ABSTRACT
Heilongjiang is the largest rice-producing province in China, with annual yield of 28.9 million tons cultivated on 3.8 million hectares (Liu et al. 2021). During field surveys from July to August (2021-2022), symptoms of wilting were observed on rice panicles across Baoqing county (46.32°N, 132.20°E), Shuangyashan city, Heilongjiang province, China. Disease incidence ranged from 10 to 35%, and yield losses were estimated to be 5 to 20% over 7 surveyed fields of 18.5 ha in total. Initially, infected panicles exhibited carmine to brownish spots at the flowering and early grain-filling stages, which gradually merged into large and irregular lesions and spread to the entire panicle surface. Eventually, panicles became wilting and decayed at the ripening stage. To identify the etiological agent, thirty-five symptomatic panicles were collected randomly from 35 plants at different positions in 7 fields. The fragments (approximately 3 mm2) were dissected from margins of individual lesions, surface-disinfested with 70% ethanol for 30 s followed by 2% sodium hypochlorite for 2 min, and rinsed three times in sterilized water. The pieces were then dried and placed onto half-strength potato dextrose agar (PDA) supplemented with 50 µg/mL of streptomycin sulfate. After incubation at 28°C for 4 days, nineteen cultures were obtained and purified using the single-spore isolation method. On PDA plates, the colonies produced fluffy and cottony aerial mycelia and were white to yellowish with deep-yellow to red-brown pigments. The microconidia were hyaline, elliptical or clavate, zero to one septum, measuring 6.3 to 19.2 × 2.6 to 5.1 µm in size (n = 50). On carnation leaf agar (CLA), the macroconidia were thick-walled, falcate to almost straight, three to five septa, apical cell hooked to tapering, basal cell foot-shaped, measuring 27.4 to 47.8 × 3.6 to 5.4 µm in size (n = 50). No chlamydospore was observed. The internal transcribed spacer (ITS) region of ribosomal RNA, translation elongation factor (TEF-1α) gene, and ß-tubulin (ß-TUB) gene were amplified and sequenced using primers ITS1/ITS4 (White et al. 1990), EF1/EF2 (O'Donnell 2000), and T1/T22 (O'Donnell and Cigelnik 1997) from three representative isolates (PJ58, PJ69 and PJ83), respectively. The obtained sequences were deposited in GenBank (accession nos. ON527509, OQ772202 and OQ777725 for ITS; ON573222, OQ784926 and OQ784927 for TEF-1α; ON573223, OQ784928 and OQ784929 for ß-TUB, respectively). BLASTn analysis revealed 99.8 to 100% homology with the corresponding sequences of Fusarium kyushuense (MH892849 for ITS, AB674297 for TEF-1α, and GQ915442 for ß-TUB, respectively) in GenBank. Maximum likelihood phylogeny based on the concatenated sequences of ITS, TEF-1α and ß-TUB grouped three representative isolates in the F. kyushuense clade. Combined with the morphological and molecular characteristics, the fungus was identified to be F. kyushuense. Pathogenicity of the three isolates of F. kyushuense was evaluated on a susceptible rice cultivar Nanjing 46 at the booting stage. The upper part of a healthy panicle was inoculated by injecting 2 ml of a conidial suspension (1 × 106 spore/ml) obtained from a 7-day-old PDA culture of each isolate. The negative control was treated with sterile distilled water. The experiment was performed thrice with ten replicated plants for each treatment. All plants were placed in a humid chamber at 25°C with a 12-h photoperiod and 80% relative humidity. Twenty days after inoculation, it was found that the inoculated panicles showed typical reddish to brownish lesions, whereas control plants remained symptomless. Pathogens were reisolated from the artificially inoculated panicles and confirmed by morphological and molecular tests, fulfilling Koch's postulates. In recent years, this species has been associated with stalk rot and ear rot of maize (Cao et al. 2021; Wang et al. 2014) and wilt of tobacco (Wang et al. 2013). Also, it was mentioned as a producer of mycotoxins, especially trichothecenes and HT-2 toxin (Varga et al. 2016). To our knowledge, this is the first report of F. kyushuense causing panicle wilting on rice in China. The appropriate control strategies should be made to reduce the risk of disease due to food security concerns and potential threats to rice production.
ABSTRACT
Fusarium proliferatum is the principal etiological agent of rice spikelet rot disease (RSRD) in China, causing yield losses and fumonisins contamination in rice. The intraspecific variability and evolution pattern of the pathogen is poorly understood. Here, we performed whole-genome resequencing of 67 F. proliferatum strains collected from major rice-growing regions in China. Population structure indicated that eastern population of F. proliferatum located in Yangtze River with the high genetic diversity and recombinant mode that was predicted as the putative center of origin. Southern population and northeast population were likely been introduced into local populations through gene flow, and genetic differentiation between them might be shaped by rice-driven domestication. A total of 121 distinct genomic loci implicated 85 candidate genes were suggestively associated with variation of fumonisin B1 (FB1) production by genome-wide association study (GWAS). We subsequently tested the function of five candidate genes (gabap, chsD, palA, hxk1, and isw2) mapped in our association study by FB1 quantification of deletion strains, and mutants showed the impact on FB1 production as compared to the wide-type strain. Together, this is the first study to provide insights into the evolution and adaptation in natural populations of F. proliferatum on rice, as well as the complex genetic architecture for fumonisins biosynthesis.
ABSTRACT
Fusarium proliferatum is the primary cause of spikelet rot disease in rice (Oryza sativa L.) in China. The pathogen not only infects a wide range of cereals, causing severe yield losses but also contaminates grains by producing various mycotoxins that are hazardous to humans and animals. Here, we firstly reported the whole-genome sequence of F. proliferatum strain Fp9 isolated from the rice spikelet. The genome was approximately 43.9 Mb with an average GC content of 48.28%, and it was assembled into 12 scaffolds with an N50 length of 4,402,342 bp. There is a close phylogenetic relationship between F. proliferatum and Fusarium fujikuroi, the causal agent of the bakanae disease of rice. The expansion of genes encoding cell wall-degrading enzymes and major facilitator superfamily (MFS) transporters was observed in F. proliferatum relative to other fungi with different nutritional lifestyles. Species-specific genes responsible for mycotoxins biosynthesis were identified among F. proliferatum and other Fusarium species. The expanded and unique genes were supposed to promote F. proliferatum adaptation and the rapid response to the host's infection. The high-quality genome of F. proliferatum strain Fp9 provides a valuable resource for deciphering the mechanisms of pathogenicity and secondary metabolism, and therefore shed light on development of the disease management strategies and detoxification of mycotoxins contamination for spikelet rot disease in rice.
