Supplementary file 2_Plasma pre-treatment and peroxynitric acid soaking as a novel oxidative seed disinfection strategy against seed-borne fungal pathogens in maize.docx

Seed viability was evaluated by germination assays.</p>Results<p>PNA exhibited rapid, concentration- and time-dependent antifungal activity against both fungal species. Strong suppression of F. fujikuroi infection was observed at PNA concentrations ≥100 mM, with complete suppression of detectable infection at 400 mM after 180 s. At 50 mM PNA, A. flavus showed substantially faster inactivation kinetics than F. fujikuroi, with estimated ET50 values of 42.3 and 95.1 s, respectively.

At 100 mM PNA, plasma pretreatment shifted the fitted time -response curves toward shorter exposure times and lower infection levels, particularly for A. flavus. However, planned plasma-versus-no-plasma comparisons were not statistically significant after adjustment for overdispersion. Neither PNA nor plasma treatment significantly affected seed germination.</p>Discussion<p>PNA demonstrated rapid and effective antifungal activity against seed-borne A. flavus and F. fujikuroi, with clear species-specific differences in susceptibility.

Plasma pretreatment appeared to accelerate early PNA-mediated fungal suppression but did not consistently improve final disinfection efficacy at prolonged exposure times. This effect may be associated with increased seed-surface hydrophilicity following plasma exposure. The results support PNA as a promising oxidative approach for maize seed disinfection and provide further insight into species-specific responses and plasma-associated kinetic effects in seed-borne filamentous fungi.</p>