Impact of chemical manipulation of tarsal liquids on attachment in the Colorado potato beetle, Leptinotarsa decemlineata

Insect tarsal attachment forces are thought to be influenced by the viscosity and surface tension of a thin film of adhesive liquid (wet adhesion). In beetles, this fluid has been shown to be composed mainly of lipophilic substances that are similar to the cuticular lipids. In this study we investigate whether and how the chemical composition of footprint lipids affects attachment forces in the Colorado potato beetle, Leptinotarsa decemlineata. After application of standardised mixtures of synthetic n-alkanes or alkenes, or a concentrated hydrocarbon extract to the surface of the elytra, we tested the beetles’ attachment performance using a beam force transducer. The results show that only the unsaturated components, but not the straight-chained alkanes reduced friction forces, confirming that attachment performance is influenced by the chemical composition of the adhesive secretion. We estimated the volume of footprint droplets and calculated a mean thickness of the liquid layer of 0.04 μm. The measured friction exceeded the viscous and capillary force expected for a film of this thickness. Therefore, alternative mechanisms (i.e. shear-thinning and solid-like behaviour) for the generation of attachment forces and their dependence on the chemical composition of the liquid are discussed.     

Locomotion in a sticky terrain

The mirid bug Pameridea roridulae lives mutalistically on the protocarnivorous plant Roridula gorgonias. The latter resembles an effective, three-dimensional flypaper trap which captures numerous flying insects. We have recently shown that P. roridulae bugs are not trapped by the plant, because they are covered with a layer of epicuticular grease, which is considerably thicker than in other insects. The present study demonstrates that the bugs’ morphology and locomotory characteristics also contribute to their specialisation for life on the adhesive plant surface. A structural analysis of the mirid bug’s attachment system, and an experimental study on its attachment ability were carried out. In traction force tests, maximum forces of 8.8 mN were measured on adaxial R. gorgonias leaves, corresponding to 126 times the bug’s body weight. On smooth surfaces, generated forces were only 47 times the bug’s body weight. Compared to closely related mirid bug species avoiding contact with plant adhesive secretion, P. roridulae is distinctly stronger and heavier, and holds its body close to the plant substrate. Two locomotion strategies on the glandular hairy plant surfaces are suggested for mirid bug species from the tribus Dicyphini: (1) avoidance strategy, characterised by the slim body held at a large distance from the plant surface by using long, slender legs, and (2) defense strategy, where trapping of the heavy bugs, situated close to the plant surface, is overcome by generating strong forces during locomotion and by having a thick anti-adhesive epicuticular greasy layer on the bugs’ cuticle.     

Structure of the integument in Paranthessius anemoniae claus,a copepod associate of the snakelocks anemone Anemonia sulcata (Pennant)

The integument of Paranthessius anemoniae has been studied with light and electron microscopy. A cuticle with clearly defined epicuticular, exocuticular and endocuticular regions overlies a cellular hypodermal layer. The distribution of carbohydrate, lipid and protein components of the cuticle were demonstrated histochemically. Parabolic striations in oblique sections of cuticle suggest that its molecular architecture fits a “twisted sheet” theory proposed for other species. Arthrodial membranes at body and limb joints have a homogeneous structure, lacking exocuticle and endocuticle. Subcuticular glands appear to secrete substances thought to be responsible for the immunity which Paranthessius seems to have to the nematocysts of its host. Small hairs, situated in cuticular cups which occur over the dorsal body surface are considered to function as rheoreceptors.     

Physical properties of insect cuticular hydrocarbons: Model mixtures and lipid interactions

Cuticular lipids include a diverse array of hydrophobic molecules that play an important role in the water economy of terrestrial arthropods. Their waterproofing abilities are believed to depend largely on their physical properties, but little is known about interactions between different surface lipids to determine the phase behavior of the total lipid mixture. I examined the biophysical properties of binary hydrocarbon mixtures, as a model for interactions between different epicuticular lipids of insects. The midpoint of the solid/liquid phase transition (T_m) for mixtures of n-alkanes differing in chain length equaled the weighted average of the T_ms of the component lipids. This was also true for n-alkane-methylalkane mixtures. However, alkane-alkene mixtures melted at temperatures up to 17°C above the temperature predicted from the weighted average of component lipid T_m values. Hydrocarbon mixtures did not exhibit biphasic melting transitions indicative of independent phase behavior of the component lipids. Instead, melting occurred continuously, over a broader temperature range than pure hydrocarbons.     

