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The bean bug Riptortus pedestris is a notorious insect pest that can damage various crops, especially soybean, in East Asia. In insects, the olfactory system plays a crucial role in host finding and feeding behaviour in which the odorant-binding proteins (OBPs) are believed to be involved in initial step in this system. In this study, we produced the R. pedestris adult antennae-expressed RpedOBP4 protein using a recombinant expression system in E. coli. Fluorescence competitive binding confirmed that RpedOBP4 has binding affinities to 7 of 20 soybean volatiles (ligands), and that a neutral condition is the best environment for it. The binding property of RpedOBP4 to these ligands was further revealed by integrating data from molecular docking, site-directed mutagenesis and ligand binding assays. This demonstrated that five amino acid residues (I30, L33, Y47, I57 and Y121) are involved in the binding process of RpedOBP4 to corresponding ligands. These findings will not only help us to more thoroughly explore the olfactory mechanism of R. pedestris during feeding on soybean, but also lead to the identification of key candidate targets for developing environmental and efficient behaviour inhibitors to prevent population expansion of R. pedestris in the future.

BACKGROUND Carboxyl/cholinesterases (CCEs) are thought to play a pivotal role in the degradation of sex pheromones and plant-derived odorants in insects, but their exact biochemistry and physiological functions remain unclear. RESULTS In this study, two paralogous antennae-enriched CCEs from Plutella xylostella (PxylCCE16a and 16c) were identified and functionally characterized. High-purity protein preparations of active recombinant PxylCCE16a and 16c have been obtained from Sf9 insect cells by Ni2+ affinity purification. Our results revealed that the purified recombinant PxylCCE016c is able to degrade two sex pheromone components Z9-14:Ac and Z11-16:Ac at 27.64 ± 0.79% and 24.40 ± 3.07%, respectively, while PxylCCE016a presented relatively lower activity. Additionally, a similar difference in activity was measured in plant-derived odorants. Furthermore, both CCEs displayed obvious preferences for the two sex pheromone components, especially on Z11-16:Ac (Km values are in the range 7.82-45.06 μmol L-1 ) which much lower than plant odorants (Km values are in the range 1290-4030 μmol L-1 ). Furthermore, the activity of the two newly identified CCEs is pH-dependent. The activity at pH 6.5 is obviously higher than that at pH 5.0. Interestingly, only PxylCCE016c can be inhibited by a common esterase inhibitor triphenyl phosphate (TPP) with LC50 of 1570 ± 520 μmol L-1 . CONCLUSION PxylCCE16c plays a more essential role in odorant degradation than PxylCCE16a. Moreover, the current study provides novel potential pesticide targets for the notorious moth Plutella xylostella. © 2021 Society of Chemical Industry.

The German cockroach Blattella germanica is an important urban insect pest worldwide. In many insects, chemosensation is essential for guiding their behaviors for survival. Although a large number of chemosensory-related genes have been identified in B. germanica, little information on tissue-specific and developmental expression patterns has not been uncovered yet. In this study, we performed transcriptome analysis of different B. germanica tissues to reveal novel chemosensory proteins (CSPs) and sensory neuron membrane proteins (SNMPs). In addition, a phylogenetic tree and gender-specific expression of multiple chemosensory gene families have been analyzed. We identified three CSPs genes (BgerCSP11, BgerCSP12, and BgerCSP13) and five SNMP genes in B. germanica. Tissue-specific expression profiling showed that CSP1, 8, and 9 exhibited significant expression levels in both adult and 5th instar nymph antennae. The results have paved the way for further functional study of the chemosensory mechanism in B. germanica and provided potential insecticide targets.

Background Athetis lepigone, a noctuid moth feeding on more than 30 different crops worldwide, has evolved a sophisticated, sensitive, and specific chemosensory system to detect and discriminate exogenous chemicals. Odorant-binding proteins (OBPs) are the most important agent in insect chemosensory systems to be explored as an alternative target for environmentally friendly approaches to pest management. Results To investigate the olfactory function of A. lepigone OBPs (AlepOBPs), AlepOBP6 was identified and expressed in Escherichia coli. The binding affinity of the recombinant OBP to 20 different ligands was then examined using a competitive binding approach. The results revealed that AlepOBP6 can bind to two sex pheromones and ten maize volatiles, and its conformation stability is pH dependent. We also carried out a structure-function study using different molecular approaches, including structure modeling, molecular docking, and a mutation functional assay to identify amino acid residues (M39, V68, W106, Q107, and Y114) involved in the binding of AlepOBP6 to both sex pheromones and maize volatiles in A. lepigone. Conclusion These results suggest that AlepOBP6 is likely involved in mediating the responses of A. lepigone to sex pheromones and maize volatiles, which may play a pivotal function in mating, feeding, and oviposition behaviors. This study not only provides new insight into the binding mechanism of OBPs to sex pheromones and host volatiles in moths, but also contributes to the discovery of novel target candidates for developing efficient behavior disruptors to control A. lepigone in the future.

