Region-specific variation in the electrophysiological responses of Spodoptera frugiperda to synthetic sex pheromone compounds

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Akinbuluma, Renée Schaijk, Peter Roessingh, Astrid T. Groot This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3778826/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 29 Feb, 2024 Read the published version in Journal of Chemical Ecology → Version 1 posted 2 You are reading this latest preprint version Abstract The fall armyworm, Spodoptera frugiperda (J.E. Smith), is a global pest that feeds on > 350 plant species and causes major yield loses. Variation in the responses of S. frugiperda males to female sex pheromone compounds affects the detection, monitoring and management of the pest. We determined geographic variation in the responses of S. frugiperda males to four different doses of synthetic sex pheromone compounds using a gas chromatography-electroantennogram detector (GC-EAD). Furthermore, we disentangled regional populations into C- and R- mitotypes via molecular analysis of the cytochrome oxidase I gene, and measured their responses to the compounds. When comparing responses of males from Florida, Benin, Nigeria and Kenya, we found some regional differences in the responses of S. frugiperda males to the major compound, Z9-14:OAc and minor component Z9-12:OAc. However, we found no differences in male responses from the different African countries. All males showed significantly higher antennal responses to Z7-12:OAc than to E7-12:OAc. When comparing the mitotypes, we found that Florida R-type males showed higher responses to Z9-14:OAc, Z7-12:OAc and Z9-12:OAc than Benin R-type males, while C-type males from both regions responded equally to Z7-12:OAc. In addition, Florida R-type males showed higher responses to E7-12:OAc than Florida C-type males. Our study thus shows some differential physiological responses of Spodoptera frugiperda males towards the known sex pheromone compounds, including E7-12:OAc, but mostly in the different mitotypes. How these differences translate to field trap catches remains to be determined. Fall armyworm electroantennogram sex pheromone compounds mitotype Figures Figure 1 Figure 2 Figure 3 Figure 4 INTRODUCTION The fall armyworm, Spodoptera frugiperda (Lepidoptera, Noctuidae) is native to the Americas and currently feeds on a large variety of agricultural crops, belonging to 76 plant families (Montezano et al. 2018 ). It was first found in Africa in 2016 (Goergen et al. 2016), likely facilitated by its high migratory behaviour (Westbrook et al. 2016 ; Baudron et al. 2019 ; Gilioli et al. 2023 ), human-assisted transport and commerce (Cock et al. 2017 ). Since then, S. frugiperda has been found in Asia in 2018 (Bhusal and Bhattarai 2019 ; Navasero et al. 2019; Sun et al. 2021 ) and Australia in 2020 (IPPC, 2021; Paudel Timilsena et al. 2022 ) and was recently found in the canary Island in Southern Europe (IPPC, 2021; Gilioli et al. 2023 ). In sub-Saharan Africa, its major host is maize ( Zea mays L.), which is the most important staple food and is critical for food security of the continent (Pardey et al. 2016 ; Badu-Apraku and Fakorede 2017 ; VIB, 2017 ). Since the arrival of S. frugiperda in Africa, it was found in almost all sub-Saharan countries within a time span of two years (Rwomushana et al. 2018 ) and causes up to 21 million tonnes annual reduction in maize yield (Abrahams et al. 2017 ), representing about 52% annual production loss (Chimweta et al. 2020 ). Thus, it is crucial to develop and implement evidence-based control measures for S. frugiperda in Africa (Prasanna et al. 2018 ). The traditional approach of pesticide use has substantial environmental and human health issues as well as causing damage to non-target organisms, including natural enemies of insect pests (Desneux et al. 2007 ; Régis Ahissou et al. 2021 ). Furthermore, success of pesticides is limited in controlling S. frugiperda as the larvae tend to conceal themselves when feeding on the host plant by hiding in the maize whorl (Harrison et al. 2022 ), and resistance has been developed against many of the cheapest and most widely used pesticides in Africa (Day et al. 2017 ; Akeme et al. 2021 ). In addition, significant expenditures are incurred by both farmers and government in controlling infestation by S. frugiperda (Kassie et al. 2020 ), thus increasing the cost of crop production. Therefore, it is essential to develop an environmentally-friendly and more effective approach to combat the improper use of pesticides. The application of synthetic species-specific sex pheromones is a helpful tool in integrated pest management (IPM) for monitoring and early detection of S. frugiperda (Prasanna et al. 2018 ; FAO. 2019b; Matova et al. 2020 ). However, for a successful monitoring system, it is essential that lures are attractive. Thus, effective, local, regional and continental species-specific detection, monitoring and pest management should be developed as solid bases of IPM strategies (Saveer et al. 2023 ). In the Western Hemisphere, S. frugiperda is subdivided into two strains, the so-called corn strain and rice strain (Pashley et al. 1992 ; Lu and Adang 1996 ; Gouin et al. 2017 ), which differ by mitochondrial markers (Nagoshi 2010 ) as well as by nuclear differences (Gouin et al. 2017 ). Even though some differences in sexual communication has been found between the strains (Unbehend et al. 2013 ), intra-strain geographic variation in response to the signals has also been found (Unbehend et al. 2014 ). Furthermore, although the effluvia and gland pheromone extracts from calling S. frugiperda females in the Western Hemisphere contain similar sex pheromone compounds as in Benin (Haenniger et al. 2020 ), the response and capture of S. frugiperda males using commercial lures with some of these compounds have been erratic (Meagher et al., 2019 ; Tepa-Yotto et al., 2022 ). The sex pheromone of S. frugiperda in the Americas consists of (Z)-9-tetradecenyl acetate (Z9-14:OAc) and (Z)-7-dodecenyl acetate (Z7-12:OA) which have been found to be important for the attraction of conspecific males (Tumlinson et al. 1986 ; Andrade et al. 2000 ; Unbehend et al. 2013 ; Unbehend et al. 2014 ; Jiang et al. 2021). Adding (Z)-11-hexadecenyl acetate (Z11-16:OAc) increased male attraction in Pennsylvania (Fleischer et al. 2015), and also elicited a small electroantennogram response to S. frugiperda males in Yunnan Province (Jiang et al. 2021). In Brazil, (E)-7-dodecenyl acetate (E7-12:OAc) was identified as an additional active sex pheromone component (Batista-Pereira et al. 2006 ; Cruz-Esteban et al. 2020). In Africa, S. frugiperda monitoring studies with pheromone lures have revealed varying results (Meagher et al. 2019 ; Koffi et al. 2021 ; Tepa-Yotto et al. 2022 ). For example, in Benin, when Tepa-Yotto et al. ( 2022 ) compared the attraction of S. frugiperda males among three blends, i.e., a four-component blend containing Z9-14:OAc, Z7-12:OAc, Z11-16:OAc and (Z)-9-dodecenyl acetate (Z9-12:OAc), a three-component blend (without Z9-12:OAc) and a two-component blend containing only Z9-14:OAc and Z7-12:OAc, they found that the 4-component blend attracted the highest number of S. frugiperda males, irrespective of the conditions of the experiment, while the 2-component blend was the most selective, as the percentage of bycatches was the lowest (Tepa-Yotto et al. 2022 ). Conversely, Koffi et al. ( 2021 ) found that the 3-component lure containing Z9-14:OAc, Z7-12:OAc, Z11-16:OAc was more attractive than the 4-component lure (adding Z9-12:OAc) in the neighboring country Togo. Thus, within African, male S. frugiperda responses to female sex pheromones may differ. To assess whether males from different populations show different physiological responses to the identified sex pheromone compounds, we investigated the antennal responses of males from three African populations and an American population to all identified synthetic sex pheromone compounds. We also determined whether male responses varied depending on the two identified mitotypes of S. frugiperda . Specifically, we evaluated variability in the antennal response of S. frugiperda males that were collected as larvae from Florida, Benin, Nigeria and Kenya and from C- and R- males from Benin and Florida, to E7-12:OAc, Z7-12:OAc, Z9-12:OAc, Z9-14:OAc and Z11-16:OAc. METHODS AND MATERIALS Insect Collection and Rearing. The S. frugiperda populations from Benin originated from larvae, collected from maize fields and alternative host plants in South and Central Benin by the International Institute of Tropical Agriculture (IITA), Benin in July 2020, from which a laboratory population was developed at the University of Amsterdam. This population was mixed with field specimens from Azowlisse, Benin, which was collected in December 2020. In July 2021 and October 2022, new specimens from the lab population from IITA were mixed into the developed lab population at the University of Amsterdam. The Kenyan population was obtained from the rearing of the International Centre of Insect Physiology and Ecology (icipe, Kenya) in December 2020 and August 2022. The Nigerian population was collected from maize fields in Southern Oyo State and from IITA, Ibadan, Nigeria, between January 2022 and May 2022. To compare responses of African S. frugiperda males to American S. frugiperda males, Florida C- and R- strains were also reared at the University of Amsterdam, with populations obtained from the Max Planck Institute of Chemical Ecology, Jena, Germany. The immature stages (eggs and larvae) of all the populations were reared on an artificial pinto bean diet in climate chambers at 25°C and humidity of 60–65%, with reversed light/dark cycle and 14:10 light/dark photoperiod at the laboratory at the University of Amsterdam (IBED). The adults were fed with 10% sugar water, ad libitum . Male and female insects were used for mitotype identification, while 2–5 day old virgin