Elimination of Curtobacterium sp. strain A7_M15, a contaminant in Prunus rootstock tissue culture production, using reduced graphene oxide-silver-copper and silver-selenium nanocomposites

Abstract

Background: Bacterial contamination poses a high risk to the successful establishment and maintenance of plant tissue cultures. The aim of this study was to identify the isolates representing the frequent bacterial contaminants of Prunus rootstock tissue cultures and to determine the most effective concentration of nanomaterials for Curtobacterium sp. strain A7_M15 elimination without a negative impact on explants. Results: Six Curtobacterium sp. strains were isolated and identified, and the whole-genome sequence was obtained for strain A7_M15. Two nanocomposites, reduced graphene oxide-copper-silver and silver-selenium, with the highest bactericidal activity were selected for elimination of Curtobacterium sp. contamination in Gisela 5 rootstock tissue cultures. Both nanocomposites showed 100% inhibition of bacterial plaque formation on culture medium at concentrations of 100, 200 and 400 mg L-1 Ag (2 x-8 x MBC). The quantity of Curtobacterium sp. on culture medium assessed using cfu enumeration was reduced by 92% and 74% in comparison to the positive control after treatment with reduced graphene oxide-silver-copper and silver-selenium at a concentration of 200 mg L-1 Ag, respectively. None of the tested concentrations resulted in a decrease in Curtobacterium sp. quantity in explants. Curtobacterium sp. was detected in donor Gisela 5 plants, indicating an endophytic character of this bacterium. The dry weight of explants was not negatively affected by the application of nanocomposites regardless of concentration, and no detrimental effect of either nanocomposite at 100 or 200 mg L-1 Ag on the surface covered by plants was observed. Conclusions: Reduced graphene oxide-silver-copper and silver-selenium nanocomposites at 200 mg L-1 Ag effectively limited the Curtobacterium sp. presence in micropropagated Prunus rootstock without causing phytotoxicity; therefore, those treatments could be offered as prevention with a high activity against bacterial contamination in plant tissue cultures.