Widespread gene flow between oceans in a pelagic seabird species complex

Global‐scale gene flow is an important concern in conservation biology as it has the potential to either increase or decrease genetic diversity in species and populations. Although many studies focus on the gene flow between different populations of a single species, the potential for gene flow and introgression between species is understudied, particularly in seabirds. The only well‐studied example of a mixed‐species, hybridizing population of petrels exists on Round Island, in the Indian Ocean. Previous research assumed that Round Island represents a point of secondary contact between Atlantic (Pterodroma arminjoniana) and Pacific species (Pterodroma neglecta and Pterodroma heraldica). This study uses microsatellite genotyping and tracking data to address the possibility of between‐species hybridization occurring outside the Indian Ocean. Dispersal and gene flow spanning three oceans were demonstrated between the species in this complex. Analysis of migration rates estimated using bayesass revealed unidirectional movement of petrels from the Atlantic and Pacific into the Indian Ocean. Conversely, structure analysis revealed gene flow between species of the Atlantic and Pacific oceans, with potential three‐way hybrids occurring outside the Indian Ocean. Additionally, geolocation tracking of Round Island petrels revealed two individuals travelling to the Atlantic and Pacific. These results suggest that interspecific hybrids in Pterodroma petrels are more common than was previously assumed. This study is the first of its kind to investigate gene flow between populations of closely related Procellariiform species on a global scale, demonstrating the need for consideration of widespread migration and hybridization in the conservation of threatened seabirds.     

Characterization of polymorphic microsatellite loci from Round Island Petrels (Pterodroma arminjoniana) and their utility in other seabird species

Six microsatellite loci were isolated from the petrels of Round Island, near Mauritius in the Indian Ocean (Pterodroma arminjoniana). Three loci were monomorphic in P. arminjoniana but were found to be polymorphic in other Procellariiforms. Cross-utility of all six loci was tested in 17 Procellariiform and 1 penguin species. In addition, 53 microsatellite loci developed for other species of birds were tested for cross-species amplification in P. arminjoniana. Six of these loci were found to be polymorphic.     

Assortative mating,sexual size dimorphism and sex determination in a seabird with plumage polymorphism

Intraspecific plumage polymorphism in seabirds is often attributed to advantages in foraging activities and escape from predators, but its role in sexual selection is not well understood. The Trindade petrel (Pterodroma arminjoniana) presents morphs varying from pale to whole dark, with no apparent sexual size dimorphism (SSD). We tested assortative mating in Trindade petrels based on plumage colours and body size. In addition, genders of Trindade petrels were identified molecularly aiming to test SSD based on morphometrics, which was also used to generate a discriminant function for sex assignment. Within-pair consistency in plumage colour (i.e. birds paired with mates of the same morph) was detected in 9 out of 10 pairs, but not in morphometric traits. Minimum bill depth and bill depth at unguis were traits significantly larger in males. The best model was adjusted with Bill depth at unguis, wing chord and body mass, with global discriminatory power of 78.4%. Our results suggest that plumage colours may be sexually selected in Trindade petrels, which brings evolutionary implications on the persistence of plumage polymorphism. Discriminatory power of the best discriminant function was similar to those found in other Procellariiformes and also among datasets obtained by distinct researchers, demonstrating its robustness.     

Phylogenetic relationships in Pterodroma petrels are obscured by recent secondary contact and hybridization

The classification of petrels (Pterodroma spp.) from Round Island, near Mauritius in the Indian Ocean, has confounded researchers since their discovery in 1948. In this study we investigate the relationships between Round Island petrels and their closest relatives using evidence from mitochondrial DNA sequence data and ectoparasites. Far from providing clear delimitation of species boundaries, our results reveal that hybridization among species on Round Island has led to genetic leakage between populations from different ocean basins. The most common species on the island, Pterodroma arminjoniana, appears to be hybridizing with two rarer species (P. heraldica and P. neglecta), subverting the reproductive isolation of all three and allowing gene flow. P. heraldica and P. neglecta breed sympatrically in the Pacific Ocean, where P. arminjoniana is absent, but no record of hybridization between these two exists and they remain phenotypically distinct. The breakdown of species boundaries in Round Island petrels followed environmental change (deforestation and changes in species composition due to hunting) within their overlapping ranges. Such multi-species interactions have implications not only for conservation, but also for our understanding of the processes of evolutionary diversification and speciation.     