male were used for gas chromatography-electroantennogram detector (GC-EAD) experiments. Mitochondrial identification of African population. Since the invasive populations of S. frugiperda are mixed in their nuclear genome, but do consist of two mitotypes (Yainnina et al. 2021; Tay et al. 2022 ), we determined whether males with different mitotypes show different electrophysiological responses. Mitochondrial identification was done by screening adults for the mitochondrial marker (COI) that is diagnostic for both strains in North and South America (Nagoshi et al. 2006 ; Nagoshi 2010 ) and generally used in other populations as well (Tay et al. 2022 ). DNA extractions were performed in a 96-well plate using Chelex 100 Resin (Bio-Rad Laboratories, Hercules, CA, USA). One adult leg was put in one well together with two metal beads and 300µl 10% Chelex (diluted in ddH 2 O). The tissue was homogenized in a tissue lyser for 4min at 30Hz. Samples were heated for 30 min at 95°C and 300 rpm spinning, after which they were frozen at − 20°C overnight. Each plate was thawed, mixed, and centrifuged at 4000 rpm for 30min. The supernatant was filtered through a fritted deep well filter plate (Thermo Fisher Scientific, Waltham, MA, USA) and used for mitotype analysis. Identification of the mitochondrial marker was performed as described by Unbehend et al. ( 2013 ) and summarized here. After amplification of the COI gene, two strain-specific digests with MspI and SacI were conducted to analyze the strain-affiliation via gel electrophoresis. To compare EAG responses between the mitotypes, we used both Florida (C) - and (R) - males and Benin (C)- and (R) - males. The Kenya males were not mitotyped and all the sampled populations from Nigeria were (R) - type. In our preliminary analysis, we observed no significant differences in the responses between Benin (R) - and Nigerian (R) - strain males. Preparation of the Multicomponent Blends. All used synthetic sex pheromone compounds of S. frugiperda were purchased from Pherobank (Wijk bij Duurstede, The Netherlands), i.e., E7-12:OAc, Z7-12:OAc, Z9-12:OAc, Z9-14:OAc and Z11-16:OAc with > 98% purity. To determine the antennal responses to these compounds, two multicomponent blends (MCBs) were prepared, one with E7-12:OAc and one with Z7-12:OAc in combination with Z9-12:OAc, Z9-14:OAc, Z11-16:OAc (Supplementary Table S1 ). This allowed us to distinguish responses between E and Z7-12:OAc. A stock solution of 10 µg/µl in hexane was made for each synthetic pheromone compounds from which the two MCBs (i.e one including E7-12:OAc and the other including Z7-12:OAc) were made by diluting 1 µl each of the four synthetic pheromone compounds into 500 µl of hexane. Each of concentrations of 1 ng/µl, 3 ng/µl, 10 ng/µl and 20 ng/µl was obtained in a serial dilution and samples were put in the vials and kept in the refrigerator till the time of chemical analysis. GC-EAD measurements. To measure the electrophysiological responses of male antennae, the two MCBs in the four different concentrations were used in random order on 2 to 4-day-old virgin males from Benin, Kenya, Nigeria and Florida (C-type and R-type). Live male insect was individually placed in a plastic pipet tip and one antenna was immobilized with a small strip of parafilm pressing the antennal base against the head. Electrical contact was made using silver wires inserted in glass microelectrodes (GC150TF-10; Warner Instruments, Hamden, CT, USA) with insect Ringer’s solution. The recording electrode was inserted at the base of the antenna and the reference electrode made contact with the cut antennal tip. The amplitude of the EAG was measured using an IDAC-4 amplifier equipped with a high impedance (> 10 9 Ohm) head stage and recorded with GC-EAD/2014 software (Syntech, Kirchzarten, Germany). An Agilent 7890B gas chromatograph (Agilent, Wilmington, DE, USA) equipped with an Agilent Cool On-Column inlet, was coupled to the EAG setup to deliver odour stimuli. Details of the GC-EAD set up are in the supplementary S1. To check the longevity of the whole insect preparation, a reference stimulus (containing Z3-6:OH, 10 − 3 v/v (4.2 µg) in paraffin oil) was delivered for 0.5 seconds from a filter paper strip in a Pasteur pipet by a CS-55 stimulus controller (Syntech). The preparations proved to be extremely stable, as no significant differences were found between the sampled responses to the reference before and after the GC-EAG run ( t -test, n = 14, P = 0.203, df = 1). Statistics and Data analysis. To analyse the antennal responses to the pheromone compounds, we built linear mixed effect models (Lindstrom and Bates 1988 ) as described by Pinheiro and Bates ( 2000 ) and implemented in R (R Core Team 2022 ) as function lme (Pinheiro et al. 2022 ). Mixed models, with individual insect as a random factor, and compound, concentration, population and (when available) mitotype as fixed factors, allowed us to separate the (random) variation between individual insect from the variation caused by the (fixed) factors of interest. We built 3 separate models (Supplementary Table S2). The first model contained compound, concentration and population as predictors, with 498 EAD responses from 25 male insects. A second model contained mitotype as explanatory variable. The model was fitted with 397 cases from 19 males for which mitotype information was available. Finally, since the results indicated that there was no significant difference between the African populations, a third model was constructed to compare EAD responses on a continental scale (Africa vs America). This dataset contained in total 697 EAD responses from 34 males. For each model, we started with a full model that contained the random factor, all the fixed factors and their interactions. Model selection of better fitting model was done using Akaike's Information criterium (AIC) (Sakamoto et al. 1986 ). All models were fit by maximum likelihood to allow model comparison using AIC. Post hoc comparisons of mean values from the final models was done with Least-squares means (Searle et al. 1980 ) as implemented in the R package emmeans (Lenth, 2023 ) with Tukey multiplicity adjustments. RESULTS Electrophysiological Responses to Synthetic Sex Pheromone Compounds . When checking the electrophysiological responses of S. frugiperda males from the different regions, we found significant effects of geographic regions, the compounds, concentrations, as well as interactions between geographic region and compounds and between concentration and compound (Fig. 1 ). The major compound, Z9-14:OAc, and Z9-12:OAc elicited significant responses among the different regions (Fig. 1 a,c), as Florida male responses were higher than male responses from Benin, Nigeria and Kenya (P 0.05). In all regions, S. frugiperda males responded similarly to Z7-12:OAc, E7-12: OAc and Z11-16:OAc (Fig. 1 b,d,e) (P > 0.05). In assessing whether males responded differently to the two synthetic isomers, Z7-12:OAc and E7-12:OAc, we plotted the responses to both compounds from each region and at different concentrations in separate boxplots (Fig. 2 ). Overall, Z7-12:OAc gave higher antennal responses than E7-12:OAc in all the regions (Fig. 2 a-d). Also, only Florida and Nigerian males responded significantly to the two compounds at 1 ng/µl, while Kenyan males responded more to Z7-12:OAc than E7-12:OAc at 3 ng/µl. At higher concentrations (10 ng/µl and 20 ng/µl), males from all the regions (Florida, Benin, Nigeria and Kenya) showed significantly higher antennal responses to Z7-12:OAc than E7-12:OAc (P < 0.001). Since there was no difference in male responses to the compounds in the three African populations (Fig. 1 ), we combined these responses to a so-called ‘African’ response and compared their overall additive responses to the Florida (American) responses. Our results revealed some differences across the two continents (Fig. 3 a-e). Specifically, EAG responses evoked by the major compound, Z9–14:OAc, the critical secondary compound, Z7–12:OAc and Z9–12:OAc were significantly higher in American males than in African (Benin, Kenyan and Nigerian) males(P < 0.001, P = 0.029 and P < 0.001, respectively). However, we found no differences in responses to E7-12:OAc and Z11-16:OAc between the continents. When analysing variation in mitotyped male responses, we found a significant effect of compound, geographic region and an interaction between compound x strain x geographic region (P < 0.001; Fig. 4 ). Specifically, Florida (C)-type males showed significantly lower response to E7-12:OAc than Florida(R)-type males (P = 0.025), while both mitotypes responded similarly to other compounds. Each of the five synthetic compounds also evoked similar responses in Benin (C)-type and (R)-type males. Between Benin (R)-type and Florida (R)-type males, we observed significant higher responses of Florida (R)-type males to Z9-14:OAc (P < 0.001), Z7-12:OAc (P = 0.006) and Z9-12:OAc (P = 0.0002) (Fig. 4 a,b,c). Also, Florida (C)-type males showed significantly higher responses than Benin (C)-type males to Z9-14:OAc and Z9-12:OAc, respectively (P < 0.001) (Fig. 4 a,c). DISCUSSION In this study, we investigated the variation in electrophysiological responses of S. frugiperda males from Florida, Benin, Nigeria and Kenya to sex pheromone compounds, and found a) some geographic variation in responses of males to Z9-14:OAc and Z9-12:OAc, b) significantly higher responses to Z7-12:OAc than E7-12:OAc in all males, c) continental differences in responses to some sex pheromones compounds, and d) some mitotype differences in responses, whereby Florida (R)-type males showed higher responses to E7-12:OAc than Florida (C)-type males and higher responses to Z7-12:OAc, Z9-14:OAc and Z9-12:OAc than Benin (R)-type males. Overall, our results show that Florida males were generally more responsive to sex pheromone compounds than the African males, while all African males were equally responsive to the pheromone compounds. Geographic variation in male