Beyond aerodynamics: The critical roles of the circulatory and tracheal systems in maintaining insect wing functionality

Insect wings consist almost entirely of lifeless cuticle; yet their veins host a complex multimodal sensory apparatus and other tissues that require a continuous supply of water, nutrients and oxygen. This review provides a survey of the various living components in insect wings, as well as the specific contribution of the circulatory and tracheal systems to provide all essential substances. In most insects, hemolymph circulates through the veinal network in a loop flow caused by the contraction of accessory pulsatile organs in the thorax. In other insects, hemolymph oscillates into and out of the wings due to the complex interaction of several factors, such as heartbeat reversal, intermittent pumping of the accessory pulsatile organs in the thorax, and the elasticity of the wall of a special type of tracheae. A practically unexplored subject is the need for continuous hydration of the wing cuticle to retain its flexibility and toughness, including the associated problem of water loss due to evaporation. Also, widely neglected is the influence of the hemolymph mass and the circulating flow in the veins on the aerodynamic properties of insect wings during flight. Ventilation of the extraordinarily long wing tracheae is probably accomplished by intricate interactions with the circulatory system, and by the exchange of oxygen via cutaneous respiration.     

Cuticle degrading proteases from insect moulting fluid and culture filtrates of entomopathogenic fungi

Insects degrade their own cuticle during moulting, a process which is catalysed by a complex mixture of enzymes. Entomopathogenic fungi infect the insect host by penetration of the cuticle, utilizing enzymatic and/or physical mechanisms. Protein is a major component of insect cuticle and a major recyclable resource for the insect and, therefore, represents a significant barrier to the invading fungus. To this end, both insects and entomopathogenic fungi produce a variety of cuticle degrading proteases. The aim of this paper is to review these proteases and to highlight their similarities, with particular reference to the tobacco hornworm, Manduca sexta, and the entomopathogenic fungus, Metarhizium anisopliae     

Extracellular hydrolytic enzymes produced by entomopathogenic fungi--role in an infection process

Entomopathogenic fungi have a great potential in biological control of insect pest population. Fungal pathogens are promising source of insecticides and notable alterative to chemical pesticides. These fungi possess a unique mechanism of insects paralysis. As natural enemies of insects they attack direct host cuticle via a combination of mechanical pressure and cuticle-degrading enzymes. Entomopathogenic fungi produce several proteo-, chitino- and lipolytic enzymes, which are accepted as key factors in insect mycosis. The role of extracellular enzymes in pathogenesis is still not well understood. Profound understanding the mechanisms of insect paralysis by entomopathogenic fungi will help in the production of safer for environment and more efficiency mycoinsecticides.     

FAR gene enables the brown planthopper to walk and jump on water in paddy field

Many insects can live on water and survive being caught in the rain. Current research has shown that insect cuticular hydrocarbons(CHC) confer desiccation resistance to maintain water balance. In this study, we identified a fatty acyl-CoA reductase gene(NlFAR) of the rice brown planthopper, Nilaparvata lugens that is essential for the production of CHCs, and found that NlFAR is essential for N. lugens to walk and jump on water when moving from one rice plant to another in paddy fields. NlFAR was mainly expressed in the integument at the beginning of each molt. Cuticular surface analysis by scanning electron microscopy and characterization of CHC extracts indicated that N. lugens with knockdown of NlFAR using RNA inference(RNAi)had a neater epicuticle layer and a significant decrease in CHC contents. Knockdown of NlFAR did not influence the desiccation resistance of N. lugens, but the ds NlFAR-treated insects were easily adhered and moistened by water droplets or their own secreted honeydew and unable to walk or jump on water. These results suggested that NlFAR is a crucial enzyme for CHC biosynthesis and cuticle waterproofing, but not for water retention of N. lugens, which may provide a potential strategy for pest management.     