Molecular evidence for the identity of the Magenta petrel

A lone petrel was shot from the decks of an Italian warship (the ‘Magenta’) while it was sailing the South Pacific Ocean in 1867, far from land. The species, unknown to science, was named the ‘Magenta petrel’ (Procellariiformes, Procellariidae, Pterodroma magentae). No other specimens of this bird were collected and the species it represented remained a complete enigma for over 100 years. We compared DNA sequence of the mitochondrial cytochrome b gene from the Magenta petrel to that of other petrels using phylogenetic methods and ancient DNA techniques. Our results strongly suggest that the Magenta petrel specimen is a Chatham Island taiko. Furthermore, given the collection location of the Magenta petrel, our finding indicates that the Chatham Island taiko forages far into the Pacific Ocean (near South America). This has implications for the conservation of the taiko, one of the world's rarest seabirds.     

Population Genetic Structure and Conservation of the Galápagos Petrel (Pterodroma phaeopygia)

The Galápagos petrel (Pterodroma phaeopygia) is endemic to the Galápagos archipelago, where it is known to breed only on five islands. The species has been listed as critically endangered due to habitat deterioration and predation by introduced mammals. Significant morphological and behavioural differences among petrels nesting on different islands suggest that island populations may differ genetically. Furthermore, nesting phenology suggests that genetically differentiated seasonal populations may exist within at least one island. We analysed variation in six microsatellite loci and part of the mitochondrial ATPase 6/8 gene in 206 Galápagos petrels sampled from all five islands. No evidence of genetic structuring within islands was found, although statistical power was low. In contrast, significant differences occurred among island populations. For the microsatellite loci, private alleles occurred at all islands, sometimes at high frequency; global and pairwise estimates of genetic differentiation were all statistically significant; Bayesian analysis of genotypes frequencies provided strong support for three genetic populations; and most estimates of migration between populations did not differ significantly from zero. Only two ATPase haplotypes were found, but the geographic distribution of haplotypes indicated significant differentiation among populations. For conservation purposes, populations from Floreana, Santa Cruz, San Cristóbal and Santiago should be regarded as separate genetic management units. Birds from Isabela appear to be derived recently from the Santiago population, and the population on San Cristóbal appears to be a mixture of birds from other populations. However, considering ecological and behavioural differences among birds from different islands, we recommend that all five populations be protected.     

Development and characterization of the first microsatellite markers in the mangrove species Pelliciera rhizophorae Triana & Planchon

Pelliciera rhizophorae is a unique Neotropical mangrove species belonging to Pelliciera genus. We isolated eight microsatellite loci from this species. All loci were polymorphic and showed three to nine alleles per locus in Colombian Pacific and Caribbean populations. Polymorphic information content ranged from 0.46 to 0.69. Two loci (PeRh‐14 and PeRh‐19) showed null alleles on the Caribbean coast, which suggest genetic differentiation between Pacific and Caribbean populations of P. rhizophorae. Development of these microsatellite loci constitutes a new molecular tool to carry out studies in the genome of the species and to evaluate its population dynamics.     

Year‐round spatial movements and trophic ecology of Trindade Petrels (Pterodroma arminjoniana)

Trindade Petrels (Pterodroma arminjoniana) are vulnerable gadfly petrels that breed on the remote Trindade Island, located ~1100 km off the Brazilian coast. Little is known about their spatial ecology, and their trophic ecology has only been described for the breeding season. We tagged four Trindade Petrels with global location sensing loggers (GLS) from October 2013 to November 2014 and sampled the blood and feathers (innermost primary and the eighth secondary) of 14 individuals to evaluate their year‐round spatial and isotopic ecology. We examined individual distributions, habitat use and suitability, activity, and isotopic values during the breeding, migration, and non‐breeding periods. Trindade Petrels used areas in the southwest Atlantic Ocean (between 10°N and 50°S in latitude) during the breeding season. They migrated through pelagic waters of the tropical Atlantic to the northwest Atlantic, where they spent the non‐breeding season. Trindade Petrels used mostly tropical to subtropical waters in areas of intermediate to high wind speeds and low marine productivity. Individuals spent more time foraging at night than during the day. During the breeding season, birds in northerly areas had higher carbon‐13 values, and birds that used more pelagic areas foraged on prey at a higher trophic level (higher nitrogen‐15 values) than those in more southern and coastal areas. Isotopic values during the breeding, migration, and non‐breeding periods differed, possibly due to differences among individuals in their at‐sea distribution throughout the year. We confirmed the non‐breeding distribution of Trindade Petrels, which was previously known only from vessel sightings and stranded birds. Our results also suggest a strong temporal segregation in the at‐sea distribution and trophic ecology between two groups of individuals, which might indicate the existence of two separate breeding populations.     

