Interspecific hybridization in Rorippa (Brassicaceae): Patterns and processes

The current knowledge on natural hybridization between European lowland Rorippa species is reviewed. Morphological, cytological and molecular markers provide substantial evidence that four Rorippa species (R. amphibia, R. austriaca, R. palustris and R. sylvestris) are involved in hybridization processes. The main factors promoting initial hybridization events between Rorippa species in Europe are (1) natural and anthropogenic habitat disturbance, (2) the invasion of non‐native R. austriaca into western central Europe and (3) the self‐incompatibility system of R. amphibia, R. austriaca and R. sylvestris coupled with clonal growth. Outcomes of hybridization between European Rorippa species show a broad range from incidental hybrids through the formation of hybrid swarms to the evolution of new hybrid taxa. Rorippa × armoracioides is certainly the most successful Rorippa hybrid. It results from hybridization between R. austriaca and R. sylvestris and is known from many localities in the native range of R. austriaca and in regions invaded by R. austriaca. Within this system hybrid zones of different age provide the rare opportunity to analyse different stages of hybrid formation and hybrid speciation in natural populations.     

The genomic consequences of adaptive divergence and reproductive isolation between species of manakins

The processes of adaptation and speciation are expected to shape genomic variation within and between diverging species. Here we analyze genomic heterogeneity of genetic differentiation and introgression in a hybrid zone between two bird species (Manacus candei and M. vitellinus) using 59 100 SNPs, a whole genome assembly, and Bayesian models. Measures of genetic differentiation () and introgression (genomic cline center [α] and rate [β]) were highly heterogeneous among loci. We identified thousands of loci with elevated parameter estimates, some of which are likely to be associated with variation in fitness in Manacus populations. To analyze the genomic organization of differentiation and introgression, we mapped SNPs onto a draft assembly of the M. vitellinus genome. Estimates of , α, and β were autocorrelated at very short physical distances (< 100 bp), but much less so beyond this. In addition, average statistical associations (linkage disequilibrium) between SNPs were generally low and were not higher in admixed populations than in populations of the parental species. Although they did not occur with a constant probability across the genome, loci with elevated , α, and β were not strongly co-localized in the genome. Contrary to verbal models that predict clustering of loci involved in adaptation and isolation in discrete genomic regions, these results are consistent with the hypothesis that genetic regions involved in adaptive divergence and reproductive isolation are scattered throughout the genome. We also found that many loci were characterized by both exceptional genetic differentiation and introgression, consistent with the hypothesis that loci involved in isolation are also often characterized by a history of divergent selection. However, the concordance between isolation and differentiation was only partial, indicating a complex architecture and history of loci involved in isolation.     

Glacial refugia of limber pine (Pinus flexilis James) inferred from the population structure of mitochondrial DNA

To make inferences about the glacial refugia that harboured the limber pine, Pinus flexilis James, we examined the range-wide population structure of mitochondrial DNA (mtDNA) with eight size variants in the second intron of nad1. The data consisted of haplotypes from 704 trees collected from 40 localities. The value of FST for these populations was 0.80, which is a much larger value than has been reported for allozymes and chloroplast DNA (cpDNA) in limber pine, and it suggests that the number of seeds moving among localities per generation is approximately 0.12. Gene flow of this magnitude would allow mutation and subsequent genetic drift to have a substantial impact on the population structure of mtDNA. The majority of the mtDNA haplotypes are restricted to minor portions of the geographical range. The data are consistent with mtDNA differentiation in seven glacial refugia, followed by dispersal out of those refugia.     

Primers designed to amplify a mitochondrial nad1 intron in ponderosa pine, Pinus ponderosa, limber pine, P. flexilis, and Scots pine, P. sylvestris

The b/c intron of the mitochondrial nad1 gene, was sequenced to characterize the indel region of ponderosa pine, Pinus ponderosa. The sequence in ponderosa pine was aligned with the sequence in Scots pine, Pinus sylvestris, to design seven primers that are useful for sequencing and for revealing size variation in amplified fragments in ponderosa pine, Scots pine, and limber pine, Pinus flexilis. These primers reveal variability in all three species, and the pattern of variability within ponderosa pine is described by a preliminary survey. The indel region of ponderosa pine contains three distinct elements with lengths of 31, 32, and 34 bp. Received: 1 March 2000 / Accepted: 14 April 2000<@head-com-p1a.lf>Communicated by P.M.A. Tigerstedt     

A set of novel DNA polymorphisms within candidate genes potentially involved in ecological divergence between Populus alba and P. tremula, two hybridizing European forest trees

We have identified 53 DNA (single nucleotide, microsatellite, and insertion-deletion) polymorphisms within 12 candidate genes potentially involved in ecological differences between Populus alba and P. tremula, two hybridizing European forest trees. The genes represent candidates for functional roles associated with abiotic or biotic stress response, cross-talk, phenology, and leaf development. Distributions within sequences, intraspecific levels of diversity, and genetic divergence (F(ST) ) between species are reported for each polymorphism, as are haplotype frequencies for each gene. The markers will be used for population genomic studies of the barrier to gene flow between these two ecologically divergent forest trees.     