THE PENETRATION OF THE INSECT CUTICLE BY ISOMERS OF BENZENE HEXACHLORIDE

The penetration of the α-, β-, γ- and δ-isomers of benzene hexachloride through the insect cuticle has been studied by a micro-colorimetric technique.
Grain weevils were exposed for varying periods to deposits of pure isomers of benzene hexachloride on filter-paper. The amounts of each isomer picked up by the insects were determined as two fractions, one 'outside' representing that which could be removed by cold methanol washing, and the other 'inside' recovered after decomposing the insects with nitric acid.
The results show that the amounts of each isomer taken up by the insects are in the approximate ratio of their solubilities in hydrocarbon solvents, and that the y-isomer penetrates through the outer layers of the insect integument much more rapidly than the other three isomers tested.
On the basis of these results it is concluded: (i) that the first stage of pick-up of insecticide by the insects is simple solution of the insecticide in the outer waxy covering of the epicuticle; and (ii) that structural effects play an important part in the penetration through the insect cuticle as well as in toxic effects at the site of action.     

培菌昆虫相关放线菌的次级代谢产物研究进展

昆虫共生微生物是一种特殊的微生物资源,其中放线菌在昆虫肠道、体表和巢穴中广泛分布。近年来,人们从培菌昆虫来源的放线菌中分离得到多种新型化合物,可以选择性抑制菌圃的致病真菌,部分还对植物致病真菌、昆虫致病真菌、人类病原菌和癌细胞有抑制活性。因此,研究培菌昆虫相关微生物不仅有助于了解宿主与微生物的共生机制,还能发掘新的活性物质,用于生物农药、生物医药的开发。本文对培菌昆虫来源放线菌次级代谢产物的研究进展进行了综述。     

Aggregation in the haematophagous bug Triatoma infestans: a novel assembling factor

Abstract. The response of Triatoma infestans (Klug) to odours left behind by walking insects was analysed. The insects aggregated on papers marked with putative chemical footprints in the absence of other assembling chemical signals. Contact of the receptor insect with the treated surface was necessary to elicit the response because the signal was not able to attract insects, only to arrest them on marked surfaces. These requirements contrast with other chemical aggregation signals of triatomines that attract the insect and involve olfactory responses. By washing the cuticle of the insects with hexane and impregnating papers with the extract, a response was evoked in test insects similar to that elicited by papers on which insects had walked. The biological role of this novel aggregation factor is discussed in relation to other assembling signals of different origin.     

A cytochrome p450 conserved in insects is involved in cuticle formation

The sequencing of numerous insect genomes has revealed dynamic changes in the number and identity of cytochrome P450 genes in different insects. In the evolutionary sense, the rapid birth and death of many P450 genes is observed, with only a small number of P450 genes showing orthology between insects with sequenced genomes. It is likely that these conserved P450s function in conserved pathways. In this study, we demonstrate the P450 gene, Cyp301a1, present in all insect genomes sequenced to date, affects the formation of the adult cuticle in Drosophila melanogaster. A Cyp301a1 piggyBac insertion mutant and RNAi of Cyp301a1 both show a similar cuticle malformation phenotype, which can be reduced by 20-hydroxyecdysone, suggesting that Cyp301a1 is an important gene involved in the formation of the adult cuticle and may be involved in ecdysone regulation in this tissue.     

The essential role of bursicon during <Emphasis Type="Italic">Drosophila</Emphasis>development

Background  

The protective external cuticle of insects does not accommodate growth during development. To compensate for this, the insect life cycle is punctuated by a series of molts. During the molt, a new and larger cuticle is produced underneath the old cuticle. Replacement of the smaller, old cuticle culminates with ecdysis, a stereotyped sequence of shedding behaviors. Following each ecdysis, the new cuticle must expand and harden. Studies from a variety of insect species indicate that this cuticle hardening is regulated by the neuropeptide bursicon. However, genetic evidence from Drosophila melanogaster only supports such a role for bursicon after the final ecdysis, when the adult fly emerges. The research presented here investigates the role that bursicon has at stages of Drosophila development which precede adult ecdysis.     

The MrCYP52 cytochrome P450 monoxygenase gene of Metarhizium robertsii is important for utilizing insect epicuticular hydrocarbons

