Parasitoids use volatiles to recognize and locate suitable hosts. Numerous studies have investigated parasitoid host location from a pest management perspective, but comparatively little is known regarding parasitoid-pollinator interactions. Previous research has shown that parasitoids of some native bees respond to volatiles emitted by host frass and cocoons. We used a Y-tube bioassay to test whether two parasitoid species (Monodontomerus torchioi and Melittobia acasta) are attracted to volatiles associated with their host, the blue orchard bee, Osmia lignaria. Specifically, we tested attraction to (1) cocooned adult O. lignaria females (2) O. lignaria frass (3) methanol and (4) acetic acid. Both parasitoid species were attracted to host frass and acetic acid. Although M. acasta showed a strong attraction to volatiles from cocooned O. lignaria females, it was repelled by methanol, a chemical constituent of Osmia cocoons. In contrast, M. torchioi showed no response to cocooned O. lignaria and only a slight attraction to methanol. Both parasitoid species appear to be differentially attracted to specific host-associated volatiles, suggesting that they may respond to distinct olfactory cues when locating potential bee hosts.

Y-tube bioassay, olfactory cues, solitary bee, Megachilidae , Melittobia acasta , Monodontomerus torchioi

Parasitoids use a variety of chemical cues to identify and locate potential hosts, including those emitted by the host’s habitat (Vet et al. 1984), host byproducts (i.e. frass, Agelopoulos et al. 1995), organisms living in association with the host (Sullivan et al. 2000), and the host itself (Wiskerke et al. 1992, Jumean et al. 2009). Numerous studies have investigated parasitoid attraction to host-associated volatiles in agroecosystems, often with the goal of developing more effective pest management strategies (Jones et al. 1973, Godfray 1994, Jumean et al. 2009). However, despite the ecological and economic importance of pollinators, few studies have investigated how parasitoids recognize and locate their bee hosts (but see Silva-Torres et al. 2005, Filella et al. 2011). A number of solitary bee species are currently used to pollinate commercial crops, including Nomia melanderi Cockrell (Halictidae) and Megachile rotundata Fabricius (Megachilidae) in alfalfa (Kemp and Bosch 2000, Cane 2008) and Osmia spp. (Megachilidae) in almonds (Bosch and Kemp 2001). Many solitary bee species are attacked by a diversity of hymenopteran parasitoids and brood parasites during development, sometimes resulting in high rates of mortality prior to adult eclosion (Vicens et al. 1994, Bosch and Kemp 2001, Wcislo and Cane 1996). Thus, building a more robust understanding of parasitoid host location is not only of interest from an ecological perspective, it may also aid efforts to control parasitism in commercially important pollinator species.

The blue orchard bee (Osmia lignaria Say, Megachilidae) is a univoltine solitary species that emerges in early spring and nests in hollow twigs and beetle burrows (Bosch and Kemp 2001). We focused on two geographically widespread ectoparasitoids commonly found in O. lignaria nests: Monodontomerus torchioi Grissell (Hymenoptera: Torymidae) and Melittobia acasta (Walker) (Hymenoptera: Eulophidae). We selected these species because they differ in both their oviposition strategy and the breadth of their host range. To parasitize the host, Monodontomerus females oviposit directly through the host cocoon onto the prepupa or pupa (Eves 1970, Bosch and Kemp 2001, Filella et al. 2011). In contrast, Melittobia parasitoids chew through nesting substrates and cocoons to feed on host hemolymph and/or oviposit (Hobbs and Krunic 1971, Cusumano et al. 2010). Unlike Monodontomerus, Melittobia spp. attack a wide diversity of insects, including Coleoptera, Diptera, and Hymenoptera (Hobbs and Krunic 1971, González et al. 2004, Silva-Torres et al. 2005, Cusumano et al. 2010).

Using M. torchioi and M. acasta reared from field-collected O. lignaria, we tested attraction to several host volatiles. Given that host bees are enclosed in a thick cocoon covered in layers of frass during the preferred life stages for parasitoid oviposition (prepupae, pupae), we expected both parasitoid species to be more attracted to volatiles associated with host frass and cocoons than volatiles emitted by the adult host. Our primary goals were (1) to determine which host-associated volatiles the two species use during host location and (2) to experimentally test parasitoid attraction to two previously-identified Osmia nest volatiles, acetic acid and methanol (Filella et al. 2011).

