Sex-specific and functional differentiation between OR23h and OR109d in aggregation pheromone detection in Riptortus pedestris

doi:10.1016/j.jia.2025.11.009

Journal of Integrative Agriculture

2026, Vol. 25

Issue (9): 3775-3785 DOI: 10.1016/j.jia.2025.11.009

Plant Protection

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Sex-specific and functional differentiation between OR23h and OR109d in aggregation pheromone detection in Riptortus pedestris

Xiaotong Zhang^1,^2,³, Jiahang Wei², Xuanpu Luan², Ian W. Keesey⁵, Xin Chen⁴, Qi Yan^1,², Shuanglin Dong^1,²^#, Jin Zhang^1,^2,³^#

^{1 Sanya Institute, Nanjing Agricultural University, Nanjing 210095, China

2 State Key Laboratory of Agricultural and Forestry Biosecurity, College of Plant Protection, Nanjing Agricultural University, Nanjing 210095, China

3 Key Laboratory of Soybean Disease and Pest Control of Ministry of Agriculture and Rural Affairs, Nanjing Agricultural University, Nanjing 210095, China

4 Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement/Institute of Industrial Crops, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China

5 School of Biological Sciences, University of Nebraska - Lincoln (UNL), Lincoln 68588, USA}

Highlights
● Riptortus pedestris detects aggregation pheromones via its distiflagellum.
● OR23h is broadly tuned to three structurally similar components, E2HE2H, E2HZ3H, and E2HH, whereas the male-biased OR109d is narrowly tuned to E2HE2H.
● Dual RNAi treatment demonstrated that OR23h and OR109d collectively mediate the response to the primary component, E2HE2H.

Abstract
References

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摘要

点蜂缘蝽（Riptortus pedestris）是东亚国家大豆的主要害虫。雄虫释放的聚集信息素能够吸引成虫和若虫，这为开发环境友好型害虫防控方法提供了潜力。然而，聚集信息素识别的分子机制尚不明确。本研究利用非洲爪蟾卵母细胞表达系统进行功能分析，发现两个气味受体（OR23h 与 OR109d）负责感知聚集信息素，其中主要组分 (E)-2-己烯基 (E)-2-己烯酸酯可被这两个受体共同识别。进一步的 qPCR 表达谱分析表明，OR109d 仅在雄虫触角中表达，而 OR23h 在雌雄个体中的表达水平相近。RNA干扰实验显示，dsOR23h 处理能显著降低雌雄个体对 (E)-2-己烯基 (Z)-3-己烯酸酯的触角电位反应。此外，同时干扰两个受体基因可显著降低雄虫对 (E)-2-己烯基 (E)-2-己烯酸酯的触角电位反应，并完全抑制雄虫对该化合物的趋向行为。这些结果与性别表达谱一致，揭示了两受体在功能及性别间的分化机制。本研究明确了调控该类信息素所引发聚集行为的关键气味受体，深化了对半翅目昆虫嗅觉信号传导机制的理解，并为优化害虫治理策略提供了重要理论依据。

Abstract

The bean bug, Riptortus pedestris, is a major pest of soybeans in East Asian countries. Male-released aggregation pheromones attract both adults and nymphs, offering potential for eco-friendly pest control. However, the molecular mechanisms underlying the detection of the aggregation pheromones remain unclear. In the present study, functional analysis using the Xenopus oocyte expression system demonstrated that two ORs (OR23h and OR109d) were responsible for sensing aggregation pheromones, with the primary component (E)-2-hexenyl (E)-2-hexenoate (E2HE2H) being shared by the two ORs. Further quantitative PCR (qPCR) profiling indicated that OR109d was expressed only in male antennae, while OR23h was expressed in both sexes at similar levels. RNA interference (RNAi) assays demonstrated that dsOR23h-treatment significantly reduced the electroantennographic (EAG) response of (E)-2-hexenyl (Z)-3-hexenoate (E2HZ3H) in both sexes. Furthermore, simultaneous RNAi knockdown of the two ORs significantly reduced the male EAG response to E2HE2H and abolished male attraction to this compound. These results were consistent with the sex expression profile, demonstrating the sex and functional differentiation between the two ORs. Taken together, this study characterizes the ORs responsible for chemical perception and the associated aggregation behaviors driven by these pheromones. Thus, this study enhances our understanding of olfactory signaling in a hemipteran insect and contributes to the knowledge required for improved pest management.

Keywords: odorant receptors Xenopus oocyte expression and two-electrode voltage clamp RNA interference synergism EAG response behavioral preference

Received: 08 August 2025 Accepted: 09 October 2025 Online: 07 November 2025

Fund:

This work was supported by grants from the National Key Research and Development Program of China (2023YFD1401000) and the National Natural Science Foundation of China (32372530).

About author: #Correspondence Jin Zhang, E-mail: jinzhang001@njau.edu.cn; Shuanglin Dong, E-mail: sldong@njau.edu.cn

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Xiaotong Zhang, Jiahang Wei, Xuanpu Luan, Ian W. Keesey, Xin Chen, Qi Yan, Shuanglin Dong, Jin Zhang. 2026. Sex-specific and functional differentiation between OR23h and OR109d in aggregation pheromone detection in Riptortus pedestris. Journal of Integrative Agriculture, 25(9): 3775-3785.