Conservation genetics of critically endangered Crepidiastrum grandicollum (Asteraceae) and two closely related woody species of the Bonin Islands,Japan

Crepidiastrum grandicollum is a critically endangered insular endemic herb found only on two oceanic islands of the Bonin Islands in Japan, namely Chichijima Island and Anijima Island. It is explicitly threatened by herbivory pressure from introduced animals. In 2009, a conservation program for C. grandicollum was begun to ensure its future persistence. To provide further information for conservation planning, we investigated the genetic diversity of C. grandicollum using 13 novel microsatellite markers in 55 individuals from four wild populations and an ex situ living collection. Two closely related woody species were also included for interspecific comparison: seven individuals of Crepidiastrum ameristophyllum and 13 of Crepidiastrum linguifolium. The 13 markers were applicable to all three species and identified 129 alleles in total. We found a clear genetic differentiation between C. grandicollum from Anijima Island and Chichijima Island. Crepidiastrum grandicollum also had low expected heterozygosity and allelic richness in populations compared to the two closely related species. Reconstructed divergence history suggested that differentiation between the islands had occurred several thousand generations ago. We suggest separate conservation units for C. grandicollum on Anijima Island and Chichijima Island given the clear (and putatively historical) genetic differentiation, which may result in speciation in the future.

     

Closely related octopus species show different spatial genetic structures in response to the Antarctic seascape

Determining whether comparable processes drive genetic divergence among marine species is relevant to molecular ecologists and managers alike. Sympatric species with similar life histories might be expected to show comparable patterns of genetic differentiation and a consistent influence of environmental factors in shaping divergence. We used microsatellite loci to quantify genetic differentiation across the Scotia Arc in three species of closely related benthic octopods, Pareledone turqueti, P. charcoti, and Adelieledone polymorpha. The relative importance of environmental factors (latitude, longitude, depth, and temperature) in shaping genetic structure was investigated when significant spatial genetic structure was uncovered. Isolated populations of P. turqueti and A. polymorpha at these species’ range margins were genetically different to samples close to mainland Antarctica; however, these species showed different genetic structures at a regional scale. Samples of P. turqueti from the Antarctic Peninsula, Elephant Island, and Signy Island were genetically different, and this divergence was associated primarily with sample collection depth. By contrast, weak or nonsignificant spatial genetic structure was evident across the Antarctic Peninsula, Elephant Island, and Signy Island region for A. polymorpha, and slight associations between population divergence and temperature or depth (and/or longitude) were detected. Pareledone charcoti has a limited geographic range, but exhibited no genetic differentiation between samples from a small region of the Scotia Arc (Elephant Island and the Antarctic Peninsula). Thus, closely related species with similar life history strategies can display contrasting patterns of genetic differentiation depending on spatial scale; moreover, depth may drive genetic divergence in Southern Ocean benthos.     

Unexpectedly complex gradation of coral population structure in the Nansei Islands,Japan

To establish effective locations and sizes of potential protected areas for reef ecosystems, detailed information about source and sink relationships between populations is critical, especially in archipelagic regions. Therefore, we assessed population structure and genetic diversity of Acropora tenuis, one of the dominant stony coral species in the Pacific, using 13 microsatellite markers to investigate 298 colonies from 15 locations across the Nansei Islands in southwestern Japan. Genetic diversity was not significant among sampling locations, even in possibly peripheral locations. In addition, our results showed that there are at least two populations of A. tenuis in the study area. The level of genetic differentiation between these populations was relatively low, but significant between many pairs of sampling locations. Directions of gene flow, which were estimated using a coalescence‐based approach, suggest that gene flow not only occurs from south to north, but also from north to south in various locations. Consequently, the Yaeyama Islands and the Amami Islands are potential northern and southern sources of corals. On the other hand, the Miyako Islands and west central Okinawa Island are potential sink populations. The Kerama Islands and the vicinity of Taketomi Island are potential contact points of genetic subdivision of coral populations in the Nansei Islands. We found that genetic population structure of A. tenuis in the Nansei Islands is more complex than previously thought. These cryptic populations are very important for preserving genetic diversity and should be maintained.     

