First Report of Austropuccinia psidii Causing Myrtle Rust on Outdoor Myrtaceae in Europe

In May 2025, rust symptoms were observed on Myrtus communis in several Lisbon gardens (Lisbon, Tropical and Ajuda Botanical Gardens, Beau-Séjour Palace, Tapada da Ajuda) and on Lophomyrtus × ralphii ‘Purpurea’ at Ajuda Botanical Garden. No symptoms were found in five other gardens (50% disease prevalence). Affected plants showed small chlorotic spots developing into bright orange pustules on young shoots, leaves and flower pedicels, with 100% of new shoots affected. Necrotic lesions were observed on older tissues. No symptoms were observed on other Myrtaceae (Agonis flexuosa, Callistemon rigidus, C. viminalis, Eucalyptus spp., Eugenia involucrata, E. uniflora, Lophostemon confertus, Melaleuca armillaris, M. elliptica, Metrosideros excelsa). Microscopy revealed bright orange uredinia containing masses of pedicellate, ovoid to globose urediniospores (18.2–21.7–26.7 × 12.1–16.6–21.7 μm, L/W ratio 1.01-1.33-1.81), with a hyaline to lightly pigmented, echinulate wall (n=30), consistent with Austropuccinia (Boufleur et al. 2023). Molecular identification was performed using the rDNA-ITS region (ITS1F/ITS4rust; Scholler et al. 2022) and LSU (LR5/LR0R; Vilgalys & Hester 1990; Cubetta et al. 1991), as recommended. For both ITS and LSU regions, all five sequenced strains showed identical sequences and a single sequence was deposited in GenBank for each: PV749118 (ITS) and PV902604 (LSU). The ITS sequence showed 0-3 bp differences to A. psidii sequences in GenBank: 627/627 bp match to MK020421 (Ho et al. 2019) and 701/704 bp to KM282146 (Machado et al. 2015). A 15 bp difference was recorded to A. licaniae (393/408 bp to OR082914; Ebinghaus et al. 2024). For LSU, a 2 bp difference (792/794 bp) was found to A. psidii (OR082909) and 28 bp to A. licaniae (766/794 bp to OR082911; Ebinghaus et al. 2024). Thus, ITS and LSU sequences confirms the fungus as A. psidii. Flow cytometry (Tavares et al. 2014) revealed a genome size of 1.144 Gb/1C, consistent with A. psidii (Tobias et al. 2020). Infected plant material was deposited in the João de Carvalho e Vasconcellos Herbarium (LISI-FUNGI-00051 to 00055). To fulfill Koch’s postulates, a spore suspension (3 × 10⁵ spores/ml in 0.02% Tween) of isolate ApTA02 was sprayed onto four M. communis plants, kept in wet chambers in darkness for 24h and incubated at 25°C under a 12h/12h day/night cycle. Symptoms appeared after eight days as leaf tumefactions, progressing to severe leaf and stem distortion, and uredinia erupting 10 days post-inoculation. Control plants (four M. communis sprayed with a 0.02% Tween solution) were asymptomatic. The experiment was repeated with identical results. Morphological and rDNA-ITS analysis of the uredinia erumpting from inoculated plants revealed identical characteristics to those of isolate ApTA02. Austropuccinia psidii is a hemicyclic rust affecting more than 480 Myrtaceae species, causing defoliation and plant mortality. Its urediniospores remain viable for up to three months and are wind-dispersed (Almeida et al. 2025). This is the first report of A. psidii on outdoor Myrtaceae in Europe, after the detection on Syzygium buxifolium bonsais in Switzerland (Ruffner et al. 2024). Austropuccinia psidii threatens native Myrtaceae, ornamentals, and plantations. Listed as a quarantine pest, it endangers biodiversity and eucalyptus (Paap et al. 2023). Early detection is essential to mitigate its spread and protect vulnerable ecosystems and economic activities.