Combined efficacy of Bacillus amyloliquefaciens CIM95 and melatonin in reducing anthracnose in strawberries (Fragaria × ananassa Duch.) by enhancing antioxidant activity and defense-related genes.

Anthracnose caused by Colletotrichum acutatum causes substantial losses in yield and fruit quality. This study aimed to investigate the combined effect of Bacillus-derived lipopeptides (LPs) and melatonin (MLT) on the suppression of anthracnose in strawberries. Compared with the control, ten different Bacillus species were screened for their antifungal activity in vitro, with Bacillus amyloliquefaciens CIM95 resulting in the greatest colony growth inhibition of C. acutatum, followed by B. subtilis LLCG89. LPs were extracted from the two strains, CIM95 and LLCG89, where CIM95 resulted in greater colony growth inhibition of the pathogen. Liquid chromatography time of flight mass spectrometry (LC-TOF/MS) analysis of CIM95 revealed the presence of eight different LPs; surfactin, bacillibactin, bacillaene, fengycin, bacillomycin D, bacilysin, iturin, and plantazolicin. The application of MLT on strawberry fruit at different concentrations revealed that the 5 mM concentration resulted in the lowest lesion diameter and disease incidence of anthracnose in strawberry fruit. Our findings revealed that, compared with individual application or no treatment, combined treatment with CIM95 LPs and MLT reduced the disease incidence and lesion diameter of the pathogen on strawberry fruit. Furthermore, CIM95 LPs and MLT presented the greatest increase in the levels of key antioxidant enzymes, viz., SOD, POD, CAT, PPO, PAL, APX, CHI, and GLU. Additionally, the expression of PR1, PR5, PPO, GLU APX, CAT, PAL, NPR1, GST, and CH-1 was up upregulated till 48 h and subsequently decreased to 96 h. These results suggest that the combination of CIM95 LPs and MLT represents a sustainable and eco-friendly approach for managing anthracnose in strawberries.