Stick insect genomes reveal natural selection's role in parallel speciation.

Natural selection can drive the repeated evolution of reproductive isolation, but the genomic basis of parallel speciation remains poorly understood. We analyzed whole-genome divergence between replicate pairs of stick insect populations that are adapted to different host plants and undergoing parallel speciation. We found thousands of modest-sized genomic regions of accentuated divergence between populations, most of which are unique to individual population pairs. We also detected parallel genomic divergence across population pairs involving an excess of coding genes with specific molecular functions. Regions of parallel genomic divergence in nature exhibited exceptional allele frequency changes between hosts in a field transplant experiment. The results advance understanding of biological diversification by providing convergent observational and experimental evidence for selection's role in driving repeatable genomic divergence.

Binding of low concentration of peptide to H-2Kd produced in insect cells requires mouse beta 2-microglobulin co-expression.

A recombinant baculovirus expressing the murine class I MHC heavy chain H-2Kd cDNA under the transcriptional control of Autografa californica nuclear polyhedrosis virus (AcNPV) polyhedrin promoter has been isolated and used to infect Sf9 lepidopteran cells either alone or in association with a previously isolated virus expressing mouse beta 2-microglobulina (beta 2-ma). When infected with the heavy chain-encoding virus alone, H-2Kd was produced in a beta 2-m-free conformation detected on the surface of infected cells by conformation-independent antibodies. When Sf9 cells were co-infected with both viruses, approximately 10% of the heavy chain pool was engaged in the formation of native heterodimeric MHC class I molecules, which were glycosylated and transported to the cell surface as demonstrated by radio-binding experiments and flow cytometry. The assembly of the recombinant class I molecule was dependent on peptide, since heterodimer formation was brought about by H-2Kd-specific peptide ligands both in vivo, upon incubation with dually infected cells, and in vitro, in cell-free detergent extracts. In addition, a change in heavy chain conformation was brought about upon incubation with high concentrations (100 microM) of an H-2Kd-restricted octapeptide epitope from Plasmodium berghei. Furthermore, using low concentrations (3 nM) of a photoaffinity label derivative of this peptide, we show direct binding to cells co-expressing class I heavy chain and mouse beta 2-m but not to cells expressing free heavy chain only.

Relaxation of selective constraints causes independent selenoprotein extinction in insect genomes

BACKGROUND: Selenoproteins are a diverse family of proteins notable for the presence of the 21st amino acid, selenocysteine. Until very recently, all metazoan genomes investigated encoded selenoproteins, and these proteins had therefore been believed to be essential for animal life. Challenging this assumption, recent comparative analyses of insect genomes have revealed that some insect genomes appear to have lost selenoprotein genes. METHODOLOGY/PRINCIPAL FINDINGS: In this paper we investigate in detail the fate of selenoproteins, and that of selenoprotein factors, in all available arthropod genomes. We use a variety of in silico comparative genomics approaches to look for known selenoprotein genes and factors involved in selenoprotein biosynthesis. We have found that five insect species have completely lost the ability to encode selenoproteins and that selenoprotein loss in these species, although so far confined to the Endopterygota infraclass, cannot be attributed to a single evolutionary event, but rather to multiple, independent events. Loss of selenoproteins and selenoprotein factors is usually coupled to the deletion of the entire no-longer functional genomic region, rather than to sequence degradation and consequent pseudogenisation. Such dynamics of gene extinction are consistent with the high rate of genome rearrangements observed in Drosophila. We have also found that, while many selenoprotein factors are concomitantly lost with the selenoproteins, others are present and conserved in all investigated genomes, irrespective of whether they code for selenoproteins or not, suggesting that they are involved in additional, non-selenoprotein related functions. CONCLUSIONS/SIGNIFICANCE: Selenoproteins have been independently lost in several insect species, possibly as a consequence of the relaxation in insects of the selective constraints acting across metazoans to maintain selenoproteins. The dispensability of selenoproteins in insects may be related to the fundamental differences in antioxidant defense between these animals and other metazoans.

