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Diorhabda rybakowi Weise is one of the dominant pests feeding on Nitraria spp., a pioneer plant used for windbreaking and sand fixation purposes, and poses a threat to local livestock and ecosystems. To clarify the key olfactory genes of D. rybakowi and provide a theoretical basis for attractant and repellent development, the optimal reference genes under two different conditions (tissue and sex) were identified, and the bioinformatics and characterization of the tissue expression profiles of two categories of soluble olfactory proteins (OBPs and CSPs) were investigated. The results showed that the best reference genes were RPL13a and RPS18 for comparison among tissues, and RPL19 and RPS18 for comparison between sexes. Strong expressions of DrybOBP3, DrybOBP6, DrybOBP7, DrybOBP10, DrybOBP11, DrybCSP2, and DrybCSP5 were found in antennae, the most important olfactory organ for D. rybakowi. These findings not only provide a basis for further in-depth research on the olfactory molecular mechanisms of host-specialized pests but also provide a theoretical basis for the future development of new chemical attractants or repellents using volatiles to control D. rybakowi.

Odorant-binding proteins (OBPs) play important roles in the insect olfactory system since they bind external odor molecules to trigger insect olfactory responses. Previous studies have identified some plant-derived volatiles that attract the pervasive insect pest Cnaphalocrocis medinalis (Lepidoptera: Pyralidae), such as phenylacetaldehyde, benzyl acetate, 1-heptanol, and hexanal. To characterize the roles of CmedOBPs in the recognition of these four volatiles, we analyzed the binding abilities of selected CmedOBPs to each of the four compounds, as well as the expression patterns of CmedOBPs in different developmental stages of C. medinalis adult. Antennaes of C. medinalis adults were sensitive to the studied plant volatile combinations. Expression levels of multiple CmedOBPs were significantly increased in the antennae of 2-day-old adults after exposure to volatiles. CmedOBP1, CmedOBP6, CmedPBP1, CmedPBP2, and CmedGOBP2 were significantly up-regulated in the antennae of volatile-stimulated female and male adults when compared to untreated controls. Fluorescence competition assays confirmed that CmedOBP1 could strongly bind 1-heptanol, hexanal, and phenylacetaldehyde; CmedOBP15 strongly bound benzyl acetate and phenylacetaldehyde; and CmedOBP26 could weakly bind 1-heptanol. This study lays a theoretical foundation for further analysis of the mechanisms by which plant volatiles can attract C. medinalis. It also provides a technical basis for the future development of efficient plant volatile attractants of C. medinalis.

Sympiezomias velatus (Chevrolat) (Coleoptera: Curculionidae) has caused serious damage on jujube trees (Ziziphus jujuba Mill) in northern China. Semiochemicals emerging from the host are essential in the process of insects identifying and localizing the host. The highly expressed odorant-binding protein 1 of S. velatus (SvelOBP1) was assumed to play a possible role in the recognition of host volatiles. In this study, SvelOBP1 was cloned based on the antennal transcriptome of S. velatus. The recombinant SvelOBP1 protein was expressed in Escherichia coli and purified by Ni-NTA resin. The predicted protein SvelOBP1 belonged to a classic OBP subfamily. The expression patterns revealed that SvelOBP1 was mainly expressed in the antennae of both males and females, whereas the expression of SvelOBP1 in other body parts could be neglected. The fluorescence binding assay indicated that SvelOBP1 displayed very strong binding affinities to dibutyl benzene-1,2-dicarboxylate and (Z)-hex-3-en-1-ol (Ki = 6.66 ± 0.03 and 7.98 ± 0.06 μM). The molecular docking results showed that residues Trp114, Phe115 and Asp110 may be involved in binding to both dibutyl benzene-1,2-dicarboxylate and (Z)-hex-3-en-1-ol and may have a great impact on odorant recognition of S. velatus. Our results provide evidence that SvelOBP1 might participate in the olfactory molecular perception of S. velatus and would promote the development of pest attractants for S. velatus control.

