Fluorescence Detection of Pyrene-Stained Bacillus amyloliquefaciens MHR24 in Tomato (Solanum lycopersicum L.) Stem Tissues.

The PGPR strain of Bacillus amyloliquefaciens MHR24 (MHR24) was recently reported as a strong biocontrol strain. In this study, MHR24 was used to investigate phyllosphere effects during inoculations of tomato leaves (Solanum lycopersicum L.). When MHR24 was inoculated on foliar tissue, it caused apical chlorosis symptoms at 3-6 days after infiltration or submersion, which suggests that the bacterium may adopt a potentially pathogenic lifestyle in the phyllosphere. In order to detect the MHR24 interaction with the plant, it was stained with the commercial fluorophore 8-hydroxypyrene-1,3,6-trisulfonic acid trisodium salt, selected from a pyrene series bearing diverse functional groups, based on several in vitro staining assays. Fluorescence used as a detection signal was observed by LSCM mainly in the vascular bundles, suggesting that rhizobacteria may preferentially colonize these tissue regions. Molecular docking, performed by analyzing the possible interactions between the outer membrane protein assembly factor BamB of the family protein B. amyloliquefaciens and the fluorophore, indicates that hydrogen bonds with serine 126 (SER126), serine 182 (SER182), isoleucine 180 (ILE180), and tryptophan 66 (TRP66), charges attraction and π-stacking with TRP66, and non-bonded attractions with leucine 224 (LEU224) can occur, which likely gives rise to a stable complex. These results are important in view of the application of MHR24 as part of a sustainable approach for increasing tomato crop production.