Climatic adaptation and ecological divergence between two closely related pine species in Southeast China

Climate is one of the most important drivers for adaptive evolution in forest trees. Climatic selection contributes greatly to local adaptation and intraspecific differentiation, but this kind of selection could also have promoted interspecific divergence through ecological speciation. To test this hypothesis, we examined intra‐ and interspecific genetic variation at 25 climate‐related candidate genes and 12 reference loci in two closely related pine species, Pinus massoniana Lamb. and Pinus hwangshanensis Hisa, using population genetic and landscape genetic approaches. These two species occur in Southeast China but have contrasting ecological preferences in terms of several environmental variables, notably altitude, although hybrids form where their distributions overlap. One or more robust tests detected signals of recent and/or ancient selection at two‐thirds (17) of the 25 candidate genes, at varying evolutionary timescales, but only three of the 12 reference loci. The signals of recent selection were species specific, but signals of ancient selection were mostly shared by the two species likely because of the shared evolutionary history. F_ST outlier analysis identified six SNPs in five climate‐related candidate genes under divergent selection between the two species. In addition, a total of 24 candidate SNPs representing nine candidate genes showed significant correlation with altitudinal divergence in the two species based on the covariance matrix of population history derived from reference SNPs. Genetic differentiation between these two species was higher at the candidate genes than at the reference loci. Moreover, analysis using the isolation‐with‐migration model indicated that gene flow between the species has been more restricted for climate‐related candidate genes than the reference loci, in both directions. Taken together, our results suggest that species‐specific and divergent climatic selection at the candidate genes might have counteracted interspecific gene flow and played a key role in the ecological divergence of these two closely related pine species.     

Contrasting patterns of hybridization in large house spiders (Tegenaria atrica group, Agelenidae)

The integrity of species is not fixed and may vary geographically. Here we investigate the geographic distributions and interactions of species in the Tegenaria atrica group (Araneae: Agelenidae). Detailed mapping of T. saeva and T. gigantea in England and Wales shows them to be broadly allopatric in southern England with a tightly defined, and possibly long-standing, narrow zone of parapatry in central southern England. In the north of England (Yorkshire), by contrast, the species are broadly sympatric as a result of recent range expansions. GIS techniques are used to map the species distributions and to quantify, we believe for the first time, the intimacy of interspecific interactions. The extent and nature of hybridization in these two areas is examined through regression and multivariate analyses of morphology. We show that the relative incidence of hybridization is much greater in Yorkshire than within the parapatric zone in the south. Clear patterns of asymmetric introgression are observed in both northern and southern England, with a greater impact of T. gigantea on T. saeva than vice versa. We find no sign of morphological reproductive character displacement at the zone of parapatry that might indicate reinforcement, although we cannot exclude more subtle effects, for example via cuticular pheromones. The integrity of these two species seems to be breaking down in northern England, a process that might gain momentum as the gene pools become more similar.     

Contrasting patterns of mitochondrial DNA and microsatellite introgressive hybridization between lineages of lake whitefish (Coregonus clupeaformis); relevance for speciation

We performed a combined analysis of mitochondrial DNA (mtDNA) and microsatellite loci among lake whitefish (Coregonus clupeaformis) populations in order to assess the levels of congruence between both types of markers in defining patterns of genetic structuring, introgressive hybridization and inferring population origins in the hybrid zone of the St. John River basin. A second objective was to test the hypothesis that secondary contact between glacial lineages always resulted in the occurrence of sympatric dwarf and normal whitefish ecotypes. Fish were sampled from 35 populations and polymorphism was screened at mtDNA and six microsatellite loci for a total of 688 and 763 whitefish, respectively. Four lakes harbouring a single whitefish population of normal ecotype admixed with mtDNA haplotypes of different lineages were found. This confirmed that secondary contact between whitefish evolutionary lineages did not always result in the persistence of reproductively isolated ecotypes. Microsatellites further supported the definition of distinct glacial lineages by identifying lineage-specific allelic size groups. They also further supported the hypothesis that ecotypes originated from either a single founding lineage (sympatric divergence) or following secondary contacts between lineages (allopatric divergence), depending on the lake. In general, however, the pattern of population differentiation and introgressive hybridization observed at microsatellites was in sharp contrast with that depicted by mtDNA variation. Both factorial correspondence analysis and analysis of admixture proportion revealed a much more pronounced pattern of introgressive hybridization than depicted by mtDNA analyses. Variable levels of introgression indicated that environmental differences may be as important as the historical contingency of secondary contact in explaining the persistence of sympatric ecotypes and the differential pattern of introgressive hybridization among lakes. Whitefish populations from the St. John River basin hybrid zone represent a rare illustration of a continuum of both morphological and genetic differentiation within a given taxon, spanning from complete introgression to possibly complete reproductive isolation, depending on lakes. Thus, each lake may be viewed as a different temporal snapshot taken throughout the gradual process of speciation.     