The tobacco cutworm Spodoptera litura (Lepidoptera: Noctuidae) is a polyphagous pest with a highly selective and sensitive chemosensory system involved in complex physiological behaviors such as searching for food sources, feeding, courtship, and oviposition. However, effective management strategies for controlling the insect pest populations under threshold levels are lacking. Therefore, there is an urgent need to formulate eco-friendly pest control strategies based on the disruption of the insect chemosensory system. In this study, we identified 158 putative chemosensory genes based on transcriptomic and genomic data for S. litura, including 45 odorant-binding proteins (OBPs, nine were new), 23 chemosensory proteins (CSPs), 60 odorant receptors (ORs, three were new), and 30 gustatory receptors (GRs, three were new), a number higher than those reported by previous transcriptome studies. Subsequently, we constructed phylogenetic trees based on these genes in moths and analyzed the dynamic expression of various genes in head capsules across larval instars using quantitative real-time polymerase chain reaction. Nine genes-SlitOBP8, SlitOBP9, SlitOBP25, SlitCSP1, SlitCSP7, SlitCSP18, SlitOR34, SlitGR240, and SlitGR242-were highly expressed in the heads of 3- to 5-day-old S. litura larvae. The genes differentially expressed in olfactory organs during larval development might play crucial roles in the chemosensory system of S. litura larvae. Our findings substantially expand the gene inventory for S. litura and present potential target genes for further studies on larval feeding in S. litura.

Background The German cockroach, Blattella germanica, is one of the most severe pests of urban and rural areas. High-throughput genetic screening approaches indicate that the olfactory system of this pest is extremely powerful because it has an extensive array of olfactory receptor genes compared with many other insect species. Several of these genes have been identified previously, but their functions have not yet been characterized. Results This study describes the sequence of five transcriptomes of B. germanica adult male antennae, female antennae, maxillary palps, legs, and fifth-instar nymph antennae to investigate expression patterns of odorant receptors (ORs). Approximately 90% of ORs were found to be the most highly expressed genes in adult or nymph antennae. Additionally, every OR requires an odorant co-receptor (Orco) to become fully functional, and this was selected and successfully inhibited by injection of the corresponding double-stranded (ds)RNA targeting the Orco. A strong RNA interference (RNAi) effect was observed in which > 75% of Orco messenger RNA (mRNA) was clearly suppressed after 72 h of treatment. Olfactory behavioral assays showed that Orco-impaired B. germanica respond more slowly and show less attraction to one volatile sex pheromone and food resources compared with a control group. Conclusion The results show that Orco plays a pivotal role in both sex pheromone and food-seeking olfactory processes, and provide an alternative genetic technique for controlling this urban pest species by olfactory disruption. © 2020 Society of Chemical Industry.

The German cockroach

Athetis lepigone is one of the most severe polyphagous pests, and it has developed resistance to different chemical insecticides. Insects primarily rely on the olfactory system to recognize various environmental chemicals, including xenobiotics such as insecticides. Here, we expressed two A. lepigone pheromone-binding proteins (AlepPBP2 and AlepPBP3), and observed they had higher binding affinities to phoxim than other insecticides, with Ki was 3.30 ± 0.38 μM and 3.27 ± 0.10 μM, respectively. Molecular dynamics simulation, binding mode analysis, and computational alanine scanning showed that six residues (Phe15, Phe39, Ile55, Leu65, Ile97, and Phe122) of AlepPBP2 and three residues (Phe12, Ile52, and Ile134) of AlepPBP3 maybe as potential residues that can change protein ability to bind an organophosphorus insecticide phoxim. Then, we used site-directed mutagenesis assay to mutate these residues into alanine, respectively. Subsequently, the binding assays displayed that Phe15, Phe39, and Ile97 of AlepPBP2, Phe12 and Ile134 of AlepPBP3 caused a significant decrease of AlepPBPs binding ability to phoxim, suggesting they should play crucial roles in the AlepPBPs/phoxim interactions. Our findings could further advance in using PBPs as unique targets to design and develop precise and environmentally-friendly pest control agents with high insecticidal potential using a computer-aided drug design (CADD) approach.

The small brown planthopper (SBPH), Laodelphax striatellus is one of the major insect pests of rice, but little is known about the molecular-level means by which it locates its hosts. SBPH host-seeking behavior heavily relies on chemosensory receptors (CRs). In this study, we utilized genome analysis of the SBPH to identify 169 CRs, including: 133 odorant receptors (ORs), 13 gustatory receptors (GRs) and 23 ionotropic receptors (IRs). The phylogenetic relationships of OR genes from three rice planthoppers and other insect species revealed that the odorant co-receptor (Orco) clade is the most conserved group. Among the candidate GRs, two sugar receptors and five fructose receptors have been identified but no carbon dioxide receptors investigated. Furthermore, we identified homologs of the three highly conserved IR co-receptors. The obtained results will provide us with precious information needed to better understand the interaction between insect pests and crop plants required for effective crop protection.