Fungal pathogens of plants and insects infect their hosts by direct penetration of the cuticle. Plant and insect cuticles are covered by a hydrocarbon-rich waxy outer layer that represents the first barrier against infection. However, the fungal genes that underlie insect waxy layer degradation have received little attention. Here we characterize the single cytochrome P450 monoxygenase family 52 (MrCYP52) gene of the insect pathogen Metarhizium robertsii, and demonstrate that it encodes an enzyme required for efficient utilization of host hydrocarbons. Expressing a green florescent protein gene under control of the MrCYP52 promoter confirmed that MrCYP52 is up regulated on insect cuticle as well as by artificial media containing decane (C10), extracted cuticle hydrocarbons, and to a lesser extent long chain alkanes. Disrupting MrCYP52 resulted in reduced growth on epicuticular hydrocarbons and delayed developmental processes on insect cuticle, including germination and production of appressoria (infection structures). Extraction of alkanes from cuticle prevented induction of MrCYP52 and reduced growth. Insect bioassays against caterpillars (Galleria mellonella) confirmed that disruption of MrCYP52 significantly reduces virulence. However, MrCYP52 was dispensable for normal germination and appressorial formation in vitro when the fungus was supplied with nitrogenous nutrients. We conclude therefore that MrCYP52 mediates degradation of epicuticular hydrocarbons and these are an important nutrient source, but not a source of chemical signals that trigger infection processes.     

THE TOXICITY OF DDT IN ABRASIVE AND NON-ABRASIVE DUSTS TO THE RICE WEEVIL, CALANDRA ORYZAE L. (COLEOPT., CURCULIONIDAE)*

The toxicity of DDT in different dust carriers to the rice weevil, Calandra oryzae , was determined under standardized conditions, using deposits large enough to ensure that the insects accumulated an excess of dust.
Some evidence was obtained that DDT is transported to the cuticle as a vapour.
At high humidity, the toxicity of DDT was not markedly affected by any carrier except charcoal which reduced the toxicity, probably by absorption of DDT vapour. Small differences in toxicity of DDT caused by other carriers could not be accounted for by differences in their average particle size, bulk density, amount adhering to insect, surface area, abrasiveness to insects or effect on behaviour of the insect.
At low humidity, abrasive dusts killed the insects by desiccation, thus adding to the toxic effect of DDT. Abrasion of the insect's cuticle did not affect the apparent rate of penetration of DDT at 50% R. H. or at 95% R. H.
Starved insects were more susceptible to DDT poisoning, and in some experiments abrasive carriers increased the toxicity of DDT by preventing the insects from feeding.     

Ecdysis period of Rhodnius prolixus head investigated using phase contrast synchrotron microtomography

Microtomography using synchrotron sources is a useful tool in biological imaging research since the phase coherence of synchrotron beams can be exploited to obtain images with high contrast resolution. This work is part of a series of works using phase contrast synchrotron microtomography in the study of Rhodnius prolixus head, the insect vector of Chagas’ disease, responsible for about 12,000 deaths per year. The control of insect vector is the most efficient method to prevent this disease and studies have shown that the use of triflumuron, a chitin synthesis inhibitor, disrupted chitin synthesis during larval development and it’s an alternative method against insect pests.The aim of this work was to investigate the biological effects of treatments with triflumuron in the ecdysis period (the moulting of the R. prolixus cuticle) using the new imaging beamline IMX at LNLS (Brazilian Synchrotron Light Laboratory). Nymphs of R. prolixus were taken from the Laboratory of Biochemistry and Physiology of Insects, Oswaldo Cruz Foundation, Brazil. Doses of 0.05 mg of triflumuron were applied directly to the abdomen on half of the insects immediately after feeding. The insects were sacrificed 25 days after feeding (intermoulting period) and fixed with glutaraldehyde.The results obtained using phase contrast synchrotron microtomography in R. prolixus showed amazing images of the effects of triflumuron on insects in the ecdysis period, and the formation of the new cuticle on those which were not treated with triflumuron. Both formation and malformation of this insect’s cuticle have never been seen before with this technique.     

Physical and physiological characteristics of Trogoderma glabrum infected with the schizogregarine pathogen Mattesia trogodermae

Physical and physiological characteristics of an infection of Trogoderma glabrum by Mattesia trogodermae were studied. Weights of infected larvae drop markedly between 10 and 20 days post-infection at 30° and 35°C. This loss is less abrupt and not as great when the incubation temperature is 25°C. Reduction of dry matter is gradual during the first 12 days of infection, but drops 70% from 12 to 20 days post-infection.Glycogen reserves in both infected and control insects drop 50% within 3 days after deprivation of food. Healthy insects recover and begin to reconstitute lost glycogen; however, infected larvae continue to deplete glycogen to 15% of prestarvation levels. Similarly, insect protein is reduced 40% within 7 days after starvation and noninfected insects apparently halt protein metabolism at this level. Diseased larvae continue to lose protein to 20% of prestarvation amounts. These losses are at least partially attributable to insect metabolism since infected insects defecate significantly more than control larvae. It is thought that defecation is an effective route of water loss which occurs during the first 20 days of infection. Relative humidities ranging from 0 to 84% had no obvious effects on mortality rates, indicating that water loss is effected through routes other than evaporation through the cuticle, e.g., failure of water retention systems and elimination of body water with feces.     