Distal leg structures of the Aculeata (Hymenoptera): A comparative evolutionary study of Sceliphron (Sphecidae) and Formica (Formicidae)

The distal parts of the legs of Sceliphron caementarium (Sphecidae) and Formica rufa (Formicidae) are documented and discussed with respect to phylogenetic and functional aspects. The prolegs of Hymenoptera offer an array of evolutionary novelties, mainly linked with two functional syndromes, walking efficiently on different substrates and cleaning the body surface. The protibial-probasitarsomeral cleaning device is almost always well-developed. A complex evolutionary innovation is a triple set of tarsal and pretarsal attachment devices, including tarsal plantulae, probasitarsomeral spatulate setae, and an arolium with an internal spring-like arcus, a dorsal manubrium, and a ventral planta. The probasitarsal adhesive sole and a complex arolium are almost always preserved, whereas the plantulae are often missing. Sceliphron has retained most hymenopteran ground plan features of the legs, and also Formica, even though the adhesive apparatus of Formicidae shows some modifications, likely linked to ground-oriented habits of most ants. Plantulae are always absent in extant ants, and the arolium is often reduced in size, and sometimes vestigial. The arolium contains resilin in both examined species. Additionally, resilin enriched regions are also present in the antenna cleaners of both species, although they differ in which of the involved structures is more flexible, the calcar in Sceliphron and the basitarsal comb in Formica. Functionally, the hymenopteran distal leg combines (a) interlocking mechanisms (claws, spine-like setae) and (b) adhesion mechanisms (plantulae, arolium). On rough substrate, claws and spine-like setae interlock with asperities and secure a firm grip, whereas the unfolding arolium generates adhesive contact on smooth surfaces. Differences of the folded arolium of Sceliphron and Formica probably correlate with differences in the mechanism of folding/unfolding.     

Patterns of biosynthesis and accumulation of hydrocarbons and contact sex pheromone in the female german cockroach,Blattella germanica

De novo synthesis of contact female sex pheromone and hydrocarbons in Blattella germanica was examined using short in vivo incubations. Accumulation of pheromone on the epicuticular surface and the internal pheromone titer were related to age-specific changes in hydrocarbon synthesis and accumulation in normal and allatectomized females. The incorporation of radiolabel from [1-¹⁴C]propionate into the cuticular methyl ketone pheromone fraction was positively related to corpora allata activity during two gonotrophic cycles. During peak pheromone production the total internal lipid fraction contained greater titers of pheromone than the cuticular surface, and it too exhibited a cycle internally, preceding the rise in external pheromone. This suggests that synthesis and accumulation of pheromone internally are followed by transport of pheromone to the epicuticular surface where it accumulates. Radiolabel was incorporated efficiently into both cuticular and internal hydrocarbons after the imaginal molt and until the peak of pheromone synthesis, but it declined to lower levels before ovulation and throughout pregnancy. The internal hydrocarbon titer decreased 58% after oviposition, suggesting deposition in the egg case. It remained relatively unchanged during pregnancy and increased again during the second gonotrophic cycle. In allatectomized females, hydrocarbon synthesis was reduced relative to control females until oviposition in the latter. However, subsequent rates of hydrocarbon synthesis in allatectomized females (without oothecae) exceeded the rates in sham-operated females (with oothecae). In the absence of ovarian uptake of hydrocarbons, the internal titer increased without the decline found in control females at oviposition. As internal hydrocarbons increased, so did cuticular hydrocarbons and both internal and cuticular methyl ketone pheromones. These patterns corresponded well with feeding patterns in sham-operated and allatectomized females, suggesting that pheromone production is normally regulated by stage-specific feeding-induced hydrocarbon synthesis (precursor accumulation internally) and juvenile hormoneinduced conversion of hydrocarbon to pheromone. They also suggest that both the cuticle and the ovaries might be target sites for hydrocarbon and possibly methyl ketone deposition. © 1994 Wiley-Liss, Inc.     

