Abstract :
[en] Maize (Zea mays L.) is one of the world’s most important cereal crops, providing a major source of food, animal feed, and industrial raw materials globally. Fusarium verticillioides, the primary causal agent of maize ear and stalk rot, threatens maize production due to its pathogenicity and fumonisin production, which poses serious risks to food safety and international trade. This study developed a field-deployable molecular diagnostic platform and characterized an effective biocontrol agent for the management of F. verticillioides.
A rapid, sensitive, and minimal-equipment assay was developed based on Chelex-100 resin-based DNA extraction, FUM1 gene targeting, and recombinase polymerase amplification coupled with CRISPR/Cas12a-based cleavage and lateral flow detection. This assay demonstrated an analytical sensitivity close to one genomic equivalent and completed the diagnostic process within 73 minutes. In field validation with diseased maize kernels from Shanxi Province, the assay identified two positive samples, which were confirmed as F. verticillioides by single-spore isolation and TEF-1α sequencing.
A promising biocontrol strain, Bacillus velezensis IFST-221, isolated from the maize rhizosphere, exhibited broad-spectrum antifungal activity, inhibiting the mycelial growth of F. verticillioides by 62.63% in plate confrontation assays. In maize kernel assays, FB1 levels decreased from 5,700 to 19.6 ppb after pretreatment with IFST-221. In greenhouse trials against cotton Verticillium wilt, pretreatment with IFST-221 reduced disease severity index from 73.21% to 8.93% and disease incidence from 85.71% to 14.29%, while also promoting better plant growth. IFST-221 additionally stimulated seedling growth in maize, tomato, and broccoli. Genome sequencing revealed a 3.86 Mb genome encoding nine secondary metabolite clusters, including surfactin, fengycin, bacillibactin, macrolactin H, difficidin, bacillaene, bacilysin, butirosin A/B, andalusicin A/B, and three unknown secondary metabolites, along with PGPR-related genes such as nitrogen fixation, indole-3-acetic acid biosynthesis, phosphate solubilization, and biofilm formation.
To elucidate the antifungal mechanism of IFST-221, liquid chromatography-tandem mass spectrometry (LC-MS/MS) and gene knockout analyses were conducted. Surfactins were detected in the crude extract, and deletion of the srfAA gene reduced the antifungal activity of mutant extracts; however, overall biocontrol efficacy remained largely unaffected in dual culture and maize kernel infection assays, suggesting that surfactin plays only a partial role. Further transcriptomic profiling of F. verticillioides under bacterial challenge revealed reprogramming of primary metabolism, ribosome-related genes, cell wall- and membrane-associated processes, and stress-adaptive pathways such as antioxidant defense, autophagy, and heat shock proteins. These findings illustrate the dynamic interaction between B. velezensis IFST-221 and its fungal target and provide a molecular framework for optimizing biocontrol efficacy.
Collectively, this study presents a robust framework for sustainable maize protection by advancing both rapid molecular diagnostic and the mechanistic characterization of a promising biocontrol agent. The findings lay a strong foundation for the application of B. velezensis IFST-221 in integrated disease management strategies.