Endo N, Takashi W, Nobuo M, Seiichi M, Rikiya S. 2005. Ambiguous response of Riptortus clavatus (Heteroptera: Alydidae) to different blends of its aggregation pheromone components. Applied Entomology and Zoology, 40, 41–45.

Guo J M, Wei Z Q, Hou J H, He Y, Luan X P, Zhang Y Y, Liu X L, Zhang X T, Zhang J, Yan Q, Dong S L. 2023. Ionotropic receptor IR75q.2 mediates avoidance reaction to nonanoic acid in the fall armyworm Spodoptera frugiperda (Lepidoptera, Noctuidae). Journal of Agricultural and Food Chemistry, 71, 20602–20612.

Guo X J, He H, Sun J H, Kang L. 2023. Plasticity of aggregation pheromones in insects. Current Opinion in Insect Science, 59, 101098.

Guo X J, Yu Q Q, Chen D F, Wei J N, Yang P C, Yu J, Wang X H, Kang L. 2020. 4-vinylanisole is an aggregation pheromone in locusts. Nature, 584, 584–588.

Hallem E A, Carlson J R. 2006. Coding of odors by a receptor repertoire. Cell, 125, 143–160.

Hallem E A, Dahanukar A, Carlson J R. 2006. Insect odor and taste receptors. Annual Review of Entomology, 51, 113–135.

Hansson B S, Stensmyr M C. 2011. Evolution of insect olfaction. Neuron, 72, 698–711.

Huh H S, Jang S A, Park C G. 2009. Variation in aggregation pheromone secretion of bean bug, Riptortus clavatus. Korean Journal of Applied Entomology, 48, 1612–1617.

Huh H S, Park K H, Seo W D, Park C G. 2005. Interaction of aggregation pheromone components of the bean bug, Riptortus clavatus (Thunberg) (Heteroptera: Alydidae). Applied Entomology and Zoology, 40, 643–648.

Huh W, Park C G. 2010. Effect of day length and temperature on the diapause termination of Riptortus pedestris (Hemiptera: Alydidae) male adults. Korean Journal of Applied Entomology, 49, 205–210.

Karlson P, Lüscher M. 1959. ‘Pheromones’: A new term for a class of biologically active substances. Nature, 183, 55–56.

Kim J, Park K C, Roh H S, Kim J, Oh H W, Kim J A, Park C G. 2016. Morphology and distribution of antennal sensilla of the bean bug Riptortus pedestris (Hemiptera: Alydidae). Microscopy Research and Technique, 79, 501–511.

Kong H L, Guo D, Zhang L, Xie D J, Wilson K, Jiang X F. 2025. Enhanced immune responses of gregarious larvae contribute to successful adult migration in the migratory oriental armyworm. Journal of Integrative Agriculture, 24, 3141–3154.

Leal W S. 2013. Odorant reception in insects: Roles of receptors, binding proteins, and degrading enzymes. Annual Review of Entomology, 58, 373–391.

Leal W S, Higuchi H, Mizutani N, Nakamori H, Kadosawa T, Ono M. 1995. Multifunctional communication in Riptortus clavatus (Heteroptera: Alydidae): Conspecific nymphs and egg parasitoid Ooencyrtus nezarae use the same adult attractant pheromone as chemical cue. Journal of Chemical Ecology, 21, 973–985.

Leal W S, Kadosawa T. 1992. (E)-2-hexenyl hexanoate, the alarm pheromone of the bean bug Riptortus clavatus (Heteroptera: Alydidae). Bioscience, Biotechnology, and Biochemistry, 56, 1004–1005.

Liu X L, Zhang J, Yan Q, Miao C L, Han W K, Hou W, Yang K, Hansson B S, Peng Y C, Guo J M, Xu H, Wang C Z, Dong S L, Knaden M. 2020. The molecular basis of host selection in a crucifer-specialized moth. Current Biology, 30, 4476–4482.e5.

Liu W, Cai C, Wu J, Wang B. 2025. A pheromone from cuticular hydrocarbons regulates mating behavior in the hoverfly Eupeodes corollae. Journal of Integrative Agriculture, 24, 4732–4743.

Love M I, Huber W, Anders S. 2014. Moderated estimation of fold change and dispersion for RNA-Seq data with DESeq2. Genome Biology, 15, 550.

Mizutani N, Wada T, Higuchi H, Ono M, Leal W S. 1997. A component of a synthetic aggregation pheromone of Riptortus clavatus (THUNBERG) (Heteroptera: Alydidae), that attracts an egg parasitoid, Ooencyrtus nezarae ISHII (Hymenoptera: Encyrtidae). Applied Entomology and Zoology, 32, 504–507.

Mizutani N, Wada T, Yasuda T, Endo N, Yamaguchi T, Moriya S. 2008a. Influence of photoperiod on attractiveness and pheromone contents of the bean bug, Riptortus pedestris (Heteroptera: Alydidae). Applied Entomology and Zoology, 43, 585–592.