responses Clearly, S. frugiperda males from the different regions responded to all synthetic sex pheromone compounds, even though to a varying degree. Our finding that Florida males showed higher responses to the major compound, Z9-14:OAc than Benin, Nigerian and Kenyan males, corresponds to previous results by Haenniger et al. ( 2020 ), where Florida males also elicited greater EAG responses to Z9-14:OAc than Benin and Nigerian males. Since it is widely agreed that Z9-14:OAc is the major sex pheromone component needed to effectively attract S. frugiperda males (Tumlinson et al. 1986 ; Andrade et al. 2000 ; Meagher et al. 2013, 2019 ; Hänniger et al. 2020), it is surprising that African male responses are lower than those from Florida. Similarly, Z9-12:OAc elicited greater EAGs in Florida males than in Benin, Nigerian and Kenyan males. The activity of Z9-12:OAc as a pheromone component has been scarcely detected, although Tepa-Yotto et al. ( 2022 ) found that the addition of Z9-12:OAc to a four-component pheromone blend improved male attraction. Possibly, adding Z9-12:OAc to the sex pheromone blend increases male attraction in Africa. A generally low EAG amplitude was observed in response to Z11-16:OAc in all the regions tested, which is consistent with previous findings (Malo et al. 2004; Unbehend et al. 2013 ; Jiang et al. 2021). Z11-16:OAc doesn’t seem to be a sex pheromone component in American fields (Unbehend et al. 2014 ) and may not increase male attraction in African fields (Tepa-Yotto et al. 2022 ). Moreover, the addition of Z11-16:OAc to lures caused large numbers of bycatches of non-target moths, particularly Mythimna loreyi (Duponchel) in West Africa (Meagher et al. 2019 ; Tabata et al. 2022 ), indicating that S. frugiperda lures in Africa should be developed without Z11-16:OAc. Variation in response to E7-12:OAc and Z7-12:OAc We found interesting variations in male responses to the two isomeric compounds, Z7-12:OAc and E7-12:OAc across the concentrations tested. Overall, all S. frugiperda males showed higher EAG responses to Z7-12:OAc than to its isomer, E7-12:OAc. So far, E7-12:OAc has been reported only within the female glands of Brazilian S. frugiperda populations (Batista-Pereira et al. 2006 ) and more interestingly, was behaviorally active on males from that region (Batista-Pereira et al. 2006 ; Cruz-Esteban et al. 2018). Whether E7-12:OAc is absent in female glands in African or other American regions is mostly still unclear, as is the male response in other regions. Unbehend et al. ( 2014 ) did find that (C)-type males from Peru are only attracted to a blend containing Z7-12:OAc, but not to a blend with E7-12:OAc, while males in North Carolina did not differentiate between the two isomers. As chromatographic separation of the two isomers is difficult, it is possible that this compound has remained undetected in other studies. However, the lower EAG responses of the males to E7-12:OAc than Z7-12:OAc from all four regions suggests that the latter may be more important than the former in male attraction in all regions. Variation in response between Africa and Florida Our finding that S. frugiperda males from Benin, Kenya and Nigeria showed similar EAD responses to all pheromone compounds, is comparable to the reports of Haenniger et al. ( 2020 ) where Benin and Nigerian S. frugiperda males exhibited similar EAG amplitudes to the five known sex pheromone synthetic compounds. These results suggest that S. frugiperda males may not show geographic variation within the Africa continent. However, geographic variation between continents seems to occur, as we found intercontinental differences in the male responses towards Z9-14:OAc, Z7-12:OAc and Z9-12:OAc. Variation in inter-type and intra-type geographic males Interestingly, when comparing the mitotypes, we found that Florida (C)- and (R)-type males differed significantly in their response to E7-12:OAc and not any other pheromone compounds. The fact that there was no type-specific differential response to all but one pheromone compound in Florida males, and no type-specific differential response to any pheromone compound in Benin males suggests that males of both mitotypes have similar response range and are not differentiated with respect to antennal response at the doses tested. Field trapping experiments with Florida population also showed that both strains were similarly attracted to pheromone lures (Unbehend et al. 2013 ; Unbehend et al. 2014 ; Kenis et al. 2023 ). Also in Kenya, both corn and rice mitotypes were equally attracted to the different commercial pheromone lures when tested in replicated field trials (Sisay et al. 2024 ). In conclusion, we found that electrophysiological responses of Spodoptera frugiperda males to sex pheromone compounds differ between Florida and Africa population, but responses do not differ within the African continent. These results suggests that pheromone lures may have to be adjusted for monitoring of Spodoptera frugiperda in Africa compared to America, but do not have to be adjusted for specific regions within the African continent. Declarations Acknowledgements: The authors thank Dr Sabine Haenniger for donating larvae from the Florida S. frugiperda colonies, Dr Olajumoke Alabi for her assistance in collecting the Nigerian specimens of the fall armyworm, Eileen Bader for rearing the insects at UvA, and Betsie Voetdijk and Peter Kuperus for their help in molecular analysis. We also thank Dr Jacques Deere and Ahmed G. Hussain for their help in data analysis. Funding This work was supported by a grant from the Tertiary Education Trust Fund of the University of Ibadan, Nigeria to MDA, and by a grant provided by Simonis BV to ATG and RAHvS, and by the University of Amsterdam, The Netherlands. Conflicts of interest/Competing interests The authors declare that they have no conflict of interest/competing interests Supplementary Information S upplementary information can be found on the online version of this article Author Contributions: MDA, RAHvS, PR and ATG planned, conceived and prepared the study. MDA and PR did electrophysiology and analysed data. All authors wrote and edited the manuscript and approved the final version of the manuscript. Availability of data and materials The authors confirm that data supporting the findings of this study are available on request. References Abrahams P, Beale T, Cock M, Corniani N, Godwin J, Murphy S, Richard JV (2017) Fall armyworm status. 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Nagoshi RN, Goergen G, Du Plessis H, van den Berg J, Meagher R (2019) Genetic comparisons of fall armyworm populations from 11 countries spanning sub-Saharan Africa provide insights in to strain composition and migratory behaviors. Sci Rep 9: 10.1038/s41598-019-44744-9. Nagoshi RN, Htain NN, Boughton D, Zhang L, Xiao Y, Nagoshi BY, Mota-Sanchez D (2020) Southeastern Asia fall armyworms are closely related to populations in Africa and India, consistent with common origin and recent migration. Sci Rep: 10:1421. doi: 10.1038/s41598-020-58249-3. Nagoshi RN, Koffi D, Agboka K, Tounou KA, Banerjee R et al (2017) Comparative molecular analyses of invasive fall armyworm in Togo reveal strong similarities to populations from the eastern United States and the Greater Antilles. PLoS One.12:e0181982 88. Nagoshi RN, Meagher R (2003) Fall armyworm FR sequences map to sex chromosomes and their distribution in the wild indicate limitations in interstrain mating. Insect Mol Biol 12: 453–58. Nagoshi RN, Meagher RL, Nuessly G, Hall DG (2006) Effects of fall armyworm (Lepidoptera: Noctuidae) interstrain mating in wild populations. Environ Entomol 35:561–668. Pardey PG, Andrade RS, Hurley TM, et al (2016) Returns to food and agricultural R&D investments in Sub-Saharan Africa, 1975–2014. Food Policy 65:1–8. https://doi.org/10.1016/j.foodpol.2016.09.009 Pashley DP, Hammond AM, Hardy TN (1992) Reproductive isolating mechanisms in fall armyworm host strains (Lepidoptera, Noctuidae). Ann Entomol Soc Am 84:400-405. https://doi.org/10.1093/aesa/85.4.400. Paudel Timilsena B, Niassy S, Kimathi E, et al (2022) Potential distribution of fall armyworm in Africa and beyond, considering climate change and irrigation patterns. Sci Rep 12:539. https://doi.org/10.1038/s41598-021-04369-3 Pinheiro J, Bates D, R Core Team (2022) _nlme: Linear and Nonlinear Mixed Effects Models_. R package version3.1-157, https://CRAN.R-project.org/package=nlme https://doi.org/10.1007/b98882. Pinheiro JC, Bates DM (2000) Mixed-Effects Models in S and S-PLUS_. Springer, New York. doi:10.1007/b98882. Prasanna BM, Huesing JE, Eddy R, Peschke VM [eds.]. 2018. Fall Armyworm in Africa: A Guide for Integrated Pest Management, First Edition. CIMMYT, El Batán, Mexico State, Mexico. R Core Team (2022). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL https://www.R-project.org/ Régis Ahissou B, Mathieu Sawadogo W, H. Bokonon-Ganta A, et al (2021) Integrated pest management options for the fall armyworm Spodoptera frugiperda in West Africa: Challenges and opportunities. A review. BASE. https://doi.org/10.25518/1780-4507.19125 Rwomushana I, Bateman M, Beale T, Beseh P, Cameron K, Chiluba M, Clottey V, Davis T, Day R, Early R et al (2018). Fall armyworm: impacts and implications for Africa. CABI. https://www.invasive-species.org/wp-content/uploads/sites/2/2019/02/FAW-Evidence-Note-October-2018.pdf Sakamoto Y, Ishiguro M, Kitagawa G (1986). Akaike Information Criterion Statistics . D. Reidel Publishing Company. Saveer AM, Hatano E, Wada-Katsumata A et al (2023) Nonanal, a new fall armyworm sex pheromone component, significantly increases the efficacy of pheromone lures. Pest Manag Sci 79:2831–2839. https://doi.org/10.1002/ps.7460. Searle S R., Speed F M, Milliken G (1980) Population Marginal Means in the Linear Model: An Alternative to Least Squares Means. Am Stat 34: 216-221. doi:10.1080/00031305.1980.10483031. Sisay B, Subramanian S, Weldon CW, et al (2024) Evaluation of pheromone lures, trap designs and placement heights for monitoring the fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae) in maize fields of Kenya. Crop Prot 176:106523. https://doi.org/10.1016/j.cropro.2023.106523. Sun X, Hu C, Jia H, et al (2021) Case study on the first immigration of fall armyworm, Spodoptera frugiperda invading into China. J Integr Agric 20:664–672. https://doi.org/10.1016/S2095-3119(19)62839-X Tabata J, Nakano R, Yasui H, et al (2022) Sex pheromone of the fall armyworm, Spodoptera frugiperda: identification of a trace component that enhances attractiveness and specificity. Entomol Exp Appl : https://doi.org/10.1111/eea.13287. Tay WT, Meagher RL, Czepak C, Groot AT (2023) Spodoptera frugiperda : Ecology, Evolution, and Management Options of an Invasive Species. Annu Rev Entomol 68:299–317. https://doi.org/10.1146/annurev-ento-120220-102548. Tay WT,Rane R, Padovan A,Walsh T, Elfekih S, et al. (2022) Global population genomic signature of fall armyworm supports complex introduction events across the Old World. Commun Biol5:297.https://doi.org/10.1038/s42003-022-03230-1 Tepa-Yotto GT, Meagher RL, Winsou JK, et al (2022) Monitoring Spodoptera frugiperda in Benin: assessing the influence of trap type, pheromone blends, and habitat on pheromone trapping. Fla Entomol 105:. https://doi.org/10.1653/024.105.0111 Tumlinson JH, Mitchell ER, Teal PEA, Heath RR and Mengelkoch LJ (1986) Sex pheromone of fall armyworm, Spodoptera frugiperda (J.E. Smith) identification of components critical to attraction in the field. J Chem Ecol 12:1909–1926. Unbehend M, Haenniger S, Meagher RL, Heckel DG, Groot AT (2013) Pheromonal divergence between two strains of Spodoptera frugiperda . J Chem Ecol 39:364–376. Unbehend M, Haenniger S, Vasquez GM, et al (2014) Geographic Variation in Sexual Attraction of Spodoptera frugiperda Corn- and Rice-Strain Males to Pheromone Lures. PLoS ONE 9:e89255. https://doi.org/10.1371/journal.pone.0089255. VIB (2017). Maize in Africa. Internationl Plant Biotechnology Outreach. https://doi.org/10.1088/0305-4470/31/34/016. Westbrook JK, Nagoshi RN, Meagher RL, et al (2016) Modeling seasonal migration of fall armyworm moths. Int J Biometeorol 60:255–267. https://doi.org/10.1007/s00484-015-1022-x. Yainna S, Nègre N, Silvie PJ, Brévault T, Tay WT, et al. (2021). Geographic monitoring of insecticide resistance mutations in native and invasive populations of the fall armyworm. Insects12:468 Additional Declarations No competing interests reported. Supplementary Files Supplementarymaterials.docx Cite Share Download PDF Status: Published Journal Publication published 29 Feb, 2024 Read the published version in Journal of Chemical Ecology → Version 1 posted Submission checks completed at journal 27 Dec, 2023 First submitted to journal 19 Dec, 2023 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-3778826","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":263677829,"identity":"2e229912-6bd9-46a1-b3d0-04e67260684a","order_by":0,"name":"Mobolade D. Akinbuluma","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA2UlEQVRIiWNgGAWjYHACAwaGAxJAkrkByLGBiD0gTgsjSEsaRCyBsBYGmJbDhLXoNjBvYPhxxiLPXCKxTepGzfnEDccPsD3Ap8XsAFsBY88NiWLLGYlt0jnHbiduOJPAboBfC48BA88HicQNN0Ba2G4nzmxIYJMgpIXxD1zLv3OJM/sfENbCzHMDqiW37UBivwQhWw6zFRyWOSORuLPnYbN1bl+ycb/Ewzb8Wo43b3z45lhd4nb25IO3c77ZybbxJx+T+IBHCwMzOFZQADiCRsEoGAWjYBRQAgDDwVLHxGW8WAAAAABJRU5ErkJggg==","orcid":"","institution":"University of Amsterdam","correspondingAuthor":true,"prefix":"","firstName":"Mobolade","middleName":"D.","lastName":"Akinbuluma","suffix":""},{"id":263677830,"identity":"b610e5de-cfc8-4b8f-8a86-862a1759e3e3","order_by":1,"name":"Renée Schaijk","email":"","orcid":"","institution":"University of Amsterdam","correspondingAuthor":false,"prefix":"","firstName":"Renée","middleName":"","lastName":"Schaijk","suffix":""},{"id":263677831,"identity":"b5c76345-1a27-445c-883f-641571c6b899","order_by":2,"name":"Peter Roessingh","email":"","orcid":"","institution":"University of Amsterdam","correspondingAuthor":false,"prefix":"","firstName":"Peter","middleName":"","lastName":"Roessingh","suffix":""},{"id":263677832,"identity":"6804d3bb-ea49-4f2e-ab85-2e07b3d6eeed","order_by":3,"name":"Astrid T. Groot","email":"","orcid":"","institution":"University of Amsterdam","correspondingAuthor":false,"prefix":"","firstName":"Astrid","middleName":"T.","lastName":"Groot","suffix":""}],"badges":[],"createdAt":"2023-12-19 22:44:13","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-3778826/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-3778826/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s10886-024-01479-w","type":"published","date":"2024-02-29T15:01:34+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":49158092,"identity":"fbcf1bd3-736e-4f79-92e6-dc7e6ad7edd5","added_by":"auto","created_at":"2024-01-04 04:52:23","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":141932,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eResponses of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eSpodoptera frugiperda\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e males to different doses of sex pheromone compounds.\u003c/strong\u003eBoxes represent mean response of males to (a) Z9-14:OAc (b) Z7-12:OAc (c) Z9-12:OAc (d) E7-12:OAc, and (e) Z11-16:OAc. Error bars indicate the range of the collected data, excluding outliers (n = 10 in Florida, except in Z7-12:OAc and Z9-12:OAc at 1 ng/µl (where n=9); n= 5 in Benin, Nigeria and Kenya). Significant difference within regions are indicated by different letters\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-3778826/v1/5523c83346c9d123d1117757.png"},{"id":49158008,"identity":"b9f6ffa3-e605-43a3-820e-256168c8dfc9","added_by":"auto","created_at":"2024-01-04 04:44:23","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":166517,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eResponses of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eSpodoptera frugiperda\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e males to Z7-12:OAc and E7-12:OAc at different doses\u003c/strong\u003e. Boxes represent mean response of males to Z7-12:OAc and E7-12:OAc. Error bars indicate the range of the collected data, excluding outliers (n=10 in Florida, except in Z7-12:OAc at 1 ng/µl (where n=9); n= 5 in Benin, Nigeria and Kenya);Within each region,* = P\u0026gt;0.05,** = P\u0026lt;0.01, ***=P\u0026lt;0.001; ns = not significant\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-3778826/v1/55a4bbff8b1dfc5dfe2ff348.png"},{"id":49158005,"identity":"36551d70-8d99-4405-822e-f8fd7e60a6a3","added_by":"auto","created_at":"2024-01-04 04:44:23","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":30992,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eThe electroantennography (EAG) dose-response curves of the antennae of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eSpodoptera frugiperda\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003emales from combined Africa\u003c/strong\u003e. Graphs show male responses to\u003cstrong\u003e (a) \u003c/strong\u003eZ9-14:OAc (b) Z7-12:OAc (c) Z9-12:OAc (d) E7-12:OAc and (e) Z11-16:OAc. Error bars indicate mean (± SEM) of EAG amplitudes (mV) (n = 25 in Africa, except in Z7-12:OAc at 1 ng/µl (where n=24); n=10 in Florida (except in Z7-12:OAc and Z9-12:OAc at 1 ng/µl, where n=9). Significant difference within continents are indicated by different letters\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-3778826/v1/8e9cb089502875d7fdfb4fb7.png"},{"id":49158091,"identity":"42189a93-1b4d-4296-a4c1-39d2b22c1ef8","added_by":"auto","created_at":"2024-01-04 04:52:23","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":132876,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eResponses of \u003c/strong\u003e\u003cem\u003e\u003cstrong\u003eSpodoptera frugiperda\u003c/strong\u003e\u003c/em\u003e\u003cstrong\u003e Florida and Benin C- and R-type males to different doses of sex pheromone compounds. \u003c/strong\u003eBoxes represent mean response of males to (a) Z9-14:OAc (b) Z7-12:OAc (c) Z9-12:OAc (d) E7-12:OAc and (e) Z11-16:OAc. Error bars indicate the range of the data, excluding the outliers (n = 5). Within each group, * = P\u0026lt;0.05,** = P\u0026gt;0.01, ***=P\u0026lt;0.001; ns = not significant\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-3778826/v1/54ca9f6d2cee209f4ee983ad.png"},{"id":51958747,"identity":"8c5a9df3-682d-40bf-a55e-6586c5a14381","added_by":"auto","created_at":"2024-03-04 15:18:05","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":790457,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-3778826/v1/115ab889-0a19-4766-a5a6-233abe1c41c1.pdf"},{"id":49158006,"identity":"dfbd73cc-db6e-41fc-8dbc-6712edb8ee63","added_by":"auto","created_at":"2024-01-04 04:44:23","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":16319,"visible":true,"origin":"","legend":"","description":"","filename":"Supplementarymaterials.docx","url":"https://assets-eu.researchsquare.com/files/rs-3778826/v1/3bb8260fd6ff495c428546a5.