Detection of the bacterium, Xylella fastidiosa, in saliva of glassy-winged sharpshooter, Homalodisca vitripennis

Homalodisca vitripennis ( Germar) ( Hemiptera: Cicadellidae), the glassy- winged sharpshooter, is one of the most important vectors of the bacterium, Xylella fastidiosa subsp. piercei ( Xanthomonadales: Xanthomonadaceae) that causes Pierce's Disease in grapevines in California. In the present study we report a new method for studying pathogen transmission or probing behavior of H. vitripennis. When confined, H. vitripennis attempt to probe the surface of sterile containers 48 hours post- acquisition of X. f. piercei. The saliva deposited during attempted feeding probes was found to contain X. f. piercei. We observed no correlation between X. f. piercei titers in the foregut of H. vitripennis that fed on Xylella- infected grapevines and the presence of this bacterium in the deposited saliva. The infection rate after a 48 h post- acquisition feeding on healthy citrus and grapevines was observed to be 77% for H. vitripennis that fed on grapevines and 81% for H. vitripennis that fed on citrus, with no difference in the number of positive probing sites from H. vitripennis that fed on either grapevine or citrus. This method is amenable for individual assessment of X. f. piercei- infectivity, with samples less likely to be affected by tissue contamination that is usually present in whole body extracts.

Organization, evolution and transcriptional profile of hexamerin genes of the parasitic wasp Nasonia vitripennis (Hymenoptera: Pteromalidae)

Hexamerins and prophenoloxidases (PPOs) proteins are members of the arthropod-haemocyanin superfamily. In contrast to haemocyanin and PPO, hexamerins do not bind oxygen, but mainly play a role as storage proteins that supply amino acids for insect metamorphosis. We identified seven genes encoding hexamerins, three encoding PPOs, and one hexamerin pseudogene in the genome of the parasitoid wasp Nasonia vitripennis. A phylogenetic analysis of hexamerins and PPOs from this wasp and related proteins from other insect orders suggests an essentially order-specific radiation of hexamerins. Temporal and spatial transcriptional profiles of N. vitripennis hexamerins suggest that they have physiological functions other than metamorphosis, which are arguably coupled with its lifestyle.

Influência do aumento do número de pupas hospedeiras de Cochliomyia macellaria (Diptera, Calliphoridae) no desenvolvimento do parasitóide Nasonia vitripennis (Hymenoptera, Pteromalidae) em laboratório

O desenvolvimento pós-embrionário, a produtividade, a taxa de parasitismo e a razão sexual de Nasonia vitripennis (Walker, 1836) criada em pupa de Cochliomyia macellaria (Fabricius, 1775) foram estudados. Densidades diferentes de hospedeiro foram usadas (proporções 1:1, 1:2, 1:3, 1:4 e 1:5) com tempo de exposição de 72 horas, em cada uma delas. Fêmeas nulíparas originárias da colônia estoque foram individualizadas em tubos de teste cobertos com algodão hidrófobo e contendo as pupas hospedeiras. Cada tratamento constituiu-se de 10 repetições. Depois da exposição, as vespas foram descartadas e as pupas hospedeiras foram individualizadas em tubos de teste até a emergência dos adultos de C. macellaria ou N. vitripennis. Amostras das pupas hospedeiras não expostas ao parasitismo e recebendo o mesmo tratamento experimental foram usadas como controle. Os parasitóides mostraram um desenvolvimento mais lento em relação ao aumento da densidade de hospedeiros. Houve uma diminuição na produção de parasitóides por hospedeiro em densidades elevadas. A razão sexual tendeu para um desvio para nascimento de fêmeas com o aumento da densidade do hospedeiro. Os índices de parasitismo mostraram uma diminuição quando o parasitóide foi exposto a mais de duas pupas hospedeiras.

Exposure of a single host (Chrysomya megacephala) (Calliphoridae) to different quantities of female parasitoids (Nasonia vitripennis) (Pteromalidae)

The aim of this study was to verify the duration of the development period, number of parasitoids produced per pupa, parasitism rate and sex ratio of Nasonia vitripennis (Hymenoptera, Pteromalidae), when they were exposed to a single host: Chrysomya megacephala (Diptera, Calliphoridae). One pupa was exposed in glass tubes to different numbers of female parasitoids (1, 3, 5, 7, 9 and 11) during 48 h. Twenty replications/treatment were used, under controlled conditions (T= 27 °C day/ 25 °C night, 60 ± 10% RH). Statistical analysis of the data was made using the ANOVA test and the "a posteriori" comparisons were made using the Tukey-HSD test (both tests with a significance level of 5%). The duration of the development period was longer in treatments where a higher density of females per host was used. When five females per host were used, the mean number of parasitoids that emerged per pupa was higher. The data showed a tendency to a decrease in the amount of parasitoids emerged per host, especially of female, when used high quantities of female per host. Higher parasitism rates were observed in the 3:1 and 5:1 treatments and an increase in the percentage of unviable pupae was observed, probably due to an increase of female densities in the treatments, possibly a consequence of superparasitism.

