Tomato brown rugose fruit virus (ToBRFV) has become a major threat to global tomato production due to its exceptional mechanical transmissibility and virion stability. Effective sanitation is essential for containment, yet the performance of commonly used disinfectants on greenhouse-relevant surfaces remains poorly characterized. This study evaluated multiple disinfectant formulations, applied by spraying or dipping, on polyethylene film, pruning shears, and human hands. After controlled inoculation with a standardized inoculum, treated surfaces were swabbed and extracts mechanically inoculated onto Nicotiana rustica L. Lesion number was visually quantified, and lesion area was measured using a computational image-analysis pipeline. Fifth-generation quaternary ammonium compounds (5°QAS) showed the highest virucidal activity on smooth, non-porous surfaces, reducing lesion numbers to fewer than 10 per leaf at 800–1000 ppm and maintaining infection severities below 1%. Glutaraldehyde at 500 ppm also performed strongly, achieving severities as low as 0.20% on plastic. Metallic pruning shears consistently retained infectious particles, with untreated controls exceeding 100 lesions per leaf and treated samples showing incomplete inactivation. Mechanical agents such as powdered milk and soap reduced infection but did not eliminate transmission. No clear dose–response trend was observed. The two most effective treatments, 5°QAS at 800–1000 ppm and glutaraldehyde at 500 ppm, significantly reduced or prevented systemic infection in tomato assays. These findings demonstrate that sanitation efficacy depends on formulation, surface type, and application method, providing operationally relevant guidelines for ToBRFV management.

Tomato brown rugose fruit virus (ToBRFV) has rapidly become a major threat to global tomato production due to its exceptional mechanical transmissibility and remarkable virion stability. Effective sanitation is therefore central to containment, yet the performance of commonly used disinfectants on greenhouse-relevant surfaces remains poorly characterized. This study systematically evaluated the efficacy of multiple disinfectant formulations, applied either by spraying or dipping, on three surfaces central to greenhouse crop management: polyethylene film, pruning shears, and human hands. Following artificial contamination, treated surfaces were swabbed and the resulting extracts mechanically inoculated onto Nicotiana rustica. Lesion number was assessed through direct visual inspection, whereas lesion area was quantified via a computational image-analysis pipeline, enabling precise measurement of foliar infection severity. Generalized linear models and correlation analyses were implemented to quantify treatment effects and to evaluate consistency across surfaces and application methods. Fifth-generation quaternary ammonium compounds and glutaraldehyde exhibited the highest virucidal activity, but only on smooth, non-porous surfaces. In contrast, metallic pruning shears repeatedly retained infectious virions regardless of formulation or application method, identifying them as a persistent reservoir in greenhouse systems. Mechanical agents such as powdered milk and soap were partially protective but failed to achieve complete inactivation. No uniform dose–response trend was observed, consistent with a saturation-type effect where increasing concentration does not guarantee improved efficacy. The most effective treatments were validated in tomato transmission assays, confirming reduced or absent systemic infection. Together, these findings demonstrate that ToBRFV sanitation efficacy is determined by the interaction among chemical formulation, surface type, and application method, rather than dose alone, and provide experimentally grounded, operationally relevant guidelines for integrated ToBRFV management in protected horticulture.