Asymmetric introgression between the M and S forms of the malaria vector, Anopheles gambiae, maintains divergence despite extensive hybridization

The suggestion that genetic divergence can arise and/or be maintained in the face of gene flow has been contentious since first proposed. This controversy and a rarity of good examples have limited our understanding of this process. Partially reproductively isolated taxa have been highlighted as offering unique opportunities for identifying the mechanisms underlying divergence with gene flow. The African malaria vector, Anopheles gambiae s.s., is widely regarded as consisting of two sympatric forms, thought by many to represent incipient species, the M and S molecular forms. However, there has been much debate about the extent of reproductive isolation between M and S, with one view positing that divergence may have arisen and is being maintained in the presence of gene flow, and the other proposing a more advanced speciation process with little realized gene flow because of low hybrid fitness. These hypotheses have been difficult to address because hybrids are typically rare (<1%). Here, we assess samples from an area of high hybridization and demonstrate that hybrids are fit and responsible for extensive introgression. Nonetheless, we show that strong divergent selection at a subset of loci combined with highly asymmetric introgression has enabled M and S to remain genetically differentiated despite extensive gene flow. We propose that the extent of reproductive isolation between M and S varies across West Africa resulting in a 'geographic mosaic of reproductive isolation'; a finding which adds further complexity to our understanding of divergence in this taxon and which has considerable implications for transgenic control strategies.     

Divergent selection and heterogeneous genomic divergence

Levels of genetic differentiation between populations can be highly variable across the genome, with divergent selection contributing to such heterogeneous genomic divergence. For example, loci under divergent selection and those tightly physically linked to them may exhibit stronger differentiation than neutral regions with weak or no linkage to such loci. Divergent selection can also increase genome‐wide neutral differentiation by reducing gene flow (e.g. by causing ecological speciation), thus promoting divergence via the stochastic effects of genetic drift. These consequences of divergent selection are being reported in recently accumulating studies that identify: (i) ‘outlier loci’ with higher levels of divergence than expected under neutrality, and (ii) a positive association between the degree of adaptive phenotypic divergence and levels of molecular genetic differentiation across population pairs [‘isolation by adaptation’ (IBA)]. The latter pattern arises because as adaptive divergence increases, gene flow is reduced (thereby promoting drift) and genetic hitchhiking increased. Here, we review and integrate these previously disconnected concepts and literatures. We find that studies generally report 5–10% of loci to be outliers. These selected regions were often dispersed across the genome, commonly exhibited replicated divergence across different population pairs, and could sometimes be associated with specific ecological variables. IBA was not infrequently observed, even at neutral loci putatively unlinked to those under divergent selection. Overall, we conclude that divergent selection makes diverse contributions to heterogeneous genomic divergence. Nonetheless, the number, size, and distribution of genomic regions affected by selection varied substantially among studies, leading us to discuss the potential role of divergent selection in the growth of regions of differentiation (i.e. genomic islands of divergence), a topic in need of future investigation.     

Direct and indirect analysis of the fitness of Chrysochus (Coleoptera: Chrysomelidae) hybrids

The chrysomelid beetles Chrysochus auratus and C. cobaltinus form a narrow hybrid zone in western North America. We used a combination of direct and indirect analyses to examine the fitness of Chrysochus hybrids. For the direct analyses, we compared the mating frequency, longevity, fecundity and fertility of hybrid females and parentals. For the indirect approach, we tested predictions of multilocus genotype frequencies at a focal site in the hybrid zone, based on the frequencies of mating combinations during the previous generation. Hybrid females produced fewer eggs than did parentals and the eggs they produced in the lab failed to hatch, in contrast to those of parental females. In addition, contrary to predictions that 15.8% of the individuals at the focal site would have multilocus genotypes other than those expected of parentals or F₁ individuals, we found no such genotypes at this site. This hybrid zone appears to be an example of a classic tension zone, with endogenous selection against hybrid individuals. We discuss the implications of low hybrid fitness for the evolution of premating barriers in this system, and argue that the integration of direct and indirect approaches is a powerful means of assessing the relative fitness of hybrids, particularly for species in which mate choices are easy to observe in the field.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 84 , 273–286.     

