Grapevine (Vitis vinifera L.) is one of the most popular and valuable fruit crops in the world, with a global vineyard surface estimated at about 7.1 million hectares (www.oiv.int). Adaptation to evolving climate conditions is currently a top priority for the grape and wine sector. Rising temperatures, prolonged drought periods, increased frequency of extreme weather events, and shifting pest and disease pressures are some of the climate change-derived factors threatening grape and wine production worldwide (Fraga et al., 2013; van Leeuwen et al., 2024). The situation is of particularly concern in regions already characterised by hot and arid climates (van Leeuwen et al., 2024), where the area suitable for grape production is expected to dramatically decrease if no action is taken (Morales-Castilla et al., 2020). Among the potential alleviating strategies, harnessing the natural genetic diversity of grapevines is essential (Wolkovich et al., 2018), and the use of more resilient varieties has emerged as a key strategy for sustaining viticulture under warming conditions (Morales-Castilla et al., 2020). Overall, the genetic diversity of V. vinifera L. is high, with an estimated six to ten thousand varieties available worldwide (Morales-Castilla et al., 2020; Wolkovich et al., 2018). This diversity is the result of independent local selection processes traditionally conducted by grape growers, who chose different grapevine varieties that matched their local climates to produce stable yields of grapes with an adequate balance of sugar, acids, and other compounds of interest for fresh fruit consumption and winemaking (Wolkovich et al., 2018). These efforts have resulted in a broad genetic diversity of traits of interest, including some that could be useful for coping with climate warming, like late-ripening (Boursiquot et al., 1995), better water use efficiency (Tortosa et al., 2016), and tolerance to extreme heat events (Coupel-Ledru et al., 2024). This situation has led to a renewed interest in the exploration of the genetic diversity preserved in old and abandoned vineyards throughout traditional viticulture regions (Maraš et al., 2020; Mendoza et al., 2022; Merheb et al., 2025; Tello et al., 2024b; Zombardo et al., 2021). In some cases, these efforts have resulted in the identification of relict plants of old varieties that may have had some relevance in the past, and which are currently being tested for their performance under current climate conditions (Maraš et al., 2020; Mendoza et al., 2022). These old varieties are also key to understanding how the local varietal assortment has changed over time in a region (Maraš et al., 2020; Tello et al., 2024b), sometimes reflecting major historical stages (El-Oualkadi et al., 2011; Tello et al., 2024b). Moreover, this information has revealed the genetic origin of some grapevine varieties of relevance (Boursiquot et al., 2009; Maraš et al., 2020; Tello et al., 2024a).

Viticulture has been practised in North Africa for centuries, with the earliest cultivated forms probably introduced by the Phoenicians by the tenth century BCE (El-Oualkadi et al., 2011; Ghaffari et al., 2014; Laiadi et al., 2009; Zinelabidine et al., 2010). Grape cultivation practices were later consolidated during the Roman and Christian periods, before undergoing a marked transformation during the Islamic period, when many pre-existing wine grape varieties were displaced in favour of table grapes (Ghaffari et al., 2014; Zinelabidine et al., 2014). Following the European grape phylloxera crisis, numerous wine grape varieties from France, Spain, and Italy were introduced to the region (Ghaffari et al., 2014; Zinelabidine et al., 2010), contributing to the present-day composition of local grapevine diversity. Moreover, this region lies within the natural range of distribution of V. vinifera ssp. sylvestris grapevines (This et al., 2006), providing an opportunity for local hybridisation events between cultivars and wild grapevines that might have shaped regional genetic diversity. Previous studies on the genetic characterisation of grapevines from the Maghreb have revealed a high level of genetic diversity (Riahi et al., 2010), and different studies have reported a considerable number of uncatalogued local genotypes (Ghaffari et al., 2014; Laiadi et al., 2009; Riahi et al., 2010; Zinelabidine et al., 2014).

