Space oddity: Absence of prezygotic-premating barriers in Eurydema lundbladi and Eurydema ornata

Space oddity: Absence of prezygotic-premating barriers in Eurydema lundbladi and Eurydema ornata | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Space oddity: Absence of prezygotic-premating barriers in Eurydema lundbladi and Eurydema ornata Mario Alamo, Diego Gil-Tapetado This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5341557/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 19 Dec, 2025 Read the published version in Journal of Insect Conservation → Version 1 posted 9 You are reading this latest preprint version Abstract Understanding the effectiveness of premating prezygotic reproductive barriers in contact zones of closely related lineages is essential for assessing hybridization risks. This study documents the first overlap zone with interspecific copulations on La Palma Island, Canary Islands, between the Macaronesian endemic Eurydema lundbladi Lindberg, 1960 and the widespread Palearctic species Eurydema ornata (Linnaeus, 1758). We analyzed morphological differences in male genitalia, climatic niches, and altitudinal distributions of both species. Notably, the differences in male genital structures do not appear sufficient to cause copulatory incompatibility, nor do size variations act as limiting factors for mating. The ecological niches of both species, while distinct, converge in certain altitudinal zones, where climatic conditions–particularly winter temperatures–significantly influence their distribution. These weak and convergent premating prezygotic reproductive barriers underscore the conservation risks faced by E. lundbladi in light of the potential expansion and competition from E. ornata . Implications for insect conservation: The genetic integrity of E. lundbladi is threatened by the encroachment of E. ornata into previously unoccupied areas. Continued monitoring of contact zones and future studies are essential to evaluate the impact of these interactions on the conservation of this endemic species. Interspecific copulation reproductive barriers Contact zone Canary Islands endemic species hybridization risks Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Introduction Prezygotic barriers play a critical role in maintaining the genetic integrity and evolutionary identity of species (Kirkpatrick and Ravigné 2002 ; Merrill et al. 2024 ). Various prezygotic isolation mechanisms act at different stages, both pre-copulatory (e.g., spatial, ecological, temporal, behavioral isolation) and post-copulatory (e.g., mechanical, gametic barriers) (Eady 2001 ). However, reproductive isolation is often maintained by a combination of barriers (Coyne and Orr 2004 ). Identifying the effective number of reproductive barriers can help determine which mechanisms disproportionately contribute to the cessation of gene flow within a lineage (Sanchez-Guillen et al. 2012 ). In parapatric contact zones, where the ranges of different species overlap, these reproductive barriers are essential to prevent hybridization and to avoid the energy costs associated with inter-lineage interactions, which have implications for species survival (Panova et al. 2006 ; Scharf and Martin 2013). The effectiveness and number of these barriers are largely influenced by the ecological context and the specific dynamics of the contact zones (Jiggins and Mallet 2000 ). Ecological barriers, often the first to arise, are crucial for speciation, as divergent selection driven by distinct habitats can restrict gene flow between populations, facilitating evolutionary divergence (Schluter 2009 ; McBride and Singer 2010 ). In closely related species that share similar ecological niches, incomplete reproductive isolation can result in gene flow within these overlap zones (Henrich and Kalbe 2016 ). This is especially the case when morphological barriers, such as genitalia differences in insects, do not provide sufficient isolation between distinct evolutionary lineages (Langton-Myers and Buckley 2019). However, when these species occupy distinct ecological niches under divergent selective pressure, niche divergence becomes key in maintaining evolutionary differentiation, even in the presence of hybrid zones (Liu et al. 2018 ). Species contact zones are not static; they can shift in response to environmental and behavioral selective pressures, which may compromise the effectiveness of reproductive barriers. Environmental changes, such as rising temperatures, can alter key behaviors like courtship (Macchiano et al. 2019 ). Geographic shifts in overlap zones can also result from anthropogenic pressures, such as land-use changes (Aguillon and Rohwer 2022 ), or from new contact zones driven by climate change (Larson et al. 2019 ). Additionally, sexual selection, particularly mate choice, may be altered in these contexts, changing reproductive dynamics and the stability of prezygotic barriers (Silva 2014 ). These dynamic processes illustrate how fluctuations in contact zones can erode isolation barriers, promote gene flow, and offer deeper insights into the mechanisms of speciation. Islands provide ideal conditions for speciation, as geographic isolation promotes genetic divergence (Gillespie and Roderick 2002 ). Geographic barriers, combined with the environmental heterogeneity generated by mountainous elevations, promote the development of endemic species of special interest (Steinbauer et al. 2012 ). However, these species, often adapted to specific altitudinal and insular niches, are particularly vulnerable to genetic introgression from introduced species or closely related lineages, which may dilute their gene pool and compromise their evolutionary integrity. This risk is evident in the introgression of genes from lowland species into thermally specialized high-mountain species (Gómez et al. 2015 ), or from widely distributed generalist species into island endemics, affecting their genetic stability and persistence (Pasachnik et al. 2009 ). In the Canary Islands, the genus Eurydema Laporte, 1833 (Heteroptera: Pentatomidae) is represented by two species: Eurydema ornata and Eurydema lundbladi . Eurydema ornata has a Western Palearctic distribution and is a generalist phytophagous species, primarily feeding on various species of the Brassicaceae family (Kívan and Kiliç 2000 ). In contrast, E. lundbladi has a much more restricted range, confined to the mountainous regions of the Macaronesian archipelagos, specifically on the islands of La Palma and Tenerife in the Canary Islands (Arechavaleta et al. 2010), as well as on Madeira and Porto Santo (Borges et al. 2008 ). However, there is some taxonomic uncertainty regarding the populations from the Canary Islands and Madeira (Lupoli 2018 ). Its diet appears to be more specialized; however, limited information is available, with existing records on Erysimum scoparium (Brouss. ex Willd.) Wetts. and species within the genus Descurainia (Aukema et al. 2013 ). Nevertheless, the role of these ecological differences in maintaining reproductive isolation between these evolutionary units remains unknown. Until recently, the existence of overlapping zones where both species coexist was largely unrecognized, raising questions about the extent of potential interspecific interactions. In the mountainous areas of the Canary Islands, E. ornata and E. lundbladi coexist within overlapping altitudinal ranges, suggesting a potential for interspecific interactions, including hybridization. Investigating these interactions can clarify the taxonomic relationships between the two species and reveal how environmental pressures and reproductive isolation mechanisms may influence the boundaries of their distributions. This research is particularly relevant in the context of conservation, as E. lundbladi , with its narrower range and ecological specialization, could be at risk if hybridization disrupts its population structure or if E. ornata displaces it from its niche. The main objective of this study is to assess the role of current prezygotic barriers in maintaining reproductive isolation between these two previously known allopatric species, focusing on spatial (geographical), morphological (genitalia), and ecological factors as they converge within an altitudinal overlap zone on La Palma, Canary Islands. Specifically, this study will: 1) analyze and compare the genital structures of E. ornata and E. lundbladi populations on La Palma. Notably, the only existing description of E. lundbladi male genitalia comes from Lindberg ( 1960 ), which covered only the pygophore. This study will provide the first detailed description and images of the internal genital structures of E. lundbladi ; 2) explore the realized climatic niches of both species; and 3) examine the altitudinal limits of their distributions to better understand how their ecological requirements converge. The ecological differentiation between E. ornata and E. lundbladi suggests that niche specialization plays a pivotal role in reinforcing reproductive isolation. By investigating both the climatic niches and the altitudinal distribution limits of these species, we aim to evaluate the degree of niche convergence. Such ecological barriers are likely key drivers of divergence, particularly for E. lundbladi , whose limited distribution and specialization make it more vulnerable to environmental fluctuations. Material and Methods Sampling and genitalia preparation of Eurydema individuals Sampling was carried out at Pico de la Nieve, part of the Caldera de Taburiente National Park in La Palma, Canary Islands (28.451N 17.531W). A total of 4 interspecific copulation events between E. ornata and E. lundbladi were recorded. The first interspecific copulation event was recorded on 3 May 2023 (Fig. S1 , Supplementary Information) at 1877 m. In this