The olfactory system plays a key role in various insect behaviors, and odorant-binding proteins participate in the first step of the olfactory process. Ophraella communa Lesage is an oligophagous phytophagous insect that is a specific biological control agent for Ambrosia artemisiifolia L. The leaf beetle must identify and locate A. artemisiifolia through olfaction; however, its odorant-binding protein (OBP) function has not yet been reported. In this study, OcomOBP7 was cloned, and its tissue expression profile and binding ability were analyzed using RT-qPCR and fluorescence binding assays, respectively. Sequence analysis demonstrated that OcomOBP7 belongs to the classical OBP family. The RT-qPCR results showed that OcomOBP7 was specifically expressed in the antennae, indicating that OcomOBP7 may be involved in chemical communication. The fluorescence binding assay showed that OcomOBP7 has an extensive binding ability to alkenes. The electroantennography experiments showed that O. communa antennal response to α-pinene and ocimene decreased significantly after interference because the two odors specifically bound to OcomOBP7. In summary, α-pinene and ocimene are odorant ligands corresponding to OcomOBP7, indicating that OcomOBP7 is involved in the chemical recognition of A. artemisiifolia. Our study lays a theoretical foundation for research into O. communa attractants, which is helpful for the better biological control of A. artemisiifolia by O. communa.

Each time an insect bites a vertebrate host, skin and vascular injury caused by piercing triggers a series of responses including hemostasis, inflammation and immunity. In place, this set of redundant and interconnected responses would ultimately cause blood coagulation, itching and pain leading to host awareness, resulting in feeding interruption in the best-case scenario. Nevertheless, hematophagous arthropod saliva contains a complex cocktail of molecules that are crucial to the success of blood-feeding. Among important protein families described so far in the saliva of blood sucking arthropods, is the D7, abundantly expressed in blood feeding Nematocera. D7 proteins are distantly related to insect Odorant-Binding Proteins (OBP), and despite low sequence identity, observation of structural similarity led to the suggestion that like OBPs, they should bind/sequester small hydrophobic compounds. Members belonging to this family are divided in short forms and long forms, containing one or two OBP-like domains, respectively. Here, we provide a review of D7 proteins structure and function, discussing how gene duplication and some modifications in their OBP-like domains during the course of evolution lead to gain and loss of function among different hematophagous Diptera species.

Drosophila melanogaster flies use their proboscis to taste and distinguish edible compounds from toxic compounds. With their proboscis, flies can detect sex pheromones at a close distance or by contact. Most of the known proteins associated with probosci’s detection belong to gustatory receptor families. To extend our knowledge of the proboscis-taste proteins involved in chemo-detection, we used a proteomic approach to identify soluble proteins from Drosophila females and males. This investigation, performed with hundreds of dissected proboscises, was initiated by the chromatographic separation of tryptic peptides, followed by tandem mass spectrometry, allowing for femtomole detection sensitivity. We found 586 proteins, including enzymes, that are involved in intermediary metabolism and proteins dedicated to various functions, such as nucleic acid metabolism, ion transport, immunity, digestion, and organ development. Among 60 proteins potentially involved in chemosensory detection, we identified two odorant-binding proteins (OBPs), i.e., OBP56d (which showed much higher expression in females than in males) and OBP19d. Because OBP56d was also reported to be more highly expressed in the antennae of females, this protein can be involved in the detection of both volatile and contact male pheromone(s). Our proteomic study paves the way to better understand the complex role of Drosophila proboscis in the chemical detection of food and pheromonal compounds.

Human and insect olfaction share many general features, but insects differ from mammalian systems in important ways. Mammalian olfactory neurons share the same overlying fluid layer in the nose, and neuronal tuning entirely depends upon receptor specificity. In insects, the olfactory neurons are anatomically segregated into sensilla, and small clusters of olfactory neurons dendrites share extracellular fluid that can be independently regulated in different sensilla. Small extracellular proteins called odorant-binding proteins are differentially secreted into this sensillum lymph fluid where they have been shown to confer sensitivity to specific odorants, and they can also affect the kinetics of the olfactory neuron responses. Insect olfactory receptors are not G-protein-coupled receptors, such as vertebrate olfactory receptors, but are ligand-gated ion channels opened by direct interactions with odorant molecules. Recently, several examples of insect olfactory neurons expressing multiple receptors have been identified, indicating that the mechanisms for neuronal tuning may be broader in insects than mammals. Finally, recent advances in genome editing are finding applications in many species, including agricultural pests and human disease vectors.