Domain shuffling in a sensor protein contributed to the evolution of insect pathogenicity in plant-beneficial Pseudomonas protegens.

Pseudomonas protegens is a biocontrol rhizobacterium with a plant-beneficial and an insect pathogenic lifestyle, but it is not understood how the organism switches between the two states. Here, we focus on understanding the function and possible evolution of a molecular sensor that enables P. protegens to detect the insect environment and produce a potent insecticidal toxin specifically during insect infection but not on roots. By using quantitative single cell microscopy and mutant analysis, we provide evidence that the sensor histidine kinase FitF is a key regulator of insecticidal toxin production. Our experimental data and bioinformatic analyses indicate that FitF shares a sensing domain with DctB, a histidine kinase regulating carbon uptake in Proteobacteria. This suggested that FitF has acquired its specificity through domain shuffling from a common ancestor. We constructed a chimeric DctB-FitF protein and showed that it is indeed functional in regulating toxin expression in P. protegens. The shuffling event and subsequent adaptive modifications of the recruited sensor domain were critical for the microorganism to express its potent insect toxin in the observed host-specific manner. Inhibition of the FitF sensor during root colonization could explain the mechanism by which P. protegens differentiates between the plant and insect host. Our study establishes FitF of P. protegens as a prime model for molecular evolution of sensor proteins and bacterial pathogenicity.

Born to be bee, fed to be worker? The caste system of a primitively eusocial insect.

ABSTRACT: INTRODUCTION: Primitively eusocial halictid bees are excellent systems to study the origin of eusociality, because all individuals have retained the ancestral ability to breed independently. In the sweat bee Halictus scabiosae, foundresses overwinter, establish nests and rear a first brood by mass-provisioning each offspring with pollen and nectar. The mothers may thus manipulate the phenotype of their offspring by restricting their food provisions. The first brood females generally help their mother to rear a second brood of males and gynes that become foundresses. However, the first brood females may also reproduce in their maternal or in other nests, or possibly enter early diapause. Here, we examined if the behavioural specialization of the first and second brood females was associated with between-brood differences in body size, energetic reserves and pollen provisions. RESULTS: The patterns of variation in adult body size, weight, fat content and food provisioned to the first and second brood indicate that H. scabiosae has dimorphic females. The first-brood females were significantly smaller, lighter and had lower fat reserves than the second-brood females and foundresses. The first-brood females were also less variable in size and fat content, and developed on homogeneously smaller pollen provisions. Foundresses were larger than gynes of the previous year, suggesting that small females were less likely to survive the winter. CONCLUSIONS: The marked size dimorphism between females produced in the first and second brood and the consistently smaller pollen provisions provided to the first brood suggest that the first brood females are channelled into a helper role during their pre-imaginal development. As a large body size is needed for successful hibernation, the mother may promote helping in her first brood offspring by restricting their food provisions. This pattern supports the hypothesis that parental manipulation may contribute to promote worker behaviour in primitively eusocial halictids.

Maternal effect on female caste determination in a social insect.

Caste differentiation and division of labor are the hallmarks of social insect colonies [1, 2]. The current dogma for female caste differentiation is that female eggs are totipotent, with morphological and physiological differences between queens and workers stemming from a developmental switch during the larval stage controlled by nutritional and other environmental factors (e.g., [3-8]). In this study, we tested whether maternal effects influence caste differentiation in Pogonomyrmex harvester ants. By conducting crossfostering experiments we identified two key factors in the process of caste determination. New queens were produced only from eggs laid by queens exposed to cold. Moreover, there was a strong age effect, with development into queens occurring only in eggs laid by queens that were at least two years old. Biochemical analyses further revealed that the level of ecdysteroids was significantly lower in eggs developing into queens than workers. By contrast, we found no significant effect of colony size or worker exposure to cold, suggesting that the trigger for caste differentiation may be independent of the quantity and quality of resources provided to larvae. Altogether these data demonstrate that the developmental fate of female brood is strongly influenced by maternal effects in ants of the genus Pogonomyrmex.

