Sugarcane orange rust, caused by Puccinia kuehnii, was first described in Java/Indonesia at the end of the nineteenth century. The disease was confined to Asia and Oceania before appearing in the Western Hemisphere at the beginning of the twenty-first century. In the Western Hemisphere, orange rust was first observed in Florida in 2007 and subsequently also diagnosed in Africa (Cameroon and Côte d'Ivoire). Although symptoms of the disease were observed in Gabon as early as 2006, the formal molecular identification of the pathogen was never performed. In this study, leaf samples were collected in Gabon in 2022 from four sugarcane varieties displaying typical symptoms of orange rust. The microscopic characteristics of the foliar lesions (uredinia or pustules) and fungal spores taken from these lesions corresponded to the description of P. kuehnii in the literature. DNA was amplified from urediniospores by multiple displacement amplification and used in a PCR assay with primer pair Pk1F/R specific for P. kuehnii. All 12 tested spore samples (three per sugarcane variety) yielded the expected¿~¿500 bp DNA fragment and forward sequenced amplicons matched with P. kuehnii in the GenBank database. DNA amplicons from two varieties (FR94129 and Q203) were cloned and sequenced in both directions. The 524–526 nucleotide sequences of eight clones matched at 99.6–100% with sequences of P. kuehnii in the GenBank database. This is the first report of occurrence of P. kuehnii in Gabon based on microscopy and molecular data.