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Region-specific variation in the electrophysiological responses of Spodoptera frugiperda to synthetic sex pheromone compounds","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eThe fall armyworm, \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e (Lepidoptera, Noctuidae) is native to the Americas and currently feeds on a large variety of agricultural crops, belonging to 76 plant families (Montezano et al. \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). It was first found in Africa in 2016 (Goergen et al. 2016), likely facilitated by its high migratory behaviour (Westbrook et al. \u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Baudron et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Gilioli et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2023\u003c/span\u003e), human-assisted transport and commerce (Cock et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Since then, \u003cem\u003eS. frugiperda\u003c/em\u003e has been found in Asia in 2018 (Bhusal and Bhattarai \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Navasero et al. 2019; Sun et al. \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) and Australia in 2020 (IPPC, 2021; Paudel Timilsena et al. \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) and was recently found in the canary Island in Southern Europe (IPPC, 2021; Gilioli et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). In sub-Saharan Africa, its major host is maize (\u003cem\u003eZea mays\u003c/em\u003e L.), which is the most important staple food and is critical for food security of the continent (Pardey et al. \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Badu-Apraku and Fakorede \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; VIB, \u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eSince the arrival of \u003cem\u003eS. frugiperda\u003c/em\u003e in Africa, it was found in almost all sub-Saharan countries within a time span of two years (Rwomushana et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2018\u003c/span\u003e) and causes up to 21\u0026nbsp;million tonnes annual reduction in maize yield (Abrahams et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), representing about 52% annual production loss (Chimweta et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Thus, it is crucial to develop and implement evidence-based control measures for \u003cem\u003eS. frugiperda\u003c/em\u003e in Africa (Prasanna et al. \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe traditional approach of pesticide use has substantial environmental and human health issues as well as causing damage to non-target organisms, including natural enemies of insect pests (Desneux et al. \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; R\u0026eacute;gis\u0026ensp;Ahissou et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). Furthermore, success of pesticides is limited in controlling \u003cem\u003eS. frugiperda\u003c/em\u003e as the larvae tend to conceal themselves when feeding on the host plant by hiding in the maize whorl (Harrison et al. \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), and resistance has been developed against many of the cheapest and most widely used pesticides in Africa (Day et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Akeme et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). In addition, significant expenditures are incurred by both farmers and government in controlling infestation by \u003cem\u003eS. frugiperda\u003c/em\u003e (Kassie et al. \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), thus increasing the cost of crop production. Therefore, it is essential to develop an environmentally-friendly and more effective approach to combat the improper use of pesticides.\u003c/p\u003e \u003cp\u003eThe application of synthetic species-specific sex pheromones is a helpful tool in integrated pest management (IPM) for monitoring and early detection of \u003cem\u003eS. frugiperda\u003c/em\u003e (Prasanna et al. \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; FAO. 2019b; Matova et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). However, for a successful monitoring system, it is essential that lures are attractive. Thus, effective, local, regional and continental species-specific detection, monitoring and pest management should be developed as solid bases of IPM strategies (Saveer et al. \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). In the Western Hemisphere, \u003cem\u003eS. frugiperda\u003c/em\u003e is subdivided into two strains, the so-called corn strain and rice strain (Pashley et al. \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e1992\u003c/span\u003e; Lu and Adang \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e1996\u003c/span\u003e; Gouin et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), which differ by mitochondrial markers (Nagoshi \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2010\u003c/span\u003e) as well as by nuclear differences (Gouin et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). Even though some differences in sexual communication has been found between the strains (Unbehend et al. \u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e2013\u003c/span\u003e), intra-strain geographic variation in response to the signals has also been found (Unbehend et al. \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Furthermore, although the effluvia and gland pheromone extracts from calling \u003cem\u003eS. frugiperda\u003c/em\u003e females in the Western Hemisphere contain similar sex pheromone compounds as in Benin (Haenniger et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), the response and capture of \u003cem\u003eS. frugiperda\u003c/em\u003e males using commercial lures with some of these compounds have been erratic (Meagher et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Tepa-Yotto et al., \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe sex pheromone of \u003cem\u003eS. frugiperda\u003c/em\u003e in the Americas consists of (Z)-9-tetradecenyl acetate (Z9-14:OAc) and (Z)-7-dodecenyl acetate (Z7-12:OA) which have been found to be important for the attraction of conspecific males (Tumlinson et al. \u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e1986\u003c/span\u003e; Andrade et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Unbehend et al. \u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Unbehend et al. \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Jiang et al. 2021). Adding (Z)-11-hexadecenyl acetate (Z11-16:OAc) increased male attraction in Pennsylvania (Fleischer et al. 2015), and also elicited a small electroantennogram response to \u003cem\u003eS. frugiperda\u003c/em\u003e males in Yunnan Province (Jiang et al. 2021). In Brazil, (E)-7-dodecenyl acetate (E7-12:OAc) was identified as an additional active sex pheromone component (Batista-Pereira et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Cruz-Esteban et al. 2020). In Africa, \u003cem\u003eS. frugiperda\u003c/em\u003e monitoring studies with pheromone lures have revealed varying results (Meagher et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Koffi et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Tepa-Yotto et al. \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). For example, in Benin, when Tepa-Yotto et al. (\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) compared the attraction of \u003cem\u003eS. frugiperda\u003c/em\u003e males among three blends, i.e., a four-component blend containing Z9-14:OAc, Z7-12:OAc, Z11-16:OAc and (Z)-9-dodecenyl acetate (Z9-12:OAc), a three-component blend (without Z9-12:OAc) and a two-component blend containing only Z9-14:OAc and Z7-12:OAc, they found that the 4-component blend attracted the highest number of \u003cem\u003eS. frugiperda\u003c/em\u003e males, irrespective of the conditions of the experiment, while the 2-component blend was the most selective, as the percentage of bycatches was the lowest (Tepa-Yotto et al. \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Conversely, Koffi et al. (\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2021\u003c/span\u003e) found that the 3-component lure containing Z9-14:OAc, Z7-12:OAc, Z11-16:OAc was more attractive than the 4-component lure (adding Z9-12:OAc) in the neighboring country Togo. Thus, within African, male \u003cem\u003eS. frugiperda\u003c/em\u003e responses to female sex pheromones may differ.\u003c/p\u003e \u003cp\u003eTo assess whether males from different populations show different physiological responses to the identified sex pheromone compounds, we investigated the antennal responses of males from three African populations and an American population to all identified synthetic sex pheromone compounds. We also determined whether male responses varied depending on the two identified mitotypes of \u003cem\u003eS. frugiperda\u003c/em\u003e. Specifically, we evaluated variability in the antennal response of \u003cem\u003eS. frugiperda\u003c/em\u003e males that were collected as larvae from Florida, Benin, Nigeria and Kenya and from C- and R- males from Benin and Florida, to E7-12:OAc, Z7-12:OAc, Z9-12:OAc, Z9-14:OAc and Z11-16:OAc.\u003c/p\u003e"},{"header":"METHODS AND MATERIALS","content":"\u003cp\u003e \u003cb\u003eInsect Collection and Rearing.\u003c/b\u003e The \u003cem\u003eS. frugiperda\u003c/em\u003e populations from Benin originated from larvae, collected from maize fields and alternative host plants in South and Central Benin by the International Institute of Tropical Agriculture (IITA), Benin in July 2020, from which a laboratory population was developed at the University of Amsterdam. This population was mixed with field specimens from Azowlisse, Benin, which was collected in December 2020. In July 2021 and October 2022, new specimens from the lab population from IITA were mixed into the developed lab population at the University of Amsterdam. The Kenyan population was obtained from the rearing of the International Centre of Insect Physiology and Ecology (icipe, Kenya) in December 2020 and August 2022. The Nigerian population was collected from maize fields in Southern Oyo State and from IITA, Ibadan, Nigeria, between January 2022 and May 2022. To compare responses of African \u003cem\u003eS. frugiperda\u003c/em\u003e males to American \u003cem\u003eS. frugiperda\u003c/em\u003e males, Florida C- and R- strains were also reared at the University of Amsterdam, with populations obtained from the Max Planck Institute of Chemical Ecology, Jena, Germany. The immature stages (eggs and larvae) of all the populations were reared on an artificial pinto bean diet in climate chambers at 25\u0026deg;C and humidity of 60\u0026ndash;65%, with reversed light/dark cycle and 14:10 light/dark photoperiod at the laboratory at the University of Amsterdam (IBED). The adults were fed with 10% sugar water, \u003cem\u003ead libitum\u003c/em\u003e. Male and female insects were used for mitotype identification, while 2\u0026ndash;5 day old virgin male were used for gas chromatography-electroantennogram detector (GC-EAD) experiments.