Discussion
Infochemical cues play an important and crucial role in parasitoid host
location and selection (Tumlinson, Lewis, et al., 1993; Vet & Dicke,
1992; Vinson, 1976). Vet and Dicke (1992) framed the use of these cues
by parasitoids as a tradeoff between reliability and detectability,
which they called the reliability-detectability problem. Host
infochemical cues were hypothesized to be a highly reliable indicator of
a host’s presence, but due to their localized nature would not be
detectable over long distances. Conversely, infochemical cues from the
plant were hypothesized to be highly detectable but poor indicators of
host presence and thus unreliable, unless they were induced by
herbivory. Vet and Dicke (1992) also hypothesized that when a parasitoid
that had one or most hosts that fed on a single plant species, plant
infochemicals could become both reliable and detectable. In a third
scenario in which the parasitoid specializes on a host species that
feeds upon multiple plant species, host infochemical cues were
hypothesized to be highly reliable while herbivore induced synomones
were not. Here, however, the synomones from each plant species could be
learned and associated with the host, thus compensating for the low
detectability of the host kairomones. Finally in a fourth scenario , in
which both the parasitoid and its hosts were extreme generalists, Vet
and Dicke (1992) hypothesized that infochemical cues would not be used.
Although Vet and Dicke (1992) acknowledged each of these four scenarios
were extremes on a continuum, and that intermediates could be found in a
nature, they did not specifically discuss cases that involved
oligophagous parasitoids and their hosts. This is the situation forEadya parasitoids, which attack multiple related beetle hosts
that feed on multiple related Eucalyptus plants. Thus, the
question remains as to how these parasitoids might use infochemicals to
locate their hosts.
For parasitoids that specialize on one host and rely heavily on host
infochemical cues, we would expect to see some degree of coevolution
leading to cospeciation between parasitoids and their hosts (Page,
2003). As Eadya are host-flexible (Peixoto et al., 2018), there
would be less expectation of coevolution and cospeciation with their
beetle hosts. Indeed, our data did not support cospeciation in the
event-based analysis, but rather provided robust evidence for
duplication and host-switching with duplication being the dominant type
of evolutionary event. Although the distance-based analysis demonstrated
strong congruence between parasitoid and beetle phylogenies indicating
cospeciation, further exploration of the data suggested that
cophylogenetic signal was restricted to the interactions betweenE. annleckieae and Pst. cloelia (Hutchinson et al., 2017).
This large disparity in cophylogentic signal between E.
annleckieae and the rest of Eadya can be attributed to sampling
error. Peixoto et al. (2018) reared E. annleckieae from P.
charybdis, Pst. cloelia, Pst. selmani, and Pst. nobilitata(Erichson, 1842) but DNA was only successfully extracted for Pst.
cloelia. As a phylogram is required to calculate the distance matrices
for the PACo distance-based analysis, only Pst. cloelia could be
included, creating a false one-to-one relationship between E.
annleckieae and Pst. cloelia . Thus, E. annleckieae is the
only parasitoid species with a single host association in the
distance-based analysis, which most likely biased the PACo analysis.
Although we cannot rule out the use of host infochemical cues inEadya as these were not directly tested here, the lack of
evidence for cospeciation between these parasitoids and their beetle
hosts as well as their known host flexibility, suggests these cues are
unlikely to be the dominant driver in the location and selection of a
suitable host.
Although Vet and Dicke (1992) discussed plant synomones and hypothesized
how the compounds are used in a tritrophic context, this was only
considered from a bottom-up perspective in which the herbivore host
breadth solely influences whether plant infochemicals could reliably be
used by the parasitoid to locate its herbivore host. For parasitoids
that specialize on a single host, the low reliability of plant cues
discussed in Vet and Dicke (1992) likely holds true, as in most cases
these signals cannot guarantee the presence of a suitable host unless
they are herbivore-induced volatiles (de Moraes et al., 1998). However,
this may not be the case for less specialized parasitoids that can
survive on a wider range of hosts, which in turn utilize a range of
closely related plant species. Here, plant infochemicals, even if not
herbivore-induced, may be both reliable and detectable for parasitoids
if they specialize on a single host tree species, or a distinct
infochemical signature, as long as they can utilize multiple hosts
associated with that species/infochemical signature. This may allow
parasitoids to reduce competition with related parasitoid species
through plant specialization rather than host specialization. Here we
find evidence that Eucalyptus infochemicals are species-specific,
corresponding to specific species of Eadya and their beetle hosts
(Fig. 5A & D). However, since we found no evidence of cospeciation
between the parasitoids and their hosts we suggest that host selection
in Eadya is influenced by Eucalyptus phytochemistry
irrespective of the specific beetle host used. Although, several
limitations to our study restrict us from considering this conclusion
definitive. 1) Euc. globulus was the intended target plant of
this study, with Euc. sp. 1 being a volunteer in a single
plantation, and therefore was not widely sampled. 2) Sampling was
limited to a small area in western Victoria and does not represent the
endemic range of Euc. globulus nor that of the beetle or
parasitoid species. 3) As only juvenile Euc. globulus plantations
were targeted, the full range of Eucalyptus utilized byEadya is still unknown.