Purification and ligand binding of a soluble class I major histocompatibility complex molecule consisting of the first three domains of H-2Kd fused to beta 2-microglobulin expressed in the baculovirus-insect cell system.

A recombinant baculovirus encoding a single-chain murine major histocompatibility complex class I molecule in which the first three domains of H-2Kd are fused to beta 2-microglobulin (beta 2-m) via a 15-amino acid linker has been isolated and used to infect lepidopteran cells. A soluble, 391-amino acid single-chain H-2Kd (SC-Kd) molecule of 48 kDa was synthesized and glycosylated in insect cells and could be purified in the absence of detergents by affinity chromatography using the anti-H-2Kd monoclonal antibody SF1.1.1.1. We tested the ability of SC-Kd to bind antigenic peptides using a direct binding assay based on photoaffinity labeling. The photoreactive derivative was prepared from the H-2Kd-restricted Plasmodium berghei circumsporozoite protein (P.b. CS) peptide 253-260 (YIPSAEKI), a probe that we had previously shown to be unable to bind to the H-2Kd heavy chain in infected cells in the absence of co-expressed beta 2-microglobulin. SC-Kd expressed in insect cells did not require additional mouse beta 2-m to bind the photoprobe, indicating that the covalently attached beta 2-m could substitute for the free molecule. Similarly, binding of the P.b. CS photoaffinity probe to the purified SC-Kd molecule was unaffected by the addition of exogenous beta 2-m. This is in contrast to H-2KdQ10, a soluble H-2Kd molecule in which beta 2-m is noncovalently bound to the soluble heavy chain, whose ability to bind the photoaffinity probe is greatly enhanced in the presence of an excess of exogenous beta 2-m. The binding of the probe to SC-Kd was allele-specific, since labeling was selectively inhibited only by antigenic peptides known to be presented by the H-2Kd molecule.

Current status of the whitefly Aleurodicus dispersus as an invasive pest in the CapeVerte Islands

In the last three decades, the spiralling whitefly (Aleurodicus dispersus) has become an important international pest. The movement of plants and parts of plants (such as fruits) in international trade and tourism, and by natural dispersal, has favoured its introduction to new areas. In common with others whiteflies of economic importance, the immature and adult stages cause direct feeding damage by piercing and sucking of sap from foliage, and indirect damage following the accumulation all over host plants of honeydew and waxy flocculent material produced by the insects. Spiralling whitefly is a pest of tropical and subtropical crops, and highly polyphagous. Up to the 1970s, it had been recorded on 44 genera of plants, belonging to 26 botanical families (Mound & Halsey, 1978). This situation changed with the dispersal of the pest to new areas. Nowadays, the spiralling whitefly is one of the major pest of vegetable, ornamental and fruit crops around the globe (Lambkin, 1999). Important host crops include: banana (Musa sapientum), Citrus spp., coconut (Cocos nocifera), eggplant (Solanum melanogena), guava (Psidium guajava), Hibiscus rosa sinensis, Indian almond (Terminalia catappa), papya (Carica papaya), Rosa sp. and tomato (Lycopersicon esculentum) (Saminathan & Jayaraj, 2001). Spiralling whitefly has its origin in the tropical Americas, including Brazil. Although the pest has been recorded only once in Brasil, in the 1920s in the state of Bahia (Bondar, 1923), it now has official quarantine status because of its economic importance. In the Cape Verte Islands, on the West African coast, the pest was initially introduced in the first half of 2000; it has since become established, reaching urban, natural and agricultural areas of the islands that constitute the archipelago. Since then, the pest has been causing damage to many native plants, ornamentals and cultivated food crops (Anon., 2001; Monteiro, 2004). The present study was done in order to produce an inventory of the most common host plants of spiralling whitefly in this new habitat.