Whole-tree sap flow is substantially diminished by leaf herbivory

Ecohydrological models consider the relationship between tree size and structure (especially leaf area index) and water use but generally treat herbivory as a source of unwanted noise in the data. Little is known of how insect damage to leaves influences whole-plant water use in trees. Water use is driven by environmental demand and the total leaf area through which transpiration can occur, but the effects of insects are expected to be complex. Different kinds of insects could have different effects; for example, chewing insects reduce leaf area, whereas sucking and tissue mining insects reduce leaf function without reducing area. Further, plants respond to herbivory in a range of ways, such as by altering leaf production or abscising leaves. We examined the effect of insects on Eucalyptus blakelyi in a woodland near Canberra, Australia, using sap flow velocity as a measure of whole-plant water use. We applied insecticide to 16 trees matched to an untreated control group. After 6 months, we examined the effects on sap flow velocity and crown condition. There was a general increase in sap flow velocity as trees produced leaves over the growing season, but the increase in sap flow for trees without insecticide protection was half that of the protected trees (increase: 4.4 vs. 9.0 cm/h, respectively). This dramatic effect on sap flow was consistent with effects on crown condition. Unprotected trees had 20% less leaf mass per unit stem in the crown. In addition, unprotected trees had a 20% greater loss of leaf functional area from necrosis. It should be noted that these effects were detected in a year in which there was not an outbreak of the psyllids (Homoptera) that commonly cause severe leaf damage to this tree species. It is predicted that the effect in a psyllid outbreak year would be even more substantial. This result underscores the significant impact that insect herbivores can have on an ecological process of significance to the ecosystem, namely, the movement of water from the soil to the atmosphere.     

Species richness of gall-forming insects in a tropical rain forest: correlations with plant diversity and soil fertility

We tested two hypotheses to explain changes in species richness ofgall-forming insects. The first hypothesis proposes that gall-forming insectspecies richness increases as more potential host–plant species areavailable. The second hypothesis implies that soil fertility affects plantcolonization by gall-forming insects. Seven sites, representing strongdifferences in vegetation and soil were chosen at the Lacandona tropical rainforest region, Chiapas, Mexico. Overall, we found 1522 individual plantsbelonging to 340 different plant species. From this, we found gall-forminginsects on 737 (43.9%) plants and on 74 (22%) of total plant species. We found asignificant negative correlation between gall-forming insect species richnessand species richness of plants, which does not support the hypothesis that plantspecies richness is an important factor in generating the radiation ofgall-forming insects. Using phosphorus as an indicator of soil fertility, wefound the lowest number of plants with gall-forming insects and the smallestgall-forming insect load per individual plant in the more fertile soil(alluvial). In contrast, the highest number of plants with galls and the highestgall-forming insect load per plant were found at a savanna-like vegetationsite, where the poorest soil was recorded. These results did not support thesoil fertility hypothesis in terms of species richness, but did with respect toabundance of plants with galls.     

Dielectric properties of insect tissues

In order to explain some effects of microwave irradiation on insects it is necessary to consider a mathematical model. The knowledge of dielectric properties of a typical insect tissue is crucial for such a model. A method based on shift of resonant frequency and of quality factor measurement in a resonator both before and after the insertion of samples was used. The method (measurements at a frequency of 2375 MHz) has been described in detail. A large number of measurements were performed on different kinds of typical insect tissues (cuticle, fat body, muscles, reproductive organs and eggs) for their dielectric properties. The values obtained compare well to those reported in the literature for some mammals. Differences seemed to depend on different water-to-fat content ratios. However, no simple dependence on the water content was found. Values obtained from insect tissue material have been discussed in detail.     