case, only this E. ornata individual was detected and recorded in the area. On 25 May 2024, three specimens of E. ornata were found, all of which were copulating with E. lundbladi , two at 1877 m (Fig. 1 a, b) and one at 2030 m (Fig. 1 c). Specimens recorded in 2024 were preserved and deposited in the entomological collection of the National Museum of Natural Sciences (MNCN), Madrid, Spain. Spain (Canary Islands), La Palma (Pico de la Nieve), 28.436N, 17.492W, 1877 m, 25 May 2024, copulation on Descurainia gilva , leg. M. Alamo; E. ornata (♂) and E. lundbladi (♀), [MNCN_Ent_395677] (Fig. 1 a). Spain (Canary Islands), La Palma (Pico de la Nieve), 28.436N, 17.492W, 1877 m, 25 May 2024, copulation on Descurainia gilva , leg. M. Alamo; E. ornata (♀) and E. lundbladi (♂), [MNCN_Ent_395678] (Fig. 1 b). Spain (Canary Islands), La Palma (Pico de la Nieve), 28.436N, 17.495W, 2033 m, 25 May 2024, copulation on Descurainia gilva , leg. M. Alamo; E. ornata (♀) and E. lundbladi (♂), [MNCN_Ent_395679] (Fig. 1 c). For the study of genitalia, we collected a total of 20 E. ornata and 10 E. lundbladi male specimens. Of the E. ornata specimens, 10 were sampled from Puntallana, La Palma, and 10 from Dehesa Boyal, San Sebastián de los Reyes (Madrid), in order to compare potential differences between island and mainland populations. The 10 E. lundbladi specimens were collected from Pico de la Nieve, La Palma. Male genitalia (pygophores) were treated with a 10% NaOH solution for 48 hours. The internal contents were cleared by thoroughly rinsing them in distilled water before dissection. Photographs were taken using a Leica DFC 450 camera mounted on a Leica M165C stereozoom microscope, and the images were processed using LAS X software. The terminology for genital morphology follows Dupuis ( 1949 ) Zhao et al. ( 2017 ) and Salini ( 2019 ). Geographical and ecological analyses We used the available records of E. lundbladi and E. ornata of the Global Biodiversity Information Facility (GBIF) to perform the altitudinal and climatic niche analyses (GBIF 2024a, 2024b, 2024c). We downloaded both the worldwide and the Canary Islands distribution of E. ornata . For the distribution of E. lundbladi in Macaronesia we have only obtained its distribution in the Canary Islands, focusing mainly on the records from La Palma and Tenerife. We consider the only record found in Las Palmas de Gran Canaria to be doubtful and prefer not to include it in the subsequent analyses. Also, we have filtered the data by removing dubious records and those that are clearly in the water or approximating those that are close to the shore. Finally, we reduced the records of the two Eurydema species on a 1x1 km grid to one record per grid. Maps were built in ArcGIS for Desktop v 10.8 (ESRI 2019 ). To calculate the altitude range, the climatic niche of both species and their overlapping areas, we used 19 bioclimatic variables and elevation data from WorldClim (Fick and Hijmans 2017 ). To account for correlations between environmental variables (both climatic and altitudinal), we conducted a hierarchical cluster analysis based on the correlation matrix. This resulted in a dendrogram showing the similarity between variables (Dormann et al. 2013), using Ward’s method (Harrell 2001). We set the distance threshold for clustering at 0.3, indicating a 70% correlation between variables (e.g., Gil-Tapetado et al. 2024 ). For each cluster, we selected the variable with the greatest ecological relevance for the species, prioritizing the more derived or seasonally precise variable in cases of uncertainty (Fig. S2, Supplementary Information). Next, we calculated the variance inflation factor (VIF) (Lin et al. 2011), to identify and eliminate variables that overestimated variance and provided redundant information (VIF > 5) (Stine 1995; Miles 2014). Finally, we selected three variables to represent the climatic niches of E. lundbladi and E. ornata in Canary Islands: temperature annual range (Bio7), mean temperature of the coldest quarter (Bio11) and precipitation seasonality (Bio15) (Fig. S2, Supplementary Information). Variable selection was performed with the R v 3.5.0 program, using RStudio software v 1.1.453 (RStudio Team 2023) with the dismo (Hijmans et al. 2017) and HH (Heiberger 2015) packages. We compared the altitudinal range of E. lundbladi and E. ornata in the Canary Islands using an ANOVA with Tukey contrasts among pairs. We also added the global elevation range of E. ornata to show if there was a difference with the island distribution. We represented these altitudinal distributions in violin plots. To quantify the climatic niche overlap, niche equivalency, and niche similarity of E. lundbladi and E. ornata in the Canary Islands, we follow the methodology of previous studies (Wiens and Graham 2005; Broennimann et al. 2012; Guisan et al. 2014). The degree of niche overlap between periods was calculated using Schoener's D index (Schoener 1970), which ranges from 0 (no overlap between niches) to 1 (complete overlap). This approach aligns with the methodology proposed by Broennimann et al. (2012). It involves calculating two key metrics: niche equivalency, which assesses whether the niches of both species overlap when occurrences are randomly reallocated, and niche similarity, which gauges whether the overlap between observed niches differs from the overlap between the observed niche and randomly selected niches. We also calculated the proportion of overlapping points between the two niches and the ratio (or relative size) of one niche to the other. This niche overlapping was represented by ellipses of the two climatic niches, which were composed of the three selected variables. The statistical analyses were conducted using the ecospat (Di Cola et al. 2017) and ellipsenm package (Cobos et al. 2022 ) in RStudio. Results Morphological description and difference of Eurydema of Canary Island Eurydema ornata male genitalia . Pygophore . Parandria deeply bilobed, ending in two sharp teeth separated by a deep incisure, forming a right angle between them. Inner parandrial tooth 2/3 the height of the outer parandrial tooth. Processus transverse bilobed, sinuate with elevated, blunt tips and slight constriction (Fig. 2 a, b). Paramere . Hook-shaped, with flattened crown forming a concave indentation. Long, sharp apex, perpendicular to the main longitudinal axis. Proximal section straight; distal section markedly curved inward, forming a pronounced angle with the straight proximal part. Short stem and weakly sclerotized apodeme (Fig. 2 c, d). Phallus . Evaginated, with a pair of basolateral conjunctival lobes strongly sclerotized, elongated, and curved towards the apex, where a soft indentation is present on the inner margin. Aedeagus . Extending from the center of the median penial plates, longer than the median plates (Fig. 2 e). No structural differences were observed between populations from Madrid and La Palma. Eurydema lundbladi male genitalia . Pygophore . Parandria deeply bilobed, ending in two sharp teeth separated by a deep incisure, causing the teeth to appear almost individualized, forming an acute angle between them. Inner parandrial tooth 1/2 the height of the outer parandrial tooth. Processus transverse bilobed, sinuate with blunt tips, less elevated and constricted than in E. ornata (Fig. 3 a, b). Paramere . Hook-shaped, with flattened crown forming a concave indentation. Long, sharp apex, perpendicular to the main longitudinal axis. Proximal section straight, continuous with the base; distal section also straight but thinner and slightly inclined, without the pronounced curve seen in E. ornata . Short stem, thinner than in E. ornata , and weakly sclerotized apodeme (Fig. 3 c, d). Phallus . Evaginated, with a pair of basolateral conjunctival lobes sclerotized, elongated, and curved towards the apex, where a soft indentation is present on the inner margin. Aedeagus . Extending from the center of the median penial plates, longer than the median plates but shorter than in E. ornata (Fig. 3 e). Altitudinal range and climatic niche of Eurydema of Canary Island We obtain significative differences among the altitudinal distribution of E. lundbladi and E. ornata in the two islands of Canary Islands (t = -30.06 p < 2e-16) (Fig. 4 ). The two species show an almost complementary distribution, with E. lundbladi being an upland species, averaging 2002.95 masl, while E. ornata is a more coastal species, averaging 555.34 masl (Fig. 5 ). There is also a significant difference in mean altitude between Canary Island and global E. ornata (228.58 masl; t = -12.79 p < 2e-16). In any case, E. ornata always seems to prefer lower altitudes to E. lundbladi . Although E. lundbladi and E. ornata showed significant differences in the mean of their altitudinal range and appear to have almost complementary distributions, there are records of E. lundbladi below the mean altitude or records of E. ornata above the mean that might appear to be outliers, but which coincide with the altitudes where we have found interspecific copulations. There are more records of E. ornata near the mean altitude of E. lundbladi than vice versa. The climatic niches of E. lundbladi and E. ornata are not equivalent and similar in addition to having a low proportion of overlapping (Table 1 ). Considering the climatic niches of these two species (Fig. 6 ), E. lundbladi can be found in different areas with annual temperature ranges, but only in areas with a limited winter temperature and rainfall range, coinciding with mountain areas in the Canary Islands; whereas E. ornata is found in different areas with variable winter temperatures, but limited rainfall range, coinciding with mountain areas in the Canary Islands; whereas E. ornata is found in different areas with variable winter temperatures, but limited rainfall