Root induction in pine (Pinus) and larch (Larix) spp. using Agrobacterium rhizogenes

Root induction on in vitro adventitious Pinus monticola Dougl. shoots from mature embryos was improved after 8 weeks co-cultivation with Agrobacterium rhizogenes Conn. strain A4 or the pRi transconjugant strain R1000 as compared to controls, and to naphthaleneacetic acid and indoleacetic acid treatments. An improvement in the number and quality of roots induced was observed on co-cultivated shoots as well as co-cultivated Pinus banksiana Lamb and Larix laricina (DuRoi) K. Koch de-rooted seedlings. Changes in substrate nutrient solution did not influence rooting as much as the presence of Agrobacterium in the rhizosphere. Rooting of cuttings from 3-year-old P. monticola trees was improved in two different seedlots with A. rhizogenes A4 strain.     

The genomics of adaptation,divergence and speciation: a congealing theory

In this issue, Flaxman et al. ( 2014 ) report the results of sophisticated whole‐genome simulations of speciation with gene flow, enhancing our understanding of the process by building on previous single‐locus, multilocus and analytical works. Their findings provide us with new insights about how genomes can diverge and the importance of statistical and chromosomal linkage in facilitating reproductive isolation. The authors characterize the conditions under which, even with high gene flow and weak divergent selection, reproductive isolation between populations can occur due to the emergent stochastic process of genomewide congealing, where numerous statistically or physically linked loci of small effect allow selection to limit effective migration rates. The initial congealing event can occur within a broad range conditions, and once initiated, the self‐reinforcing process leads to rapid divergence and ultimately two reproductively isolated populations. Flaxman et al.'s ( 2014 ) work is a valuable contribution to our understanding of speciation with gene flow and in making a more predictive field of evolutionary genomics and speciation.     

Mortality dynamics of Daphnia in contrasting habitats and their role in ecological divergence

1. The Daphnia pulex‐pulicaria species complex has been proposed as an example of rapid ecological speciation, associated with divergence along the gradient of waterbody size from temporary ponds to deep, stratified lakes. However, this divergence is incomplete, and thus represents an opportunity to study ecological divergence as it is occurring. 2. Dynamics of twelve populations of Daphnia in the pulex‐pulicaria species complex were monitored over 1 year. Six temporary pond populations and six permanent lake populations were compared to evaluate demographic differences that may contribute to ecological divergence in this complex. 3. Pond populations experienced greater changes in density, which were reflected in more extreme growth rates, higher birth rates and higher mortality rates than those of lake populations. 4. Mitochondrial DNA was isolated from up to three clones from each population, the D‐loop of the control region was sequenced, and a phylogenetic tree was constructed. This tree revealed two strongly supported clades. The clades were not congruent with habitat type and nominal status, indicating that interhabitat gene flow occurs easily and that the nominal taxa are incompletely diverged. 5. Published reports of genetic life history differences in the D. pulex‐pulicaria complex are consistent with the demographic differences reported here. This suggests that ecological differences between the habitats are selectively maintaining trait differences despite the possibility for genetic exchange. Thus, these taxa may be at the inception of ecological speciation.     

Molecular and morphological evidence of hybridization between newts Triturus vulgaris and T. montandoni (Caudata: Salamandridae) in Slovakia

Natural hybridization and introgression is recorded between the newt species Triturus vulgaris and T. montandoni in Slovakia. To confirm a hybrid status of two putative hybrids, morphological and Randomly Amplified Polymorphic DNA (RAPD) markers specific for T. vulgaris and T. montandoni were used. The individuals found in Zbojská (Veporské vrchy Mts, Slovakia; analyzed morphologically and genetically) and Zubrohlava (Oravská kotlina basin, Slovakia; analyzed only morphologically) possessed markers of both species. Segregation of RAPD markers together with a model-based Bayesian analysis revealed that the specimen from Zbojská belonged to later generation hybrids.     