Agroforestry coffee soils increase the insect‐suppressive potential offered by entomopathogenic fungi over full‐sun soils: A case proposing a “bait survival technique”

Entomopathogenic fungi are important natural enemies of insects. However, there is little information on the insect‐suppressive potential of these fungi and possible effects of farming management on this. Meanwhile, changes in natural landscapes due to agricultural intensification have caused considerable biodiversity loss and consequent decay of ecosystem services. However, the adoption of practices such as agroforestry in agroecosystems can foster abiotic and biotic conditions that conserve biodiversity, consequently restoring the provision of ecosystems services. Here, we assessed the effect of management systems (agroforestry or full‐sun) on the pest‐suppressive potential of entomopathogenic fungi in Brazilian coffee plantations. We used the insect bait method coupled with survival analyses to assess the speed of kill by entomopathogenic fungi and their presence in soil samples from both farming systems. We found that insects exposed to agroforestry soils died more quickly than insects exposed to full‐sun soils. Of the fungi isolated from the bait insects, Metarhizium was found most frequently, followed by Beauveria. Meanwhile, Fusarium was frequently isolated as primary or secondary infections. We propose that the differential survival of insects is indicative of a greater suppressive potential by entomopathogenic fungi in agroforestry, and that this could be promoted by the diversified landscape, microclimatic stability, and reduced soil disturbance in agroforestry systems. Furthermore, our results provide a useful demonstration of the potential use of the insect bait method to investigate pest‐suppressive potential through bait insect mortality, and we term this the “bait survival technique.”     

昆虫海藻糖酶的基因特性及功能研究进展

海藻糖酶(Treh)是昆虫能量代谢必不可少的一类酶, 亦是昆虫体内几丁质合成通路的第一个酶。其基因表达和酶活性直接与正常发育、 蜕皮、 变态以及繁殖等昆虫重要生理过程密切相关。目前已有多种昆虫的海藻糖酶基因被成功克隆, 从而发现昆虫海藻糖酶基因家族由多个成员组成。海藻糖酶基因所编码的蛋白大多数具有一个信号肽前导区, 部分蛋白拥有1~2个跨膜结构域, 根据是否具有跨膜结构, 可将其分为可溶性海藻糖酶(Treh1)和膜结合型海藻糖酶(Treh2)两类, 膜结合型海藻糖酶具有2个特有的标签序列, 即“PGGRFREFYYWDSY”和“QWDYPNAWPP”。海藻糖酶的主要功能是将胞外和胞内的海藻糖降解成葡萄糖, 为昆虫的生命活动提供能量。具体表现为两个方面, 一是参与昆虫几丁质合成途径, 从而调控表皮、 中肠等处的几丁质合成; 二是通过与激素的协同作用, 调控昆虫体内海藻糖和葡萄糖等糖类物质的浓度变化, 从而有效保护体内细胞的适应并渡过相应的逆境环境, 并提高其抗逆能力。鉴于海藻糖酶的重要功能, 其已成为害虫控制的潜在新靶标。不同类型海藻糖酶的功能研究及酶抑制剂的研发与应用将进一步推动害虫生物防治的发展。     

Sticky plant captures prey for symbiotic bug: is this digestive mutualism?

Many plants capture and kill insects but, until relatively recently, only carnivorous plants with digestive enzymes were known to gain directly from the nutrients of those insects. Recent studies show that some carnivorous plants lack digestive enzymes and have evolved digestive mutualisms with symbiotic insects that digest their prey for them. Rhododendron macrosepalum, a plant with sticky leaves that captures insects, has an association with symbiotic Mirid bugs that consume the insects captured. Here, we determine what the nature of the relationship is between Mirid and plant. We find that R. macrosepalum has no digestive enzymes of its own but that it does not seem to have the ability to absorb hemipteran faeces through its leaf cuticle. Naturally occurring levels of ¹⁵N and ¹⁴N were used to determine that R. macrosepalum gains no nitrogen through its association with the Mirid bugs and that it obtains all of its nitrogen from the soil. The Mirids, on the other hand, seem to obtain nitrogen from insects captured by the plant, as well as from plant tissues. The relationship between plant and Mirid is not a digestive mutualism but more likely an antagonistic relationship. This study adds to our understanding of how digestive mutualisms evolve and shows that insect capture alone, or in combination with a symbiotic insect relationship does not necessarily make a plant ‘carnivorous’.