Non-wounding contact-based Inoculation of fruits with fungal pathogens in postharvest

Fungal pathogens significantly impact the quality of fruits and vegetables at different stages of the supply chain, leading to substantial food losses. Understanding how these persistent fungal infections occur and progress in postharvest conditions is essential to developing effective control strategies. In this study, we developed a reliable and consistent inoculation protocol to simulate disease spread from infected fruits to adjacent healthy fruits during postharvest storage. We tested different combinations of relevant fruit commodities, including oranges, tomatoes, and apples, against impactful postharvest pathogens such as Penicillium digitatum, Penicillium italicum, Botrytis cinerea, and Penicillium expansum. We assessed the efficacy of this protocol using fruits treated with various postharvest methods and multiple isolates for each pathogen. We optimized the source of infected tissue and incubation conditions for each fruit-pathogen combination. Disease incidence and severity were quantitatively evaluated to study infection success and progression. At the final evaluation point, 80% or higher disease incidence rates were observed in all trials except for the fungicide-treated oranges inoculated with fungicide-susceptible Penicillium spp. isolates. Although disease incidence was lower in that particular scenario, it is noteworthy that the pathogen was still able to establish itself under unfavorable conditions, indicating the robustness of our methodology. Finally, we used multispectral imaging to detect early P. digitatum infections in oranges before the disease became visible to the naked eye but after the pathogen was established. We developed a non-invasive inoculation strategy that can be used to recreate infections caused by contact or nesting in postharvest. The observed high disease incidence and severity values across fruit commodities and fungal pathogens demonstrate the robustness, efficacy, and reproducibility of the developed methodology. The protocol has the potential to be tailored for other pathosystems. Additionally, this approach can facilitate the study of fruit-pathogen interactions and the assessment of innovative control strategies.