Experimental fertilization increases amino acid content in floral nectar, fruit set and degree of selfing in the orchid Gymnadenia conopsea

Floral traits evolved to maximize reproductive success by attracting pollinators and facilitating pollination. Highly attractive floral traits may, however, also increase the degree of self-pollination, which could become detrimental for plant fitness through inbreeding depression. Floral nectar is a trait that is known to strongly mediate pollinator attraction and plant reproductive success, but the particular role of the nectar amino acid (AA) composition is poorly understood. Therefore, we experimentally manipulated the nectar AA composition and abundance of the Lepidoptera pollinated orchid Gymnadenia conopsea through soil fertilization, and we quantified AA content and AA composition through high performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD). Mixed models were then used to evaluate differences in pollinia removal, fruit set, seed set and degree of selfing between fertilized and control individuals. Selfing rates were estimated using microsatellite markers. We found that fertilized individuals had a significantly higher nectar AA content and an altered AA composition, whereas plant height, number of flowers, nectar volume and sugar concentration remained unchanged. Fertilized individuals also had significantly more pollinia removed and a higher fruit set, whereas control plants that did not receive the fertilization treatment had significantly fewer selfed seeds, and more viable seeds. Although we cannot exclude a role of changes in floral scent following the fertilization treatment, our results strongly suggest a relation among nectar AA composition, fruiting success and selfing rates. Our results also indicate potential consequences of nutrient pollution for plant reproductive success, through the induced changes in nectar AA composition.status: publishe