Background

Fusarium crown rot (FCR) is a soil-borne disease caused by multiple Fusarium species, including F. pseudograminearum, F. graminearum, and F. culmorum [1]. Among these, F. pseudograminearum is the predominant causal agent in the Huang-Huai-Hai wheat region of China, a critical area for national wheat production [2]. FCR can infect wheat throughout its entire growth cycle. At the seedling stage, typical symptoms include browning of the leaf sheaths and stem bases, with severe infections leading to chlorosis and seedling death [3]. As plants advance to jointing and heading stages, stem base necrosis intensifies, often accompanied by pink mycelial growth at nodes under high humidity [4]. By maturity, severely infected plants exhibit premature senescence, whiteheads, and grain defects such as shriveled kernels or sterility, collectively leading to significant reductions in both yield and grain quality [5–6].

Currently, breeding resistant cultivars remains the cornerstone for effective management of FCR [7]. Chemical control strategies are constrained by the emergence of fungicide resistance and concerns over environmental contamination, which are also inconsistent with China’s Dual Reduction policy aimed at reducing pesticide and fertilizer use [8]. In this context, biological control has emerged as a promising alternative due to its sustainability and compatibility with ecological agriculture practices [9].

A major bottleneck in FCR research is the lack of reliable phenotyping tools. The terrace method can detect adult plant resistance, but it does not distinguish the very high level of susceptibility observed in durums in the field [8]. In current greenhouse resistance screening methods, glasshouse seedling bioassay is a good predictor of crown rot resistance in adult plants under field conditions, but crown rot severity is assessed 35 days after inoculation [10]. To overcome these limitations, we developed a standardized greenhouse inoculation protocol for FCR by optimizing pathogen preparation and the inoculation method. This system enables accurate quantification of F. pseudograminearum pathogenicity and high-throughput screening of resistant wheat germplasm under controlled environmental conditions.