Rust disease on Nymphoides peltata caused by Puccinia sp. in Korea

Nymphoides peltata (Asteraceae), commonly known as 'yellow floating heart', is an aquatic plant with buoyant leaves. It is native to East Asia and commonly cultivated in garden ponds and lakes for ornamental purposes. In other regions, for example in North America, N. peltata is an unwanted species, posing a high ecological risk due to its ability to densely cover water surface, shade submersed plants and reduce gas exchange (Plant Epidemiology and Risk Analysis Laboratory, 2012). In June 2023, during surveys for candidate biological control agents for Nymphoides, typical aecial symptoms of rust disease were found on N. peltata leaves at the Kumho River in Daegu (35°53'10"N, 128°30'01"E) and Mankyong river in Wanju (35°54'21.4"N, 127°07'32.6"E) in Korea. Disease incidence was more than 90% (Figure 1). Symptoms included round, yellowish, discoloured spots on the upper surface of the leaves. Subsequently, these spots developed into yellow to orange aecia. A representative sample was deposited at the Kunsan National University Herbarium (acc. no. KSNUH2120), Gunsan, Korea. The rust-infected leaves and the causal agent were examined under a dissecting microscope (M205C; Leica, Germany), a differential interference contrast microscope (Axio Imager 2; Carl Zeiss, Germany) and a scanning electron microscope (S-4800+EDS; Hitachi, Japan). Spermogonia were epiphyllous, subepidermal, centrally surrounded by aecia, bright yellow to brown, round, and 100–150 μm in diameter (Figure 2). Aecia were epiphyllous, densely grouped, yellow to orange, cup-shaped with peridia, and 150–300 μm in diameter (Figure 3). The peridial cells were hyaline, rhomboidal with a verrucose, thick-walled and 18.8–34.5 × 15.1–21.6 μm. Aeciospores were yellowish, subglobose or suboblate and (15.0–)16.9–19.4(–21.3) × (13.0–)14.0–16.4(–17.6) μm (av. 18.2 × 15.2 μm) (Figure 4). The Aeciospore wall was verrucose with large granules and 0.5–1.2 μm thick (Figure 5). These morphological characteristics are similar to those of Puccinia scirpi found on Nymphoides indica in Brazil (Soares et al., 2006; Table 1). To confirm the morphological identification, genomic DNA was extracted from one specimen (KSNUH2120) using a MagListo 5M plant Genomic DNA Extraction Kit (Bioneer; Daejeon, Korea). The internal transcribed spacer (ITS) and large subunit (LSU) regions of the rDNA, and the cytochrome oxidase 3 (cox3) mtDNA region were amplified and sequenced with primers ITS5u/ ITS4rust, LRust1R/LRust3, and CO3-F1/CO3-R1, respectively (Vialle et al., 2009). The resulting sequences were deposited in GenBank (Accession. Nos. OR821997, OR821998 and OR830028) and compared with other reference sequences using BLASTn analysis. The ITS and LSU sequences of the Korean specimen had ten and eight nucleotide differences, respectively, with those of P. scirpi on N. indica (KX999892). The cox3 sequence differed at two nucleotides with P. scirpi on N. indica (KX999929). In maximum-likelihood phylogenetic trees inferred from a concatenated alignment of ITS, LSU, and cox3 sequences (Figure 6), the Korean specimen formed a clade with the reference sequence of P. scirpi. However, due to poor support and multiple sequence differences, we report this rust fungus here as Puccinia sp. It is possible that it represents a new species or one of the Puccinia species that have no publicly available sequence data. To our knowledge, this is the first report of Puccinia sp. on Nymphoides peltata in Korea. Given its high disease incidence, this pathogen could be a potential concern for N. peltata in Korea and further research is required to determine its identity. This work was supported by the US Army Engineer Research and Development Center, Aquatic Plant Control Research Program, and the Ministry of Environment, Republic of Korea (grant 2021002270004).