Introduction
Olfactory perception of food cues and sex signals is intimately
interconnected in insects (Reddy and Guerrero 2004; Varela et al.2011; Rouyar et al. 2015; Lebreton et al. 2017;
Borrero-Echeverry et al. 2018; Conchou et al. 2019). Deciphering
the chemicals encoding food and mates is basic to understanding insect
ecology and evolution. Moreover, the knowledge of such
behaviour-modifying chemicals can be applied for detection and
environmentally safe control of insects (Ridgway et al. 1990; Witzgall
et al. 2008, 2010a; Reddy and Guerrero 2010; Suckling et al. 2014;
Evenden and Silk 2016; Gregg et al. 2018).
New tools for insect management are needed in the wake of a changing
climate that accelerates insect invasions and outbreaks, aggravating
food insecurity (Deutsch et al. 2018). Recent efforts to deregulate the
most toxic compounds has left growers with few efficient insecticides
(Chandler et al. 2011; Jactel et al. 2019). The overwhelming majority of
insect species, however, does not feed on human food crops. Including
pollination services, insects are integral to all terrestrial food webs.
The overuse of synthetic pesticides affects non-target and beneficial
insects and other arthropods, and is a contributing cause of the
biodiversity apocalypse. This has been a point of debate since DDT
(Carson 1962) and nonethless, evidence is accumulating for severe side
effects of the currently most widely used family of insecticides, the
neonicotinoids (Seibold et al. 2019; Yamamuro et al. 2019; Chmiel et al.
2019; Longing et al. 2020; Wagner 2020).
The establishment of pheromones and other semiochemicals as a
species-specific and environmentally safe alternative to conventional
insecticides has therefore always been an outstanding rationale for
chemical ecology research. Air-permation with synthetic pheromone, for
disruption of premating sexual communication, is used against a few key
orchard and forest insects (Reddy and Guerrero 2010; Witzgall et
al. 2010; Evenden and Silk 2015). Pheromone lures for specific and
sensitive detection are available for hundreds of species. Such lures,
in combination with traps, insect pathogens or insecticides, may even
achieve population control, when the female sex becomes attracted
(Ridgway et al. 1990; El-Sayed et al. 2009; Sucklinget al. 2014). In stark contrast to pheromones attracting insects
for mating, only few semiochemicals have been identified that attract
gravid females for oviposition. Designing female or bisexual lures is
therefore a current main challenge towards a more widespread use of
behaviour-modifying chemicals for insect control.
Identification of many hundreds of sex pheromones, across all insect
orders (El-Sayed et al. 2016), has been facilitated by a mutual
coordination of production and response in both sexes. Pheromones are
produced in dedicated glands, produce strong antennal responses and
immediately trigger a sequence of distinctive behaviours.
Identification of semiochemicals, or kairomones, that mediate
oviposition behaviour meets substantial methodological difficulties.
Synthetic plant volatile blends that have been found to attract insect
herbivores typically build on compounds found across many plant species
(Najar-Rodriguez et al. 2010; Tasin et al. 2010; Bruce and
Pickett 2011; Lu et al. 2015). The attractant power of such
ubiquitous plant volatiles is sometimes faint, compared with sex
pheromones.
In comparison, plant compounds that are unique or characteristic for
larval food plants have been found to mediate significant attractancy.
One such key host plant compound is ethyl (E,Z)-2,4-decadienoate, pear
ester, a bisexual attractant for codling moth Cydia pomonella(Lepidoptera, Tortricidae) (Light et al. 2001; Light and Knight
2005). Pear ester is efficient for population monitoring (Knightet al . 2013, 2019) and for behavioural disruption of codling moth
larvae and adults, alone or combined with sex pheromone (Light and
Knight 2011; Knight et al. 2012; Light and Beck 2012; Knight and
Light 2013). The discovery of pear ester demonstrates the potential of
kairomones to both improve pheromone-based techniques and to design
stand-alone applications. That pear ester is released only in trace
amounts from green apples (Gonzalez et al. 2020) underlines that the
abundance of volatiles in plant headspace does not correlate with their
behavioural saliency. Compounds released in large amounts often stem
from main biosynthetic pathways shared by many plants, and cannot encode
specific host plant finding.
