Odorant-Binding Protein HvarOBP5 in Ladybird Hippodamia variegata Regulates the Perception of Semiochemicals from Preys and Habitat Plants.

A total of 12 OBPs were identified in the antennae of ladybird Hippodamia variegata. HvarOBP1/2/5/6/10/11 were highly expressed in adults, whereas HvarOBP3/4/7/8/9/12 had higher expression levels in larvae. In adults, HvarOBP1/2/4/5/6/7/8/11/12 were highly expressed in antennae. Among these, recombinant HvarOBP5 strong bound with (E)-ß-farnesene (EßF), (R)-(+)-limonene, (E,E)-4,8,12-trimethyltrideca-l,3,7,11-tetraene (TMTT), (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), hexyl hexanoate, and geranyl acetate. Molecular docking indicated that Leu42, Lys43, and His64 were the key binding sites of HvarOBP5. All six ligands evoked electroantennography (EAG) responses in ladybirds. Moreover, (R)-(+)-limonene and hexyl hexanoate were attractive to both sexes. After RNA interference for 72 h, the EAGs of dsRNA-injected ladybirds to DMNT and hexyl hexanoate were significantly decreased by 73.8 and 78.6%, respectively. Both dsRNA-injected males and females showed significantly lower behavioral preferences for DMNT and hexyl hexanoate. These findings suggest that HvarOBP5 in H. variegata plays an important role in the perception of semiochemical cues from preys and habitat plants.

Volatiles from cotton aphid (Aphis gossypii) infested plants attract the natural enemy Hippodamia variegata.

The Aphis gossypii is a major threat of cotton worldwide due to its short life cycle and rapid reproduction. Chemical control is the primary method used to manage the cotton aphid, which has significant environmental impacts. Therefore, prioritizing eco-friendly alternatives is essential for managing the cotton aphid. The ladybird, Hippodamia variegata, is a predominant predator of the cotton aphid. Its performance in cotton plantation is directly linked to chemical communication, where volatile compounds emitted from aphid-infested plants play important roles in successful predation. Here, we comprehensively studied the chemical interaction between the pest, natural enemy and host plants by analyzing the volatile profiles of aphid-infested cotton plants using gas chromatography-mass spectrometry (GC-MS). We then utilized the identified volatile compounds in electrophysiological recording (EAG) and behavioral assays. Through behavioral tests, we initially demonstrated the clear preference of both larvae and adults of H. variegata for aphid-infested plants. Subsequently, 13 compounds, namely α-pinene, cis-3-hexenyl acetate, 4-ethyl-1-octyn-3-ol, ß-ocimene, dodecane, E-ß-farnesene, decanal, methyl salicylate, ß-caryophyllene, α-humulene, farnesol, DMNT, and TMTT were identified from aphid-infested plants. All these compounds were electrophysiologically active and induced detectable EAG responses in larvae and adults. Y-tube olfactometer assays indicated that, with few exceptions for larvae, all identified chemicals were attractive to H. variegata, particularly at the highest tested concentration (100 mg/ml). The outcomes of this study establish a practical foundation for developing attractants for H. variegata and open avenues for potential advancements in aphid management strategies by understanding the details of chemical communication at a tritrophic level.

Identification and Tissue Expression Profiles of Odorant Receptor Genes in the Green Peach Aphid Myzus persicae.

The green peach aphid Myzus persicae (Hemiptera: Aphididae) relies heavily on its olfactory system to locate plant hosts, find mates, and avoid parasitoids or predators. The insect odorant receptors (ORs) have been proven to play a critical role in the perception of odorants from the environment. In the present study, 33 odorant receptor candidate genes including the Orco gene were identified from the antennal, head, legs and body transcriptomes of M. persicae. Phylogenetic analysis of ORs from seven different orders of insect species suggests that ORs from different insect species are highly divergent and most ORs from the same species formed monophyletic groups. In addition, the aphid ORs were clustered into six different sub-clades in the same clade. Furthermore, the genomic structure of the OR genes also tends to be consistent, suggesting that ORs from the family Aphididae have a relatively close evolutionary relationship. Reads per kilobase per million (RPKM) and tissue expression profiles analyses revealed that 27 out of the 33 MperORs were uniquely or primarily expressed in the antennae, indicating their putative roles in chemoreception. This work provides a foundation to further investigate the molecular and ecological functions of MperORs in the aphid-aphid, aphid-plant and aphid-natural enemy interactions.

Antenna-Biased Odorant Receptor HvarOR25 in Hippodamia variegata Tuned to Allelochemicals from Hosts and Habitat Involved in Perceiving Preys.