Comparação dos sistemas de cultivo nativo e adensado de erva mate, Ilex paraguariensis St. Hil., quanto à ocorrência e flutuação populacional de insetos

This research was carried out in order to compare the occurrence of insects in two maté cultivation systems, native and high tree density. It was performed from August/2000 to September/2001, in a private property in São Mateus do Sul county, in Paraná State, Brazil. Visual inspections of trees and light traps were used to evaluate insect populations in both areas. For Hedypathes betulinus (Klug) (Coleoptera, Cerambycidae), only six adults were observed in the dense area. Based on presence of sawdust at the basis of the trunk, it was obtained that the number of attacked trees did not surpass 11% in either area. For Gyropsylla spegazziniana (Lizer y Trelles) (Hemiptera, Psyllidae), the number of galls per tree was counted and it was observed that the population peak occurred from November to January. For Hylesia spp. (Lepidoptera, Saturniidae) and Thelosia camina Schaus (Lepidoptera, Eupterotidae), the presence of caterpillars on the trees was noticed from September to February, with the population peak in November and December. Adults of Hylesia spp. were more numerous in February and March. Two species that were not previously recorded for Brazil on maté were identified: Hylesia paulex Dognin (83%) and Hylesia remex Dyer (17%), collected with light traps. The maté caterpillar, T. camina was not collected with these traps. Nymphs and adults of Ceroplastes grandis Hempel (Hemiptera, Coccidae) were observed along the year on the branches, with population peak between April and June for the nymphs and from September to November for the adults. It should be considered that despite higher insect incidence in the dense area compared to the native area, the first presents higher yield, and that with a good pest management program the insect problems can be minimized.

Aspectos biológicos de Zabrotes subfasciatus (Bohemann, 1833) (Coleoptera, Bruchidae) em Phaseolus vulgaris L., cv. Carioca (Fabaceae), sob condições de laboratório

Zabrotes subfasciatus is a serious pest of common beans, P. vulgaris L.. In Brazil there are several studies dealing with resistance of bean genotypes to this insect, while other studies have emphasized the utilization of oils and powders from plants to repel their attack. In this paper, fecundity, fertility, pattern of oviposition, life cycle and longevity were evaluated for a Brazilian stock from the Goiás State on P. vulgaris cv. Carioca, at 30ºC and 70% R.H. The mean fecundity was 38 eggs per female and 73% of viability. Egg laying showed an aggregated pattern. Males and females lived an average of 13 and 9 days, respectively. The total life cycle lasted for about 28 days.

Development of a control alternative for the citrus fruit borer, Ecdytolopha aurantiana (Lepidoptera, Tortricidae): from basic research to the grower

All research steps, developed from 1995 to 2000, to synthesize the sex pheromone of Ecdytolopha aurantiana (Lima, 1927) are described, in order to monitoring this pest that causes losses in the order of 50 million dollars per year to citriculture in the State of São Paulo. The basic researches conducted are described, including the development of an artificial diet for the insect, the study of its temperature and humidity requirements, behavioral studies, and synthesis of the male-attracting substance up to the formulation and distribution of the pheromone to the grower, by means of its commercialization. It is a case of success, at a cost of 50 thousand dollars, involving inter- and multidisciplinary researches, which can be adopted to other insect pests in the country.

Ants and their effects on an insect herbivore community associated with the inflorescences of Byrsonima crassifolia (Linnaeus) H.B.K. (Malpighiaceae)

The effects of ants on the insect community on inflorescences of Byrsonima crassifolia (Malpighiaceae) were tested in an ant exclusion experiment in a cerrado vegetation in southeastern Brazil. Forty-four species of insects (23 families) and nine species of ants (6 genera and 3 subfamilies) were found on the inflorescences of B. crassifolia. The exclusion of ants, primarily Camponotus sericeiventris and Camponotus spp., reduced the treehopper population to 20% of the original abundance. Ant exclusion and time influenced the abundance of chewing (Exclusion, P<0.001; Time, P<0.002), and sucking insects (Exclusion, P<0.02; Time, P<0.01). Twice as many chewing and sucking insects were found on ant-excluded inflorescences as compared to control inflorescences (P<0.001). One and half more sucking insects were found on ant-excluded than on control inflorescences. Only time significantly influenced the richness of chewing and sucking insects associated with B. crassifolia inflorescences. Inflorescences on control branches were significantly less attacked by herbivores than inflorescences on ant-excluded branches (P<0.001). Therefore, these results suggest that the presence of ants alters the structure of insect herbivore community associated with B. crassifolia.