The most widely employed tool for studying plant compounds mediating
host attraction is gas chromatography coupled to electroantennographic
detection. GC-EAD measures the response of the entire antenna to
odorants (Arn et al. 1975), and biases compounds occurring in large
amounts in headspace collections. GC-EAD was designed as an efficient
and reliable tool for detection and identification of trace amounts of
sex pheromones. GC-EAD suffers, however, from serious bias and produces
false positives when screening plant headspace. Ubiquitous compounds
present in large amounts, for example short aliphatic acetates or
alcohols, farnesenes, linalools and caryophyllenes, invariably elicit an
antennal response, generated by the ensemble of olfactory sensory
neurons (OSNs) on the antenna, expressing the entire olfactory receptor
(OR) repertoire. Typically, ORs respond to large amounts of volatiles
that are structurally similar to their cognate ligands. A diffuse signal
from many OSNs on the antenna is sufficient to produce
electroantennograms, but its behavioural relevance is uncertain. An
active compound such as pear ester, on the other hand, has been
overlooked in GC-EAD recordings due to its low abundance.
The discovery of the genetic code of insect ORs (Clyne et al. 1999)
enables a new approach. The ligand binding specificity of ORs determines
the spectrum of volatile chemicals transmitted by OSNs from the antenna
to olfactory centers in the brain. Sequencing antennal RNA extracts and
annotation provides OR expression data and a first functional
differentiation, between pheromone receptors (PRs) and ordinary ORs,
responding to environmental odorants. Subsequent phylogenetic analysis
groups orthologous ORs from related species and provides leads on
putative ligands, through comparison with an accumulating database of
deorphaned insect ORs (Fleischer et al. 2018, Robertson 2019). Single
ORs are accordingly a tool of choice to interrogate the plant odorscape
for bioactive compounds. A powerful experimental approach is to express
ORs singly in defined sensilla of the antenna of Drosophila
melanogaster (Dobritsa et al. 2003; Hallem et al. 2004), where they can
be addressed with single sensillum electrophysiological recordings,
coupled to gas chromatography (GC-SSR).
In codling moth Cydia pomonella (Lepidoptera, Tortricidae), Cpom
OR3 has been deorphaned, following transcriptome analysis (Bengtsson et
al. 2012, Walker et al. 2016) and heterologous expression
(Bengtsson et al. 2014, Cattaneo et al. 2017, Wan et al. 2019). The main
ligand of CpomOR3, which belongs to the PR clade, is the plant volatile
pear ester (Light et al. 2001; Light and Knight 2005, Bengtsson
et al. 2014). A recent assembly of the codling moth genome reveals
presence of two copies of CpomOR3, which, according to functional
characterization in Xenopus oocytes, respond to a lesser extent
also to codling moth sex pheromone, codlemone (Wan et al. 2019). A
seemingly conserved response in a closely related species underscores
this deeply rooted interconnection of pheromone and plant volatiles.
Green budworm moth Hedya nubiferana (Lepidoptera, Tortricidae) is
attracted to codlemone (Arn et al. 1974, El-Sayed, 2019) and to pear
ester (Schmidt et al. 2007, Jósvai et al. 2016).
We have investigated the response of green budworm moth H .nubiferana to codling moth sex pheromone and to pear ester, in
laboratory and field bioassays. Comparative phylogenetic analysis of ORs
in the antennal transcriptome of green budworm and codling moth confirm
the behavioural evidence and suggest the presence of a conserved
olfactory channel dedicated to these compounds, in both species. This
demonstrates how functional characterization of ORs in model species
such as codling moth (Bengtsson et al. 2014; Gonzalez et
al. 2016), followed by in silico studies of antennal transcriptomes in
the taxonomically related species will advance the identification of
insect kairomones, and the development of insect management.