Genetic analyses have revealed some differentiation between local Moroccan grapevines and those from other Maghrebi and European genetic pools (El-Oualkadi et al., 2011; Riahi et al., 2010). Prospection efforts across the country have led to the identification of diverse unknown or unclassified cultivated grapevines (Zinelabidine et al., 2010), suggesting the presence of potentially valuable varieties that are at high risk of disappearing. The Beni Mellal–Khénifra region, located in central Morocco, encompasses a wide range of landscapes, from the mountainous peaks of the Middle Atlas (900–3,890 masl) to extensive plains and plateaus at around 600 masl (Eddoughri et al., 2022). This altitudinal gradient generates pronounced climatic variability, with conditions ranging from humid in the high mountains to semi-arid in the plains, and marked seasonality characterised by intensely cold winters and very hot summers (Lebrini et al., 2019). This region is one of the main agricultural areas of the country, with land-use predominantly devoted today to rainfed agriculture, primarily cereals, citrus, and olive trees (Eddoughri et al., 2022). However, technical reports provide documentary evidence of traditional viticultural practices in the region (Protectorat de la République Française au Maroc, 1930), where relict plants still growing in areas no longer devoted to viticulture can be found (Zinelabidine et al., 2010). In light of the relevance of detecting novel grapevine genetic resources to face current challenges in viticulture, in this work we conducted a prospection of grapevines across the Beni Mellal–Khénifra region in the Atlas region of central Morocco. This effort led to the detection of 67 relict grapevines, which were analysed with a combined SSR (simple sequence repeat) and SNP (single nucleotide polymorphism) genotyping strategy for further genetic analyses. We proved that most of the prospected grapevines belong to the sativa subspecies, while some genotypes likely resulted from natural crosses between sylvestris and sativa individuals. The abundance of some unidentified genotypes suggests that they may have been varieties of local relevance in the past, warranting preservation and future characterisation.

Today, the agricultural sector of the Beni Mellal–Khénifra region in central Morocco focuses on cereals, citrus and olive trees (Eddoughri et al., 2022). However, an important tradition of grapevine cultivation used to exist, and grape production played a relevant role in the local economy decades ago (Protectorat de la République Française au Maroc, 1930). Previous works have revealed that, despite severe genetic erosion, a few relict grapevines have survived throughout the region as isolated plants (Zinelabidine et al., 2010). The identification of 67 relict grapevines in this work highlights the relevance of this region for the traditional production of grapes in Morocco, and the identification of up to 20 different grapevine genetic profiles indicates that a remarkable level of genetic diversity was cultivated in the recent past. Interestingly, the study of these genotypes has provided an opportunity to uncover how local diversity originated and was shaped. Comparative analyses with a core panel of V. vinifera ssp. sativa and V. vinifera ssp. sylvestris grapevines of Western Mediterranean origin revealed three distinct sources of genetic diversity underlying the origin of these traditional varieties: i) local sativa varieties of autochthonous Maghrebi origin, ii) introduced sativa varieties from the Iberian Peninsula, and iii) cultivated varieties originating from hybridisation events between sativa varieties and local sylvestris individuals. Together, these three sources of genetic diversity have shaped the traditional local assortment of the region explored in this work, contributing to the overall heterogeneity of the Moroccan germplasm available today, which reflects a complex history of grapevine cultivation.

