The isolation of populations of C. americanum in multiple refugia has led to a degree of phylogeographic structure greater than that found in most previously studied plants in eastern North America, which may be attributable to its short generation time. Reproductively isolated populations of C. americanum belong to divergent lineages, which suggests that survival in multiple glacial refugia contributed to the development of reproductive isolation in this species.

Genetic incompatibility is a hallmark of speciation. Cytonuclear incompatibilities are proposed to be among the first genetic barriers to arise during speciation. Accordingly, reproductive isolation (RI) within species should be heavily influenced by interactions between the organelle and nuclear genomes. However, there are few clear examples of cytonuclear incompatibility within a species. Here, we show substantial postzygotic RI in first-generation hybrids between differentiated populations of an herbaceous plant (up to 92% reduction in fitness). RI was primarily due to germination and survival, with moderate RI for pollen viability. RI for survival was asymmetric and caused by cytonuclear incompatibility, with the strength of incompatibility linearly related to chloroplast genetic distance. This cytonuclear incompatibility may be the result of a rapidly evolving plastid genome. Substantial asymmetric RI was also found for germination, but was not associated with cytonuclear incompatibility, indicating endosperm or maternal-zygote incompatibilities. These results demonstrate that cytonuclear incompatibility contributes to RI within species, suggesting that initial rates of speciation could be influenced by rates of organelle evolution. However, other genetic incompatibilities are equally important, indicating that even at early stages, speciation can be a complex process involving multiple genes and incompatibilities.

BackgroundAlthough the plastid genome is highly conserved across most angiosperms, multiple lineages have increased rates of structural rearrangement and nucleotide substitution. These lineages exhibit an excess of nonsynonymous substitutions (i.e., elevated dN/dS ratios) in similar subsets of plastid genes, suggesting that similar mechanisms may be leading to relaxed and/or positive selection on these genes. However, little is known regarding whether these mechanisms continue to shape sequence diversity at the intraspecific level.ResultsWe examined patterns of interspecific divergence and intraspecific polymorphism in the plastid genome of Campanulastrum americanum, and across plastid genes found a significant correlation between dN/dS and pN/pS (i.e., the within-species equivalent of dN/dS). A number of genes including ycf1, ycf2, clpP, and ribosomal protein genes exhibited high dN/dS ratios. McDonald-Kreitman tests detected little evidence for positive selection acting on these genes, likely due to the presence of substantial intraspecific divergence.ConclusionsThese results suggest that mechanisms leading to increased nucleotide substitution rates in the plastid genome are continuing to act at the intraspecific level. Accelerated plastid genome evolution may increase the likelihood of intraspecific cytonuclear genetic incompatibilities, and thereby contribute to the early stages of the speciation process.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-014-0268-y) contains supplementary material, which is available to authorized users.

Although organelle inheritance is predominantly maternal across animals and plants, biparental chloroplast inheritance has arisen multiple times in the angiosperms. Biparental inheritance has the potential to impact the evolutionary dynamics of cytonuclear incompatibility, interactions between nuclear and organelle genomes that are proposed to be among the earliest types of genetic incompatibility to arise in speciation. We examine the interplay between biparental inheritance and cytonuclear incompatibility in Campanulastrum americanum, a plant species exhibiting both traits. We first determine patterns of chloroplast inheritance in genetically similar and divergent crosses, and then associate inheritance with hybrid survival across multiple generations. There is substantial biparental inheritance in C. americanum. The frequency of biparental inheritance is greater in divergent crosses and in the presence of cytonuclear incompatibility. Biparental inheritance helps to mitigate cytonuclear incompatibility, leading to increased fitness of F hybrids and recovery in the F generation. This study demonstrates the potential for biparental chloroplast inheritance to rescue cytonuclear compatibility, reducing cytonuclear incompatibility's contribution to reproductive isolation and potentially slowing speciation. The efficacy of rescue depended upon the strength of incompatibility, with a greater persistence of weak incompatibilities in later generations. These findings suggest that incompatible plastids may lead to selection for biparental inheritance.

Phenology is the timing of life cycle events of an organism. Alterations in phenology can have profound effects on individual fitness, population growth, and community dynamics. Recent changes in climate have altered the phenology of many organisms, which may result in selection to shift phenological traits. Understanding the relationship between local climates and population differentiation in phenology will allow us to anticipate responses to novel selective environments caused by global climate change. We evaluated population differentiation in the number of days to germination, first flower, and fruit maturation for 33 populations throughout the range of Campanulastrum americanum (American Bellflower). Germination and fruit maturation had geographical clines with earlier timing in populations from northern latitudes. Northern sites were cooler and drier, suggesting potential adaptive differentiation of the shorter life cycle associated with earlier phenology. Similarly, higher elevations were cooler and had earlier fruit maturation. However, seed germination was later in higher elevation populations. Although there was substantial variation in the day to first flower, ranging 40 days between population means, it was idiosyncratic and not related to latitude, suggesting differentiation in response to selective factors distinct from those on germination and fruit maturation. Thus, germination and fruit maturation in C. americanum may shift in response to selection by rising temperatures. However, such changes are not expected for flowering time, a typical indicator of climate change.