Pollen adaptation to ant pollination: a case study from the Proteaceae.

BACKGROUND AND AIMS: Ant-plant associations are widely diverse and distributed throughout the world, leading to antagonistic and/or mutualistic interactions. Ant pollination is a rare mutualistic association and reports of ants as effective pollinators are limited to a few studies. Conospermum (Proteaceae) is an insect-pollinated genus well represented in the south-western Australia biodiversity hotspot, and here we aimed to evaluate the role of ants as pollinators of C. undulatum. METHODS: Pollen germination after contact with several species of ants and bees was tested for C. undulatum and five co-flowering species for comparison. We then sampled the pollen load of floral visitors of C. undulatum to assess whether ants carried a pollen load sufficient to enable pollination. Lastly, we performed exclusion treatments to assess the relative effect of flying- and non-flying-invertebrate floral visitors on the reproduction of C. undulatum. For this, we measured the seed set under different conditions: ants exclusion, flying-insects exclusion and control. KEY RESULTS: Pollen of C. undulatum, along with the other Conospermum species, had a germination rate after contact with ants of ~80 % which did not differ from the effect of bees; in contrast, the other plant species tested showed a drop in the germination rate to ~10 % following ant treatments. Although ants were generalist visitors, they carried a pollen load with 68-86 % of suitable grains. Moreover, ants significantly contributed to the seed set of C. undulatum. CONCLUSIONS: Our study highlights the complexity of ant-flower interactions and suggests that generalizations neglecting the importance of ants as pollinators cannot be made. Conospermum undulatum has evolved pollen with resistance to the negative effect of ant secretions on pollen grains, with ants providing effective pollination services to this threatened species.

Floral display and habitat fragmentation: Effects on the reproductive success of the threatened mass-flowering Conospermum undulatum (Proteaceae).

Fragmentation of natural vegetation is currently one of the largest threats to plant populations and their interactions with pollinators. Plant reproductive susceptibility to habitat fragmentation has been investigated in many species; however, the response of wild mass-flowering species is poorly known, with research limited to mainly boreal plant species.Here, we studied twelve remnant populations of the threatened mass-flowering shrub Conospermum undulatum in the southwest Australian biodiversity hotspot, each presenting different population size, level of isolation, and floral display. We assessed the impact of fragmentation on (a) fruit and seed production; and (b) seed germination. To gain a deeper understanding of factors influencing the reproductive success of C. undulatum, we performed pollinator exclusion and self-pollination treatments to experimentally assess the mating system of this threatened shrub.We found C. undulatum to be strictly self-incompatible and totally reliant on pollinators visiting with an outcrossed pollen load to complete the reproductive cycle. Further, we found that fruit production dropped from 35% to <20% as a result of decreasing floral display. A reduction in population size from 880 to 5 plants and from ~700 to 0.21 in the floral display index led to a decrease in seed output, while a similar reduction in seed output, from 6% to 3%, was observed as a result of increasing isolation index from -21.41 to -0.04. Overall, seed germination was positively related to population size, and a negative relationship was found between germination and isolation. Synthesis and applications. Our results demonstrate the important relationship between pollinators and floral morphology in plants of southwest Australia that have coevolved with native pollinators and developed characteristic flower morphologies over long time frames. Indeed, due to its characteristic pollination mechanism, the self-incompatible C. undulatum can only rely on specialized native pollinators for pollen flow and cannot rely on its mass-flowering trait to attract generalist pollinators from coflowering species; neither can it compensate for the lack of visitors by promoting geitonogamy. Consequently, fragmentation has a significant effect on the reproductive output of C. undulatum, and size, isolation, and floral display of populations are important factors to be considered when planning conservation actions for the species.

Isolation, characterization, and cross-amplification of 20 microsatellite markers for Conospermum undulatum (Proteaceae).

PREMISE: Recent habitat fragmentation is posing a risk to the wavy-leaved smokebush, Conospermum undulatum (Proteaceae), a rare plant species endemic to southwestern Western Australia. Microsatellite markers are required to characterize the genetic diversity and structure of the species for conservation purposes and to facilitate ecological studies. METHODS AND RESULTS: Illumina MiSeq high-throughput sequencing was used to develop 20 novel microsatellite markers for C. undulatum. Polymorphism at each locus was assessed using 72 individuals from three natural populations. Nineteen markers were polymorphic, with the number of alleles per locus ranging from two to 21, and observed and expected heterozygosity ranging from 0.000 to 1.000 and 0.117 to 0.919, respectively. All markers successfully amplified in three congeneric species (C. stoechadis, C. canaliculatum and C. triplinervium). CONCLUSIONS: The microsatellite markers will be useful for revealing patterns of genetic diversity, dispersal dynamics, and hybridization events for C. undulatum to inform future conservation efforts.