\u003c/p\u003e \u003cp\u003e \u003cb\u003eMitochondrial identification of African population.\u003c/b\u003e Since the invasive populations of \u003cem\u003eS. frugiperda\u003c/em\u003e are mixed in their nuclear genome, but do consist of two mitotypes (Yainnina et al. 2021; Tay et al. \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), we determined whether males with different mitotypes show different electrophysiological responses. Mitochondrial identification was done by screening adults for the mitochondrial marker (COI) that is diagnostic for both strains in North and South America (Nagoshi et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Nagoshi \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2010\u003c/span\u003e) and generally used in other populations as well (Tay et al. \u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). DNA extractions were performed in a 96-well plate using Chelex 100 Resin (Bio-Rad Laboratories, Hercules, CA, USA). One adult leg was put in one well together with two metal beads and 300\u0026micro;l 10% Chelex (diluted in ddH\u003csub\u003e2\u003c/sub\u003eO). The tissue was homogenized in a tissue lyser for 4min at 30Hz. Samples were heated for 30 min at 95\u0026deg;C and 300 rpm spinning, after which they were frozen at \u0026minus;\u0026thinsp;20\u0026deg;C overnight. Each plate was thawed, mixed, and centrifuged at 4000 rpm for 30min. The supernatant was filtered through a fritted deep well filter plate (Thermo Fisher Scientific, Waltham, MA, USA) and used for mitotype analysis. Identification of the mitochondrial marker was performed as described by Unbehend et al. (\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e2013\u003c/span\u003e) and summarized here. After amplification of the COI gene, two strain-specific digests with MspI and SacI were conducted to analyze the strain-affiliation via gel electrophoresis.\u003c/p\u003e \u003cp\u003eTo compare EAG responses between the mitotypes, we used both Florida (C) - and (R) - males and Benin (C)- and (R) - males. The Kenya males were not mitotyped and all the sampled populations from Nigeria were (R) - type. In our preliminary analysis, we observed no significant differences in the responses between Benin (R) - and Nigerian (R) - strain males.\u003c/p\u003e \u003cp\u003e \u003cb\u003ePreparation of the Multicomponent Blends.\u003c/b\u003e All used synthetic sex pheromone compounds of \u003cem\u003eS. frugiperda\u003c/em\u003e were purchased from Pherobank (Wijk bij Duurstede, The Netherlands), i.e., E7-12:OAc, Z7-12:OAc, Z9-12:OAc, Z9-14:OAc and Z11-16:OAc with \u0026gt;\u0026thinsp;98% purity. To determine the antennal responses to these compounds, two multicomponent blends (MCBs) were prepared, one with E7-12:OAc and one with Z7-12:OAc in combination with Z9-12:OAc, Z9-14:OAc, Z11-16:OAc (Supplementary Table \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e). This allowed us to distinguish responses between E and Z7-12:OAc.\u003c/p\u003e \u003cp\u003eA stock solution of 10 \u0026micro;g/\u0026micro;l in hexane was made for each synthetic pheromone compounds from which the two MCBs (i.e one including E7-12:OAc and the other including Z7-12:OAc) were made by diluting 1 \u0026micro;l each of the four synthetic pheromone compounds into 500 \u0026micro;l of hexane. Each of concentrations of 1 ng/\u0026micro;l, 3 ng/\u0026micro;l, 10 ng/\u0026micro;l and 20 ng/\u0026micro;l was obtained in a serial dilution and samples were put in the vials and kept in the refrigerator till the time of chemical analysis.\u003c/p\u003e \u003cp\u003e \u003cb\u003eGC-EAD measurements.\u003c/b\u003e To measure the electrophysiological responses of male antennae, the two MCBs in the four different concentrations were used in random order on 2 to 4-day-old virgin males from Benin, Kenya, Nigeria and Florida (C-type and R-type). Live male insect was individually placed in a plastic pipet tip and one antenna was immobilized with a small strip of parafilm pressing the antennal base against the head. Electrical contact was made using silver wires inserted in glass microelectrodes (GC150TF-10; Warner Instruments, Hamden, CT, USA) with insect Ringer\u0026rsquo;s solution. The recording electrode was inserted at the base of the antenna and the reference electrode made contact with the cut antennal tip. The amplitude of the EAG was measured using an IDAC-4 amplifier equipped with a high impedance (\u0026gt;\u0026thinsp;10\u003csup\u003e9\u003c/sup\u003e Ohm) head stage and recorded with GC-EAD/2014 software (Syntech, Kirchzarten, Germany). An Agilent 7890B gas chromatograph (Agilent, Wilmington, DE, USA) equipped with an Agilent Cool On-Column inlet, was coupled to the EAG setup to deliver odour stimuli. Details of the GC-EAD set up are in the supplementary S1.\u003c/p\u003e \u003cp\u003eTo check the longevity of the whole insect preparation, a reference stimulus (containing Z3-6:OH, 10\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e v/v (4.2 \u0026micro;g) in paraffin oil) was delivered for 0.5 seconds from a filter paper strip in a Pasteur pipet by a CS-55 stimulus controller (Syntech). The preparations proved to be extremely stable, as no significant differences were found between the sampled responses to the reference before and after the GC-EAG run (\u003cem\u003et\u003c/em\u003e-test, n\u0026thinsp;=\u0026thinsp;14, P\u0026thinsp;=\u0026thinsp;0.203, df\u0026thinsp;=\u0026thinsp;1).\u003c/p\u003e \u003cp\u003e \u003cb\u003eStatistics and Data analysis.\u003c/b\u003e To analyse the antennal responses to the pheromone compounds, we built linear mixed effect models (Lindstrom and Bates \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e1988\u003c/span\u003e) as described by Pinheiro and Bates (\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2000\u003c/span\u003e) and implemented in R (R Core Team \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) as function lme (Pinheiro et al. \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Mixed models, with individual insect as a random factor, and compound, concentration, population and (when available) mitotype as fixed factors, allowed us to separate the (random) variation between individual insect from the variation caused by the (fixed) factors of interest.\u003c/p\u003e \u003cp\u003eWe built 3 separate models (Supplementary Table S2). The first model contained compound, concentration and population as predictors, with 498 EAD responses from 25 male insects. A second model contained mitotype as explanatory variable. The model was fitted with 397 cases from 19 males for which mitotype information was available. Finally, since the results indicated that there was no significant difference between the African populations, a third model was constructed to compare EAD responses on a continental scale (Africa vs America). This dataset contained in total 697 EAD responses from 34 males.\u003c/p\u003e \u003cp\u003eFor each model, we started with a full model that contained the random factor, all the fixed factors and their interactions. Model selection of better fitting model was done using Akaike's Information criterium (AIC) (Sakamoto et al. \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e1986\u003c/span\u003e). All models were fit by maximum likelihood to allow model comparison using AIC. Post hoc comparisons of mean values from the final models was done with Least-squares means (Searle et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e1980\u003c/span\u003e) as implemented in the R package emmeans (Lenth, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2023\u003c/span\u003e) with Tukey multiplicity adjustments.\u003c/p\u003e"},{"header":"RESULTS","content":"\u003cp\u003e \u003cb\u003eElectrophysiological Responses to Synthetic Sex Pheromone Compounds\u003c/b\u003e. When checking the electrophysiological responses of \u003cem\u003eS. frugiperda\u003c/em\u003e males from the different regions, we found significant effects of geographic regions, the compounds, concentrations, as well as interactions between geographic region and compounds and between concentration and compound (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The major compound, Z9-14:OAc, and Z9-12:OAc elicited significant responses among the different regions (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea,c), as Florida male responses were higher than male responses from Benin, Nigeria and Kenya (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001). However, Benin, Nigerian and Kenyan males responded equally to Z9-14:OAc and Z9-12:OAc (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05). In all regions, \u003cem\u003eS. frugiperda\u003c/em\u003e males responded similarly to Z7-12:OAc, E7-12: OAc and Z11-16:OAc (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb,d,e) (P\u0026thinsp;\u0026gt;\u0026thinsp;0.05).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn assessing whether males responded differently to the two synthetic isomers, Z7-12:OAc and E7-12:OAc, we plotted the responses to both compounds from each region and at different concentrations in separate boxplots (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Overall, Z7-12:OAc gave higher antennal responses than E7-12:OAc in all the regions (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea-d). Also, only Florida and Nigerian males responded significantly to the two compounds at 1 ng/\u0026micro;l, while Kenyan males responded more to Z7-12:OAc than E7-12:OAc at 3 ng/\u0026micro;l. At higher concentrations (10 ng/\u0026micro;l and 20 ng/\u0026micro;l), males from all the regions (Florida, Benin, Nigeria and Kenya) showed significantly higher antennal responses to Z7-12:OAc than E7-12:OAc (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eSince there was no difference in male responses to the compounds in the three African populations (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), we combined these responses to a so-called \u0026lsquo;African\u0026rsquo; response and compared their overall additive responses to the Florida (American) responses. Our results revealed some differences across the two continents (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea-e). Specifically, EAG responses evoked by the major compound, Z9\u0026ndash;14:OAc, the critical secondary compound, Z7\u0026ndash;12:OAc and Z9\u0026ndash;12:OAc were significantly higher in American males than in African (Benin, Kenyan and Nigerian) males(P\u0026thinsp;\u0026lt;\u0026thinsp;0.001, P\u0026thinsp;=\u0026thinsp;0.029 and P\u0026thinsp;\u0026lt;\u0026thinsp;0.001, respectively). However, we found no differences in responses to E7-12:OAc and Z11-16:OAc between the continents.