Abstract
Speciation is typically accompanied by the formation of isolation barriers between lineages. Commonly, reproductive barriers are separated into pre‐ and post‐zygotic mechanisms that can evolve with different speed. In this study, we measured the strength of different reproductive barriers in two closely related, sympatric orchids of the Ophrys insectifera group, namely Ophrys insectifera and Ophrys aymoninii to infer possible mechanisms of speciation. We quantified pre‐ and post‐pollination barriers through observation of pollen flow, by performing artificial inter‐ and intraspecific crosses and analyzing scent bouquets. Additionally, we investigated differences in mycorrhizal fungi as a potential extrinsic factor of post‐zygotic isolation. Our results show that floral isolation mediated by the attraction of different pollinators acts apparently as the sole reproductive barrier between the two orchid species, with later‐acting intrinsic barriers seemingly absent. Also, the two orchids share most of their fungal mycorrhizal partners in sympatry, suggesting little or no importance of mycorrhizal symbiosis in reproductive isolation. Key traits underlying floral isolation were two alkenes and wax ester, present predominantly in the floral scent of O. aymoninii. These compounds, when applied to flowers of O. insectifera, triggered attraction and a copulation attempt of the bee pollinator of O. aymoninii and thus led to the (partial) breakdown of floral isolation. Based on our results, we suggest that adaptation to different pollinators, mediated by floral scent, underlies species isolation in this plant group. Pollinator switches may be promoted by low pollination success of individuals in dense patches of plants, an assumption that we also confirmed in our study.
Highlights
In the last decades, an increasing number of studies have focused on the strength and evolution of reproductive isolating barriers among co-occurring species (Coyne & Orr, 1989, 2004; Ramsey, Bradshaw,& Schemske, 2003; Schemske, 2010; Scopece, Widmer, & Cozzolino, 2008)
We focused on the following specific questions: (1) Which reproductive barriers maintain species boundaries? (2) Which plant traits underlie reproductive isolation? (3) Is population density negatively associated with fecundity?
Using species delineation based on 3% intergenic transcribed spacer (ITS) divergence is unlikely to have masked cryptic Tulasnellaceae species: first, this is a usual threshold and ITS species delineation is validated in Tulasnellaceae by the fact that it is congruent with other genes (Linde, Phillips, Crisp, & Peakall, 2014); second, lowering the threshold to 1.5% did not change Operational taxonomic units (OTUs) delineation in our work
Summary
An increasing number of studies have focused on the strength and evolution of reproductive isolating barriers among co-occurring species Pre-zygotic barriers (e.g., floral isolation) often predate post-zygotic barriers, and are, thought to play a more critical role during speciation (Coyne & Orr, 2004; Grant, 1994; Kirkpatrick & Ravigne, 2002; Lowry, Modliszewski, Wright, Wu, & Willis, 2008; Moyle, Olson, & Tiffin, 2004; Rieseberg & Willis, 2007; Widmer, Lexer, & Cozzolino, 2009) An example for this are plant adaptations to different pollinators, with reduced gene flow between individuals attracting different pollen vectors (Kay, 2006; Ramsey et al, 2003; Sun, Schlüter, Gross, & Schiestl, 2015; Van der Niet, Peakall, & Johnson, 2014; Waelti, Muhlemann, Widmer, & Schiestl, 2008; Widmer et al, 2009). We focused on the following specific questions: (1) Which reproductive barriers maintain species boundaries? (2) Which plant traits underlie reproductive isolation? (3) Is population density negatively associated with fecundity?
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