Root-knot nematodes (RKNs) Meloidogyne spp. are extremely polyphagous pests and four species severely affect grapevines throughout the world: M. arenaria, M. incognita, M. javanica and M. ethiopica. Californian populations of M. arenaria and M. incognita are reported to be virulent to widely used rootstocks and to the rootstock ‘Harmony’ in particular. Breeding RKNs-resistant grape rootstocks is a promising alternative to highly toxic nematicides. Muscadine (Vitis rotundifolia syn. Muscadinia rotundifolia) is a resistance (R) source with undercharacterised genetics. To this end, we used a segregating progeny between the RKN-resistant Vitis x Muscadinia accession ‘VRH8771’ from the muscadine source ‘NC184-4’ and the RKN-susceptible V. vinifera cv. Cabernet-Sauvignon. We first phenotyped its resistance to isolates of the i) M. arenaria, ii) M. incognita and iii) M. javanica species, and then to iv) two mixed Harmony-virulent Californian populations of M. arenaria and M. incognita. Finally, we created an isolate of M. arenaria and M. incognita from these Harmony populations and phenotyped the progeny to each of them [v) and vi)], and to vii) an isolate of M. ethiopica. The resistance phenotype of all the progeny’s individuals was independent of the RKN isolates or populations used. Resistance was mapped in a region of chromosome 18 in VRH8771, supporting the hypothesis that it is conferred by a single gene with an unprecedented wide spectrum in grapevine, including Harmony-virulent isolates. This dominant gene, referred to as MsppR1, is linked to the telomeric QTL XiR4 for X. index resistance from the same source. Additionally, plant mortality data showed that MsppR1-resistant material expressed a high-level resistance to the Harmony-virulent isolates. Our results are a first step towards the development of marker-assisted breeding using SSR and SNP markers for resistance to RKNs in accession VRH8771. © 2023, International Viticulture and Enology Society. All rights reserved.< Réduire