Across all trials, 70–90% of M. torchioi and 73–93% of M. acasta made a choice. Both species showed strong attraction to several of the volatiles tested, suggesting that they respond to multiple olfactory cues when searching for a host. Significantly more M. torchioi and M. acasta females were attracted to host frass and its primary chemical component, acetic acid, than the control (Table 1). These results empirically confirm previous work suggesting these volatiles act as chemical cues for parasitoid females (Agelopoulos et al. 1995, Sullivan et al. 2000, Filella et al. 2011). Specifically, frass and its associated volatile components may indicate host suitability. In general, Monodontomerus and Melittobia parasitize host pre-pupae and pupae (Eves 1970, Dahms 1984). In this system, O. lignaria defecate as fifth instar larvae, later incorporating this frass into the cocoon with salivary strands (Torchio 1989). Thus, frass-associated volatiles may allow parasitoids to discriminate between host life stages.

In contrast, the parasitoid species showed substantially different reactions to methanol. We expected both species to be attracted to methanol because it is a primary chemical component of host cocoons (Filella et al. 2011) and a natural byproduct of wood rot (Arantes and Goodell 2014). Megachilid bees often nest in standing dead trees (Macivor and Salehi 2014), therefore we hypothesized that methanol may act as a general cue that allows parasitoids to locate host nests. Although a higher number of M. torchioi females selected methanol over the control, this difference was not statistically significant (Table 1a). Using exact binomial testing with a larger sample size would be needed to assess the significance of these trends. Contrary to our expectations, M. acasta appeared to be repelled by methanol (Table 1b). It is possible M. acasta females avoided the methanol treatment because the concentration was too high. An alternative explanation for this behavior is that methanol acts as a repellent when isolated from other host-associated volatiles. The ratio-specific odor recognition hypothesis presented in Bruce et al. (2005) stresses that correct ratios and combinations of common plant volatiles are more generally used by phytophagous insects than any one species-specific volatile source. Furthermore, Takemoto and Takabayashi (2015) observed repellent effects of an isolated compound, linalool, regardless of concentration. However, the parasitoid was attracted to linalool when in combination with other herbivore-induced plant volatile components. Additional studies that test attraction to multiple methanol concentrations and blends of host-associated volatiles are necessary to test this hypothesis.

Lastly, M. acasta was strongly attracted to cocooned O. lignaria females, but M. torchioi was not (Table 1). Our results for M. acasta are consistent with previous research showing Melittobia rely primarily on indirect host-associated chemical cues, rather than cues produced by the appropriate host stage (Cusumano et al. 2010). Because parasitoids in this group employ a ‘sit-and-wait’ strategy in the host nest (Cusumano et al. 2010) and are limited in their dispersal ability (Matthews 2009), a strong attraction to the adult host may also allow them to locate nests at short distances. Surprisingly, M. torchioi was not attracted to volatiles from cocooned host bees (Table 1). Given that the closely-related M. aeneus (Fonscolombe) does not respond to volatiles from uncocooned Osmia host prepupae but is attracted to host cocoons and frass (Filella et al. 2011), it is possible that Monodontomerus spp. rely on a combination of volatiles emitted by the host at the preferred life stage. To disentangle parasitoid response to adult hosts and cocoons, future studies could examine attraction to uncocooned Osmia adults.

Our study is among the first of its kind to test parasitoid attraction to pollinator-associated volatiles. Collectively, our results suggest that both M. torchioi and M. acasta use a number of volatile cues to locate hosts. By simultaneously comparing parasitoid attraction to multiple volatiles and combinations of volatiles, future studies may allow us to rank the relative importance of each cue and further elucidate the complexities of parasitoid host location.

SG contributed to data collection and writing. SF contributed to data collection and analysis, writing, and project design.

K. Eaton for use of the olfactometer, G. Wardell for assistance x-raying specimens, S. Heydon for parasitoid identification, and L. Yang for manuscript feedback.