Mizutani N, Yasuda T, Yamaguchi T, Moriya S. 2007. Individual variation in the amounts of pheromone components in the male bean bug, Riptortus pedestris (Heteroptera: Alydidae) and its attractiveness to the same species. Applied Entomology and Zoology, 42, 629–636.

Mizutani N, Yasuda T, Yamaguchi T, Moriya S. 2008b. Pheromone contents and physiological conditions of adult bean bugs, Riptortus pedestris (Heteroptera: Alydidae), attracted to conspecific males during non-diapause and diapause periods in fields. Applied Entomology and Zoology, 43, 331–339.

Morishima M, Tabuchi K, Ito K, Mizutani N, Moriya S. 2005. Effect of feeding on the attractiveness of Riptortus clavatus (Thunberg) (Heteroptera: Alydidae) males to conspecific individuals. Japanese Journal of Applied Entomology and Zoology, 49, 262–265.

Numata H, Kon M, Hidaka T. 1990. Male adults attract conspecific adults in the bean bug, Riptortus clavatus THUNBERG: Heteroptera: Alydidae. Applied Entomology and Zoology, 25, 144–145.

Rahman M M, Lim U T. 2017. Evaluation of aggregation and alarm pheromones of Riptortus pedestris (Hemiptera: Alydidae) as a push–pull strategy in soybean fields. Applied Entomology and Zoology, 52, 469–479.

Rahman M M, Kim E, Kim D, Bhuyain M M, Lim U T. 2018. Use of aggregation pheromone traps increases infestation of adult Riptortus pedestris (Hemiptera: Alydidae) in soybean fields. Pest Management Science, 74, 2578–2588.

Rani P U, Madhavendra S S. 2005. External morphology of antennal and rostral sensillae in four hemipteran insects and their possible role in host plant selection. International Journal of Tropical Insect Science, 25, 198–207.

Roh G H, Cha D H, Park C G. 2021. Olfactory attraction to aggregation pheromone is mediated by disti-flagellum of antennal segments in Riptortus pedestris. Journal of Asia-Pacific Entomology, 24, 415–420.

Sims C, Birkett M A, Withall D M. 2022. Enantiomeric discrimination in insects: The role of OBPs and ORs. Insects, 13, 368.

Sun D D, Zhang Y T, Cao S, Wang X Q, Cao Q, Zhang S, Liu Y. 2025. A compound produced by Helicoverpa armigera male genitalia activates a conserved pheromone receptor. Journal of Integrative Agriculture, 24, 1892–1904.

Tabuchi K, Moriya S, Mizutani N. 2005. Seasonal catches of the bean bug, Riptortus clavatus (Thunberg) (Heteroptera: Alydidae), in water-pan traps with synthetic attractants. Japanese Journal of Applied Entomology and Zoology, 49, 99–104.

Tateishi K, Nishimura Y, Sakuma M, Yokohari F, Watanabe H. 2020. Sensory neurons that respond to sex and aggregation pheromones in the nymphal cockroach. Scientific Reports, 10, 1995.

Vickers N J. 2017. Animal communication: When I’m calling you, will you answer too? Current Biology, 27, R713–R715.

Wertheim B, Allemand R, Vet L E M, Dicke M. 2006. Effects of aggregation pheromone on individual behaviour and food web interactions: A field study on Drosophila. Ecological Entomology, 31, 216–226.

Wertheim B, Baalen E J A V, Dicke M, Vet L E M. 2005. Pheromone-mediated aggregation in nonsocial arthropods: An evolutionary ecological perspective. Annual Review of Entomology, 50, 321–346.

Yasuda T, Mizutani N, Endo N, Fukuda T, Matsuyama T, Ito K, Moriya S, Sasaki R. 2007a. A new component of attractive aggregation pheromone in the bean bug, Riptortus clavatus (Thunberg) (Heteroptera: Alydidae). Applied Entomology and Zoology, 42, 1–7.

Yasuda T, Mizutani N, Honda Y, Endo N, Yamaguchi T, Moriya S, Fukuda T, Sasaki R. 2007b. A supplemental component of aggregation attractant pheromone in the bean bug Riptortus clavatus (Thunberg) (Heteroptera: Alydidae), related to food exploitation. Applied Entomology and Zoology, 42, 161–166.

Yew J Y, Chung H. 2015. Insect pheromones: An overview of function, form, and discovery. Progress in Lipid Research, 59, 88–105.

Yuvaraj J K, Roberts R E, Sonntag Y, Hou X Q, Grosse-Wilde E, Machara A, Zhang D D, Hansson B S, Johanson U, Löfstedt C, Andersson M N. 2021. Putative ligand binding sites of two functionally characterized bark beetle odorant receptors. BMC Biology, 19, 16.

Zhang X T, Luan X P, Wei J H, Zhang P P, Guo J M, Keesey I W, Gao Y, Yan Q, Zhang J, Dong S L. 2024. Identification of a soybean volatile attractive for Riptortus pedestris using reverse chemical ecology approach. Journal of Agricultural and Food Chemistry, 72, 27084–27093.

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