Desempenho do parasitóide Nasonia vitripennis (Walker)(Hymenoptera, Pteromalidae) utilizando como hospedeiro Cochliomyia macellaria (Fabricius) (Diptera, Calliphoridae), sob diferentes tempos de exposição

Desempenho do parasitóide Nasonia vitripennis (Walker) (Hymenoptera, Pteromalidae) utilizando como hospedeiro Cochliomyia macellaria (Fabricius) (Diptera, Calliphoridae), sob diferentes tempos de exposição. Foram estudados o desenvolvimento pós-embrionário, a produtividade da pupa hospedeira, a taxa de parasitismo e a razão sexual de Nasonia vitripennis (Walker) criadas em pupas hospedeiras de Cochliomyia macellaria (Fabricius). Analisaram-se diferentes tempos de exposição: 24, 48, 72 e 96 horas. Fêmeas nulíparas foram individualizadas em tubos de ensaio contendo as pupas hospedeiras e tampados com algodão hidrófobo. Cada tratamento foi constituído de 10 repetições. Após a exposição, os himenópteros foram descartados e as pupas hospedeiras foram mantidas individualizadas. O controle consistiu de pupas de C. macellaria não expostas ao parasitismo utilizando-se o mesmo delineamento experimental. Os parasitóides apresentaram um desenvolvimento pós-embrionário mais lento com aumento do tempo de exposição. A exposição mais produtiva foi a de 72 horas. A razão sexual apresentou uma tendência ao nascimento de fêmeas após a exposição de 48 horas. A taxa de parasitismo apresentou crescimento até a exposição de 72 horas. A taxa de nascimento de dípteros também apresentou queda até o tempo de exposição de 72 horas.

Unique features of odorant-binding proteins of the parasitoid wasp Nasonia vitripennis revealed by genome annotation and comparative analyses

Insects are the most diverse group of animals on the planet, comprising over 90% of all metazoan life forms, and have adapted to a wide diversity of ecosystems in nearly all environments. They have evolved highly sensitive chemical senses that are central to their interaction with their environment and to communication between individuals. Understanding the molecular bases of insect olfaction is therefore of great importance from both a basic and applied perspective. Odorant binding proteins (OBPs) are some of most abundant proteins found in insect olfactory organs, where they are the first component of the olfactory transduction cascade, carrying odorant molecules to the olfactory receptors. We carried out a search for OBPs in the genome of the parasitoid wasp Nasonia vitripennis and identified 90 sequences encoding putative OBPs. This is the largest OBP family so far reported in insects. We report unique features of the N. vitripennis OBPs, including the presence and evolutionary origin of a new subfamily of double-domain OBPs (consisting of two concatenated OBP domains), the loss of conserved cysteine residues and the expression of pseudogenes. This study also demonstrates the extremely dynamic evolution of the insect OBP family: (i) the number of different OBPs can vary greatly between species; (ii) the sequences are highly diverse, sometimes as a result of positive selection pressure with even the canonical cysteines being lost; (iii) new lineage specific domain arrangements can arise, such as the double domain OBP subfamily of wasps and mosquitoes.

Suppression subtractive hybridization analysis reveals expression of conserved and novel genes in male accessory glands of the ant Leptothorax gredleri