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWhen analysing variation in mitotyped male responses, we found a significant effect of compound, geographic region and an interaction between compound x strain x geographic region (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). Specifically, Florida (C)-type males showed significantly lower response to E7-12:OAc than Florida(R)-type males (P\u0026thinsp;=\u0026thinsp;0.025), while both mitotypes responded similarly to other compounds. Each of the five synthetic compounds also evoked similar responses in Benin (C)-type and (R)-type males. Between Benin (R)-type and Florida (R)-type males, we observed significant higher responses of Florida (R)-type males to Z9-14:OAc (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001), Z7-12:OAc (P\u0026thinsp;=\u0026thinsp;0.006) and Z9-12:OAc (P\u0026thinsp;=\u0026thinsp;0.0002) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea,b,c). Also, Florida (C)-type males showed significantly higher responses than Benin (C)-type males to Z9-14:OAc and Z9-12:OAc, respectively (P\u0026thinsp;\u0026lt;\u0026thinsp;0.001) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea,c).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eIn this study, we investigated the variation in electrophysiological responses of \u003cem\u003eS. frugiperda\u003c/em\u003e males from Florida, Benin, Nigeria and Kenya to sex pheromone compounds, and found a) some geographic variation in responses of males to Z9-14:OAc and Z9-12:OAc, b) significantly higher responses to Z7-12:OAc than E7-12:OAc in all males, c) continental differences in responses to some sex pheromones compounds, and d) some mitotype differences in responses, whereby Florida (R)-type males showed higher responses to E7-12:OAc than Florida (C)-type males and higher responses to Z7-12:OAc, Z9-14:OAc and Z9-12:OAc than Benin (R)-type males. Overall, our results show that Florida males were generally more responsive to sex pheromone compounds than the African males, while all African males were equally responsive to the pheromone compounds.\u003c/p\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eGeographic variation in male responses\u003c/h2\u003e \u003cp\u003eClearly, \u003cem\u003eS. frugiperda\u003c/em\u003e males from the different regions responded to all synthetic sex pheromone compounds, even though to a varying degree. Our finding that Florida males showed higher responses to the major compound, Z9-14:OAc than Benin, Nigerian and Kenyan males, corresponds to previous results by Haenniger et al. (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2020\u003c/span\u003e), where Florida males also elicited greater EAG responses to Z9-14:OAc than Benin and Nigerian males. Since it is widely agreed that Z9-14:OAc is the major sex pheromone component needed to effectively attract \u003cem\u003eS. frugiperda\u003c/em\u003e males (Tumlinson et al. \u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e1986\u003c/span\u003e; Andrade et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Meagher et al. 2013, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; H\u0026auml;nniger et al. 2020), it is surprising that African male responses are lower than those from Florida.\u003c/p\u003e \u003cp\u003eSimilarly, Z9-12:OAc elicited greater EAGs in Florida males than in Benin, Nigerian and Kenyan males. The activity of Z9-12:OAc as a pheromone component has been scarcely detected, although Tepa-Yotto et al. (\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) found that the addition of Z9-12:OAc to a four-component pheromone blend improved male attraction. Possibly, adding Z9-12:OAc to the sex pheromone blend increases male attraction in Africa.\u003c/p\u003e \u003cp\u003eA generally low EAG amplitude was observed in response to Z11-16:OAc in all the regions tested, which is consistent with previous findings (Malo et al. 2004; Unbehend et al. \u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Jiang et al. 2021). Z11-16:OAc doesn\u0026rsquo;t seem to be a sex pheromone component in American fields (Unbehend et al. \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) and may not increase male attraction in African fields (Tepa-Yotto et al. \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Moreover, the addition of Z11-16:OAc to lures caused large numbers of bycatches of non-target moths, particularly \u003cem\u003eMythimna loreyi\u003c/em\u003e (Duponchel) in West Africa (Meagher et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Tabata et al. \u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), indicating that \u003cem\u003eS. frugiperda\u003c/em\u003e lures in Africa should be developed without Z11-16:OAc.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eVariation in response to E7-12:OAc and Z7-12:OAc\u003c/h2\u003e \u003cp\u003eWe found interesting variations in male responses to the two isomeric compounds, Z7-12:OAc and E7-12:OAc across the concentrations tested. Overall, all \u003cem\u003eS. frugiperda\u003c/em\u003e males showed higher EAG responses to Z7-12:OAc than to its isomer, E7-12:OAc. So far, E7-12:OAc has been reported only within the female glands of Brazilian \u003cem\u003eS. frugiperda\u003c/em\u003e populations (Batista-Pereira et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2006\u003c/span\u003e) and more interestingly, was behaviorally active on males from that region (Batista-Pereira et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Cruz-Esteban et al. 2018). Whether E7-12:OAc is absent in female glands in African or other American regions is mostly still unclear, as is the male response in other regions. Unbehend et al. (\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e2014\u003c/span\u003e) did find that (C)-type males from Peru are only attracted to a blend containing Z7-12:OAc, but not to a blend with E7-12:OAc, while males in North Carolina did not differentiate between the two isomers. As chromatographic separation of the two isomers is difficult, it is possible that this compound has remained undetected in other studies. However, the lower EAG responses of the males to E7-12:OAc than Z7-12:OAc from all four regions suggests that the latter may be more important than the former in male attraction in all regions.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eVariation in response between Africa and Florida\u003c/h2\u003e \u003cp\u003eOur finding that \u003cem\u003eS. frugiperda\u003c/em\u003e males from Benin, Kenya and Nigeria showed similar EAD responses to all pheromone compounds, is comparable to the reports of Haenniger et al. (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2020\u003c/span\u003e) where Benin and Nigerian \u003cem\u003eS. frugiperda\u003c/em\u003e males exhibited similar EAG amplitudes to the five known sex pheromone synthetic compounds. These results suggest that \u003cem\u003eS. frugiperda\u003c/em\u003e males may not show geographic variation within the Africa continent. However, geographic variation between continents seems to occur, as we found intercontinental differences in the male responses towards Z9-14:OAc, Z7-12:OAc and Z9-12:OAc.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eVariation in inter-type and intra-type geographic males\u003c/h2\u003e \u003cp\u003eInterestingly, when comparing the mitotypes, we found that Florida (C)- and (R)-type males differed significantly in their response to E7-12:OAc and not any other pheromone compounds. The fact that there was no type-specific differential response to all but one pheromone compound in Florida males, and no type-specific differential response to any pheromone compound in Benin males suggests that males of both mitotypes have similar response range and are not differentiated with respect to antennal response at the doses tested. Field trapping experiments with Florida population also showed that both strains were similarly attracted to pheromone lures (Unbehend et al. \u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Unbehend et al. \u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Kenis et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Also in Kenya, both corn and rice mitotypes were equally attracted to the different commercial pheromone lures when tested in replicated field trials (Sisay et al. \u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn conclusion, we found that electrophysiological responses of \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e males to sex pheromone compounds differ between Florida and Africa population, but responses do not differ within the African continent. These results suggests that pheromone lures may have to be adjusted for monitoring of \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e in Africa compared to America, but do not have to be adjusted for specific regions within the African continent.