Transcriptome wide identification of neuropeptides and G protein-coupled receptors (GPCRs) in Sunn pest, Eurygaster integriceps Puton.

Sunn pest (Eurygaster integriceps Puton) is major wheat pest causing economic damage. Neuropeptides and their receptors, G protein-coupled receptors (GPCRs), are involved in the regulation of insect physiology and behavior. Herein, a transcriptome-wide analysis was conducted in order to identify genes encoding neuropeptides, and putative GPCRs to gain insight into neuropeptide-modulated processes. De novo transcriptome assembly was undertaken using paired-end sequence reads derived from RNA samples collected from whole adults and yielded 582,398 contigs. In total, 46 neuropeptides have been identified, encompassing various known insect neuropeptide families. In addition, we discovered four previously uncharacterized neuroparsin peptides, which contributes to our understanding of the neuropeptide landscape. Furthermore, 85 putative neuropeptide GPCRs were identified, comprising three classes of GPCRs, A, B, C, and LGR, of which class C is not widely reported in insects. In addition, the identified GPCRs exhibited a remarkable 80% homology with the GPCRs found in the brown marmorated stink bug. It is noteworthy that these GPCRs displayed only a 20% homology to GPCRs from many other insect species. This information may be used to understand the neuropeptide-modulated physiology and behavior of Eurygaster integriceps, and to develop specific neuropeptide-based pest management strategies.

Abiotic and biotic factors influence diapause induction in sexual and asexual strains of Trichogramma brassicae (Hym: Trichogrammatidae).

The effects of some abiotic (maternal photoperiod and offspring developmental temperature) and biotic (host quality during both maternal and offspring generations) factors on diapause induction were investigated for two sympatric strains of Trichogramma brassicae Bezdenko (Hym: Trichogrammatidae) differing by infection status with regard to Wolbachia. The mode of reproduction, developmental temperature, maternal photoperiod and the quality of the host significantly affected diapause induction. The highest percentage of diapausing individuals were observed with the sexual strain reared at 14 °C using a "high quality" host and after a long photophase during the maternal generation. Environment-by-Environment, as well as Genotype-by-Environment interactions, was observed. All these patterns were discussed with the goal of providing relevant protocols for the commercial mass-rearing of such biocontrol agents. A successful diapause in beneficial insects could affect the efficiency of mass rearing by increasing the duration of storage conditions based on a high percent emergence and providing a large number of individuals at the appropriate time in the field season.

The effect of Wolbachia on diapause, fecundity, and clock gene expression in Trichogramma brassicae (Hymenoptera: Trichogrammatidae).

The short day lengths of late summer in moderate regions are used to induce diapause in various insects. Many studies have shown the maternal effect of photoperiod on diapause induction of Trichogramma wasps, but there is no study to show the relationship between photoperiodic regimes and clock genes in these useful biological control agents. Here, we investigated the role of photoperiods on diapause, fecundity, and clock gene expression (clk, cyc, cry2, per, and timeout) in asexual and sexual Trichogramma brassicae as a model insect to find any differences between two strains. Asexual strain was infected by Wolbachia, an endosymbiont bacterium. The diapause percentage was significantly higher under short days (8 h in sexual and 12 h in the asexual T. brassicae), although the diapause percentage of the sexual strain was significantly higher than the asexual one in all the photoperiods. The ANOVA revealed no significant changes between different photoperiods in the clock gene expression in the sexual strain but significant photoperiodic changes in clk, cyc, and timeout in the asexual strain. Our results showed that the mRNA levels of clock genes of asexual T. brassicae were significantly lower than those of sexual strain. The fecundity was significantly higher in the asexual strain. These results suggest that Wolbachia infection makes disturbance on the clock gene expression which consequently reduces the percentage of diapause but increases the fecundity in asexual T. brassicae.

Demographic traits of Tetranychus urticae (Acari: Tetranychidae) on leaf discs and whole leaves.

Life tables of twospotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), were studied on leaf discs and whole leaves of the common bean plant, Phaseolus vulgaris L. Data were analyzed based on the age-stage, two-sex life table theory. Durations of deutonymph stage, total preadult stage, and preoviposition period of mites reared on whole leaves (1.22, 7.6, and 0.29 d, respectively) were significantly shorter than those reared on leaf discs (1.73, 8.2, and 0.89 d, respectively). The lifetime fecundities were 22.81 and 12.05 offspring on whole leaves and leaf discs, respectively. Although hooked trichomes on the lower surface of the bean leaf contribute to adult mortality, spider mites successfully survive and reproduce on bean plants. The intrinsic rate of increase on whole leaves (0.235 d(-1)) was higher than that on leaf discs (0.159 d(-1). Higher intrinsic rate and fecundity found in mites reared on whole leaves may be due to the higher nutritional quality of whole leaves over leaf discs. We suggest that whole leaves should be used in future studies to better simulate realistic life history characteristics. The advantages of using age-stage, two-sex life tables over female age-specific life tables are discussed.

Digestive proteolytic activity in the Sunn pest, Eurygaster integriceps.

The Sunn pest, Eurygaster integriceps Puton (Heteroptera: Scutelleridae), is one of the most important pests of wheat and causes considerable damage to this valuable crop annually. Digestive proteinase activity of adult insects was investigated using general and specific substrates and inhibitors. Proteolytic activity was low when the common conventional substrates, azoalbumin, azocasein and hemoglobin were used to assay salivary glands and midguts. Using the fluorescent casein substrate (BODIPY FL casein), total proteolytic activity was measured at different pH. Maximum proteolytic activity was detected at pH 7 (100%) and 8(65%) which suggested the presence of serine proteinases in the salivary glands. There was no detectable proteolytic activity in midgut extracts. The inhibitors; PMSF (inhibitor of serine proteinases) and TPCK (a specific chymotrypsin inhibitor) showed greater than 50% inhibitory effect on total proteolytic activity, however, TLCK (specific trypsin inhibitor) and E-64(specific cysteine proteinase inhibitor) did not inhibit total proteolytic activity. Using fluorescent specific substrates for serine and cysteine proteinases (Z-Arg-AMC, Z-Arg-Arg-AMC, Z-Arg-Phe-AMC and Suc-Ala-Ala-Pro-Phe-AMZ) revealed the presence of tryptic and chymotryptic activity in the salivary gland extract. Zymogram analysis under non-reducing SDS-PAGE conditions and using the substrate APNE showed at least 8 tryptic and chymotryptic activity bands in salivary gland extracts. A single high molecular weight band with tryptic activity (165 kDa) was detected using the substrate BApNA in a zymogram analysis using native-PAGE. Kinetic studies showed a k(m) value of 0.6 mM for this enzyme against the substrate BApNA .The inhibitor TLCK decreased activity of the trypsin-like enzyme up to 73% and almost completely eliminated the only band related to this proteinase in the zymogram. Soybean Kunitz type trypsin inhibitor showed no effect on proteolytic activity of the trypsin-like serine proteinase. In general, the results revealed the presence of chymotrypsin- and trypsin-like serine proteinases in the salivary gland of E. integriceps, and it seems that the major total proteolytic activity is due to chymotrypsin proteinases.