Background: During mating, insect males eject accessory gland proteins (Acps) into the female genital tract. These substances are known to affect female post-mating behavior and physiology. In addition, they may harm the female, e. g., in reducing its lifespan. This is interpreted as a consequence of sexual antagonistic co-evolution. Whereas sexual conflict abounds in non-social species, the peculiar life history of social insects (ants, bees, wasps) with lifelong pair-bonding and no re-mating aligns the reproductive interests of the sexes. Harming the female during mating would negatively affect male fitness and sexual antagonism is therefore not expected. Indeed, mating appears to increase female longevity in at least one ant species. Acps are presumed to play a role in this phenomenon, but the underlying mechanisms are unknown. In this study, we investigated genes, which are preferentially expressed in male accessory glands of the ant Leptothorax gredleri, to determine which proteins might be transferred in the seminal fluid. Results: By a suppression subtractive hybridization protocol we obtained 20 unique sequences (USs). Twelve had mutual best matches with genes predicted for Apis mellifera and Nasonia vitripennis. Functional information (Gene Ontology) was available only for seven of these, including intracellular signaling, energy-dependent transport and metabolic enzyme activities. The remaining eight USs did not match sequences from other species. Six genes were further analyzed by quantitative RT-PCR in three life cycle stages of male ants. A gene with carboxy-lyase activity and one of unpredicted function were significantly overexpressed in accessory glands of sexually mature males. Conclusions: Our study is the first one to investigate differential gene expression in ants in a context related to mating. Our findings indicate that male accessory glands of L. gredleri express a series of genes that are unique to this species, possibly representing novel genes, in addition to conserved ones for which functions can be predicted. Identifying differentially expressed genes might help to better understand molecular mechanisms involved in reproductive processes in eusocial Hymenoptera. While the novel genes could account for rapidly evolving ones driven by intra-sexual conflict between males, conserved genes imply that rather beneficial traits might get fixed by a process described as inter-sexual cooperation between males and females.

Functional and Evolutionary Insights from the Genomes of Three Parasitoid Nasonia Species

We report here genome sequences and comparative analyses of three closely related parasitoid wasps: Nasonia vitripennis, N. giraulti, and N. longicornis. Parasitoids are important regulators of arthropod populations, including major agricultural pests and disease vectors, and Nasonia is an emerging genetic model, particularly for evolutionary and developmental genetics. Key findings include the identification of a functional DNA methylation tool kit; hymenopteran-specific genes including diverse venoms; lateral gene transfers among Pox viruses, Wolbachia, and Nasonia; and the rapid evolution of genes involved in nuclear-mitochondrial interactions that are implicated in speciation. Newly developed genome resources advance Nasonia for genetic research, accelerate mapping and cloning of quantitative trait loci, and will ultimately provide tools and knowledge for further increasing the utility of parasitoids as pest insect-control agents.

Microhimenópteros parasitóides de Chrysomya megacephala

O objetivo do estudo foi levantar os principais parasitóides de Chrysomya megacephala, na cidade do Rio de Janeiro, RJ, devido a importância dessa mosca como vetor de patógenos para o ambiente urbano. As coletas foram realizadas semanalmente, no período de agosto de 1999 a julho de 2000, por meio da exposição de larvas e pupas da mosca em carne putrefata. Foram identificadas três espécies de microhimenópteros no local: Tachinaephagus zealandicus (Encyrtidae), Pachycrepoideus vindemiae (Pteromalidae) e Nasonia vitripennis (Pteromalidae), cujos testes posteriores poderão mostrar seu potencial para utilização em futuros programas de controle.

Parasitóides de Chrysomya megacephala (Fabricius) coletados em Itumbiara, Goiás

Foram determinadas as espécies de parasitóides associadas com Chrysomya megacephala, coletados com isca de rins de bovino, em Itumbiara, Estado de Goiás. As pupas foram obtidas pelo método de flutuação, individualizadas em cápsulas de gelatina e mantidas até a emergência das moscas e/ou dos parasitóides. A prevalência total de parasitismo foi de 18,6%. Brachymeria podagrica, Nasonia vitripennis e Pachycrepoideus vindemiae apresentaram uma freqüência de 8,6%, 8,6% e 1,4%, respectivamente. Relata-se a primeira ocorrência de Brachymeria podagrica em pupas de Chrysomya megacephala.

Draft genome of the globally widespread and invasive Argentine ant (Linepithema humile).

Ants are some of the most abundant and familiar animals on Earth, and they play vital roles in most terrestrial ecosystems. Although all ants are eusocial, and display a variety of complex and fascinating behaviors, few genomic resources exist for them. Here, we report the draft genome sequence of a particularly widespread and well-studied species, the invasive Argentine ant (Linepithema humile), which was accomplished using a combination of 454 (Roche) and Illumina sequencing and community-based funding rather than federal grant support. Manual annotation of >1,000 genes from a variety of different gene families and functional classes reveals unique features of the Argentine ant's biology, as well as similarities to Apis mellifera and Nasonia vitripennis. Distinctive features of the Argentine ant genome include remarkable expansions of gustatory (116 genes) and odorant receptors (367 genes), an abundance of cytochrome P450 genes (>110), lineage-specific expansions of yellow/major royal jelly proteins and desaturases, and complete CpG DNA methylation and RNAi toolkits. The Argentine ant genome contains fewer immune genes than Drosophila and Tribolium, which may reflect the prominent role played by behavioral and chemical suppression of pathogens. Analysis of the ratio of observed to expected CpG nucleotides for genes in the reproductive development and apoptosis pathways suggests higher levels of methylation than in the genome overall. The resources provided by this genome sequence will offer an abundance of tools for researchers seeking to illuminate the fascinating biology of this emerging model organism.