Reduced genetic variation and strong genetic population structure in the freshwater killifish Valencia letourneuxi (Valenciidae) based on nuclear and mitochondrial markers

The genetic variation of the critically endangered Corfu killifish (Valencia letourneuxi), an endemic freshwater fish species of the western Balkans, was assessed for nine populations sampled in eight water systems in western continental Greece, the Peloponnese and the Ionian Island of Corfu, using mitochondrial and microsatellite markers. The analyses were based on data from three mtDNA regions (D‐loop, COI and 16S rRNA sequences) and 14 microsatellite loci. Samples from the congeneric species Valencia hispanica and the phylogenetically closely related species Aphanius fasciatus were also used in the study as outgroups. Both the mitochondrial and the microsatellite analyses revealed three distinct population groupings associated with the geographical distribution of the populations: one southern group occupying rivers draining to the Patraikos Gulf, the second one including the populations flowing into the Amvrakikos Gulf and the third, more northern group, including the other populations from rivers in Corfu Island and Epirus flowing into the Ionian Sea. Within these groupings there is limited genetic differentiation between populations; in addition, there is reduced intrapopulation genetic variation, evidenced by low heterozygosity values, number of alleles and haplotype diversity. In terms of taxonomic implications and appropriate management actions for conservation, our data suggest that the major population groups should be regarded at least as three distinct conservation units (CUs), with translocation and restocking actions to take place only within the geographical range of the CU concerned. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 334–349.     

Population divergence and gene flow in an endangered and highly mobile seabird

Seabirds are highly vagile and can disperse up to thousands of kilometers, making it difficult to identify the factors that promote isolation between populations. The endemic Hawaiian petrel (Pterodroma sandwichensis) is one such species. Today it is endangered, and known to breed only on the islands of Hawaii, Maui, Lanai and Kauai. Historical records indicate that a large population formerly bred on Molokai as well, but this population has recently been extirpated. Given the great dispersal potential of these petrels, it remains unclear if populations are genetically distinct and which factors may contribute to isolation between them. We sampled petrels from across their range, including individuals from the presumably extirpated Molokai population. We sequenced 524 bp of mitochondrial DNA, 741 bp from three nuclear introns, and genotyped 18 microsatellite loci in order to examine the patterns of divergence in this species and to investigate the potential underlying mechanisms. Both mitochondrial and nuclear data sets indicated significant genetic differentiation among all modern populations, but no differentiation was found between historic samples from Molokai and modern birds from Lanai. Population-specific nonbreeding distribution and strong natal philopatry may reduce gene flow between populations. However, the lack of population structure between extirpated Molokai birds and modern birds on Lanai indicates that there was substantial gene flow between these populations and that petrels may be able to overcome barriers to dispersal prior to complete extirpation. Hawaiian petrel populations could be considered distinct management units, however, the dwindling population on Hawaii may require translocation to prevent extirpation in the near future.     

Genetic differentiation of the Thorn‐tailed Rayadito Aphrastura spinicauda (Furnariidae: Passeriformes) revealed by ISSR profiles suggests multiple palaeorefugia and high recurrent gene flow

Nucleotide sequence data (cytochrome b) and ISSR genomic fingerprints were used to analyse the genetic variation and population differentiation in Thorn‐tailed Rayadito, a widespread Patagonian forest bird. We included samples from eight populations of Thorn‐tailed Rayadito covering most of the distribution range of the species: from fragmented patches of Olivillo forest in northern Chile to Isla Navarino forests in the extreme south of South America. Low levels of genetic diversity were found among populations, with a large within‐population molecular variance indicating high levels of gene flow. The multivariate and cluster analyses based on ISSR markers show that the subspecies bullocki (from Mocha Island) differs significantly from all other populations. The subspecies fulva (Chiloé Island) shows less differentiation than bullocki, sharing several alleles with continental populations. Bayesian analyses suggest that the Mocha Island population contributes most to the total genetic diversity observed in the species. Mantel tests revealed no significant correlation between geographical distance and pairwise genetic distance and cytochrome b sequence analyses failed to detect differentiation among subspecies. Mocha Island might have been a palaeorefuge and this population may have diversified by genetic drift after the last glacial maximum. There is also the possibility of a postglacial colonization of the Thorn‐tailed Rayadito from an austral palaeorefugium, supporting a multiple refugia hypothesis. This study illustrates the usefulness of the rarely used ISSR genomic fingerprint method in avian phylogeography.     

