninsula with respect to those of Costa Rica. Based on our findings, we conclude that the origin of M. beecheii from Cuba is more likely the Yucatan peninsula (Mexico). We found evidence that isolation has not been large enough to result in a new genetic lineage.

Strong genetic structure revealed by microsatellite variation in Callicarpa species endemic to the Bonin (Ogasawara) Islands.

Adaptive radiation is the diversification of a founding population into multiple taxa that are differentially adapted to diverse ecological niches. The three Callicarpa (Lamiaceae) species endemic to the Bonin Islands are considered to represent an example of adaptive radiation on oceanic islands. All three species are distributed in the Chichijima Island Group and grow in different habitats, while only one species, C. subpubescens, is distributed among other island groups. Particularly, in the Hahajima and Mukojima Island Groups, C. subpubescens grows in various habitats and shows relatively high morphological variation. We investigated genotypes of the three Callicarpa species at 14 microsatellite markers to elucidate genetic differentiation within and between species or island groups and between different habitats or morphologies. We found that genetic differentiation within C. subpubescens in the Hahajima and Mukojima Island Groups was equally as high as that between the three species in the Chichijima Island Group, while differentiation within C. subpubescens in the Chichijima Island Group was much lower. Analyses such as a Bayesian clustering analysis showed that genetically distinct groups were associated with the three species in the Chichijima Island Group, whereas they showed strong genetic structure within C. subpubescens in the Hahajima and Mukojima Island Groups among different habitats and morphologies. These results indicated that ecological diversification occurred in the Hahajima and Mukojima Island Groups. Meanwhile, high genetic differentiation among different island groups was also observed, reflecting isolation by distance. It implies that non-ecological factors such as geographic isolation also played important roles in genetic differentiation in Callicarpa species in the Bonin Islands. These findings suggest that the Callicarpa species in the Bonin Islands are differentiated into multiple genetic groups by both ecological and non-ecological factors.