Seasonal changes in pollinator activity influence pollen dispersal and seed production of the alpine shrub Rhododendron aureum (Ericaceae).

In alpine ecosystems, microscale variation in snowmelt timing often causes different flowering phenology of the same plant species and seasonal changes in pollinator activity. We compared the variations in insect visitation, pollen dispersal, mating patterns, and sexual reproduction of Rhododendron aureum early and late in the flowering season using five microsatellites. Insects visiting the flowers were rare early in the flowering season (mid-June), when major pollinators were bumblebee queens and flies. In contrast, frequent visitations by bumblebee workers were observed late in the season (late July). Two-generation analysis of pollen pool structure demonstrated that quality of pollen-mediated gene flow was more diverse late in the season in parallel with the high pollinator activity. The effective number of pollen donors per fruit (N(ep)) increased late in the season (N(ep) = 2.2-2.7 early, 3.4-4.4 late). However, both the outcrossing rate (t(m)) and seed-set ratio per fruit were smaller late in the season (t(m) = 0.89 and 0.71, seed-set ratio = 0.52 and 0.18, early and late in the season, respectively). In addition, biparental inbreeding occurred only late in the season. We conclude that R. aureum shows contrasting patterns of pollen movement and seed production between early and late season: in early season, seed production can be high but genetically less diverse and, during late season, be reduced, possibly due to higher inbreeding and inbreeding depression, but have greater genetic diversity. Thus, more pollinator activity does not always mean more pollen movement.

Effect of gene flow on spatial genetic structure in the riparian canopy tree Cercidiphyllum japonicum revealed by microsatellite analysis.

Few studies have analyzed pollen and seed movements at local scale, and genetic differentiation among populations covering the geographic distribution range of a species. We carried out such a study on Cercidiphyllum japonicum; a dioecious broad-leaved tree of cool-temperate riparian forest in Japan. We made direct measurement of pollen and seed movements in a site, genetic structure at the local scale, and genetic differentiation between populations covering the Japanese Archipelago. Parentage analysis of seedlings within a 20-ha study site indicated that at least 28.8% of seedlings were fertilized by pollen from trees outside the study site. The average pollination distance within the study site was 129 m, with a maximum of 666 m. The genotypes of 30% of seedlings were incompatible with those of the nearest female tree, and the maximum seed dispersal distance within the study site was over 300 m. Thus, long-distance gene dispersal is common in this species. The correlation between genetic relatedness and spatial distance among adult trees within the population was not significant, indicating an absence of fine-scale genetic structure perhaps caused by high levels of pollen flow and overlapping seed shadows. Six populations sampled throughout the distribution of C. japonicum in Japan showed significant isolation-by-distance but low levels of genetic differentiation (F(ST) = 0.043), also indicating long-distance gene flow in C. japonicum. Long-distance gene flow had a strong influence on the genetic structure at different spatial scales, and contributes to the maintenance of genetic diversity in C. japonicum.

Variation in pollen dispersal between years with different pollination conditions in a tropical emergent tree.

We examined differences in pollen dispersal efficiency between 2 years in terms of both spatial dispersal range and genetic relatedness of pollen in a tropical emergent tree, Dipterocarpus tempehes. The species was pollinated by the giant honeybee (Apis dorsata) in a year of intensive community-level mass-flowering or general flowering (1996), but by several species of moths in a year of less-intensive general flowering (1998). We carried out paternity analysis based on six DNA microsatellite markers on a total of 277 mature trees forming four spatially distinct subpopulations in a 70 ha area, and 147 and 188 2-year-old seedlings originating from seeds produced in 1996 and 1998 (cohorts 96 and 98, respectively). Outcrossing rates (0.93 and 0.96 for cohorts 96 and 98, respectively) did not differ between years. Mean dispersal distances (222 and 192 m) were not significantly different between the 2 years but marginally more biased to long distance in 1996. The mean relatedness among cross-pollinated seedlings sharing the same mothers in cohort 96 was lower than that in cohort 98. This can be attributed to the two facts that the proportion of intersubpopulations pollen flow among cross-pollination events was marginally higher in cohort 96 (44%) than in cohort 98 (33%), and that mature trees within the same subpopulations are genetically more related to each other than those between different subpopulations. We conclude that D. tempehes maintained effective pollen dispersal in terms of outcrossing rate and pollen dispersal distance in spite of the large difference in foraging characteristics between two types of pollinators. In terms of pollen relatedness, however, a slight difference was suggested between years in the level of biparental inbreeding.

Patterns and levels of gene flow in Rhododendron metternichii var. hondoense revealed by microsatellite analysis.

Parentage analysis was conducted to elucidate the patterns and levels of gene flow in Rhododendron metternichii Sieb. et Zucc. var. hondoense Nakai in a 150 x 70 m quadrant in Hiroshima Prefecture, western Japan. The population of R. metternichii occurred as three subpopulations at the study site. Seventy seedlings were randomly collected from each of three 10 x 10 m plots (S1, S2, and S3) on the forest floor of each subpopulation (A1, A2, and A3). Almost all parents (93.8%) of the 70 seedlings were unambiguously identified by using 12 pairs of microsatellite markers. Within the quadrant, adult trees less than 5 m from the centre of the seedling bank (plots S1, S2, and S3) produced large numbers of seedlings. The effects of tree height and distance from the seedling bank on the relative fertilities of adult trees were highly variable among subpopulations because of the differences in population structure near the seedling bank: neither distance nor tree height had any significant effect in subpopulation A1; distance from the seedling bank had a significant effect in subpopulation A2; and tree height had a significant effect in subpopulation A3. Although gene flow within each subpopulation was highly restricted to less than 25 m and gene flow among the three subpopulations was extremely small (0-2%), long-distance gene flow from outside the quadrant reached 50%. This long-distance gene flow may be caused by a combination of topographical and vegetational heterogeneity, differences in flowering phenology, and genetic substructuring within subpopulations.

Microsatellite analysis of the regeneration process of Magnolia obovata Thunb.

We analysed the regeneration process of Magnolia obovata using polymorphic microsatellite markers. Eighty-three adult trees standing in a watershed covering an area of 69 ha, and saplings collected from a smaller research plot (6 ha) located at the centre of the watershed were genotyped using microsatellite markers. Among 91 saplings analysed, 24 (26%) had both parents, 31 (34%) had one parent and 36 (40%) had no parent within the watershed. The proportion of genes in saplings inherited from the adults within the watershed was 43%, and therefore 57% were from outside the site, indicating active gene exchange across the watershed area. Average distance between parents and saplings (264.6 +/- 135.3 (SD) m) was significantly smaller than that of pairs randomly chosen between adults and saplings (436.7 +/- 203.0 (SD) m). The distance of pollen movement inferred from the distance between the two parents of each sapling ranged from 3.2 m to 540 m with an average of 131.1 m +/- 121.1 m (SD). Because 34% ( = 31/91) of saplings had only one parent within the watershed, the estimate of average pollen movement must be smaller than the actual one. Long-distance seed dispersal by birds, inbreeding depression and limitation in acceptance of pollen because of the difference of phenology in each individual flower were considered to be the probable causes of large gene exchange across the watershed.