Two sides of a leaf blade: <Emphasis Type="Italic">Blumeria graminis</Emphasis> needs chemical cues in cuticular waxes of <Emphasis Type="Italic">Lolium perenne</Emphasis> for germination and differentiation

Plant surface characteristics were repeatedly shown to play a pivotal role in plant–pathogen interactions. The abaxial leaf surface of perennial ryegrass (Lolium perenne) is extremely glossy and wettable compared to the glaucous and more hydrophobic adaxial surface. Earlier investigations have demonstrated that the abaxial leaf surface was rarely infected by powdery mildew (Blumeria graminis), even when the adaxial surface was densely colonized. This led to the assumption that components of the abaxial epicuticular leaf wax might contribute to the observed impairment of growth and development of B. graminis conidia on abaxial surfaces of L. perenne. To re-assess this hypothesis, we analyzed abundance and chemical composition of L. perenne ab- and adaxial epicuticular wax fractions. While the adaxial epicuticular waxes were dominated by primary alcohols and esters, the abaxial fraction was mainly composed of n-alkanes and aldehydes. However, the major germination and differentiation inducing compound, the C₂₆-aldehyde n-hexacosanal, was not present in the abaxial epicuticular waxes. Spiking of isolated abaxial epicuticular Lolium waxes with synthetically produced n-hexacosanal allowed reconstituting germination and differentiation rates of B. graminis in an in vitro germination assay using wax-coated glass slides. Hence, the absence of the C₂₆-aldehyde from the abaxial surface in combination with a distinctly reduced surface hydrophobicity appears to be primarily responsible for the failure of normal germling development of B. graminis on the abaxial leaf surfaces of L. perenne.     

Histochemical and ultrastructural analyses of adhesive setae of lizards indicate that they contain lipids in addition to keratins

We studied the distribution of lipid material and organelles in the epidermal layers of toe pads from two species of lizards representing the two main lizard families in which adhesive scansors are found (gekkonids and polychrotids), the dull day gecko, Phelsuma dubia and the green anole, Anolis carolinensis. Although lipids are a conspicuous component of the mesos layer of squamate reptiles and function in reducing cutaneous water loss, their distribution has not been specifically studied in the highly elaborated epidermal surface of adhesive toe pads. We found that, in addition to the typical cutaneous water loss‐resistant mesos and alpha‐layer lipids, the Oberhäutchen (including the setae) on the most exterior layers of the epidermis in P. dubia and A. carolinensis also contain lipid material. We also present detailed histochemical and ultrastructural analyses of the toe pads of P. dubia, which indicate that lipid material is closely associated spatially with maturing setae as they branch during the renewal phase of epidermal regeneration. This lipid material appears associated with the packing of keratin within setae, possibly affecting permeability to water loss in the pad lamella, where the surface area is from 4–60‐fold greater compared with normal scales. J. Morphol., 2011. © 2011 Wiley‐Liss, Inc.     

STRUCTURAL DIMORPHISM OF STIGMATIC PAPILLAE IN DISTYLOUS LINUM SPECIES

Intermorph differences in the wall structure and constituents of stigmatic papillae are described for distylous Linum pubescens and L. grandiflorum. In the long-styled morph of both species the wall portion around the apex of the papilla has a thickened cellulose-pectin layer. In the short-styled morph of L. pubescens a cap zone is interposed between the cuticle and apical portion of the papilla wall. The subcuticular cap space contains pectins and lipid particles. Similar particles are also present in deposits on the cuticle surface. Papillae of the short-styled morph in L. grandiflorum lack a cap zone and have only epicuticular lipid deposits. Other distylous Linum species in which the two morphs differ in wall structure of the papillae are L. mucronatum, L. flavum, L. perenne, L. austriacum, and L. maritimum. Studies of stigma dimorphism can help to elucidate evolutionary relations between dimorphic and monomorphic Linum species.     