Patterns of genetic,phenotypic, and acoustic variation across a chiffchaff (Phylloscopus collybita abietinus/tristis) hybrid zone

Characterizing patterns of evolution of genetic and phenotypic divergence between incipient species is essential to understand how evolution of reproductive isolation proceeds. Hybrid zones are excellent for studying such processes, as they provide opportunities to assess trait variation in individuals with mixed genetic background and to quantify gene flow across different genomic regions. Here, we combine plumage, song, mtDNA and whole‐genome sequence data and analyze variation across a sympatric zone between the European and the Siberian chiffchaff (Phylloscopus collybita abietinus/tristis) to study how gene exchange between the lineages affects trait variation. Our results show that chiffchaff within the sympatric region show more extensive trait variation than allopatric birds, with a large proportion of individuals exhibiting intermediate phenotypic characters. The genomic differentiation between the subspecies is lower in sympatry than in allopatry and sympatric birds have a mix of genetic ancestry indicating extensive ongoing and past gene flow. Patterns of phenotypic and genetic variation also vary between regions within the hybrid zone, potentially reflecting differences in population densities, age of secondary contact, or differences in mate recognition or mate preference. The genomic data support the presence of two distinct genetic clades corresponding to allopatric abietinus and tristis and that genetic admixture is the force underlying trait variation in the sympatric region—the previously described subspecies (“fulvescens”) from the region is therefore not likely a distinct taxon. In addition, we conclude that subspecies identification based on appearance is uncertain as an individual with an apparently distinct phenotype can have a considerable proportion of the genome composed of mixed alleles, or even a major part of the genome introgressed from the other subspecies. Our results provide insights into the dynamics of admixture across subspecies boundaries and have implications for understanding speciation processes and for the identification of specific chiffchaff individuals based on phenotypic characters.     

Hybridization among sympatric species of Rhododendron (Ericaceae) in Turkey: morphological and molecular evidence

Rhododendron (Ericaceae) is a large genus in which barriers to hybridization are especially weak, but many species are maintained in sympatry. Hybridization among four species of Rhododendron subsect. Pontica, which occur in sympatry in Turkey, was investigated. Material of R. ponticum, R. smirnovii, R. ungernii, and R. caucasicum and their putative hybrids was collected from the wild. Based on morphology, chloroplast DNA and nuclear ribosomal DNA restriction fragment length polymorphism (RFLP) profiles, each accession was identified as a species or hybrid combination. Five of the six possible hybrid combinations among the four species were detected. Rhododendron ponticum × R. smirnovii was represented by a single individual and R. caucasicum × R. smirnovii by one small group of hybrid plants. The combinations R. ponticum × R. ungernii and R. ungernii × R. smirnovii showed evidence of frequent backcrossing, while R. ponticum × R. caucasicum appeared unusual in that an intermediate hybrid type was abundant, whereas hybrids with phenotypes approaching either parent were rare. Possible explanations of this latter situation are discussed. The results suggest that natural hybridization among Rhododendron species is common and that ecological factors are important in maintaining integrity when species occur in sympatry.     

Patterns of genetic diversity and candidate genes for ecological divergence in a homoploid hybrid sunflower, Helianthus anomalus

Natural hybridization accompanied by a shift in niche preference by hybrid genotypes can lead to hybrid speciation. Natural selection may cause the fixation of advantageous alleles in the ecologically diverged hybrids, and the loci experiencing selection should exhibit a reduction in allelic diversity relative to neutral loci. Here, we analyzed patterns of genetic diversity at 59 microsatellite loci associated with expressed sequence tags (ESTs) in a homoploid hybrid sunflower species, Helianthus anomalus. We used two indices, ln RV and ln RH, to compare variation and heterozygosity (respectively) at each locus between the hybrid species and its two parental species, H. annuus and H. petiolaris. Mean values of ln RV and ln RH were significantly lower than zero, which implies that H. anomalus experienced a population bottleneck during its recent evolutionary history. After correcting for the apparent bottleneck, we found six loci with a significant reduction in variation or with heterozygosity in the hybrid species, compared to one or both of the parental species. These loci should be viewed as a ranked list of candidate loci, pending further sequencing and functional analyses. Sequence data were generated for two of the candidate loci, but population genetics tests failed to detect deviations from neutral evolution at either locus. Nonetheless, a greater than eight-fold excess of nonsynonymous substitutions was found near a putative N-myristoylation motif at the second locus (HT998), and likelihood-based models indicated that the protein has been under selection in H. anomalus in the past and, perhaps, in one or both parental species. Finally, our data suggest that selective sweeps may have united populations of H. anomalus isolated by a mountain range, indicating that even low gene-flow species may be held together by the spread of advantageous alleles.