Medicago sativa, commonly known as “zi hua mu xu,” is a leguminosae plant known for its pasture properties and is distributed throughout China. In July 2025, root rot was observed on M. sativa plants in Zhangjiakou (39.55°N, 113.56°E), Hebei Province, China (incidence rate 55%). Plants in 300 ha were observed to be severely affected by a disease in the field, causing a yield loss of 60%. Infected roots displayed brown necrotic spots, progressing to deep brown rot, accompanied by leaves turning yellow and withering, ultimately leading to plant mortality. To isolate the causal agent, tissues (5×5 mm) from four symptomatic plants were removed from the border of lesions, surface sterilized in 75% ethanol for 30 s and 0.1% HgCl2 for 1 min, then rinsed three times with sterile distilled water, plated on potato dextrose agar (PDA) at 25℃, and incubated in the dark for 7 days. Two representative isolates, 202528 and 202529, were cultivated on PDA. Macroconidia of the isolates had three to five septa and were relatively slender; the apical cells were tapered, and the basal cells were poorly developed. Microconidia were oval or club-shaped with a flattened base and 0-to 1-septate. Aerial mycelium were false heads and short to medium-length chains. Conidiogenous cells were polyphialides and monphialides (Leslie and Summerell 2006). The translation elongation factor (TEF-1α) and partial RNA polymerase second-largest subunit (RPB2) were amplified (accession nos. TEF-1α: PX926509 and RPB2: PX926511) (O'Donnell et al. 2022). When compared with other Fusarium species in GenBank, the isolate exhibited 100% (TEF-1α, KX656206.1) and 100% (RBP2, ON934680.1) similarity with Fusarium fujikuroi (query coverage=100%, per-centage identity=100%). A phylogenetic tree was constructed in MEGA software (version 11.0.10) (Tamura et al. 2021) using the partial concatenated gene sequences. Maximum-likelihood analysis revealed that the isolates were closely related to F. fujikuroi (99% bootstrap support). To test pathogenicity, conidial suspensions of 202528 and 202529 at a concentration of 1×106 conidia/mL were inoculated via root wounding on 36 plants per isolate. For the control treatment, 36 plants were treated with an equivalent volume of sterile water. The experiment was repeated three times. The plants were placed in a greenhouse at 26-30℃ and 95% relative humidity. The typical symptoms were observed 10 days after inoculation, except in the control samples, where no symptoms were observed. The same fungus was successfully reisolated from the symptomatic plant tissue and was reidentified as F. fujikuroi through morphological characteristics, TEF-1α and RBP2 sequencing analysis (accession nos. TEF-1α: PX926510 and RPB2: PX926513), thereby fulfilling Koch’s postulates. While F. fujikuroi has been reported to cause Juglans sigillata stem rot, plum fruit blotch, Zanthoxylum bungeanum black stem rot and maize ear rot in China (Han et al. 2021; Long et al. 2021; Luo et al. 2021; Duan et al. 2020), this is the first study to report that F. fujikuroi can infect M. sativa in China. The finding is greatly significant for preventing and controlling root rot in M. sativa and reducing economic losses for farmers.
Zhang et al. (Sun,) studied this question.