Etude ultrastructurale de l'évolution du tégument de la Sangsue Hirudo medicinalis L. (Annélide,Hirudinée) au cours d'un cycle de mue / Developmental changes in the integument of the leech Hirudo medicinalis L. (Annelida,Hirudinea) during a molting cycle. An ultrastructural study

Summary

The integument of the leech Hirudo medicinalis is mainly composed of a single layer of cuticle-secreting epidermal cells. The cuticle is made up of collagen fibers which support a layer of membrane-bound epicuticular projections.

Shedding of the old cuticle is preceded by the formation of a new cuticle. The epicuticular projections are the first to develop: they originate from the tips of numerous microvilli of the epidermal cells. As soon as it appears, the newly-formed collagen layer is firmly attached to the epidermal cells by numerous hemidesmo-somes, whereas the old cuticle is no longer connected with the epidermal surface. The epidermal cells exhibit marked characteristics of secretory activity during the laying down of the new cuticle.

The observations are discussed in connexion with recent findings of high ecdysteroid levels in leeches at the beginning of the molting cycle.     

Biosynthesis of pentacyclic triterpenoids in leaves of Ilex aquifolium L.

Ilex leaves can utilize sucrose and mevalonate for the synthesis of triterpene esters. Mevalonate is also used for the synthesis of free triterpenoid alcohols, but sucrose is not. The selectivity of precursor utilization indicates separate sites for triterpenol and triterpenol-ester synthesis. The sites of synthesis are not found at the main locations of accumulation of triterpenols (the epicuticular wax) or triterpene esters (the cytoplasmatic lipid globules). Transport from the site of synthesis to the lipid globules, and especially towards the epicuticular wax, is slow.Abbreviations MVA mevalonic acid lactone - TLC thin layer chromatography     

Structure of the tarsi in some Stenus species (Coleoptera,Staphylinidae): external morphology,ultrastructure, and tarsal secretion

SEM studies show that the differentiation among Stenus species with respect to the formation of the tarsi (wide bilobed vs. slender tarsomeres) takes place with a considerable augmentation of tarsal ventral setae in wide bilobed tarsomeres. The structural diversity of ventral tarsal setae among and within species is discussed with respect to 1) their different roles as mechanosensilla and tenent setae, respectively, and 2) the different selection pressures in terms of adhesive requirements along the longitudinal tarsus axis. The tarsi are provided with four groups of tarsal mechanosensilla, comprising hair and bristle sensilla, campaniform sensilla, and scolopidia. The tarsus wall is supported by an epidermis, which forms three different types of glands pouring their secretion via different exit paths onto the outer cuticle. The organization and ultrastructure of each of these glands is described. Only one (unicellular) gland is directly associated with the ventral tenent setae and is thus considered to form the main part of the adhesive secretion. The beetles appear to release the tarsal secretion through mediation of the tenent setae, which contains a lipid and a proteinaceous fraction. I propose that the secretion is discharged to the outside via a system of very fine pore canals in the wall of the setal shaft. Gas chromatography and infrared spectroscopy revealed that the lipid fraction of the secretion is a mixture of unsaturated fatty acid glycerides and aliphatic hydrocarbons whose spectra are similar to those of extractions of the superficial lipid coating of the body surface.     

The structure of a model membrane in relation to the viscoelastic properties of the red cell membrane

The molecular arrangement within a lamellar structure composed of human erythrocyte lipids is determined. The 45 A thick lipid layer, in water, is filled in the interior with a liquid-like configuration of the hydrocarbon chains of phospholipid molecules and is covered on both sides by their hydrophilic polar groups. Cholesterol is located so that part of its steroid nucleus is between the polar groups of the phospholipid molecules while the rest of the molecule extends into the inner hydrocarbon layer. This lipid leaflet would be expected to have the mechanical properties of a purely liquid surface, as other authors have shown for the "black" lipid membranes. Data are presented which demonstrate that the intact erythrocyte membrane is a tough viscoelastic substance with a Young''s modulus of 10⁶–10⁸ dynes/cm² and a viscosity of 10⁷–10¹⁰ poises. The parameters and the kinetics of membrane breakdown are incompatible with the model system of pure lipid. Caution must be exercised in applying various data on the model systems to intact membranes.     

