First Report of Fusarium pseudograminearum Causing Root Rot on Soybean (Glycine max) in Henan, China

Abstract

Fusarium pseudograminearum has previously been reported as a pathogen of wheat and barley in Henan Province of China (Li et al. 2012; Xu et al. 2017), but not of soybean (Glycine max L.). In 2017, soybean plants exhibiting symptoms of root rot characterized by brown lesions on the tap and lateral roots at the beginning pod (R3) growth stage were collected from 15 fields representing eight counties in Henan Province. The symptomatic root tissues were surface-sterilized with 70% ethanol for 50 s and rinsed in sterilized water three times. Small pieces of symptomatic tissue were then placed on potato dextrose agar (PDA) and incubated at 25°C for 5 days in the dark. A total of 132 Fusarium-like colonies were isolated, and hyphal tips were transferred to fresh PDA plates to obtain pure cultures. Single-conidia isolates were then produced by dilution plating conidia from pure cultures and then subculturing from colonies that developed from the single-conidia isolations. The average mycelial growth rate was 5.3 to 8.7 mm/day at 25°C on PDA, and the colonies produced aerial mycelium varying from rose to yellow white, and rose to burgundy pigment in agar. Macroconidia of the isolates were produced on carnation leaf agar (CLA) incubated under black light and observed to be abundant, relatively slender, almost straight, commonly 3- to 7-septate, averaged 38.6 × 4.1 μm, matching that reported for F. pseudograminearum (Aoki and O’Donnell 1999). No perithecia were observed. DNA of isolates YB17SN07 and YB17SN08 from Yuanyang County and isolate YB17SN09 from Yongcheng County was extracted using the CTAB method. The translation elongation factor 1 alpha (EF1-α) gene was amplified and sequenced (O’Donnell et al. 1998). Results of sequences were deposited in GenBank (accession nos. MG189598 to MG189600). BLASTn analysis of the EF1-α gene shared 99 to 100% similarity with those of F. pseudograminearum (HBZX1675 and CBS131261). The pathogen identity was further confirmed using species-specific PCR primers, Fp1-1 and Fp1-2 (Aoki and O’Donnell 1999). To test pathogenicity, inoculum was grown for 10 days at 25°C on 100 g of sterilized boiled wheat grain and sand (3:1 v/v) after infesting with a 1-ml spore suspension (10⁸ conidia/ml) of either isolate YB17SN07, YB17SN08, or YB17SN09. Five grams of this inoculum was placed 3 cm below the surface of autoclaved soil in 15 × 20 cm pots prior to planting pregerminated soybean seeds (cv. Wandou 35). Six plants were grown per pot, five pots were used per isolate, and the experiment was repeated three times. Noninoculated plants were grown in autoclaved soil amended with sterile wheat/sand mix and served as controls. Plants were grown in the greenhouse with a 14:10 h day/night cycle. After 42 days, the root system of all inoculated soybean plants exhibited dark brown lesions over the entire taproot. Noninoculated plants did not exhibit symptoms. The fungus was reisolated and identified from infected plants for each isolate as described above. This is the first report of F. pseudograminearum causing root rot on soybean in China. Considering that soybean is a major crop in China covering 7.8 million ha in 2017, root rot caused by F. pseudograminearum could develop into a disease with major economic impacts.