range, coinciding with mountain areas in the Canary Islands; whereas E. ornata is found in different areas with variable winter temperatures, but limited rainfall range. Table 1 Calculated parameters and values of overlapping climatic niche of E. lundbladi and E. ornata Parameter Value Total points 7486 Overlapped points 172 Overlap 0.023 D' Schoener 0.095 Proportion niche ornata vs lundbladi 8.36 Proportion niche lundbladi vs ornata 0.12 Similarity (p) 1.0 Equivalency (p) 0.495 Overlapping areas niche of both species of Eurydema is characterized by a relatively high precipitation seasonality (~ 85 mm), a relatively low mean temperature in winter (~ 4–8 ºC) and also a relatively low temperature range (~ 15 ºC). This niche characterizes the intermediate altitude areas where we have indeed found the interspecific copulations of E. lundbladi and E. ornata . Discussion Reproductive barriers are crucial for maintaining genetic isolation within contact zones of closely related lineages. This study documents the first known contact zone between the endemic E. lundbladi of the Canary Islands and the widespread E. ornata , emphasizing the reproductive barriers that may inhibit gene flow between these two evolutionary lineages. The occurrence of copulation events involving individuals from both species underscores the potential risk of genetic exchange, threatening the genetic integrity of the endemic species. Subtle morphological differences have been identified in the male genitalia of both species. While their ecological niches are relatively well-defined, they exhibit convergence that results in overlapping altitudinal distribution limits (i.e.: the spatial prezygotic barriers have disappeared). In this context, we assess the effectiveness of the pre-copulatory prezygotic barriers and the potential risks to the future integrity and maintenance of both lineages. Differences in male genital structures between E. lundbladi and E. ornata do not appear to be sufficient to cause copulatory incompatibility, nor do variations in body size between males and females of both species act as limiting factors. Interspecific copulation among heteropterans is relatively common (e.g. Kon et al. 1993 ; Shapiro et al. 2010 ) and can sometimes lead to successful fertilization (Hamel et al. 2018 ) and hybrid formation (Bugaj-Nawrocka 2020; Pinotti et al. 2021 ), highlighting the potential weakness of prezygotic-precopulating barriers, such as mechanical incompatibilities or species recognition, in closely related lineages. In some species, traits such as pronotum width are used as cues for species recognition during mating, helping to distinguish between conspecific and heterospecific individuals (Hickman et al. 2021). However, for E. lundbladi and E. ornata , pronotum width does not appear to function as a species recognition barrier, as conspicuous differences in size (e.g.: see Fig. 1 B) are insufficient to prevent interspecific mating attempts. Consequently, postcopulatory prezygotic and postzygotic barriers may play a more significant role in maintaining reproductive isolation between these species. To assess the risk of hybridization and the type of contact zone formed—where hybrids may be rare and a bimodal hybrid zone could emerge (Jiggins and Mallet 2000 )—it is essential to investigate whether effective interspecific copulation occurs, explore the functionality of the spermatheca, and evaluate reproductive success to determine the full effectiveness of prezygotic and postzygotic barriers. The ecological niches of E. lundbladi and E. ornata are distinct, though they overlap in certain altitudinal zones. Winter temperatures, the annual temperature range, and precipitation seasonality play significant roles in shaping the realized niches of both species. Eurydema species undergo diapause during colder months (Benedek 1967 ), a process regulated by winter conditions, particularly minimum temperatures, which affect female reproductive success and offspring survival in overwintering heteropterans, such as Nezara viridula (Musolin 2007 ). This thermal dependence suggests that E. ornata may be constrained to lower-altitude environments, where E. lundbladi , as a thermal specialist, exhibits greater adaptability. While ecological divergence has historically contributed to genetic isolation between these species (Jiggins et al. 2000), climate change could reduce these ecological barriers, potentially increasing the likelihood of gene flow between the two lineages. Although the altitudinal distributions of the two species appear to be complementary, there are overlapping areas at the lower distributional limits of E. lundbladi and the upper limits of E. ornata , where interspecific copulations may occur. Given the wide distribution and abundance of E. ornata in thermophilic zones (Stankevych et al. 2021 ), sexual competition could arise in these intermediate contact zones, where females may be subject to mating attempts from both species, regardless of whether effective copulation occurs. This could reduce the number of successful mating pairs for E. lundbladi , potentially displacing the endemic species to higher elevations where E. ornata is absent. Similar cases have been documented in the genus Nezara , where more thermophilic and abundant species expand their ranges in response to increases in temperatures during the colder months, leading to the displacement of less competitive species and competition for mating partners (Yukawa et al. 2007 ). The altitudinal distribution representation of the two Eurydema species (see Figure X) shows a tendency for E. ornata individuals to inhabit higher altitudes than E. lundbladi at lower altitudes. This, coupled with the generalist character of E. ornata , suggests that this species may be undergoing an altitudinal expansion, occupying areas previously inhabited only by E. lundbladi . Additionally, in a scenario of climate change, it is possible that regions that were previously unsuitable for E. ornata due to their low temperatures may now become suitable as temperatures have risen. This phenomenon appears to be occurring with other insect species. This phenomenon has been observed in other insect species (Gil-Tapetado et al. 2023 , 2024 ). In phytophagous insects, fidelity to the host plant can act as a barrier to encounter and reproductive isolation (Matsubayashi et al. 2010 ), where attraction to chemical cues (Piersanti et al. 2020 ) and nutritional requirements influence phytophagous survival (Auclair 1969 ). However, for a trophic generalist species like E. ornata , the host plant might not be a limiting factor or a reproductive barrier. Nonetheless, it would be necessary to assess the viability of E. ornata under the nutritional resources provided by the host plants of E. lundbladi . Additionally, in a climate change scenario, the alpine and subalpine host plants of E. lundbladi could experience a regression, potentially lead a reduction in the distribution of E. lundbladi , as has been observed with the sensitivity of Erysimum scoparium to high temperatures (González-Rodríguez et al. 2021 ; Jackson and Chapman 2024). Recent phylogenetic analyses have shown that E. ornata and E. lundbladi belong to the tribe Strachiini; however, their phylogenetic relationships remain unresolved, resulting in a polytomy involving E. maracandica (Roca-Cusachs et al. 2022 ). This evolutionary ambiguity contrasts with their ecological differentiation, as both species exhibit distinct niches and reproductive barriers. Nonetheless, under the influence of climate change, the prezygotic barriers currently maintaining genetic isolation could weaken, potentially facilitating gene flow between these lineages. To conserve the Macaronesian endemic E. lundbladi , it is crucial to conduct future-focused studies on distribution limits, particularly in light of the recent emergence of sympatry with E. ornata . Long-term monitoring of their contact zones is essential to evaluate the progression of interspecific copulations and the risk of hybridization. Although an increase in interspecific encounters has been observed during the two sampling periods, further research is needed to analyze the viability and reproductive success of immigrant individuals and to understand the potential consequences for the genetic integrity of E. lundbladi . Declarations Author Contribution All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by M.A. and D.GT. The first draft of the manuscript was written by M.A. and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. Acknowledgement We would like to extend our sincere thanks to the professors and the director of the Master's Program in Conservation of Tropical Areas at the Menéndez Pelayo International University and the Consejo Superior de Investigaciones Científicas (CSIC), whose organization of the field trips to the Canary Islands made these findings possible. It is important to note that the observations presented here were made during fieldwork conducted as part of a university master’s program, which significantly enhanced the observational and sampling efforts in underexplored yet common areas. We would also like to thank all the generators of georeferenced biogeographic data and information who contribute data altruistically to the GBIF repository. We are also grateful to Mercedes Paris for her assistance in the examination of genital structures, and to Manuel Baena for his contributions and expertise to the Hemiptera group. The title of this article was inspired in the song Space Oddity of David Bowie because these interspecific copulations are found only in a part of the altitudinal range and seem to be a singularity of these areas and because these areas were found on Pico de la Nieve, near Roque de los Muchachos, the astronomical observatory where different European solar and nocturnal telescopes are located. 