We identified a diverse array of plant metabolites that could accurately
distinguish between the two groups of Eadya (Figure 5D, Table
S3). Among top contributing compounds to axes 1 & 2, we identified two
general groups of compounds: (1) fatty acid derivatives (Docosanoic
acid, phenylmethyl ester (C106), 4H-1-Benzopyran-4-one,
5-hydroxy-7-methoxy-2-(4-methoxyphenyl)-6,8-dimethyl- (C105), and
Octadecanoic acid, 2,3-dihydroxypropyl ester) and (2) compounds with
known larvicide activity (agarospirol, ascaridole epoxide, alpha-pinene,
alpha-phellandrene, phytol, and eucalyptol) (de Castro et al., 2016;
Johnson & Singh, 2017; Kaczmarek, Wrońska, Kazek, & Boguś, 2020;
Mohamed & Jong, 2014)
Docosanoic acid has the potential to function as either a kairomone or
synomone in both insects and plants. Docosanoic acid is a derivative ofEucalyptus cuticular wax, a known Coleopteran attractant, and
changes in Docosanoic acid correspond with larval maturation in certain
insects (Courtney, Lassak, & Speirs, 1983; Gosney et al., 2016;
Kaczmarek et al., 2020; Sarkar & Barik, 2015). Docosanoic acid is
commercially manufactured and presents as a strong candidate for
attractants for Eadya in New Zealand P. charybdisbiological control efforts (T. Withers, Todoroki, Allen, Pugh, &
Gresham, 2019) with low potential for toxic effects on non-target
insects (Kaczmarek et al., 2020; Sarkar & Barik, 2015). Although we can
discern which Eucalyptus infochemicals are associated with host
use and that compounds can separate the two groups of Eadya ,
future work should aim at directly testing for attractant responses
across the genus Eadya . Further work to identify compounds that
attract host flexible parasitoids should consider volatile compounds
that are present in both the host and associated plant species, with
foci on fatty acid derivatives based on this study’s results.
We did not find evidence for an herbivore-induced response, as no
overall pattern could be discerned between herbivore damaged and
undamaged E. globulus leaves from the PCA (Fig. 5B). However,
these inconclusive results may be due to the methodology of our study.
As herbivore damaged leaves were collected from the field, it was not
possible to determine when the leaves were damaged. The production of
volatile compounds is metabolically expensive (Cipollini, Walters, &
Voelckel, 2018) and is likely not maintained indefinitely after
herbivore damage. Based on our data, we were unable to determine whether
herbivore damage induced the production of volatile compounds inEuc. globulus , but cannot rule out the possibility until a study
can be undertaken using methodology better suited for this question
(Materić et al., 2015).
With this evidence we propose an amendment to Vet and Dicke’s (1992)
reliability-detectability hypothesis to include how infochemicals are
used when oligophagous parasitoids utilize oligophagous hosts. In this
ecological context, we expect little to no response to host kairomones
and no evidence of cospeciation between the parasitoid and their hosts,
but a strong response to plant synomones and potentially
herbivore-induced plant synomones. Although host kairomones are the most
reliable indicators of host presence, especially for specialists, plant
synonomes, whether induced or not, may play a larger role for
oligophagous parasitoids. The ability to successfully utilize multiple
hosts is an adaptive advantage in instances when hosts have a patchy
distribution or small population size, or when multiple parasitoids are
competing for a limited number of hosts (Price, 1971). Further, if hosts
feed on multiple plants, specializing on one plant where multiple plant
species are present may limit competition between parastioids.Eadya annleckieae and E. daenerys ’ wide niche breadth and
preference for hosts that feed on plants with different chemoprofiles
may aid in reducing competition between the two species. This study
greatly expands our understanding of the complex interactions within the
tri-trophic Eadya system and how they utilize plant infochemicals
for host location and potentially to limit intra-specific competition.
To further understand these interactions a more robust taxon sampling ofEucalyptus and Eadya across their native range is required
to definitively establish the adaptive advantages of host flexibility
and the use of plant synomones for host location in similar tri-trophic
systems. Subsequently no-choice competition assays between Eadyaspecies using distinct infochemical signatures should be conducted under
a controlled environment to establish the competitive advantages of
plant specialization.