Variability in space and time: contrasting fruit distribution patterns in the deceptive orchid Orchis militaris.

In angiosperms, a decrease in fruit production towards the apex of individual inflorescences is usually observed. Orchids are thought to be primarily pollination-limited species, and non-uniform pollination could cause this decrease pattern in several species. Fruit production was investigated in relation to flower position and floral display size in Orchis militaris (Orchidaceae), a deceptive species. Over 2 years, eight populations of O. militaris were studied and fruit position along the inflorescence was recorded. Generalised linear models were performed to examine the effect of population, year, flower position and floral display size on fruit production. The dominant pattern was characterised by a higher fruit set in the middle part of the inflorescence (parabolic pattern). A non-directional pattern of fruit production was also detected in some populations. Within a given population, patterns were generally consistent among years. In one of the two study years and in one of the eight populations specifically, the proximal-to-distal decrease in fruit production was dramatic in plants with a large floral display but weak or absent in small displays. Our study demonstrates the intraspecific diversity of fruit distribution patterns in O. militaris. Non-uniform pollination along the inflorescence is likely to be responsible for the parabolic pattern, while irregular visitation could explain the non-directional pattern of fruit production. Pattern variation among years and between populations could arise from spatiotemporal variation in pollinator assemblages. Resource competition effects could explain the interaction effect between display size and flower position.

Frequency and spatial patterning of clonal reproduction in Louisiana iris hybrid populations.

The plant genera in which natural hybridization is most prevalent tend to be outcrossing perennials with some mechanism for clonal (i.e., asexual) reproduction. Although clonal reproduction in fertile, sexually reproducing hybrid populations could have important evolutionary consequences, little attention has been paid to quantifying this parameter in such populations. In the present study, we examined the frequency and spatial patterning of clonal reproduction in two Louisiana iris hybrid populations. Allozyme analysis of both populations revealed relatively high levels of genotypic diversity. However, a considerable amount of clonality was apparent. Nearly half of all genets (47%) in one population and more than half (61%) in the other had multiple ramets. Furthermore, both populations exhibited relatively high levels of genetic structuring, a pattern that resulted from the aggregation of clonal ramets. The occurrence of clonal reproduction in hybrid populations could not only facilitate introgression through an increase in the number of flowering ramets per genet and/or the survivorship of early generation hybrids, but might also influence the mating system of such populations. Any potential increase in the selfing rate due to cross-pollination among ramets of the same genet may, in turn, increase the likelihood of homoploid hybrid speciation.

Outcrossing rates in two self-compatible, hybridising Rhinanthus species: implications for hybrid formation.

The congeners Rhinanthus angustifolius and Rhinanthus minor, two annual hemiparasites pollinated by bumblebees, are known to hybridise in the wild. Both species are self-compatible, but the capacity for autonomous selfing is higher in R. minor. This suggests a difference in realized outcrossing rates, which have not been determined before in these species. Using microsatellites, both species turned out to have mixed mating systems, but with a much lower multilocus outcrossing rate (0.13) for R. minor compared to R. angustifolius (0.76). We hypothesised that a higher outcrossing rate should lead to a higher chance of heterospecific pollination, and we therefore determined the rate of hybrid formation on each species in an artificial mixed population. Hybrid seeds were produced at low frequency (4.5%), and no significant difference was found between the species. It is therefore likely that post-pollination processes influence hybrid seed formation to counteract the expected difference in heterospecific pollen deposition. We checked fruit set, seed set and the rate of autonomous selfing in controlled crosses in the greenhouse in 2 years, and found that fruit set (2003) or seed set (2010) were lower in R. angustifolius × R. minor crosses relative to the reciprocal cross. Hybrid seeds produced on R. angustifolius also had a much lower germination rate, so most of the established F1 hybrid plants have the R. minor cytoplasm. The formation of advanced hybrids depends on pollinator preference, which is biased towards R. angustifolius if present in sufficient numbers, because it offers more rewards.

PERMANENT GENETIC RESOURCES: Development of microsatellite markers in Rhinanthus angustifolius and cross-species amplification.

Fifteen polymorphic microsatellite loci were developed from an enriched genomic library of the annual plant Rhinanthus angustifolius and characterized using 36 individuals. These markers provided high polymorphism ranging from two to 15 alleles per locus. Four loci showed significant departure from Hardy-Weinberg equilibrium, probably because of the occurrence of null alleles. No significant linkage disequilibrium was detected between pairs of loci. Tests of cross-species transferability were performed on four congeners with a success rate of 100% in Rhinanthus minor, 93% in R. mediterraneus and R. glacialis, and 80% in R. alectorolophus. These microsatellite loci will be useful tools to study mating system, gene flow and hybridization in the genus Rhinanthus.

Intimately linked or hardly speaking? The relationship between genotype and environmental gradients in a Louisiana Iris hybrid population.

Several models of hybrid zone evolution predict the same spatial patterns of genotypic distribution whether or not structuring is due to environment-dependent or -independent selection. In this study, we tested for evidence of environment-dependent selection in an Iris fulva x Iris brevicaulis hybrid population by examining the distribution of genotypes in relation to environmental gradients. We selected 201 Louisiana Iris plants from within a known hybrid population (80 m x 80 m) and placed them in four different genotypic classes (I. fulva, I. fulva-like hybrid, I. brevicaulis-like hybrid and I. brevicaulis) based on seven species-specific random amplified polymorphic DNA (RAPD) markers and two chloroplast DNA haplotypes. Environmental variables were then measured. These variables included percentage cover by tree canopy, elevation from the high water mark, soil pH and percentage soil organic matter. Each variable was sampled for all 201 plants. Canonical discriminant analysis (CDA) was used to infer the environmental factors most strongly associated with the different genotypic groups. Slight differences in elevation (-0.5 m to +0.4 m) were important for distinguishing habitat distributions described by CDA, even though there were no statistical differences between mean elevations alone. I. brevicaulis occurred in a broad range of habitats, while I. fulva had a narrower distribution. Of all the possible combinations, I. fulva-like hybrids and I. brevicaulis-like hybrids occurred in the most distinct habitat types relative to one another. Each hybrid class was not significantly different from its closest parent with regard to habitat occupied, but was statistically unique from its more distant parental species. Within the hybrid genotypes, most, but not all, RAPD loci were individually correlated with environmental variables. This study suggests that, at a very fine spatial scale, environment-dependent selection contributed to the genetic structuring of this hybrid zone.