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Dissertation, Universidade de Lisboa. http://hdl.handle.net/10451/15315 Stankevych S, Zabrodina I, Yushchuk D, Dolya M, Balan H, Yakovlev R, Kosylovych H, Holiachuk Y, Zakharchuk N, Galagan T, Nemerytska L, Zhuravska I, Romanov O, Romanova T, Bragin O, Hudym O, Hordiienko I (2021) Eurydema bugs: Review of distribution, ecology, harmfulness, and control. Ukrainian J Ecol 11(9):131–149 Steinbauer MJ, Otto R, Naranjo-Cigala A, Beierkuhnlein C, Fernández‐Palacios JM (2012) Increase of island endemism with altitude–speciation processes on oceanic islands. Ecography 35(1):23–32. https://doi.org/10.1111/j.1600-0587.2011.07064.x Yukawa J, Kiritani K, Gyoutoku N, Uechi N, Yamaguchi D, Kamitani S (2007) Distribution range shift of two allied species, Nezara viridula and N. antennata (Hemiptera: Pentatomidae), in Japan, possibly due to global warming. Appl Entomol Zool 42(2):205–215. https://doi.org/10.1303/aez.2007.205 Zhao W, Zhao Q, Li M, Wei J, Zhang X, Zhang H (2017) DNA barcoding of Chinese species of the genus Eurydema Laporte, 1833 (Hemiptera: Pentatomidae). Zootaxa 4286(2):151–175. https://doi.org/10.11646/zootaxa.4286.2.1 Additional Declarations No competing interests reported. Supplementary Files SupplementaryInformation.docx Cite Share Download PDF Status: Published Journal Publication published 19 Dec, 2025 Read the published version in Journal of Insect Conservation → Version 1 posted Editorial decision: Revision requested 30 Mar, 2025 Reviews received at journal 07 Mar, 2025 Reviewers agreed at journal 23 Feb, 2025 Reviews received at journal 18 Feb, 2025 Reviewers agreed at journal 01 Feb, 2025 Reviewers invited by journal 03 Nov, 2024 Editor assigned by journal 28 Oct, 2024 Submission checks completed at journal 28 Oct, 2024 First submitted to journal 27 Oct, 2024 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. 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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-5341557","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":373500560,"identity":"a978f58e-2ab6-4d2d-beaa-97192f7edaf0","order_by":0,"name":"Mario Alamo","email":"data:image/png;base64,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","orcid":"","institution":"Menéndez Pelayo International University","correspondingAuthor":true,"prefix":"","firstName":"Mario","middleName":"","lastName":"Alamo","suffix":""},{"id":373500561,"identity":"0e0e7011-029a-4af4-a919-832cf372215e","order_by":1,"name":"Diego Gil-Tapetado","email":"","orcid":"","institution":"Menéndez Pelayo International University","correspondingAuthor":false,"prefix":"","firstName":"Diego","middleName":"","lastName":"Gil-Tapetado","suffix":""}],"badges":[],"createdAt":"2024-10-27 13:53:09","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5341557/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5341557/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s10841-025-00742-z","type":"published","date":"2025-12-19T15:57:56+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":68457453,"identity":"f0cb1f4e-79c8-448e-8b56-55370fccb97c","added_by":"auto","created_at":"2024-11-07 12:55:45","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":10910105,"visible":true,"origin":"","legend":"\u003cp\u003eInterspecific copulation recorded at Pico de la Nieve, La Palma, Canary Islands: A. \u003cem\u003eE. lundbladi\u003c/em\u003e ♀ and \u003cem\u003eE. ornata\u003c/em\u003e ♂; B. \u003cem\u003eE. lundbladi\u003c/em\u003e ♂ and \u003cem\u003eE. ornata\u003c/em\u003e ♀; C. \u003cem\u003eE. lundbladi\u003c/em\u003e ♂ and \u003cem\u003eE. ornata\u003c/em\u003e ♀. Photographs by A.M.\u003c/p\u003e","description":"","filename":"Fig1.png","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/e92b1c6116ceb15c42b2eeb4.png"},{"id":68457454,"identity":"86b96e45-c031-49f8-beb2-bd0bc1ede195","added_by":"auto","created_at":"2024-11-07 12:55:45","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":10744348,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eEurydema ornata\u003c/em\u003e male genitalia. \u003cstrong\u003eA\u003c/strong\u003e. Pygophore dorsal view; \u003cstrong\u003eB\u003c/strong\u003e. Pygophore dorsal view; \u003cstrong\u003eC\u003c/strong\u003e. Paramere dorso-lateral view; \u003cstrong\u003eD\u003c/strong\u003e. Paramere ventro-lateral view; \u003cstrong\u003eE\u003c/strong\u003e. Phallus ventral view. Photographs by A.M.\u003c/p\u003e","description":"","filename":"Fig2.png","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/2db38cef12660946b9662763.png"},{"id":68456130,"identity":"1c5c178e-33ba-4d92-88f8-64ea09cde9b4","added_by":"auto","created_at":"2024-11-07 12:47:45","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":9881795,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eEurydema lundbladi\u003c/em\u003e male genitalia. \u003cstrong\u003eA\u003c/strong\u003e. Pygophore dorsal view; \u003cstrong\u003eB\u003c/strong\u003e. Pygophore dorsal view; \u003cstrong\u003eC\u003c/strong\u003e. Paramere dorso-lateral view; \u003cstrong\u003eD\u003c/strong\u003e. Paramere ventro-lateral view; \u003cstrong\u003eE\u003c/strong\u003e. Phallus ventral view. Photographs by M.A.\u003c/p\u003e","description":"","filename":"Fig3.png","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/51b7584cc570a4e510fa0148.png"},{"id":68457451,"identity":"ff79efdf-5ad6-4fb2-b76b-d055376d4507","added_by":"auto","created_at":"2024-11-07 12:55:45","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":2174321,"visible":true,"origin":"","legend":"\u003cp\u003eAltitudinal distribution map of \u003cem\u003eEurydema ornata\u003c/em\u003e (red) and \u003cem\u003eEurydema lundbladi\u003c/em\u003e (blue) in the Canary Islands. Altitude is shown on the map as low (purple), medium (yellow) and high (green)\u003c/p\u003e","description":"","filename":"Fig4.png","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/2e04f62f77b6134d63a4ccf1.png"},{"id":68457853,"identity":"1be9592c-b70e-4d2c-8f98-e39f9f02d2d6","added_by":"auto","created_at":"2024-11-07 13:03:45","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":285400,"visible":true,"origin":"","legend":"\u003cp\u003eViolin plots representing the density of occurrences across altitudinal distribution ranges for \u003cem\u003eEurydema lundbladi\u003c/em\u003e (grey) and \u003cem\u003eEurydema ornata\u003c/em\u003e in the Canary Islands (red), alongside the global altitudinal range of \u003cem\u003eEurydema ornata\u003c/em\u003e (orange)\u003c/p\u003e","description":"","filename":"Fig5.png","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/b0cabbc428a0fbbd14b34fc4.png"},{"id":68456124,"identity":"fffcf50d-e94f-48f7-b136-26587e3ae13c","added_by":"auto","created_at":"2024-11-07 12:47:45","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":915615,"visible":true,"origin":"","legend":"\u003cp\u003eSpatial representation of the realized niches (ellipsoids) of \u003cem\u003eEurydema ornata\u003c/em\u003e (red) and \u003cem\u003eEurydema lundbladi\u003c/em\u003e (blue) based on the climatic variables BIO11 (Mean Temperature of Coldest Quarter), BIO7 (Temperature Annual Range), and BIO15 (Precipitation Seasonality). The niche overlap zone is depicted in yellow and green\u003c/p\u003e","description":"","filename":"Fig6.png","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/eb015987e8b0733e5ae3326a.png"},{"id":98814803,"identity":"ce190384-d12d-489f-9e00-66bab7dc997b","added_by":"auto","created_at":"2025-12-22 16:12:50","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":31935566,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/e63a65ec-30fb-4a99-99b5-0ff21e8160a1.pdf"},{"id":68456128,"identity":"540c3458-8e8a-4bb8-9b6b-a8120c2d0129","added_by":"auto","created_at":"2024-11-07 12:47:45","extension":"docx","order_by":9,"title":"","display":"","copyAsset":false,"role":"supplement","size":2774732,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryInformation.docx","url":"https://assets-eu.researchsquare.com/files/rs-5341557/v1/e0697f4984594c94d230a06f.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Space oddity: Absence of prezygotic-premating barriers in Eurydema lundbladi and Eurydema ornata","fulltext":[{"header":"Introduction","content":"\u003cp\u003ePrezygotic barriers play a critical role in maintaining the genetic integrity and evolutionary identity of species (Kirkpatrick and Ravign\u0026eacute; \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2002\u003c/span\u003e; Merrill et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Various prezygotic isolation mechanisms act at different stages, both pre-copulatory (e.g., spatial, ecological, temporal, behavioral isolation) and post-copulatory (e.g., mechanical, gametic barriers) (Eady \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2001\u003c/span\u003e). However, reproductive isolation is often maintained by a combination of barriers (Coyne and Orr \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2004\u003c/span\u003e). Identifying the effective number of reproductive barriers can help determine which mechanisms disproportionately contribute to the cessation of gene flow within a lineage (Sanchez-Guillen et al. \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2012\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn parapatric contact zones, where the ranges of different species overlap, these reproductive barriers are essential to prevent hybridization and to avoid the energy costs associated with inter-lineage interactions, which have implications for species survival (Panova et al. \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Scharf and Martin 2013). The effectiveness and number of these barriers are largely influenced by the ecological context and