A high phyletic affinity was found between 13 of the 20 differentiated genotypes and the sativa varieties from the Maghreb included in the core panel of genetic profiles designed for comparative purposes (Figure 3A). This finding indicates that the traditional Maghrebi sativa germplasm was the main source of diversity that contributed to the current traditional local assortment of the Atlas region explored in this work. These varieties include five cultivated grapevines (‘Abouhou’, ‘Agueyer’, ‘Bou Choukka’, ‘Hadari’, and ‘Outanat’) that are commonly found across distinct viticultural regions of the country (El-Oualkadi et al., 2011; Zinelabidine et al., 2010; Zinelabidine et al., 2014), which corroborates their alleged Moroccan origin and their historical relevance for grape cultivation in the country. Interestingly, pedigree analyses revealed ‘Hadari’ as a relevant founder of local varieties (Table 2), indicating an important role in the historical development of the Moroccan grapevine germplasm. This group of autochthonous grapevine varieties also contained ‘Dedo de Dama’ (Figure 3A), an important genitor in North Africa (Lacombe et al., 2013) that is known in Morocco as ‘Bouqseb’/‘Bougseb’ (El-Oualkadi et al., 2011; Zinelabidine et al., 2014)). ‘Dedo de Dama’ is a table grape variety with elongated berries, which is considered today to be of Portuguese origin (www.vivc.de). However, its region of origin is controversial (El-Oualkadi et al., 2011). Identifying its true region of origin has been hindered by the lack of a complete pedigree, as well as by the identification of descendants of ‘Dedo de Dama’ in the Iberian Peninsula (e.g., ‘Montua’ and ‘Boal Branco’) and in the Maghreb (e.g., ‘Azougah’, ‘Bezoul el Aouda’, and ‘Bouzouga’) (Cunha et al., 2020; El-Oualkadi et al., 2011; Lacombe et al., 2013; Zinelabidine et al., 2015). In fact, the ubiquity of its offspring suggests that ‘Dedo de Dama’ might have been among the varieties involved in the long history of interaction and traditional exchange of plant material between the Iberian Peninsula and the Maghreb (Zinelabidine et al., 2010). While this bidirectional movement of plant material might have obscured the geographic origin of ‘Dedo de Dama’, our results point to a plausible Maghrebi origin for this variety (Figure 3A). Thus, ‘Dedo de Dama’ could have been introduced into the Iberian Peninsula during the Arab period when table grape cultivation was favoured (Zinelabidine et al., 2010). It then might have been adopted by local Iberian grape growers, further contributing to the development of new cultivars. Moreover, we found eight genetic profiles with no matches in the consulted databases clustering within this group of varieties. This result suggests that the traditional Moroccan sativa germplasm exhibits greater diversity than has previously been assumed, with numerous varieties of former relevance persisting as relict plants in areas that are no longer dedicated to viticulture. The fact that some of these genetic profiles were repeatedly found across different sites of the Beni Mellal–Khénifra region (Table 1) not only supports their historical relevance, but suggests that they harbour beneficial traits that at some point promoted their vegetative propagation (This et al., 2006). In addition, the parentage relationships obtained between some of these unknown genotypes in grapevines from different locations (Table 2) indicate that they could have originated from the human-mediated propagation of seeds, a common process observed in other Maghrebi regions (El-Oualkadi et al., 2011; Zinelabidine et al., 2010). Remarkably, we identified the unknown variety “MAR-02” as the likely father of ‘Abouhou’ (Table 2), which is itself the mother of several Moroccan varieties (‘Azougah’, ‘Bezoul el Aouda’, ‘Bouzouga’ (Lacombe et al., 2013)). Although finding a compatible duo between two genotypes does not necessarily correspond to a parent–offspring relationship (Augusto et al., 2021), our results suggest that a distinctive lineage of Moroccan grapevine varieties originated from the unknown variety “MAR-02”.

This study was useful for evaluating the additional impact of most recent exogenous introductions on the composition of the Moroccan grapevine germplasm. The traditional Moroccan grapevine germplasm, initiated by Phoenicians and Romans (El-Oualkadi et al., 2011; Ghaffari et al., 2014; Zinelabidine et al., 2010; Zinelabidine et al., 2014), was further modified during the Arab period and later as a result of French influence (El-Oualkadi et al., 2011). Additional changes occured as a result of the introduction of wine grape varieties from France, Spain, and Italy following the European grape phylloxera crisis (Ghaffari et al., 2014; Zinelabidine et al., 2010). The two relict plants that we identified as being the alleged Spanish variety ‘Negra Rayada’ (Table 1) are likely linked to these historical introductions. Although this variety lacks a full pedigree and thus cannot be fully assigned a region of origin, ‘Negra Rayada’ has been recently identified in old Spanish vineyards (Tello et al., 2024a), and pedigree analyses indicate some parent-offspring relationships with other Spanish cultivars (Lacombe et al., 2013; Tello et al., 2024a). Based on these previous results, an Iberian origin can be assigned to ‘Negra Rayada’, which is reinforced by its genetic similarity to other Iberian varieties as observed in this work (Figure 3A). Thus, ‘Negra Rayada’ might have been introduced into the Maghreb from the Iberian Peninsula through vegetative propagation by cuttings, a practice that was subsequently maintained to preserve its varietal characteristics. This strategy would have complemented the commonly documented historical approach of grapevine multiplication by seeds in the Maghreb (El-Oualkadi et al., 2011; Zinelabidine et al., 2010). In fact, as observed in previous work (El-Oualkadi et al., 2011), we obtained few parentage relationships between Maghrebi and European varieties, pointing to seed propagation as an important method for the traditional multiplication of grapevines in Morocco. The use of seeds for grapevine propagation might have derived in a differentiated local germplasm that has a unique genetic identity (El-Oualkadi et al., 2011) while being residually genetically related to old varieties from abroad. Thus, “MAR-06” and “MAR-13” might be two examples of local varieties derived from exogenous introductions, as observed from their genetic affinity with varieties from Portugal and Spain, respectively (Figure 3A). In fact, our findings indicate that “MAR-13” could have resulted from a self-pollination event of ‘Beba’, a variety of Spanish origin, but which is commonly found in the Maghreb (Laiadi et al., 2009; Zinelabidine et al., 2010). The identification of grape varieties derived from selfing is rare (Maraš et al., 2020; Tello et al., 2024a) due to the negative effects of inbreeding depression on agronomic performance (Cattonaro et al., 2014). In fact, “MAR-13” was only found once, so it may be not a variety but a single plant that had not been subjected to vegetative propagation.