Nature, evolution and characterisation of rhizospheric chemical exudates affecting root herbivores

Similar to aboveground herbivores, root-feeding insects must locate and identify suitable resources. In the darkness of soil, they mainly rely on root chemical exudations and, therefore, have evolved specific behaviours. Because of their impact on crop yield, most of our knowledge in belowground chemical ecology is biased towards soil-dwelling insect pests. Yet the increasing literature on volatile-mediated interactions in the ground underpins the great importance of chemical signalling in this ecosystem and its potential in pest control. Here, we explore the ecology and physiology of these chemically based interactions. An evolutionary approach reveals interesting patterns in the response of insects to particular classes of volatile or water-soluble organic compounds commonly emitted by roots. Food web analyses reasonably support that volatiles are used as long-range cues whereas water-soluble molecules serve in host acceptance/rejection by the insect; however, data are still scarce. As a case study, the chemical ecology of Diabrotica virgifera virgifera is discussed and applications of belowground signalling in pest management are examined. Soil chemical ecology is an expanding field of research and will certainly be a hub of our understanding of soil communities and subsequently of the management of belowground ecosystem services.

Combination of diatomaceous earth and powder deltamethrin for insect control in stored corn

The use of diatomaceous earth (DE) is a very efficient insect control measure in stored grain IPM due to its low cost, easy application, reduction of active ingredient residues, lower environmental contamination and operator safety. The objective of this research was to evaluate the efficacy of different dosages of DE mixed with powder deltamethrin for controlling Sitophilus zeamais in stored corn. Samples of 100 g of clean and dry corn, in three replicates, were submitted to the following treatments: DE (Keepdry®), at the dosages of 500, 750 and 1000 g/t; powder deltamethrin (K-Obiol®) at 0,5 g a.i. /t and 1,0 g a.i. /t; and combinations of the lowest and highest DE dosages with the two dosages of deltamethrin. Thirty adults of S. zeamais were placed in each vial with the treated grains and kept in environment chambers at 25ºC. Mortality was evaluated from the 1st to the 28th day. In the treatments mixing DE with deltamethrin or deltamethrin alone, the mortalyti was registered since the first day. In the treatments using only DE, the first dead insects were recorded after the 3rd day, especially in the highest dosages. After the 7th day, however, there was no statistical difference among all treatments, except for the lowest dosage of DE which reached a satisfactory control level only by the 14th day. It was concluded that treatments using DE combined with low dosages of powder deltamethrin represent an efficient control measure against S. zeamais in stored corn because insect mortality is faster than in treatments using DE alone and residues of active ingredients are much lower than using the insecticide in high dosages.

Regulation of the expression of Fit insect toxin locus genes in the root-associated biocontrol pseudomonad CHA0

The root-colonizing Pseudomonas fluorescens strain CHA0 is a biocontrol agent of soil-borne plant diseases caused by fungal and oomycete pathogens. Remarkably, this plant-beneficial pseudomonad is also endowed with potent insecticidal activity that depends on the production of a large protein toxin termed Fit (for P. fluorescens insecticidal toxin). In our present work, the genomic locus encoding the P. fluorescens insect toxin is subjected to a detailed molecular analysis. The Fit toxin gene fitD is flanked upstream by the fitABC genes and downstream by the fitE gene that encode the ABC transporter, membrane fusion, and outer membrane efflux components of a type I protein secretion system predicted to function in toxin export. The fitF, fitG, and fitH genes located downstream of fitE code for regulatory proteins having domain structures typical of signal transduction histidine kinases, LysR-type transcriptional regulators, and response regulators, respectively. The role of these insect toxin locus-associated control elements is being investigated with mutants defective for the regulatory genes and with GFP-based reporter fusions to putative promoter regions upstream of the transporter genes fitA and fitE, the toxin gene fitD, and the regulatory genes fitF and fitH. Our preliminary findings suggest that the three regulators interact with known global regulators of biocontrol factor expression to control Fit toxin expression and secretion.