Corrigendum: Shear adhesive performance of leaf-cutting ant workers (Atta cephalotes)

In our previous article, Shear adhesive performance of leaf-cutting ant workers (Atta cephalotes), a technical error occurred when calculating shear adhesive performance divided by tarsal pad area of worker ants carrying leaves (Stark, A. Y., Davis, H. R., & Harrison, W. K. (2019). Biotropica, 51(4), 572–580). Shear adhesive performance of worker ants carrying leaves was mistakenly divided by the tarsal pad area of workers riding on leaves. This has slightly changed our results. After correcting for this error, there is now no statistically significant difference in shear adhesive performance divided by tarsal pad area of worker ants riding on leaves and those carrying leaves. Shear adhesive performance of these groups remains statistically different than soldiers, and our original results (shear adhesive performance of all three groups differ from one another) are consistent with the reported trend in the original article using true pad area for each individual (rather than estimated). We list below the pages and figures in the original article that this correction applies to. All authors agree with this change, apologize for their error, and appreciate the opportunity granted by the editors and publisher to make this correction. Our original conclusion, that there is variation in shear adhesive performance per unit area among worker roles of Atta cephalotes, is still supported by these corrected results.     

Changes in emergence phenology,fatty acid composition,and xenobiotic-metabolizing enzyme expression is associated with increased insecticide resistance in the Colorado potato beetle

The Colorado potato beetle (Leptinotarsa decemlineata) is a major agricultural pest of solanaceous crops. An effective management strategy employed by agricultural producers to control this pest species is the use of systemic insecticides. Recent emphasis has been placed on the use of neonicotinoid insecticides. Despite efforts to curb resistance development through integrated pest management approaches, resistance to neonicotinoids in L. decemlineata populations continues to increase. One contributing factor may be alterations in insect fatty acids, which have multiple metabolic functions and are associated with the synthesis of xenobiotic-metabolizing enzymes to mitigate the effects of insecticide exposure. In this study, we analyzed the fatty acid composition of L. decemlineata populations collected from an organic production field and from a commercially managed field to determine if fatty acid composition varied between the two populations. We demonstrate that a population of L. decemlineata that has a history of systemic neonicotinoid exposure (commercially managed) has a different lipid composition and differential expression of known metabolic detoxification mechanisms relative to a population that has not been exposed to neonicotinoids (organically managed). The fatty acid data indicated an upregulation of Δ⁶ desaturase in the commercially managed L. decemlineata population and suggest a role for eicosanoids and associated metabolic enzymes as potential modulators of insecticide resistance. We further observed a pattern of delayed emergence within the commercially managed population compared with the organically managed population. Variations in emergence timing together with specific fatty acid regulation may significantly influence the capacity of L. decemlineata to develop insecticide resistance.     

Chemical composition of the epicuticular and intracuticular wax layers on adaxial sides of Rosa canina leaves