the specific dynamics of the contact zones (Jiggins and Mallet \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Ecological barriers, often the first to arise, are crucial for speciation, as divergent selection driven by distinct habitats can restrict gene flow between populations, facilitating evolutionary divergence (Schluter \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; McBride and Singer \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). In closely related species that share similar ecological niches, incomplete reproductive isolation can result in gene flow within these overlap zones (Henrich and Kalbe \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). This is especially the case when morphological barriers, such as genitalia differences in insects, do not provide sufficient isolation between distinct evolutionary lineages (Langton-Myers and Buckley 2019). However, when these species occupy distinct ecological niches under divergent selective pressure, niche divergence becomes key in maintaining evolutionary differentiation, even in the presence of hybrid zones (Liu et al. \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2018\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eSpecies contact zones are not static; they can shift in response to environmental and behavioral selective pressures, which may compromise the effectiveness of reproductive barriers. Environmental changes, such as rising temperatures, can alter key behaviors like courtship (Macchiano et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Geographic shifts in overlap zones can also result from anthropogenic pressures, such as land-use changes (Aguillon and Rohwer \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), or from new contact zones driven by climate change (Larson et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Additionally, sexual selection, particularly mate choice, may be altered in these contexts, changing reproductive dynamics and the stability of prezygotic barriers (Silva \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). These dynamic processes illustrate how fluctuations in contact zones can erode isolation barriers, promote gene flow, and offer deeper insights into the mechanisms of speciation.\u003c/p\u003e \u003cp\u003eIslands provide ideal conditions for speciation, as geographic isolation promotes genetic divergence (Gillespie and Roderick \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2002\u003c/span\u003e). Geographic barriers, combined with the environmental heterogeneity generated by mountainous elevations, promote the development of endemic species of special interest (Steinbauer et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). However, these species, often adapted to specific altitudinal and insular niches, are particularly vulnerable to genetic introgression from introduced species or closely related lineages, which may dilute their gene pool and compromise their evolutionary integrity. This risk is evident in the introgression of genes from lowland species into thermally specialized high-mountain species (G\u0026oacute;mez et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), or from widely distributed generalist species into island endemics, affecting their genetic stability and persistence (Pasachnik et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2009\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn the Canary Islands, the genus \u003cem\u003eEurydema\u003c/em\u003e Laporte, 1833 (Heteroptera: Pentatomidae) is represented by two species: \u003cem\u003eEurydema ornata\u003c/em\u003e and \u003cem\u003eEurydema lundbladi\u003c/em\u003e. \u003cem\u003eEurydema ornata\u003c/em\u003e has a Western Palearctic distribution and is a generalist phytophagous species, primarily feeding on various species of the \u003cem\u003eBrassicaceae\u003c/em\u003e family (K\u0026iacute;van and Kili\u0026ccedil; \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). In contrast, \u003cem\u003eE. lundbladi\u003c/em\u003e has a much more restricted range, confined to the mountainous regions of the Macaronesian archipelagos, specifically on the islands of La Palma and Tenerife in the Canary Islands (Arechavaleta et al. 2010), as well as on Madeira and Porto Santo (Borges et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). However, there is some taxonomic uncertainty regarding the populations from the Canary Islands and Madeira (Lupoli \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). Its diet appears to be more specialized; however, limited information is available, with existing records on \u003cem\u003eErysimum scoparium\u003c/em\u003e (Brouss. ex Willd.) Wetts. and species within the genus \u003cem\u003eDescurainia\u003c/em\u003e (Aukema et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Nevertheless, the role of these ecological differences in maintaining reproductive isolation between these evolutionary units remains unknown. Until recently, the existence of overlapping zones where both species coexist was largely unrecognized, raising questions about the extent of potential interspecific interactions.\u003c/p\u003e \u003cp\u003eIn the mountainous areas of the Canary Islands, \u003cem\u003eE. ornata\u003c/em\u003e and \u003cem\u003eE. lundbladi\u003c/em\u003e coexist within overlapping altitudinal ranges, suggesting a potential for interspecific interactions, including hybridization. Investigating these interactions can clarify the taxonomic relationships between the two species and reveal how environmental pressures and reproductive isolation mechanisms may influence the boundaries of their distributions. This research is particularly relevant in the context of conservation, as \u003cem\u003eE. lundbladi\u003c/em\u003e, with its narrower range and ecological specialization, could be at risk if hybridization disrupts its population structure or if \u003cem\u003eE. ornata\u003c/em\u003e displaces it from its niche.\u003c/p\u003e \u003cp\u003eThe main objective of this study is to assess the role of current prezygotic barriers in maintaining reproductive isolation between these two previously known allopatric species, focusing on spatial (geographical), morphological (genitalia), and ecological factors as they converge within an altitudinal overlap zone on La Palma, Canary Islands. Specifically, this study will: 1) analyze and compare the genital structures of \u003cem\u003eE. ornata\u003c/em\u003e and \u003cem\u003eE. lundbladi\u003c/em\u003e populations on La Palma. Notably, the only existing description of \u003cem\u003eE. lundbladi\u003c/em\u003e male genitalia comes from Lindberg (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e1960\u003c/span\u003e), which covered only the pygophore. This study will provide the first detailed description and images of the internal genital structures of \u003cem\u003eE. lundbladi\u003c/em\u003e; 2) explore the realized climatic niches of both species; and 3) examine the altitudinal limits of their distributions to better understand how their ecological requirements converge. The ecological differentiation between \u003cem\u003eE. ornata\u003c/em\u003e and \u003cem\u003eE. lundbladi\u003c/em\u003e suggests that niche specialization plays a pivotal role in reinforcing reproductive isolation. By investigating both the climatic niches and the altitudinal distribution limits of these species, we aim to evaluate the degree of niche convergence. Such ecological barriers are likely key drivers of divergence, particularly for \u003cem\u003eE. lundbladi\u003c/em\u003e, whose limited distribution and specialization make it more vulnerable to environmental fluctuations.\u003c/p\u003e"},{"header":"Material and Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eSampling and genitalia preparation of Eurydema individuals\u003c/h2\u003e \u003cp\u003eSampling was carried out at Pico de la Nieve, part of the Caldera de Taburiente National Park in La Palma, Canary Islands (28.451N 17.531W). A total of 4 interspecific copulation events between \u003cem\u003eE. ornata\u003c/em\u003e and \u003cem\u003eE. lundbladi\u003c/em\u003e were recorded. The first interspecific copulation event was recorded on 3 May 2023 (Fig. \u003cspan refid=\"MOESM1\" class=\"InternalRef\"\u003eS1\u003c/span\u003e, Supplementary Information) at 1877 m. In this case, only this \u003cem\u003eE. ornata\u003c/em\u003e individual was detected and recorded in the area. On 25 May 2024, three specimens of \u003cem\u003eE. ornata\u003c/em\u003e were found, all of which were copulating with \u003cem\u003eE. lundbladi\u003c/em\u003e, two at 1877 m (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea, b) and one at 2030 m (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec). Specimens recorded in 2024 were preserved and deposited in the entomological collection of the National Museum of Natural Sciences (MNCN), Madrid, Spain.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eSpain\u003c/strong\u003e \u003cp\u003e(Canary Islands), La Palma (Pico de la Nieve), 28.436N, 17.492W, 1877 m, 25 May 2024, copulation on \u003cem\u003eDescurainia gilva\u003c/em\u003e, leg. M. Alamo; \u003cem\u003eE. ornata\u003c/em\u003e (♂) and \u003cem\u003eE. lundbladi\u003c/em\u003e (♀), [MNCN_Ent_395677] (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea).\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eSpain\u003c/strong\u003e \u003cp\u003e(Canary Islands), La Palma (Pico de la Nieve), 28.436N, 17.492W, 1877 m, 25 May 2024, copulation on \u003cem\u003eDescurainia gilva\u003c/em\u003e, leg. M. Alamo; \u003cem\u003eE. ornata\u003c/em\u003e (♀) and \u003cem\u003eE. lundbladi\u003c/em\u003e (♂), [MNCN_Ent_395678] (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb).\u003c/p\u003e \u003c/p\u003e \u003cp\u003e \u003cstrong\u003eSpain\u003c/strong\u003e \u003cp\u003e(Canary Islands), La Palma (Pico de la Nieve), 28.436N, 17.495W, 2033 m, 25 May 2024, copulation on \u003cem\u003eDescurainia gilva\u003c/em\u003e, leg. M. Alamo; \u003cem\u003eE. ornata\u003c/em\u003e (♀) and \u003cem\u003eE. lundbladi\u003c/em\u003e (♂), [MNCN_Ent_395679] (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec).\u003c/p\u003e \u003c/p\u003e \u003cp\u003eFor the study of genitalia, we collected a total of 20 \u003cem\u003eE. ornata\u003c/em\u003e and 10 \u003cem\u003eE. lundbladi\u003c/em\u003e male specimens. Of the \u003cem\u003eE. ornata\u003c/em\u003e specimens, 10 were sampled from Puntallana, La Palma, and 10 from Dehesa Boyal, San Sebasti\u0026aacute;n de los Reyes (Madrid), in order to compare potential differences between island and mainland populations. The 10 \u003cem\u003eE. lundbladi\u003c/em\u003e specimens were collected from Pico de la Nieve, La Palma.\u003c/p\u003e \u003cp\u003eMale genitalia (pygophores) were treated with a 10% NaOH solution for 48 hours. The internal contents were cleared by thoroughly rinsing them in distilled water before dissection. Photographs were taken using a Leica DFC 450 camera mounted on a Leica M165C stereozoom microscope, and the images were processed using LAS X software. The terminology for genital morphology follows Dupuis (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1949\u003c/span\u003e) Zhao et al. (\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e2017\u003c/span\u003e) and Salini (\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eGeographical and ecological analyses\u003c/h3\u003e\n\u003cp\u003eWe used the available records of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e of the Global Biodiversity Information Facility (GBIF) to perform the altitudinal and climatic niche analyses (GBIF 2024a, 2024b, 2024c). We downloaded both the worldwide and the Canary Islands distribution of \u003cem\u003eE. ornata\u003c/em\u003e. For the distribution of \u003cem\u003eE. lundbladi\u003c/em\u003e in Macaronesia we have only obtained its distribution in the Canary Islands, focusing mainly on the records from La Palma and Tenerife. We consider the only record found in Las Palmas de Gran Canaria to be doubtful and prefer not to include it in the subsequent analyses. Also, we have filtered the data by removing dubious records and those that are clearly in the water or approximating those that are close to the shore. Finally, we reduced the records of the two \u003cem\u003eEurydema\u003c/em\u003e species on a 1x1 km grid to one record per grid. Maps were built in ArcGIS for Desktop v 10.8 (ESRI \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2019\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eTo calculate the altitude range, the climatic niche of both species and their overlapping areas, we used 19 bioclimatic variables and elevation data from WorldClim (Fick and Hijmans \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2017\u003c/span\u003e). To account for correlations between environmental variables (both climatic and altitudinal), we conducted a hierarchical cluster analysis based on the correlation matrix. This resulted in a dendrogram showing the similarity between variables (Dormann et al. 2013), using Ward\u0026rsquo;s method (Harrell 2001). We set the distance threshold for clustering at 0.3, indicating a 70% correlation between variables (e.g., Gil-Tapetado et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). For each cluster, we selected the variable with the greatest ecological relevance for the species, prioritizing the more derived or seasonally precise variable in cases of uncertainty (Fig. S2, Supplementary Information). Next, we calculated the variance inflation factor (VIF) (Lin et al. 2011), to identify and eliminate variables that overestimated variance and provided redundant information (VIF\u0026thinsp;\u0026gt;\u0026thinsp;5) (Stine 1995; Miles 2014). Finally, we selected three variables to represent the climatic niches of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e in Canary Islands: temperature annual range (Bio7), mean temperature of the coldest quarter (Bio11) and precipitation seasonality (Bio15) (Fig. S2, Supplementary Information). Variable selection was performed with the R v 3.5.0 program, using RStudio software v 1.1.453 (RStudio Team 2023) with the \u003cem\u003edismo\u003c/em\u003e (Hijmans et al. 2017) and \u003cem\u003eHH\u003c/em\u003e (Heiberger 2015) packages.\u003c/p\u003e \u003cp\u003eWe compared the altitudinal range of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e in the Canary Islands using an ANOVA with Tukey contrasts among pairs. We also added the global elevation range of \u003cem\u003eE. ornata\u003c/em\u003e to show if there was a difference with the island distribution. We represented these altitudinal distributions in violin plots. To quantify the climatic niche overlap, niche equivalency, and niche similarity of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e in the Canary Islands, we follow the methodology of previous studies (Wiens and Graham 2005; Broennimann et al. 2012; Guisan et al. 2014). The degree of niche overlap between periods was calculated using Schoener's D index (Schoener 1970), which ranges from 0 (no overlap between niches) to 1 (complete overlap). This approach aligns with the methodology proposed by Broennimann et al. (2012). It involves calculating two key metrics: niche equivalency, which assesses whether the niches of both species overlap when occurrences are randomly reallocated, and niche similarity, which gauges whether the overlap between observed niches differs from the overlap between the observed niche and randomly selected niches. We also calculated the proportion of overlapping points between the two niches and the ratio (or relative size) of one niche to the other. This niche overlapping was represented by ellipses of the two climatic niches, which were composed of the three selected variables. The statistical analyses were conducted using the \u003cem\u003eecospat\u003c/em\u003e (Di Cola et al. 2017) and \u003cem\u003eellipsenm\u003c/em\u003e package (Cobos et al. \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2022\u003c/span\u003e) in RStudio.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e\u003cem\u003eMorphological description and difference of Eurydema of Canary Island\u003c/em\u003e\u003c/h2\u003e \u003cp\u003e \u003cb\u003eEurydema ornata\u003c/b\u003e \u003cb\u003emale genitalia\u003c/b\u003e. \u003cb\u003ePygophore\u003c/b\u003e. Parandria deeply bilobed, ending in two sharp teeth separated by a deep incisure, forming a right angle between them. Inner parandrial tooth 2/3 the height of the outer parandrial tooth. \u003cem\u003eProcessus transverse\u003c/em\u003e bilobed, sinuate with elevated, blunt tips and slight constriction (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea, b). \u003cb\u003eParamere\u003c/b\u003e. Hook-shaped, with flattened crown forming a concave indentation. Long, sharp apex, perpendicular to the main longitudinal axis. Proximal section straight; distal section markedly curved inward, forming a pronounced angle with the straight proximal part. Short stem and weakly sclerotized apodeme (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec, d). \u003cb\u003ePhallus\u003c/b\u003e. Evaginated, with a pair of basolateral conjunctival lobes strongly sclerotized, elongated, and curved towards the apex, where a soft indentation is present on the inner margin. \u003cem\u003eAedeagus\u003c/em\u003e. Extending from the center of the median penial plates, longer than the median plates (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ee). No structural differences were observed between populations from Madrid and La Palma.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eEurydema lundbladi\u003c/b\u003e \u003cb\u003emale genitalia\u003c/b\u003e. \u003cb\u003ePygophore\u003c/b\u003e. Parandria deeply bilobed, ending in two sharp teeth separated by a deep incisure, causing the teeth to appear almost individualized, forming an acute angle between them. Inner parandrial tooth 1/2 the height of the outer parandrial tooth. \u003cem\u003eProcessus transverse\u003c/em\u003e bilobed, sinuate with blunt tips, less elevated and constricted than in \u003cem\u003eE. ornata\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea, b). \u003cb\u003eParamere\u003c/b\u003e. Hook-shaped, with flattened crown forming a concave indentation. Long, sharp apex, perpendicular to the main longitudinal axis. Proximal section straight, continuous with the base; distal section also straight but thinner and slightly inclined, without the pronounced curve seen in \u003cem\u003eE. ornata\u003c/em\u003e. Short stem, thinner than in \u003cem\u003eE. ornata\u003c/em\u003e, and weakly sclerotized apodeme (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ec, d). \u003cb\u003ePhallus\u003c/b\u003e. Evaginated, with a pair of basolateral conjunctival lobes sclerotized, elongated, and curved towards the apex, where a soft indentation is present on the inner margin. \u003cem\u003eAedeagus\u003c/em\u003e. Extending from the center of the median penial plates, longer than the median plates but shorter than in \u003cem\u003eE. ornata\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ee).