Aphids are one of the most damaging agricultural pests. For the sake of novel eco-friendly compounds with good activity for aphid control, a series of novel geranic acid esters containing substituted aromatic rings were designed by inverting ester groups of lead compounds. All compounds were characterized by HRMS, 1H-NMR, and 13C-NMR. In order to identify the effect of inversion ester groups on activity, a bioassay was conducted. The results showed that the repellent activity against Acyrthosiphon pisum (A. pisum) and the binding affinity with the odorant-binding protein 9 from A. pisum (ApisOBP9) of the compounds were increased after inversion of the ester groups. Particularly, 5f showed the best repellent activity (repellency proportion: 55.6%) and binding affinity (1/Ki: 0.49 µM). Meanwhile, the structure–activity relationships revealed that the introduction of meta-substitution of the benzene ring and halogen atoms, such as Cl and Br, facilitated the biological activity. The further molecular docking results demonstrated that hydrogen bonding interactions and hydrophobic interactions were vital for the binding affinity with ApisOBP9. Additionally, all compounds were predicted to be eco-friendly and their volatile physicochemical properties have been enhanced compared to the leads. The present results provide valuable clues for the further rational design of aphids’ behavioral control agents.

Entomopathogenic fungi such as Beauveria bassiana are extensively used for the control of insect pests worldwide. They infect mostly by adhesion to the insect surface and penetration through the cuticle. However, some insects, such as the red flour beetle Tribolium castaneum (Herbst), have evolved resistance by embedding their cuticle with antifungal compounds. Thus, they avoid fungal germination on the cuticle, which result in low susceptibility to entomopathogenic fungi. In adult T. castaneum, these antifungals are the well-known defensive compounds methyl-1,4- and ethyl-1,4-benzoquinone. In this study, we added B. bassiana conidia on the diet of adult beetles to study the effect of the entomopathogen on the secretion and detection of the beetle volatile blend containing both benzoquinones. The compounds were analyzed by solid phase microextraction coupled to gas chromatography–flame ionization detection, and were detected by electroantennography. In addition, we measured the expression level of four genes encoding for two odorant-binding proteins (OBP), one chemosensory protein (CSP), and one odorant receptor (OR) in both healthy and fungus-treated insects. Significant alterations in the secretion of both benzoquinones, as well as in the perception of methyl-1,4-benzoquinone, were found in fungus-treated insects. TcOBP7D, TcOBP0A and TcCSP3A genes were down-regulated in insects fed conidia for 12 and 48 h, and the latter gene was up-regulated in 72 h samples. TcOR1 expression was not altered at the feeding times studied. We conclude that fungus-treated insects alter both secretion and perception of benzoquinones, but additional functional and genetic studies are needed to fully understand the effects of fungal infection on the insect chemical ecology.

Olfaction is a complex recognition process that is critical for chemical communication in insects. Though some insect species are capable of discrimination between compounds that are structurally similar, little is understood about how this high level of discrimination arises. Some insects rely on discriminating between enantiomers of a compound, demonstrating an ability for highly selective recognition. The role of two major peripheral olfactory proteins in insect olfaction, i.e., odorant-binding proteins (OBPs) and odorant receptors (ORs) has been extensively studied. OBPs and ORs have variable discrimination capabilities, with some found to display highly specialized binding capability, whilst others exhibit promiscuous binding activity. A deeper understanding of how odorant-protein interactions induce a response in an insect relies on further analysis such as structural studies. In this review, we explore the potential role of OBPs and ORs in highly specific recognition, specifically enantiomeric discrimination. We summarize the state of research into OBP and OR function and focus on reported examples in the literature of clear enantiomeric discrimination by these proteins.

Odorant-binding proteins (OBPs) play a key role in the olfactory system and are essential for mating and oviposition host selection. Tirathaba rufivena, a serious lepidopterous insect pest of the palm area in recent years, has threatened cultivations of Areca catechu in Hainan. Female-biased odorant-binding protein 4 of T. rufivena (TrufOBP4) expression was hypothesized to participate in the process of oviposition host recognition and localization. In this study, we cloned and analyzed the cDNA sequence of TrufOBP4. The predicted mature protein TrufOBP4 is a small, soluble, secretory protein and belongs to a classic OBP subfamily. Fluorescence binding assay results showed that TrufOBP4 had high binding abilities with the host plant volatiles, octyl methoxycinnamate, dibutyl phthalate, myristic acid and palmitic acid. These four components tend to dock in the same binding pocket based on the molecular docking result. The interactions and contributions of key amino acid residues were also characterized. This research provides evidence that TrufOBP4 might participate in the chemoreception of volatile compounds from inflorescences of A. catechu and can contribute to the integrated management of T. rufivena.