The Kuroshio Current influences genetic diversity and population genetic structure of a tropical seagrass,Enhalus acoroides

Information on genetic diversity and differentiation of seagrass populations is essential for the conservation of coastal ecosystems. However, little is known about the seagrasses in the Indo‐West Pacific Ocean, where the world's highest diversity of seagrasses occurs. The influence of sea currents on these populations is also unknown. We estimated the genetic diversity and population genetic structure and identified reproductive features in Enhalus acoroides populations from the Yaeyama Islands, Hainan Island and the Philippines. The Philippines are situated at the centre of the E. acoroides range, Yaeyama and Hainan are peripheral populations, and the Yaeyama population is at the northern limit of the species range. The powerful Kuroshio Current flows from the Philippines to Yaeyama. Genetic analyses using nine microsatellite markers indicated that reproduction of E. acoroides is mostly sexual. Clonal diversity does not decrease in northern populations, although genetic diversity does. However, the genetic diversity of the Yaeyama populations is greater than that of the Hainan populations. Significant genetic differentiation among most populations was evident; however, the Yaeyama and north‐east Philippines populations were genetically similar, despite being separated by ~1100 km. An assignment test suggested that recruitment occurs from the north‐east Philippines to Yaeyama. The strong current in this region is probably responsible for the extant genetic diversity and recruitment patterns.     

Population structure of island‐associated dolphins: Evidence from mitochondrial and microsatellite markers for common bottlenose dolphins (Tursiops truncatus) around the main Hawaiian Islands

We used mitochondrial and nuclear genetic markers to investigate population structure of common bottlenose dolphins, Tursiops truncatus, around the main Hawaiian Islands. Though broadly distributed throughout the world's oceans, bottlenose dolphins are known to form small populations in coastal waters. Recent photo‐identification data suggest the same is true in Hawaiian waters. We found genetic differentiation among (mtDNA Φ_ST= 0.014–0.141, microsatellite F’_ST= 0.019–0.050) and low dispersal rates between (0.17–5.77 dispersers per generation) the main Hawaiian Island groups. Our results are consistent with movement rates estimated from photo‐identification data and suggest that each island group supports a demographically independent population. Inclusion in our analyses of samples collected near Palmyra Atoll provided evidence that the Hawaiian Islands are also occasionally visited by members of a genetically distinct, pelagic population. Two of our samples exhibited evidence of partial ancestry from Indo‐Pacific bottlenose dolphins (T. aduncus), a species not known to inhabit the Hawaiian Archipelago. Our findings have important implications for the management of Hawaiian bottlenose dolphins and raise concerns about the vulnerability to human impacts of pelagic species in island ecosystems.     

Environmental Variables Explain Genetic Structure in a Beetle-Associated Nematode

The distribution of a species is a complex expression of its ecological and evolutionary history and integrating population genetic, environmental, and ecological data can provide new insights into the effects of the environment on the population structure of species. Previous work demonstrated strong patterns of genetic differentiation in natural populations of the hermaphroditic nematode Pristionchus pacificus in its La Réunion Island habitat, but gave no clear understanding of the role of the environment in structuring this variation. Here, we present what is to our knowledge the first study to statistically evaluate the role of the environment in shaping the structure and distribution of nematode populations. We test the hypothesis that genetic structure in P. pacificus is influenced by environmental variables, by combining population genetic analyses of microsatellite data from 18 populations and 370 strains, with multivariate statistics on environmental data, and species distribution modelling. We assess and quantify the relative importance of environmental factors (geographic distance, altitude, temperature, precipitation, and beetle host) on genetic variation among populations. Despite the fact that geographic populations of P. pacificus comprise vast genetic diversity sourced from multiple ancestral lineages, we find strong evidence for local associations between environment and genetic variation. Further, we show that significantly more genetic variation in P. pacificus populations is explained by environmental variation than by geographic distances. This supports a strong role for environmental heterogeneity vs. genetic drift in the divergence of populations, which we suggest may be influenced by adaptive forces.     

