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Eucalyptus spp. are widely cultivated in India and worldwide for their versatile uses. Thus, sustainable disease management requires critical knowledge of Eucalyptus pathogens. In December 2022, a leaf blight disease was observed in Eucalyptus plantations in Haryana, northern India. Isolations from the symptomatic leaves yielded Botryosphaeria -like fungal isolates. Megablast homology search and phylogenetic analysis utilizing internal transcribed spacer (ITS), partial translation elongation factor-1α (tef1), and partial β-tubulin (tub2) regions of two isolates confirmed the species as Botryosphaeria dothidea. Leaf inoculations with the two B. dothidea isolates produced disease symptoms on Eucalyptus camaldulensis clone 2023. Finally, Koch's postulates were established by re-isolation and re-identification of B. dothedia isolates from the inoculated leaves. To the best of our knowledge, this is the first report of B. dothidea causing foliar disease symptoms on Eucalyptus in India. These findings will contribute to developing disease management strategies for Eucalyptus pathogens in northern India. [Display omitted] • This is the first report of Botryosphaeria dothidea causing leaf blight disease of Eucalyptus in India. • The pathogen was identified based on microscopic features, multilocus DNA sequence analysis, and pathogenicity tests. • This is an important addition to the molecular taxonomy of Botryosphaericeae affecting Eucalyptus in India. • This study warrants screening of Eucalyptus genotypes/clones for the identification of disease-resistant sources. [ABSTRACT FROM AUTHOR]