First report of cucumber damping‐off caused by Globisporangium ultimum var. ultimum in Japan

Globisporangium ultimum (Trow) var. ultimum Uzuhashi, Tojo, and Kakish. (syn. Pythium ultimum Trow var. ultimum) can significantly reduce cucumber (Cucumis sativus) production (Keinath et al., 2017). This study is the first to demonstrate that damping-off in cucumber is caused by G. ultimum var. ultimum in Japan. Damping-off disease was observed in a vegetable garden in May 2022 in Osaka, Japan. Five of ten cucumber seedlings showed the symptom at the growth stage of 2-2.5 true leaves (Fig. 1a). The damaged tissues were softened, thinned, darkly browned, and had a fluffy white cobweb-like growth on their surface. Sexual aplerotic oospores and hyphal swelling were found in the affected tissues (Fig. 1b). Three Globisporangium-like isolates were obtained from diseased tissues by incubating on a Globisporangium and Pythium selective NARM (nystatin + ampicillin + rifampicin + miconazole) agar (Morita & Tojo, 2007) for one to three days at 25°C. All three isolates were cultivated on a bentgrass-leaf culture (van der Plaats-Niterink, 1981) for three days for morphological observations, while potato carrot agar was utilised to measure hyphal growth response to temperatures for one to two days according to van der Plaats-Niterink (1981). Since all three isolates displayed an identical morphology and hyphal growth response, a representative strain (KS1) was used to determine the rDNA-ITS sequence according to Kawasumi et al. (2022). The morphological characteristics (Fig. 2a-c) and hyphal growth temperature matched those of G. ultimum var. ultimum (Table 1; van der Plaats-Niterink, 1981). A BLASTn search showed 99.88% identity with the sequence of G. ultimum var. ultimum strain CBS 398.51 (GenBank Accession No. AY598657). The rDNA-ITS sequence of strain KS1 was deposited in GenBank (OQ352816). The living culture was deposited as accession MAFF 247832 in the Genebank Project, NARO, Tsukuba, Japan. A pathogenicity test of strain KS1 was performed as follows. Both inoculated and uninoculated plants were grown in 300 mL plastic pots using commercial nursery soil (Takii Tanemaki Baido, Takii Co. Ltd., Japan) for one week in a growth chamber (30°C, 12 hr day/25°C, 12 hr night). The experiment was conducted in triplicate, with three pots per replication and six plants per pot. Inoculum of the isolate was cultured at 28°C for seven days using bentgrass-seed agar containing 0.15% (w/w) agar and 0.5% (w/w) of bentgrass seeds per litre of distilled water. (Kawasumi et al., 2022), and 1 mL of the inoculum was placed on the basal part of the plants. The plants were grown subsequently for an additional five days in the chamber. All inoculated plants developed damping-off symptoms at a growth stage of <0.5 true leaves (Fig. 3a). Oospores and hyphal swellings were observed in the infected tissue (Fig. 3b-c). Globisporangium ultimum var. ultimum was re-isolated from the inoculated seedlings, thereby satisfying Koch's postulates. Uninoculated control plants remained asymptomatic and G. ultimum var. ultimum could not be isolated. Cucumber has been reported as a host plant of G. splendens (Hans Braun) Uzuhashi, Tojo & Kakishima in Japan (Sugawara et al., 2015) but there is no record of G. ultimum var. ultimum on cucumber (NARO Genebank, 2024). This study represents the first report of G. ultimum var. ultimum causing damping-off cucumber in Japan. The current findings also indicate that G. ultimum var. ultimum can induce damping-off in mature cucumber seedlings. This underscores the importance of implementing preventative measures throughout the entire cucumber nursery stage, from early to late development.