Species composition, distribution, and pathogenicity of Botryosphaeriaceae species associated with canker, twig dieback, and fruit rot of walnut in Central Anatolia, Türkiye

Background Botryosphaeriaceae species are associated with important diseases such as canker, dieback, and fruit rot in walnut ( Juglans regia L.) trees. In this study, the prevalence, species composition, associations with symptoms and geographical distribution, pathogenicity, and temperature-dependent mycelial growth of the walnut disease complex associated with Botryosphaeriaceae were investigated in an integrated manner across eight provinces in the Central Anatolia Region of Türkiye (Aksaray, Kayseri, Kırıkkale, Kırşehir, Nevşehir, Niğde, Sivas, and Yozgat). Results Within the scope of the survey, 30,501 trees in 265 walnut orchards were examined; the prevalence of orchards containing at least one symptomatic tree was 87.9%, and the pooled overall incidence of symptomatic trees was 16.0%. The incidence of symptomatic trees varied significantly among provinces. Out of 1283 tissue samples taken from perennial canker, twig dieback, and fruit rot symptoms, 666 Botryosphaeriaceae isolates were obtained (51.9%); Diplodia seriata De Not. (37.4%), Neofusicoccum parvum (Pennycook & Samuels) Crous, Slippers & A.J.L. Phillips (29.0%), Botryosphaeria dothidea (Moug.) Ces. & De Not. (17.4%), Lasiodiplodia theobromae (Pat.) Griffon & Maubl. (8.6%), and Lasiodiplodia pseudotheobromae A.J.L. Phillips, A. Alves & Crous (7.7%) were identified. Isolate-level species composition showed significant associations with both symptom type and province. Multilocus phylogenetic analyses of 56 representative isolates based on ITS, TUB2, and TEF1-α loci supported the five species-level groups identified in the collection. In pathogenicity tests conducted on detached and attached walnut branches using 15 multilocus-confirmed isolates (three per species), all species caused disease; the highest pathogenicity was generally determined in L. theobromae and L. pseudotheobromae isolates, and the lowest in D. seriata isolates. Mycelial growth varied depending on temperature and species; L. pseudotheobromae showed its highest growth at 30 °C, while L. theobromae and L. pseudotheobromae maintained higher growth at 35 °C compared to other species. Conclusions The findings indicate that the Botryosphaeriaceae disease complex in Central Anatolian walnut orchards is widespread and heterogeneous in terms of species and symptoms, with spatial variation in isolate composition; in particular, the high pathogenicity of the tested Lasiodiplodia isolates and their comparatively strong in vitro growth at the upper end of the tested temperature range warrant further investigation.