Phylogeography of a pearl oyster (Pinctada maxima) across the Indo‐Australian Archipelago: evidence of strong regional structure and population expansions but no phylogenetic breaks

This study investigates the genetic structure and phylogeography of a broadcast spawning bivalve mollusc, Pinctada maxima, throughout the Indo‐West Pacific and northern Australia. DNA sequence variation of the mitochondrial cytochrome oxidase subunit I (COI) gene was analysed in 367 individuals sampled from nine populations across the Indo‐West Pacific. Hierarchical AMOVA indicated strong genetic structuring amongst populations (Φ_ST = 0.372, P < 0.001); however, sequence divergence between the 47 haplotypes detected was low (maximum 1.8% difference) and no deep phylogenetic divergence was observed. Results suggest the presence of genetic barriers isolating populations of the South China Sea and central Indonesian regions, which, in turn, show patterns of historical separation from northern Australian regions. In P. maxima, historical vicariance during Pleistocene low sea levels is likely to have restricted planktonic larval transport, causing genetic differentiation amongst populations. However, low genetic differentiation is observed where strong ocean currents are present and is most likely due to contemporary larval transport along these pathways. Geographical association with haplotype distributions may indicate signs of early lineage sorting arising from historical population separations, yet an absence of divergent phylogenetic clades related to geography could be the consequence of periodic pulses of high genetic exchange. We compare our results with previous microsatellite DNA analysis of these P. maxima populations, and discuss implications for future conservation management of this species. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 107 , 632–646.     

Contrasting patterns of nuclear microsatellite genetic structure of Fraxinus mandshurica var. japonica between northern and southern populations in Japan

Aim The aim of this study is to detect extant patterns of population genetic structure of Fraxinus mandshurica var. japonica in Japan, and to provide insights into the post‐glacial history of this species during the Holocene. Location Hokkaido and Honshu islands, Japan (including the Oshima and Shimokita peninsulas). Methods We examined nine polymorphic nuclear microsatellite loci to assess genetic variation within and among 15 populations across almost the entire range of the species in Japan. Extant patterns of geographical structure were analysed using Bayesian clustering, Monmonier’s algorithm, analysis of molecular variance, Mantel tests and principal coordinates analysis. Recent bottlenecks within populations and regional genetic variation were also assessed. Results Northern populations (Hokkaido Island and the Shimokita Peninsula) formed a single homogeneous deme, maintaining the highest level of allelic diversity on the Oshima Peninsula. By contrast, southern populations (Honshu Island) demonstrated strong substructure on both coasts. Specifically, populations on the Pacific side of Honshu exhibited significant bottlenecks and erosion of allelic diversity but preserved distinct subclusters diverging from widespread subclusters on the Japan Sea side of this island. Main conclusions Genetic evidence and life history traits suggest that F. mandshurica occupied cryptic northern refugia on the Oshima Peninsula during the Last Glacial Maximum, which is reflected in the species’ extant northern distribution. Strong geographical structure in southern populations, in agreement with fossil pollen records, suggests geographical isolation by mountain ranges running north–south along Honshu. Given that this tree species is cold‐adapted and found in riparian habitats, populations on the Pacific side of Honshu probably contracted into higher‐elevation swamps during warm post‐glacial periods, leading to a reduction of effective population sizes and rare allelic richness.     

Consequences of multiple mating‐system shifts for population and range‐wide genetic structure in a coastal dune plant

Evolutionary transitions from outcrossing to selfing can strongly affect the genetic diversity and structure of species at multiple spatial scales. We investigated the genetic consequences of mating‐system shifts in the North American, Pacific coast dune endemic plant Camissoniopsis cheiranthifolia (Onagraceae) by assaying variation at 13 nuclear (n) and six chloroplast (cp) microsatellite (SSR) loci for 38 populations across the species range. As predicted from the expected reduction in effective population size (N_e) caused by selfing, small‐flowered, predominantly selfing (SF) populations had much lower nSSR diversity (but not cpSSR) than large‐flowered, predominantly outcrossing (LF) populations. The reduction in nSSR diversity was greater than expected from the effects of selfing on N_e alone, but could not be accounted for by indirect effects of selfing on population density. Although selfing should reduce gene flow, SF populations were not more genetically differentiated than LF populations. We detected five clusters of nSSR genotypes and three groups of cpSSR haplotypes across the species range consisting of parapatric groups of populations that usually (but not always) differed in mating system, suggesting that selfing may often initiate ecogeographic isolation. However, lineage‐wide genetic variation was not lower for selfing clusters, failing to support the hypothesis that selection for reproductive assurance spurred the evolution of selfing in this species. Within three populations where LF and SF plants coexist, we detected genetic differentiation among diverged floral phenotypes suggesting that reproductive isolation (probably postzygotic) may help maintain the striking mating‐system differentiation observed across the range of this species.