Odorant receptors (ORs) of ladybird Hippodamia variegata play vital chemosensory roles in searching and locating preys. In the current study, 37 ORs were initially identified from the antennal transcriptome of H. variegata. The quantitative polymerase chain reaction demonstrated that several HvarORs including HvarOR25 were specific or enriched in ladybird antennae. In two-electrode voltage clamp recordings, recombinant HvarOR25 was narrowly tuned to six chemical ligands including aphid-induced, aphid-derived, and plant-derived volatiles. In electroantennogram assays, all six volatiles elicited electrophysiological responses. Among the six volatiles, cis-3-hexenyl acetate, hexyl butyrate, hexyl hexanoate, and 3-methyl-3-buten-1-ol were attractive for both sexes of H. variegata. Additionally, molecular docking indicated that HvarOR25 was bound to all ligands with high binding affinities. Taken together, HvarOR25 facilitates perception of preys by recognizing relevant allelochemicals from hosts and habitat. Our findings provide valuable insights into understanding biological functions of HvarORs and help to develop a novel biocontrol strategy based on olfactory-active compounds.

Identification and Evaluation of Suitable Reference Genes for RT-qPCR Analysis in Hippodamia variegata (Coleoptera: Coccinellidae) Under Different Biotic and Abiotic Conditions.

Reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) is an accurate and convenient technique for quantifying expression levels of the target genes. Selection of the appropriate reference gene is of the vital importance for RT-qPCR analysis. Hippodamia variegata is one of the most important predatory natural enemies of aphids. Recently, transcriptome and genome sequencings of H. variegata facilitate the gene functional studies. However, there has been rare investigation on the detection of stably expressed reference genes in H. variegata. In the current study, by using five analytical tools (Delta Ct, geNorm, NormFinder, BestKeeper, and RefFinder), eight candidate reference genes, namely, Actin, EF1α, RPL7, RPL18, RPS23, Tubulin-α, Tubulin-ß, and TufA, were evaluated under four experimental conditions including developmental stages, tissues, temperatures, and diets. As a result, a specific set of reference genes were recommended for each experimental condition. These findings will help to improve the accuracy and reliability of RT-qPCR data, and lay a foundation for further exploration on the gene function of H. variegata.

Glomerular Organization of the Antennal Lobes of the Diamondback Moth, Plutella xylostella L.

The antennal lobe of the moth brain is the primary olfactory center processing information concerning pheromones and plant odors. Plutella xylostella is a major worldwide pest of cruciferous vegetables and its behavior is highly dependent on their olfactory system. However, detailed knowledge of the anatomy and function of the P. xylostella olfactory system remains limited. In the present study, we present the 3-Dimentional (3-D) map of the antennal lobe of P. xylostella, based on confocal microscopic analysis of glomerular segmentation and Neurobiotin backfills of Olfactory Receptor Neurons (ORNs). We identified 74-76 ordinary glomeruli and a macroglomerular complex (MGC) situated at the entrance of the antennal nerve in males. The MGC contained three glomeruli. The volumes of glomeruli in males ranged from 305.83 ± 129.53 to 25440.00 ± 1377.67 µm3. In females, 74-77 glomeruli were found, with the largest glomerulus ELG being situated at the entrance of the antennal nerve. The volumes of glomeruli in females ranged from 802.17 ± 95.68 to 8142.17 ± 509.46 µm3. Sexual dimorphism was observed in anomalous supernumerary, anomalous missing, shape, size, and array of several of the identified glomeruli in both sexes. All glomeruli, except one in the antennal lobe (AL), received projections of antennal ORNs. The glomeruli PV1 in both sexes received input from the labial palp nerve and was assumed as the labial pit organ glomerulus (LPOG). These results provide a foundation for better understanding of coding mechanisms of odors in this important pest insect.

Distribution patterns and morphology of sensilla on the antennae of Plutella xylostella (L.)-A scanning and transmission electron microscopic study.

The antennal morphology, types of antennal sensilla, fine structures and distributions of the sensilla in Plutella xylostella L. (Lepidoptera: Plutellidae) were studied by scanning (SEM) and transmission (TEM) electron microscopy. The antenna, scape, pedicel and flagellum were all longer in males than in females. A total of seventeen types of sensilla were identified on the antennae: trichodea (two subtypes), basiconica, coeloconica (three subtypes), Böhm's bristles (two subtypes), styloconica (two subtypes), squamiformia, auricillica, furcatea (three subtypes), cupuliform organs and terminal sensory pegs. Their numbers and distributions were studied in both male and female, and we found some of the sensilla exhibited various degrees of sexual dimorphisms. Sensilla trichodea were the most abundant of all sensillum types whereas terminal peg was present only once per antenna. Sensilla trichodea in males were bigger (subtype I) and more abundant than in females, however, sensilla basiconica and squamiformia were significantly smaller and less abundant in males than in females. Sensilla styloconica II was only found in females. Seven common sensillum types were studied with TEM to reveal its fine internal structure providing morphological evidences of their sensory functions. Sensilla trichodea I, basiconica and coeloconica III have porous walls suggesting olfactory functions. Combined with the sexual dimorphism, sensilla trichodea male P. xylostella might be involved in detecting sexual pheromones and sensilla basiconica of female might respond to host plant volatiles. Whereas sensilla coeloconica (subtype I and II) and Böhm's bristles have nonporous walls suggesting non-olfactory functions. The study presented a thorough inventory of sensilla on the antennae and laid a solid foundation for future functional studies of these sensilla in this important economical pest.