*2.2. Detached in-vitro Tissue Assay*

### 2.2.1. Surface Disinfection of Seeds

Maize seeds were surface disinfected by sonication (ultrasonic bath 2.8L, Fisher Scientific) in sterile distilled water with Tween 20 (0.1% *v*/*v*) for 5 min. Subsequently, seeds were immersed in 1.5% (*v*/*v*) sodium hypochlorite (NaOCl) at 52◦C for 20 min (Thermobath FE-377, Felisa), followed by rinsing three times in sterile distilled water, and air dried in a Class II Type A2 Biological Safety Cabinet (Herasafe KS, Thermo Scientific, Langenselbold, Germany) and grown in culture room in Magenta ™ vessel (no. cat V8505, Sigma, MO, USA) with sterile sand.

### 2.2.2. Conidia Suspensions and Inoculation of Tissue by *Fusarium*

The *Fusarium* strains (Table 1) were cultivated in the SNA medium with a 1-cm<sup>2</sup> filter paper [8] supplemented with neomycin (0.12 mg/mL) and streptomycin (1 μg/mL), and cultured at 25 ± 2◦C for 7 days [23]. Conidia were harvested by adding 5 mL of sterile saline solution (0.8% ( *w*/*v*) sodium chloride) to the culture medium with gentle shaking. The conidia quantification was performed using a Neubauer chamber (Hausser Scientific, Horsham, PA, USA) and a light microscope (B-383-M11, Optika, Ponteranica, Italy) and by CFU on PDA plates. The conidia suspension was prepared with the final concentration of inoculation of 1 × 10<sup>6</sup> CFU/mL. Leaves and roots were collected from 2 weeks old in-vitro grown maize plants with no visible fungi contamination [24] for detached tissue assays. The tissues were inoculated with 200 μL of conidia solution and incubated at 25◦C in a wet chamber and photographed after 5 days post-inoculation (dpi) (Figure 1).