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003e\u003c/h3\u003e\n\u003cdiv class=\"Heading\"\u003e\u003cem\u003eAltitudinal range and climatic niche of Eurydema of Canary Island\u003c/em\u003e\u003c/div\u003e \u003cp\u003eWe obtain significative differences among the altitudinal distribution of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e in the two islands of Canary Islands (t = -30.06 p\u0026thinsp;\u0026lt;\u0026thinsp;2e-16) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). The two species show an almost complementary distribution, with \u003cem\u003eE. lundbladi\u003c/em\u003e being an upland species, averaging 2002.95 masl, while \u003cem\u003eE. ornata\u003c/em\u003e is a more coastal species, averaging 555.34 masl (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). There is also a significant difference in mean altitude between Canary Island and global \u003cem\u003eE. ornata\u003c/em\u003e (228.58 masl; t = -12.79 p\u0026thinsp;\u0026lt;\u0026thinsp;2e-16). In any case, \u003cem\u003eE. ornata\u003c/em\u003e always seems to prefer lower altitudes to \u003cem\u003eE. lundbladi\u003c/em\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eAlthough \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e showed significant differences in the mean of their altitudinal range and appear to have almost complementary distributions, there are records of \u003cem\u003eE. lundbladi\u003c/em\u003e below the mean altitude or records of \u003cem\u003eE. ornata\u003c/em\u003e above the mean that might appear to be outliers, but which coincide with the altitudes where we have found interspecific copulations. There are more records of \u003cem\u003eE. ornata\u003c/em\u003e near the mean altitude of \u003cem\u003eE. lundbladi\u003c/em\u003e than vice versa.\u003c/p\u003e \u003cp\u003eThe climatic niches of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e are not equivalent and similar in addition to having a low proportion of overlapping (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Considering the climatic niches of these two species (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e), \u003cem\u003eE. lundbladi\u003c/em\u003e can be found in different areas with annual temperature ranges, but only in areas with a limited winter temperature and rainfall range, coinciding with mountain areas in the Canary Islands; whereas \u003cem\u003eE. ornata\u003c/em\u003e is found in different areas with variable winter temperatures, but limited rainfall range, coinciding with mountain areas in the Canary Islands; whereas \u003cem\u003eE. ornata\u003c/em\u003e is found in different areas with variable winter temperatures, but limited rainfall range, coinciding with mountain areas in the Canary Islands; whereas \u003cem\u003eE. ornata\u003c/em\u003e is found in different areas with variable winter temperatures, but limited rainfall range.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eCalculated parameters and values of overlapping climatic niche of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"2\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eParameter\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eValue\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eTotal points\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e7486\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eOverlapped points\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e172\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eOverlap\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.023\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eD' Schoener\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.095\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eProportion niche ornata vs lundbladi\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e8.36\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eProportion niche lundbladi vs ornata\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.12\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSimilarity (p)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1.0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eEquivalency (p)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0.495\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eOverlapping areas niche of both species of \u003cem\u003eEurydema\u003c/em\u003e is characterized by a relatively high precipitation seasonality (~\u0026thinsp;85 mm), a relatively low mean temperature in winter (~\u0026thinsp;4\u0026ndash;8 \u0026ordm;C) and also a relatively low temperature range (~\u0026thinsp;15 \u0026ordm;C). This niche characterizes the intermediate altitude areas where we have indeed found the interspecific copulations of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eReproductive barriers are crucial for maintaining genetic isolation within contact zones of closely related lineages. This study documents the first known contact zone between the endemic \u003cem\u003eE. lundbladi\u003c/em\u003e of the Canary Islands and the widespread \u003cem\u003eE. ornata\u003c/em\u003e, emphasizing the reproductive barriers that may inhibit gene flow between these two evolutionary lineages. The occurrence of copulation events involving individuals from both species underscores the potential risk of genetic exchange, threatening the genetic integrity of the endemic species. Subtle morphological differences have been identified in the male genitalia of both species. While their ecological niches are relatively well-defined, they exhibit convergence that results in overlapping altitudinal distribution limits (i.e.: the spatial prezygotic barriers have disappeared). In this context, we assess the effectiveness of the pre-copulatory prezygotic barriers and the potential risks to the future integrity and maintenance of both lineages.\u003c/p\u003e \u003cp\u003eDifferences in male genital structures between \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e do not appear to be sufficient to cause copulatory incompatibility, nor do variations in body size between males and females of both species act as limiting factors. Interspecific copulation among heteropterans is relatively common (e.g. Kon et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e1993\u003c/span\u003e; Shapiro et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2010\u003c/span\u003e) and can sometimes lead to successful fertilization (Hamel et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2018\u003c/span\u003e) and hybrid formation (Bugaj-Nawrocka 2020; Pinotti et al. \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), highlighting the potential weakness of prezygotic-precopulating barriers, such as mechanical incompatibilities or species recognition, in closely related lineages. In some species, traits such as pronotum width are used as cues for species recognition during mating, helping to distinguish between conspecific and heterospecific individuals (Hickman et al. 2021). However, for \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e, pronotum width does not appear to function as a species recognition barrier, as conspicuous differences in size (e.g.: see Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB) are insufficient to prevent interspecific mating attempts. Consequently, postcopulatory prezygotic and postzygotic barriers may play a more significant role in maintaining reproductive isolation between these species. To assess the risk of hybridization and the type of contact zone formed\u0026mdash;where hybrids may be rare and a bimodal hybrid zone could emerge (Jiggins and Mallet \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2000\u003c/span\u003e)\u0026mdash;it is essential to investigate whether effective interspecific copulation occurs, explore the functionality of the spermatheca, and evaluate reproductive success to determine the full effectiveness of prezygotic and postzygotic barriers.\u003c/p\u003e \u003cp\u003eThe ecological niches of \u003cem\u003eE. lundbladi\u003c/em\u003e and \u003cem\u003eE. ornata\u003c/em\u003e are distinct, though they overlap in certain altitudinal zones. Winter temperatures, the annual temperature range, and precipitation seasonality play significant roles in shaping the realized niches of both species. \u003cem\u003eEurydema\u003c/em\u003e species undergo diapause during colder months (Benedek \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e1967\u003c/span\u003e), a process regulated by winter conditions, particularly minimum temperatures, which affect female reproductive success and offspring survival in overwintering heteropterans, such as \u003cem\u003eNezara viridula\u003c/em\u003e (Musolin \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). This thermal dependence suggests that \u003cem\u003eE. ornata\u003c/em\u003e may be constrained to lower-altitude environments, where \u003cem\u003eE. lundbladi\u003c/em\u003e, as a thermal specialist, exhibits greater adaptability. While ecological divergence has historically contributed to genetic isolation between these species (Jiggins et al. 2000), climate change could reduce these ecological barriers, potentially increasing the likelihood of gene flow between the two lineages.