Olfaction is orchestrated at different stages and involves various proteins at each step. For example, odorant-binding proteins (OBPs) are soluble proteins found in sensillum lymph that might encounter odorants before reaching the odorant receptors. In tsetse flies, the function of OBPs in olfaction is less understood. Here, we investigated the role of OBPs in Glossina fuscipes fuscipes olfaction, the main vector of sleeping sickness, using multidisciplinary approaches. Our tissue expression study demonstrated that GffLush was conserved in legs and antenna in both sexes, whereas GffObp44 and GffObp69 were expressed in the legs but absent in the antenna. GffObp99 was absent in the female antenna but expressed in the male antenna. Short odorant exposure induced a fast alteration in the transcription of OBP genes. Furthermore, we successfully silenced a specific OBP expressed in the antenna via dsRNAi feeding to decipher its function. We found that silencing OBPs that interact with 1-octen-3-ol significantly abolished flies’ attraction to 1-octen-3-ol, a known attractant for tsetse fly. However, OBPs that demonstrated a weak interaction with 1-octen-3-ol did not affect the behavioral response, even though it was successfully silenced. Thus, OBPs’ selective interaction with ligands, their expression in the antenna and their significant impact on behavior when silenced demonstrated their direct involvement in olfaction.

Odour perception has been the object of fast growing research interest in the last three decades. Parallel to the study of the corresponding biological systems, attempts are being made to model the olfactory system with electronic devices. Such projects range from the fabrication of individual sensors, tuned to specific chemicals of interest, to the design of multipurpose smell detectors using arrays of sensors assembled in a sort of artificial nose. Recently, proteins have attracted increasing interest as sensing elements. In particular, soluble olfaction proteins, including odorant-binding proteins (OBPs) of vertebrates and insects, chemosensory proteins (CSPs) and Niemann-Pick type C2 (NPC2) proteins possess interesting characteristics for their use in sensing devices for odours. In fact, thanks to their compact structure, their soluble nature and small size, they are extremely stable to high temperature, refractory to proteolysis and resistant to organic solvents. Moreover, thanks to the availability of many structures solved both as apo-proteins and in complexes with some ligands, it is feasible to design mutants by replacing residues in the binding sites with the aim of synthesising proteins with better selectivity and improved physical properties, as demonstrated in a number of cases.

Since the first attempts to mimic the human nose with artificial devices, a variety of sensors have been developed, ranging from simple inorganic and organic gas detectors to biosensing elements incorporating proteins of the biological olfactory system. In order to design a device able to mimic the human nose, two major issues still need to be addressed regarding the complexity of olfactory coding and the extreme sensitivity of the biological system. So far, only 50 of the approximately 300–400 functioning olfactory receptors have been de-orphanized, still a long way from breaking the human olfactory code. On the other hand, the exceptional sensitivity of the human nose is based on amplification mechanisms difficult to reproduce with electronic circuits, and perhaps novel approaches are required to address this issue. Here, we review the recent literature on chemical sensing both in biological systems and artificial devices, and try to establish the state-of-the-art towards the design of an electronic nose.

In the following we give a short summary of examples for biosensor concepts in areas in which reduced graphene oxide-based electronic devices can be developed into new classes of biosensors, which are highly sensitive, label-free, disposable and cheap, with electronic signals that are easy to analyze and interpret, suitable for multiplexed operation and for remote control, compatible with NFC technology, etc., and in many cases a clear and promising alternative to optical sensors. The presented areas concern sensing challenges in medical diagnostics with an example for detecting general antibody-antigen interactions, for the monitoring of toxins and pathogens in food and feed stuff, exemplified by the detection of aflatoxins, and the area of smell sensors, which are certainly the most exciting development as there are very few existing examples in which the typically small and hydrophobic odorant molecules can be detected by other means. The example given here concerns the recording of a honey flavor (and a cancer marker for neuroblastoma), homovanillic acid, by the odorant binding protein OBP 14 from the honey bee, immobilized on the reduced graphene oxide gate of an FET sensor.