Results
Sex pheromone
identification
Analysis of green budworm moth H . nubiferana pheromone
gland extracts by GC and GC-MS showed eight further compounds, in
addition to the previously identified acetates (Frerot et al. 1979). The
major compound E 8,E 10-12Ac was accompanied by the
monounsaturated 8- and 10-dodecenyl acetates, its three geometric
isomers (EZ , ZE , and Z 8,Z 10-12Ac) as well as
the analogous alcohol codlemone, E 8,E 10-12OH (Table 1).
Field attraction of H. nubiferana males to compounds identified
from the female gland confirms that the sex pheromone of H.
nubiferana is a blend of E 8,E 10-12Ac and Z 8-12Ac
(Table 2; Frerot et al. 1979). The main compound,E 8,E 10-12Ac by itself was not attractive, while addition
of Z 8-12Ac had a strong synergistic effect (F(7,72)=61.95,P <0.0001). Addition of E 8-12Ac further increased
male attraction in untreated apple orchards, but the difference was not
significant. Blends of E 8,E 10-12Ac and the ∆ 10-12
monoenes or the analogous alcohol, codlemone, did not produce
significant captures. Adding these compounds to the 3-component acetate
blend slightly diminished trap catch (Table 2).
The gland compounds identified from female glands with no apparent
effect on attraction may be biosynthetic by-products or precursors. A
study of the female effluvium will show whether they are released at
all, and at which ratio. The full blend of compounds may also carry
information that cannot be revealed by a field trapping test.
Attraction to codlemone and pear
ester
A trap test in an apple orchard adjacent to a pea field corroborates
that codlemone acetate E 8,E 10-12Ac as a single compound
does not attract green budworm moth. Attraction of pea moth confirms
that the trap lures released E 8,E 10-12Ac at high isomeric
purity (Table 3; Witzgall et al. 1993, 1996). In comparison, traps
baited with codlemone alone regularly captured few green budworm moth
males, in addition to codling moth. Blends of codlemone and codlemone
acetate attract far fewer codling moths and no green budworm moths at
all (Table 3).
Interestingly, a blend of codlemone and its three geometric isomers
significantly increased green budworm moth captures over codlemone alone
(Table 4; F(7,72)=2.62, P =0.04413). In contrast, this isomer
blend captured fewer codling moth males (Table 4; F(7,72)=4.22,P =0.02135; El-Sayed et al . 1998).
Green budworm moth has also been reported to respond to pear ester
(Schmidt et al. 2007; Jósvai et al. 2016). A further field
test in Hungary confirmed this and showed that addition of codlemone to
pear ester does not enhance attraction of either sex (Table 5).
Orchard mating disruption treatments with codlemone strongly diminished
attraction of H. nubiferana males to pheromone traps (Table 2),
corroborating a behavioural effect of codlemone via a dedicated
olfactory channel. This supports the idea that communication disruption
in moths may be achieved with single pheromonal compounds or incomplete
pheromone blends (Cardé and Minks 1995; Porcel et al. 2015), which is of
practical importance for the implementation of pheromonal control of
codling moth and leafrollers in European orchards.
Phylogenetic analysis and antennal
expression
Hedya nubiferana Haworth and Hedya dimidioalba Retzius are
synonymous taxonomic names for green budworm moth. The National Center
for Biotechnology Information (NCBI) lists OR sequences (including PRs)
as ”HnubOR##”.
Predicted putative PRs from H. nubiferana clustered in 4
different subfamily clades when compared with PRs from other tortricid
species (Figure 2A). Several of these displayed homology to receptors inCydia pomonella (CpomOR3, CpomOR6 and CpomOR22) and these
relationships were supported by boostrap values above 95.
Notably, HnubOR6 was >50% similar to CpomOR6. Sequence
comparison analysis revealed that CpomOR1 and HnubOR2 shared 49% amino
acid identity and 66% similarity, while the OR3 orthologs of both
species shared 64% and 76% identity and similarity, respectively.
Amino acid differences between these putative PRs are observed across
the entire length of the protein sequences (Figure 3).
Abundance estimation of the predicted sequences showed that the most
highly expressed were HnubOR6 and HnubOR2. The other 3 putative PRs
detected in male antennae were one or two orders of magnitude lower
(Figure 2B).