BACKGROUND AND AIMS: The waxy cuticle is the first point of contact for many herbivorous and pathogenic organisms on rose plants. Previous studies have reported the average composition of the combined wax extract from both sides of rose leaves. Recently, the compositions of the waxes on the adaxial and abaxial surfaces of Rosa canina leaves were determined separately. In this paper, a first report is made on the compositions of the epicuticular and intracuticular wax layers of Rosa canina leaves. The methods described enable the determination of which compounds are truly available at the surface for plant-organism interactions. METHODS: An adhesive was used to mechanically strip the epicuticular wax from the adaxial leaf surface and the removal was visually confirmed using scanning electron microscopy. After the epicuticular wax had been removed, the intracuticular wax was then isolated using standard chemical extraction. Gas chromatography, flame ionization detection and mass spectrometry were used to identify and quantify compounds in the separated wax mixtures. KEY RESULTS: The epicuticular wax contained higher concentrations of alkanes and alkyl esters but lower concentrations of primary alcohols and alkenols when compared to the intracuticular wax. In addition, the average chain lengths of these compound classes were higher in the epicuticular wax. Secondary alcohols were found only in the epicuticular layer while triterpenoids were restricted mainly to the intracuticular wax. CONCLUSIONS: A gradient exists between the composition of the epi- and intracuticular wax layers of Rosa canina leaves. This gradient may result from polarity differences, in part caused by differences in chain lengths. The outer wax layer accessible to the phyllosphere showed a unique composition of wax compounds. The ecological consequences from such a gradient may now be probed.     

Hydrocarbons from epicuticular waxes of Citrus peels

The hydrocarbon fraction of the epicuticular wax layer of peel from 5 cultivars of Citrus was monitored at various stages of fruit development. GLC     

What do microbes encounter at the plant surface? Chemical composition of pea leaf cuticular waxes

In the cuticular wax mixtures from leaves of pea (Pisum sativum) cv Avanta, cv Lincoln, and cv Maiperle, more than 70 individual compounds were identified. The adaxial wax was characterized by very high amounts of primary alcohols (71%), while the abaxial wax consisted mainly of alkanes (73%). An aqueous adhesive of gum arabic was employed to selectively sample the epicuticular wax layer on pea leaves and hence to analyze the composition of epicuticular crystals exposed at the outermost surface of leaves. The epicuticular layer was found to contain 74% and 83% of the total wax on adaxial and abaxial surfaces, respectively. The platelet-shaped crystals on the adaxial leaf surface consisted of a mixture dominated by hexacosanol, accompanied by substantial amounts of octacosanol and hentriacontane. In contrast, the ribbon-shaped wax crystals on the abaxial surface consisted mainly of hentriacontane (63%), with approximately 5% each of hexacosanol and octacosanol being present. Based on this detailed chemical analysis of the wax exposed at the leaf surface, their importance for early events in the interaction with host-specific pathogenic fungi can now be evaluated. On adaxial surfaces, approximately 80% of Erysiphe pisi spores germinated and 70% differentiated appressoria. In contrast, significantly lower germination efficiencies (57%) and appressoria formation rates (49%) were found for abaxial surfaces. In conclusion, the influence of the physical structure and the chemical composition of the host surface, and especially of epicuticular leaf waxes, on the prepenetration processes of biotrophic fungi is discussed.     

Chemical composition of epicuticular wax crystals on the slippery zone in pitchers of five <Emphasis Type="Italic">Nepenthes</Emphasis> species and hybrids

Plants of the carnivorous genus Nepenthes efficiently trap insects in leaf pitchers, mostly employing epicuticular wax crystals on the pitcher walls to make them slippery for the prey. In the present study, the compositions and micromorphologies of the wax crystals of five Nepenthes species and hybrids were analysed in order to test whether the chemical principles underlying this ecological function are widespread within the genus. Three wax layers could be distinguished within the Nepenthes pitcher cuticles: (1) the outermost part of the crystals forming the platelets visible in standard scanning electron microscopy, (2) the bottom portion of the epicuticular wax crystals, and (3) an intracuticular wax layer. The composition of the intracuticular wax differed significantly from that of the neighbouring epicuticular layer. The compositions of corresponding wax mixtures from all five Nepenthes species and hybrids were very similar, with almost equal amounts of very long chain aldehydes and primary alcohols. While triacontanal (C₃₀ aldehyde) was prevailing in the epicuticular crystals of Nepenthes albomarginata and Nepenthes x intermedia, Nepenthes x superba and Nepenthes x henriana were found to have especially high percentages of dotriacontanal (C₃₂ aldehyde). Nepenthes “khasiana” had an intermediate aldehyde composition with almost equal amounts of both chain lengths.