Temporal changes in colonization levels of Ustilago esculenta in different parts of Zizania latifolia plants competent and incompetent for stem gall formation

Development of a real-time loop-mediated isothermal amplification method with toothpick sampling for non-destructive detection of Ustilago esculenta in Zizania latifolia

“…For this purpose, we applied this method to seedlings from a Shirakawa-derived variant designated as the male-like type. Recently, we have shown by qPCR that seedlings from this type of plants are colonized by U. esculenta at very low levels and did not produce makomotake (Yamato et al 2024). The LAMP reaction did not show an increase in uorescence in seedlings of male-like types that were pricked with toothpicks at depths of 1, 2, and 3 mm (Fig.…”

“…DNA was extracted from the colony of U. esculenta, Athelia rolfsii or Fusarium oxysporum f.sp. conglutinans using the cetyltrimethylammonium bromide (CTAB) method essentially as described previously (Yamato et al 2024) except that the centrifugation speed was changed from 1,900 g to 10,000 g. The colony of the MAT-2 haploid strain, Sn-1-(1)-F1, was grown on YEPS agar medium as described previously (Chigira et al 2023). The extracted DNA was 10-fold serially diluted from 1.0 × 10 2 to 1.0 × 10 − 4 ng/µL to evaluate the detection limit.…”

“…In Z. latifolia cultivation, the challenge of makomotake production is the decrease in makomotake yield caused by the occasional appearance of plants without stem gall formation. In our paddy elds in 2019, in addition to the "normal" Z. latifolia plants that form makomotake during the harvesting period, we identi ed two types of variant plants that neither produced ears nor galls, or produced ears but failed to form galls (Yamato et al 2024). We designated the former type as the "non-gall-forming type".…”

“…Meanwhile, the latter type was designated as the "male-like type", as in China, Z. latifolia plants that retain the ability to produce ears due to the disappearance of U. esculenta infection are referred to as "male" plants ( Guo et al 2007). Recently, our quantitative PCR (qPCR) analyses showed that fungal colonization levels remained signi cantly low in both the non-gall-forming and male-like types from the seedling to harvesting stages (Yamato et al 2024). In contrast, normal plants that developed stem galls showed higher fungal colonization levels than the two types at the seedling stage.…”

“…In contrast, normal plants that developed stem galls showed higher fungal colonization levels than the two types at the seedling stage. In addition, the normal plants also showed an increasing pattern of the fungal levels in the culm and leaf during vegetative growth and stem gall formation (Yamato et al 2024). The ndings indicate that the stem gall development in Z. latifolia may be in uenced by the colonization level of the fungus at the seedling stage.…”