Composition and Variability of Epicuticular Lipids of Azaleas and their Relationship to Azalea Lace Bug Resistance

Epicuticular lipids were extracted from the foliage of six deciduous and one evergreen azalea genotypes (Rhododendron sp.) and identified by gas chromatography-mass spectrometry. The relationship of leaf-surface lipid composition with measures of resistance to azalea lace bug, Stephanitis pyrioides Scott, was evaluated. Each genotype had a distinct epicuticular lipid composition. The major surface lipid components from all test taxa were n-alkanes and triterpenoids. In the most resistant genotypes [R. canescens Michaux and R. periclymenoides (Michaux) Shinners] ursolic acid, n-hentriacontane, and n-nonacosane were the most abundant epicuticular lipids. The lipids present in largest proportion among all susceptible deciduous genotypes tested were α-amyrin, β-amyrin, and n-nonacosane. The proportions of the lipid components from the same plant of each genotype varied between spring and fall samples. Among classes of lipids, n-alkanes, n-1-alkanols, and triterpenoids had significant correlations with azalea lace bug behavior on host plants. Among individual components, heptadecanoic acid, n-hentriacontane, oleanolic acid, ursolic acid and one unknown compound (with major mass spectra 73/179/192/284/311) were significantly negatively correlated with host plant susceptibility to azalea lace bug, as measured by oviposition, leaf area damaged, egg and nymphal development, and nymphal survivorship. Triacontanol, α-amyrin, β-amyrin, and three unknowns were significantly positively correlated with host plant susceptibility. Acceptance or rejection by azalea lace bug to a particular plant may be mediated by a balance of positively and negatively interpreted sensory signals evoked by plant chemicals. This study indicated that the high levels of resistance observed in R. canescens and R. periclymenoides may be due to the lesser amount or the absence of attractants and stimulants for feeding or oviposition.

Factors contributing to seasonal increases in inoculative freezing resistance in overwintering fire-colored beetle larvae dendroides canadensis

The insects and microarthropods that vary seasonally in susceptibility to cross-cuticular inoculation by external ice (inoculative freezing) represent a phylogenetically diverse group; however, few studies have explored possible mechanisms experimentally. This study documents seasonally variable inoculative freezing resistance in Dendroides canadensis beetle larvae and combines immunofluorescence, in vivo removal of epicuticular lipids and in vitro chamber studies to explore the roles of seasonal modification in the cuticle and in epidermal and hemolymph antifreeze proteins (AFPs). Seasonal cuticular modifications contribute to the inhibition of inoculative freezing since more cold-hardy larvae froze inoculatively when epicuticular waxes were removed with hexane and, in in vitro chamber experiments, cuticle patches (with the underlying epidermis removed) from winter larvae provided greater protection from inoculative freezing than did cuticle patches from summer larvae. The results indicate that seasonal modifications in epidermal and hemolymph AFPs contribute most strongly to the inhibition of inoculative freezing. Subcuticular epidermal AFPs were present in immunocytochemically labeled transverse sections of winter larvae but were absent in summer ones. Winter integument patches (cuticle with epidermis) were more resistant to inoculative freezing than were summer integument patches. Integument patches resisted inoculative freezing as well as live winter-collected larvae only when hemolymph AFP was added. The results also suggest that some integumentary ice nucleators are removed in cold-hardy larvae and that AFP promotes supercooling by inhibiting the activity of these nucleators.