Trees of the genus Glochidion (Phyllanthaceae) are pollinated by females of Epicephala moths (Gracillariidae) whose larvae consume the seeds of the flowers that they pollinate. Each Epicephala moth species is specific locally to a single host species, although two to four Glochidion hosts often cooccur. To investigate the role of olfactory signals in maintaining the plant-moth specificity, we analyzed floral scent composition of five Glochidion species by using gas chromatography-mass spectrometry (GC-MS) and conducted Y-tube olfactometer bioassays with Epicephala moths and their host flowers. The GC-MS analysis showed that the floral scents of the five Glochidion species are dominated by (R)-(-)- and (S)-(+)-linalool, and (E)- and (Z)-beta-ocimene, and that each species produces 6-20 compounds. Transformation of scent profiles by using chord-normalized expected species shared distances and analysis of the data with nonmetric multidimensional scaling showed that floral volatiles of cooccurring Glochidion species can be distinguished by relative chemical composition, especially that of minor compounds. The bioassay with pollinators of Glochidion lanceolatum and Glochidion ruburm further indicated that Epicephala moths are capable of discriminating their hosts by using floral odor. The results suggest that the floral scent of Glochidion is one of the important key signals that mediate the encounters of the species-specific partners in the Glochidion-Epicephala mutualism.

The evolutionary stability of mutualisms is enhanced when partners possess mechanisms to prevent overexploitation by one another. In obligate pollination-seed consumption mutualisms, selective abortion of flowers containing excessive eggs represents one such mechanism, but empirical tests have long been limited to the yucca-yucca moth mutualism. We present evidence for selective abortion in the recently discovered mutualism between Glochidion trees and Epicephala moths. In Glochidion acuminatum, proportion of aborted flowers progressively increased both with higher egg load and increased ovule damage. Selective abortion resulted in a 16% seed production increase compared with expectations under random abortion, and moths suffered fitness losses as high as 62% when ovipositing into pre-infested flowers. Moth eggs were laid singly more often than expected under random oviposition, thus avoiding potential disadvantages from multiple infestations. As new pollination mutualisms are being discovered, selective abortion mechanisms may prove to be more widespread than previously thought.

Summary• Obligate mutualisms involving actively pollinating seed predators are among the most remarkable insect-plant relationships known, yet almost nothing is known about the chemistry of pollinator attraction in these systems. The extreme species specificity observed in these mutualisms may be maintained by specific chemical compounds through 'private channels'. Here, we tested this hypothesis using the monoecious Breynia vitis-idaea and its host-specific Epicephala pollinator as a model.• Headspace samples were collected from both male and female flowers of the host. Gas chromatography with electroantennographic detection (GC-EAD), coupled gas chromatography-mass spectrometry, and olfactometer bioassays were used to identify the floral compounds acting as the pollinator attractant.• Male and female flowers of B. vitis-idaea produced similar sets of general floral compounds, but in different ratios, and male flowers emitted significantly more scent than female flowers. A mixture of 2-phenylethyl alcohol and 2-phenylacetonitrile, the two most abundant compounds in male flowers, was as attractive to female moths as the male flower sample, although the individual compounds were slightly less attractive when tested separately.• Data on the floral scent signals mediating obligate mutualisms involving active pollination are still very limited. We show that system-specific chemistry is not necessary for efficient host location by exclusive pollinators in these tightly coevolved mutualisms.

Zoophilous flowers often transmit olfactory signals to attract pollinators. In plants with unisexual flowers, such signals are usually similar between the sexes because attraction of the same animal to both male and female flowers is essential for conspecific pollen transfer. Here, we present a remarkable example of sexual dimorphism in floral signal observed in reproductively highly specialized clades of the tribe Phyllantheae (Phyllanthaceae). These plants are pollinated by species-specific, seed-parasitic Epicephala moths (Gracillariidae) that actively collect pollen from male flowers and pollinate the female flowers in which they oviposit; by doing so, they ensure seeds for their offspring. We found that Epicephala-pollinated Phyllanthaceae plants consistently exhibit major qualitative differences in scent between male and female flowers, often involving compounds derived from different biosynthetic pathways. In a choice test, mated female Epicephala moths preferred the scent of male flowers over that of female flowers, suggesting that male floral scent elicits pollen-collecting behaviour. Epicephala pollination evolved multiple times in Phyllantheae, at least thrice accompanied by transition from sexual monomorphism to dimorphism in floral scent. This is the first example in which sexually dimorphic floral scent has evolved to signal an alternative reward provided by each sex, provoking the pollinator's legitimate altruistic behaviour.

Coevolved mutualisms often exhibit high levels of partner specificity. Obligate pollination mutualisms, such as the fig -fig wasp and yucca -yucca moth systems, represent remarkable examples of such highly species-specific associations; however, the evolutionary processes underlying these patterns are poorly understood. The prevailing hypothesis suggests that the high degree of specificity in pollinating seed parasites is the fortuitous result of specialization in their ancestors because these insects are derived from endophytic herbivores that are themselves highly host-specific. Conversely, we show that in the Glochidion -Epicephala obligate pollination mutualism, pollinators are more host-specific than are closely related endophytic leaf-feeding taxa, which co-occur with Epicephala on the same Glochidion hosts. This difference is probably not because of shifts in larval diet (i.e. from leaf-to seed-feeding), because seed-eating lepidopterans other than Epicephala do not show the same degree of host specificity as Epicephala. Species of a tentative sister group of Epicephala each attack several distantly related plants, suggesting that the evolution of strict host specificity is tied to the evolution of pollinator habit. These results suggest that mutualists can attain higher host specificity than that of their parasitic ancestors and that coevolutionary selection can be a strong promoter of extreme reciprocal specialization in mutualisms.