\u003c/p\u003e \u003cp\u003eAlthough the altitudinal distributions of the two species appear to be complementary, there are overlapping areas at the lower distributional limits of \u003cem\u003eE. lundbladi\u003c/em\u003e and the upper limits of \u003cem\u003eE. ornata\u003c/em\u003e, where interspecific copulations may occur. Given the wide distribution and abundance of \u003cem\u003eE. ornata\u003c/em\u003e in thermophilic zones (Stankevych et al. \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2021\u003c/span\u003e), sexual competition could arise in these intermediate contact zones, where females may be subject to mating attempts from both species, regardless of whether effective copulation occurs. This could reduce the number of successful mating pairs for \u003cem\u003eE. lundbladi\u003c/em\u003e, potentially displacing the endemic species to higher elevations where \u003cem\u003eE. ornata\u003c/em\u003e is absent. Similar cases have been documented in the genus \u003cem\u003eNezara\u003c/em\u003e, where more thermophilic and abundant species expand their ranges in response to increases in temperatures during the colder months, leading to the displacement of less competitive species and competition for mating partners (Yukawa et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). The altitudinal distribution representation of the two \u003cem\u003eEurydema\u003c/em\u003e species (see Figure X) shows a tendency for \u003cem\u003eE. ornata\u003c/em\u003e individuals to inhabit higher altitudes than \u003cem\u003eE. lundbladi\u003c/em\u003e at lower altitudes. This, coupled with the generalist character of \u003cem\u003eE. ornata\u003c/em\u003e, suggests that this species may be undergoing an altitudinal expansion, occupying areas previously inhabited only by \u003cem\u003eE. lundbladi\u003c/em\u003e. Additionally, in a scenario of climate change, it is possible that regions that were previously unsuitable for \u003cem\u003eE. ornata\u003c/em\u003e due to their low temperatures may now become suitable as temperatures have risen. This phenomenon appears to be occurring with other insect species. This phenomenon has been observed in other insect species (Gil-Tapetado et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2023\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2024\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eIn phytophagous insects, fidelity to the host plant can act as a barrier to encounter and reproductive isolation (Matsubayashi et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), where attraction to chemical cues (Piersanti et al. \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2020\u003c/span\u003e) and nutritional requirements influence phytophagous survival (Auclair \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e1969\u003c/span\u003e). However, for a trophic generalist species like \u003cem\u003eE. ornata\u003c/em\u003e, the host plant might not be a limiting factor or a reproductive barrier. Nonetheless, it would be necessary to assess the viability of \u003cem\u003eE. ornata\u003c/em\u003e under the nutritional resources provided by the host plants of \u003cem\u003eE. lundbladi\u003c/em\u003e. Additionally, in a climate change scenario, the alpine and subalpine host plants of \u003cem\u003eE. lundbladi\u003c/em\u003e could experience a regression, potentially lead a reduction in the distribution of \u003cem\u003eE. lundbladi\u003c/em\u003e, as has been observed with the sensitivity of \u003cem\u003eErysimum scoparium\u003c/em\u003e to high temperatures (Gonz\u0026aacute;lez-Rodr\u0026iacute;guez et al. \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2021\u003c/span\u003e; Jackson and Chapman 2024).\u003c/p\u003e \u003cp\u003eRecent phylogenetic analyses have shown that \u003cem\u003eE. ornata\u003c/em\u003e and \u003cem\u003eE. lundbladi\u003c/em\u003e belong to the tribe Strachiini; however, their phylogenetic relationships remain unresolved, resulting in a polytomy involving \u003cem\u003eE. maracandica\u003c/em\u003e (Roca-Cusachs et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). This evolutionary ambiguity contrasts with their ecological differentiation, as both species exhibit distinct niches and reproductive barriers. Nonetheless, under the influence of climate change, the prezygotic barriers currently maintaining genetic isolation could weaken, potentially facilitating gene flow between these lineages. To conserve the Macaronesian endemic \u003cem\u003eE. lundbladi\u003c/em\u003e, it is crucial to conduct future-focused studies on distribution limits, particularly in light of the recent emergence of sympatry with \u003cem\u003eE. ornata\u003c/em\u003e. Long-term monitoring of their contact zones is essential to evaluate the progression of interspecific copulations and the risk of hybridization. Although an increase in interspecific encounters has been observed during the two sampling periods, further research is needed to analyze the viability and reproductive success of immigrant individuals and to understand the potential consequences for the genetic integrity of \u003cem\u003eE. lundbladi\u003c/em\u003e.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eAll authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by M.A. and D.GT. The first draft of the manuscript was written by M.A. and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eWe would like to extend our sincere thanks to the professors and the director of the Master's Program in Conservation of Tropical Areas at the Men\u0026eacute;ndez Pelayo International University and the Consejo Superior de Investigaciones Cient\u0026iacute;ficas (CSIC), whose organization of the field trips to the Canary Islands made these findings possible. It is important to note that the observations presented here were made during fieldwork conducted as part of a university master\u0026rsquo;s program, which significantly enhanced the observational and sampling efforts in underexplored yet common areas. We would also like to thank all the generators of georeferenced biogeographic data and information who contribute data altruistically to the GBIF repository. We are also grateful to Mercedes Paris for her assistance in the examination of genital structures, and to Manuel Baena for his contributions and expertise to the Hemiptera group. The title of this article was inspired in the song Space Oddity of David Bowie because these interspecific copulations are found only in a part of the altitudinal range and seem to be a singularity of these areas and because these areas were found on Pico de la Nieve, near Roque de los Muchachos, the astronomical observatory where different European solar and nocturnal telescopes are located.\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eThe data is available in the reference section as part of the download package from the GBIF platform.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAguillon SM, Rohwer VG (2022) Revisiting a classic hybrid zone: Movement of the northern flicker hybrid zone in contemporary times. 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This study documents the first overlap zone with interspecific copulations on La Palma Island, Canary Islands, between the Macaronesian endemic \u003cem\u003eEurydema lundbladi\u003c/em\u003e Lindberg, \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e1960\u003c/span\u003e and the widespread Palearctic species \u003cem\u003eEurydema ornata\u003c/em\u003e (Linnaeus, 1758). We analyzed morphological differences in male genitalia, climatic niches, and altitudinal distributions of both species. Notably, the differences in male genital structures do not appear sufficient to cause copulatory incompatibility, nor do size variations act as limiting factors for mating. The ecological niches of both species, while distinct, converge in certain altitudinal zones, where climatic conditions\u0026ndash;particularly winter temperatures\u0026ndash;significantly influence their distribution. These weak and convergent premating prezygotic reproductive barriers underscore the conservation risks faced by \u003cem\u003eE. lundbladi\u003c/em\u003e in light of the potential expansion and competition from \u003cem\u003eE. ornata\u003c/em\u003e. Implications for insect conservation: The genetic integrity of \u003cem\u003eE. lundbladi\u003c/em\u003e is threatened by the encroachment of \u003cem\u003eE. ornata\u003c/em\u003e into previously unoccupied areas. Continued monitoring of contact zones and future studies are essential to evaluate the impact of these interactions on the conservation of this endemic species.\u003c/p\u003e","manuscriptTitle":"Space oddity: Absence of prezygotic-premating barriers in Eurydema lundbladi and Eurydema ornata","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-11-07 12:47:40","doi":"10.21203/rs.3.rs-5341557/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-03-30T09:02:21+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-03-07T19:30:09+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"310444060420602621200043213332410542474","date":"2025-02-23T15:41:58+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-02-18T07:25:20+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"175970050458113287879348501224386698599","date":"2025-02-01T17:54:23+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-11-03T15:17:35+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-10-29T02:23:43+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-10-29T02:22:46+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Insect Conservation","date":"2024-10-27T13:42:30+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-insect-conservation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jico","sideBox":"Learn more about [Journal of Insect Conservation](http://link.springer.com/journal/10841)","snPcode":"10841","submissionUrl":"https://submission.nature.com/new-submission/10841/3","title":"Journal of Insect Conservation","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"a9a2d4df-ce42-464c-bfb1-4330f20e6239","owner":[],"postedDate":"November 7th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-12-22T16:08:24+00:00","versionOfRecord":{"articleIdentity":"rs-5341557","link":"https://doi.org/10.1007/s10841-025-00742-z","journal":{"identity":"journal-of-insect-conservation","isVorOnly":false,"title":"Journal of Insect Conservation"},"publishedOn":"2025-12-19 15:57:56","publishedOnDateReadable":"December 19th, 2025"},"versionCreatedAt":"2024-11-07 12:47:40","video":"","vorDoi":"10.1007/s10841-025-00742-z","vorDoiUrl":"https://doi.org/10.1007/s10841-025-00742-z","workflowStages":[]},"version":"v1","identity":"rs-5341557","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5341557","identity":"rs-5341557","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}