Draft genome of the red harvester ant Pogonomyrmex barbatus.

We report the draft genome sequence of the red harvester ant, Pogonomyrmex barbatus. The genome was sequenced using 454 pyrosequencing, and the current assembly and annotation were completed in less than 1 y. Analyses of conserved gene groups (more than 1,200 manually annotated genes to date) suggest a high-quality assembly and annotation comparable to recently sequenced insect genomes using Sanger sequencing. The red harvester ant is a model for studying reproductive division of labor, phenotypic plasticity, and sociogenomics. Although the genome of P. barbatus is similar to other sequenced hymenopterans (Apis mellifera and Nasonia vitripennis) in GC content and compositional organization, and possesses a complete CpG methylation toolkit, its predicted genomic CpG content differs markedly from the other hymenopterans. Gene networks involved in generating key differences between the queen and worker castes (e.g., wings and ovaries) show signatures of increased methylation and suggest that ants and bees may have independently co-opted the same gene regulatory mechanisms for reproductive division of labor. Gene family expansions (e.g., 344 functional odorant receptors) and pseudogene accumulation in chemoreception and P450 genes compared with A. mellifera and N. vitripennis are consistent with major life-history changes during the adaptive radiation of Pogonomyrmex spp., perhaps in parallel with the development of the North American deserts.

Dinâmica populacional e parasitismo de Himenópteros parasitóides de Chrysomya megacephala (Fabricius) (Diptera, Calliphoridae), no Rio de Janeiro, RJ

Este estudo foi realizado no período de agosto de 1999 a julho de 2000 com o objetivo de conhecer os parasitóides de Chrysomya megacephala (Fabricius, 1794) (Diptera, Calliphoridae) e avaliar sua dinâmica populacional no Instituto Oswaldo Cruz (IOC/ FIOCRUZ) e Jardim Zoológico, na cidade do Rio de Janeiro, RJ. As coletas foram realizadas semanalmente através da exposição de larvas de terceiro instar da mosca e seu substrato de criação, carne bovina moída em putrefação. Foram identificadas três espécies de himenópteros parasitóides: Tachinaephagus zealandicus Ashmead, 1904 (Encyrtidae), Pachycrepoideus vindemiae (Rondani, 1875) (Pteromalidae) and Nasonia vitripennis (Walker, 1836) (Pteromalidae). Nos dois locais, T. zealandicus foi a espécie com maior taxa de parasitismo de C. megacephala, seguida por P. vindemiae e N. vitripennis. A população de parasitóides diminuiu drasticamente no verão (temperatura média = 28°C; precipitação = 6,5 mm).O pico populacional desses insetos foi verificado no final do outono e durante todo o inverno. No Jardim Zoológico, o lixo acumulado no local de coleta exerceu forte influência na dinâmica populacional dos himenópteros parasitóides identificados nesse estudo e o pico populacional de parasitismo foi verificado em junho e julho de 2000.

Ocorrência de multiparasitismo em larvas de terceiro ínstar e pupas de Chrysomya megacephala (Fabricius) em condições de campo

Durante experimento de pesquisa envolvendo o parasitismo de larvas de terceiro ínstar e pupas de Chrysomya megacephala (Fabricius, 1794) em dois locais da cidade do Rio de Janeiro, RJ, observou-se a ocorrência de multiparasitismo em 1,83% das pupas expostas no Instituto Oswaldo Cruz (IOC) e em 2,16% expostas no Jardim Zoológico (ZOO). O experimento foi conduzido semanalmente de agosto de 1999 a julho de 2000. Em ambos os locais, houve a co-ocorrência de duas espécies por pupa e os parasitóides encontrados foram os himenópteros Tachinaephagus zealandicus Ashmead, 1904, Pachycrepoideus vindemiae (Rondani, 1875) e Nasonia vitripennis (Walker, 1836). 72,73% do multiparasitismo ocorreu no ZOO em julho de 2000. Em condições de multiparasitismo, T. zealandicus e P. vindemiae mantiveram seu hábito gregário e solitário, respectivamente, mas N. vitripennis foi encontrado solitário em algumas pupas.