Lastly, we analysed the potential contribution of the local sylvestris genetic pool to the Moroccan germplasm explored in this study. The region included in this study lies within the natural range of wild grapevine distribution (This et al., 2006; Zinelabidine et al., 2010), providing an opportunity for natural hybridisation between grapevines of both subspecies. Our results suggest intrasubspecific sativa-sylvestris hybridisation as the origin of three of the unknown genotypes differentiated in this work: “MAR-04”, “MAR-11”, and “MAR-12” (Figure 3C). The presence of cultivated varieties with similar sativa-sylvestris genome ancestries has been documented in different viticultural regions where commercial vineyards are in close proximity to wild populations (Maraš et al., 2020; Schneider et al., 2024), including the Maghreb (Riahi et al., 2010). In fact, some renowned cultivated varieties like ‘Cabernet franc’, ‘Pinot’, or ‘Savagnin’ are known to share an ancestry percentage of around 50 % with the sylvestris subspecies (Maraš et al., 2020; Schneider et al., 2024), which is probably the result of the low number of sexual generations separating these varieties from the sylvestris genetic pool (Cattonaro et al., 2014). Our results add ‘Alvarinho’ and ‘Ondarrabi Beltza’ to this group of varieties with high sylvestris ancestry, which is in agreement with previous studies that had identified them as potential first-generation descendants of sylvestris individuals (D'Onofrio, 2020; Lopes et al., 2009). Interestingly, and in contrast to previous studies where intrasubspecific hybrid genotypes had been identified only once (Maraš et al., 2020), we identified the genetic profile named “MAR-04” in four plants from Oued Srou, a location next to Khénifra (Table 1). This result indicates that, due to some likely desirable traits, intrasubspecific sativa-sylvestris hybrids generated in the Beni Mellal–Khénifra region in central Morocco were selected by humans for vegetative propagation in order to adapt viticultural practices to local needs.

This study demonstrates that a previously underestimated reservoir of grapevine genetic diversity has been preserved in the Beni Mellal–Khénifra region of Morocco, despite severe erosion of traditional viticulture systems having taken place in the region. The identification of numerous relict grapevines that have uncatalogued genetic profiles (some of them family-related) has revealed a complex genetic landscape shaped by autochthonous Maghrebi varieties, historical introductions from the Iberian Peninsula, and local hybridisation events involving wild grapevines. These findings not only provide new insight into the historical dynamics that has shaped Moroccan grapevine germplasm, but they also highlight the importance of characterising the agronomic and adaptive value of these old varieties in the context of climate change. Preserving and characterising this genetic heritage is therefore essential, to both safeguard an important component of Mediterranean viticultural history and expand the pool of genetic resources available for the development of more resilient grapevine varieties.

This research was supported by the Spanish CSIC through the I-COOP+ 2020 call (Project COOPB20562) and by the research project PID2023-152513OB-I00 funded by MICIU/AEI/10. 13039/501100011033 and by ERDF/EU. JT acknowledges funding from the Ramón y Cajal Programme (RYC2022-037758-I), supported by MCIU/AEI/10.13039/501100011033 and the European Social Fund Plus (ESF+). We gratefully acknowledge Silvia Hernáiz (ICVV) for her technical support in the laboratory. We also thank SGIker (UPV/EHU/ERDF, Spain) and Centro de Investigación Biomédica de La Rioja (CIBIR, Spain) for their technical and human support in SNP and SSR genotyping, respectively. Finally, we extend our appreciation to all collaborators whose contributions helped to expand the number of genotypes included in the ICVV-DNA database.