Molecular species delimitation of a symbiotic fig-pollinating wasp species complex reveals extreme deviation from reciprocal partner specificity

Abstract

BackgroundSymbiotic relationships have contributed to major evolutionary innovations, the maintenance of fundamental ecosystem functions, and the generation and maintenance of biodiversity. However, the exact nature of host/symbiont associations, which has important consequences for their dynamics, is often poorly known due to limited understanding of symbiont taxonomy and species diversity. Among classical symbioses, figs and their pollinating wasps constitute a highly diverse keystone resource in tropical forest and savannah environments. Historically, they were considered to exemplify extreme reciprocal partner specificity (one-to-one host-symbiont species relationships), but recent work has revealed several more complex cases. However, there is a striking lack of studies with the specific aims of assessing symbiont diversity and how this varies across the geographic range of the host.ResultsHere, we use molecular methods to investigate cryptic diversity in the pollinating wasps of a widespread Australian fig species. Standard barcoding genes and methods were not conclusive, but incorporation of phylogenetic analyses and a recently developed nuclear barcoding gene (ITS2), gave strong support for five pollinator species. Each pollinator species was most common in a different geographic region, emphasising the importance of wide geographic sampling to uncover diversity, and the scope for divergence in coevolutionary trajectories across the host plant range. In addition, most regions had multiple coexisting pollinators, raising the question of how they coexist in apparently similar or identical resource niches.ConclusionOur study offers a striking example of extreme deviation from reciprocal partner specificity over the full geographical range of a fig-wasp system. It also suggests that superficially identical species may be able to co-exist in a mutualistic setting albeit at different frequencies in relation to their fig host’s range. We show that comprehensive sampling and molecular taxonomic techniques may be required to uncover the true structure of cryptic biodiversity underpinning intimate ecological interactions.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-014-0189-9) contains supplementary material, which is available to authorized users.

Local coexistence and genetic isolation of three pollinator species on the same fig tree species

“…The cytb gene was amplified and sequenced in all samples using CB1/CB2 primers and the protocol described in (Jermiin and Crozier, 1994). Although COI is the standard barcoding gene in animals, most barcoding efforts for this species complex have utilised the cytb gene (Haine et al, 2006;Darwell et al, 2014) since COI amplifies inconsistently and is compromised as a marker by nuclear pseudogenes in fig wasps. DNA sequences were edited by eye in Sequencher v4.10.1 (Gene Codes, Ann Arbor, MI, USA) and aligned in Clustal X v2.0 (Larkin et al, 2007).…”

“…A cytb haplotype phylogeny was constructed in MrBayes v3.2 (Ronquist et al, 2012) to assign individuals to lineages according to Darwell et al (2014). Reference sequences for each of the three lineages (Haine et al, 2006;Darwell et al, 2014) were included in the phylogeny, along with outgroup sequences for Pleistodontes nigriventris (Girault; Lopez-Vaamonde et al, 2001) and another fig wasp, Ceratosolen galili Wiebes (Kerdelhue et al, 1999).…”

“…Haine et al (2006) found support for four species based on mitochondrial cytochrome b (cytb); however, the nuclear 28S rDNA and wingless genes only resolved two lineages. With wider sampling, Darwell et al (2014), subsequently showed support for five species using nuclear ITS2 and mitochondrial COI and cytb.…”

“…Recently, there have been several reports of cryptic pollinator 'species' using the same host fig species both in sympatry and allopatry (Molbo et al, 2003;Haine et al, 2006;Sun et al, 2011;Chen et al, 2012;Darwell et al, 2014). Where morphologically cryptic pollinator species coexist locally, their persistence requires ecological and/or behavioural explanation (Zhang et al, 2004).…”

“…), which is pollinated by five 'species' of the Pleistodontes imperialis Saunders species complex along the Australian east coast (Haine et al, 2006;Darwell et al, 2014). Following taxonomic revision in 2002, these were considered a single species (Lopez-Vaamonde et al, 2002), but were later found to resolve into distinct lineages (Haine et al, 2006;Darwell et al, 2014). Haine et al (2006) found support for four species based on mitochondrial cytochrome b (cytb); however, the nuclear 28S rDNA and wingless genes only resolved two lineages.…”