\u003c/p\u003e \u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements:\u0026nbsp;\u003c/strong\u003eThe authors thank Dr Sabine Haenniger for donating larvae from the Florida \u003cem\u003eS. frugiperda\u003c/em\u003e colonies, Dr Olajumoke Alabi for her assistance in collecting the Nigerian specimens of the fall armyworm, Eileen Bader for rearing the insects at UvA, and Betsie Voetdijk and Peter Kuperus for their help in molecular analysis. We also thank Dr Jacques Deere and Ahmed G. Hussain for their help in data analysis.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003eThis work was supported by a grant from the Tertiary Education Trust Fund of the University of Ibadan, Nigeria to MDA, and by a grant provided by Simonis BV to ATG and RAHvS, and by the University of Amsterdam, The Netherlands.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflicts of interest/Competing interests\u0026nbsp;\u003c/strong\u003eThe authors declare that they have no conflict of interest/competing interests\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSupplementary Information S\u003c/strong\u003eupplementary information can be found on the online version of this article\u003c/p\u003e\n\u003cp\u003eAuthor Contributions: MDA, RAHvS, PR and ATG planned, conceived and prepared the study. MDA and PR did electrophysiology and analysed data. All authors wrote and edited the manuscript and approved the final version of the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u0026nbsp;\u003c/strong\u003eThe authors confirm that data supporting the findings of this study are available on request.\u003c/p\u003e\n"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eAbrahams P, Beale T, Cock M, Corniani N, Godwin J, Murphy S, Richard JV (2017) Fall armyworm status. Impacts and control options in Africa: Preliminary Evidence Note. https://www.cabi.org/Uploads/isc/Dfid%20Faw%20Inception%20Report04may2017final.pdf.\u003c/li\u003e\n\u003cli\u003eAkeme CN, Ngosong C, Sumbele SA, et al (2021) Different controlling methods of fall armyworm (\u003cem\u003eSpodoptera frugiperda\u003c/em\u003e) in maize farms of small-scale producers in Cameroon. 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Fall armyworm: impacts and implications for Africa. CABI. https://www.invasive-species.org/wp-content/uploads/sites/2/2019/02/FAW-Evidence-Note-October-2018.pdf\u003c/li\u003e\n\u003cli\u003eSakamoto Y, Ishiguro M, Kitagawa G (1986).\u003cem\u003eAkaike Information Criterion Statistics\u003c/em\u003e. D. Reidel Publishing Company.\u003c/li\u003e\n\u003cli\u003eSaveer AM, Hatano E, Wada-Katsumata A et al (2023) Nonanal, a new fall armyworm sex pheromone component, significantly increases the efficacy of pheromone lures. Pest Manag Sci 79:2831\u0026ndash;2839. https://doi.org/10.1002/ps.7460.\u003c/li\u003e\n\u003cli\u003eSearle S R., Speed F M, Milliken G (1980) Population Marginal Means in the Linear Model: An Alternative to Least Squares Means. Am Stat 34: 216-221. doi:10.1080/00031305.1980.10483031.\u003c/li\u003e\n\u003cli\u003eSisay B, Subramanian S, Weldon CW, et al (2024) Evaluation of pheromone lures, trap designs and placement heights for monitoring the fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae) in maize fields of Kenya. Crop Prot 176:106523. https://doi.org/10.1016/j.cropro.2023.106523.\u003c/li\u003e\n\u003cli\u003eSun X, Hu C, Jia H, et al (2021) Case study on the first immigration of fall armyworm, \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e invading into China. J Integr Agric 20:664\u0026ndash;672. https://doi.org/10.1016/S2095-3119(19)62839-X\u003c/li\u003e\n\u003cli\u003eTabata J, Nakano R, Yasui H, et al (2022) Sex pheromone of the fall armyworm, Spodoptera frugiperda: identification of a trace component that enhances attractiveness and specificity. Entomol Exp Appl : https://doi.org/10.1111/eea.13287.\u003c/li\u003e\n\u003cli\u003eTay WT, Meagher RL, Czepak C, Groot AT (2023) \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e : Ecology, Evolution, and Management Options of an Invasive Species. Annu Rev Entomol 68:299\u0026ndash;317. https://doi.org/10.1146/annurev-ento-120220-102548.\u003c/li\u003e\n\u003cli\u003eTay WT,Rane R, Padovan A,Walsh T, Elfekih S, et al. (2022) Global population genomic signature of fall armyworm supports complex introduction events across the Old World. Commun Biol5:297.https://doi.org/10.1038/s42003-022-03230-1 \u003c/li\u003e\n\u003cli\u003eTepa-Yotto GT, Meagher RL, Winsou JK, et al (2022) Monitoring \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e in Benin: assessing the influence of trap type, pheromone blends, and habitat on pheromone trapping. Fla Entomol 105:. https://doi.org/10.1653/024.105.0111\u003c/li\u003e\n\u003cli\u003eTumlinson JH, Mitchell ER, Teal PEA, Heath RR and Mengelkoch LJ (1986) Sex pheromone of fall armyworm, \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e (J.E. Smith) identification of components critical to attraction in the field. J Chem Ecol 12:1909\u0026ndash;1926. \u003c/li\u003e\n\u003cli\u003eUnbehend M, Haenniger S, Meagher RL, Heckel DG, Groot AT (2013) Pheromonal divergence between two strains of\u0026ensp;\u003cem\u003eSpodoptera frugiperda\u003c/em\u003e. J Chem Ecol 39:364\u0026ndash;376.\u003c/li\u003e\n\u003cli\u003eUnbehend M, Haenniger S, Vasquez GM, et al (2014) Geographic Variation in Sexual Attraction of Spodoptera frugiperda Corn- and Rice-Strain Males to Pheromone Lures. PLoS ONE 9:e89255. https://doi.org/10.1371/journal.pone.0089255.\u003c/li\u003e\n\u003cli\u003eVIB (2017). Maize in Africa. Internationl Plant Biotechnology Outreach. https://doi.org/10.1088/0305-4470/31/34/016.\u003c/li\u003e\n\u003cli\u003eWestbrook JK, Nagoshi RN, Meagher RL, et al (2016) Modeling seasonal migration of fall armyworm moths. Int J Biometeorol 60:255\u0026ndash;267. https://doi.org/10.1007/s00484-015-1022-x.\u003c/li\u003e\n\u003cli\u003eYainna S, N\u0026egrave;gre N, Silvie PJ, Br\u0026eacute;vault T, Tay WT, et al. (2021). Geographic monitoring of insecticide resistance mutations in native and invasive populations of the fall armyworm. Insects12:468\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-chemical-ecology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"joce","sideBox":"Learn more about [Journal of Chemical Ecology](https://www.springer.com/journal/10886)","snPcode":"10886","submissionUrl":"https://submission.nature.com/new-submission/10886/3","title":"Journal of Chemical Ecology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Fall armyworm, electroantennogram, sex pheromone compounds, mitotype","lastPublishedDoi":"10.21203/rs.3.rs-3778826/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-3778826/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThe fall armyworm, \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e (J.E. Smith), is a global pest that feeds on \u0026gt;\u0026thinsp;350 plant species and causes major yield loses. Variation in the responses of \u003cem\u003eS. frugiperda\u003c/em\u003e males to female sex pheromone compounds affects the detection, monitoring and management of the pest. We determined geographic variation in the responses of \u003cem\u003eS. frugiperda\u003c/em\u003e males to four different doses of synthetic sex pheromone compounds using a gas chromatography-electroantennogram detector (GC-EAD). Furthermore, we disentangled regional populations into C- and R- mitotypes via molecular analysis of the cytochrome oxidase I gene, and measured their responses to the compounds. When comparing responses of males from Florida, Benin, Nigeria and Kenya, we found some regional differences in the responses of \u003cem\u003eS. frugiperda\u003c/em\u003e males to the major compound, Z9-14:OAc and minor component Z9-12:OAc. However, we found no differences in male responses from the different African countries. All males showed significantly higher antennal responses to Z7-12:OAc than to E7-12:OAc. When comparing the mitotypes, we found that Florida R-type males showed higher responses to Z9-14:OAc, Z7-12:OAc and Z9-12:OAc than Benin R-type males, while C-type males from both regions responded equally to Z7-12:OAc. In addition, Florida R-type males showed higher responses to E7-12:OAc than Florida C-type males. Our study thus shows some differential physiological responses of \u003cem\u003eSpodoptera frugiperda\u003c/em\u003e males towards the known sex pheromone compounds, including E7-12:OAc, but mostly in the different mitotypes. How these differences translate to field trap catches remains to be determined.\u003c/p\u003e","manuscriptTitle":"Region-specific variation in the electrophysiological responses of Spodoptera frugiperda to synthetic sex pheromone compounds","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-01-04 04:44:19","doi":"10.21203/rs.3.rs-3778826/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"checksComplete","content":"","date":"2023-12-27T11:43:52+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Chemical Ecology","date":"2023-12-19T22:33:28+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-chemical-ecology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"joce","sideBox":"Learn more about [Journal of Chemical Ecology](https://www.springer.com/journal/10886)","snPcode":"10886","submissionUrl":"https://submission.nature.com/new-submission/10886/3","title":"Journal of Chemical Ecology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"59b7d9cd-288d-4606-ab43-28b88f0d18b5","owner":[],"postedDate":"January 4th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2024-03-04T15:15:08+00:00","versionOfRecord":{"articleIdentity":"rs-3778826","link":"https://doi.org/10.1007/s10886-024-01479-w","journal":{"identity":"journal-of-chemical-ecology","isVorOnly":false,"title":"Journal of Chemical Ecology"},"publishedOn":"2024-02-29 15:01:34","publishedOnDateReadable":"February 29th, 2024"},"versionCreatedAt":"2024-01-04 04:44:19","video":"","vorDoi":"10.1007/s10886-024-01479-w","vorDoiUrl":"https://doi.org/10.1007/s10886-024-01479-w","workflowStages":[]},"version":"v1","identity":"rs-3778826","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3778826","identity":"rs-3778826","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}