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Abstract:

“Crazy roots” (also known as “hairy root disease”) caused by rhizogenic Agrobacterium biovar 1 strains has become a major problem in hydroponics greenhouses, especially in the cultivation of tomato and cucumber (Weller et al., 2006). Hairy root disease is characterized by excessive root development, leading to a strong vegetative growth and severe losses in marketable yield. Importantly, infections are often persistent as many Agrobacterium strains are able to form biofilms in the irrigation system. Prevention and removal of biofilms are considered to be crucial in management of the disease. To this end, hydrogen peroxide (H2O2) is frequently used in hydroponics systems as a disinfecting agent. However, in a recent study it was demonstrated that many rhizogenic Agrobacterium biovar 1 strains show catalase activity, enabling them to survive exposure to H2O2 (Bosmans et al., 2015). Therefore, the effectiveness of H2O2 to control Agrobacterium containing biofilms can be questioned. The objective of this study was to evaluate the effectiveness of H2O2 to control biofilms that harbour rhizogenic agrobacteria using a pilot-scale system. Material and Methods Experiments were performed using the pilot-scale system described by Vankerckhoven et al., (2011). Briefly, the system consists of four subsystems (allowing to test different H2O2 concentrations at once), each containing a water container. From each container water was pumped through a piping system harboring a biofilm sampling device consisting of polycarbonate rings. Following inoculation with Agrobacterium (106 cells per ml), biofilms were allowed to develop on the polycarbonate rings for 65 h. Subsequently, H2O2 was applied at different concentrations, including 25 ppm, 50 ppm and 100 ppm. A treatment in which no H2O2 was applied was used as negative control. Next, every 24 h bulk water and biofilm samples were taken to quantify the number of Agrobacterium cells as well as the total number of culturable bacteria. Quantification was performed by quantitative PCR (rhizogenic agrobacteria; Bosmans et al., 2016), and traditional culture-plate enumeration on Plate Count Agar, respectively. The experiment was performed for a catalase-positive and catalase-negative Agrobacterium biovar 1 strain. Results and Conclusions While application of 25 ppm H2O2 was ineffective for both the catalase-positive and catalase-negative strain, substantial differences between both strains were observed for the other concentrations tested. More particularly, while treatment of the catalase-negative strain with 50 ppm resulted in a significant reduction in Agrobacterium cells, the same treatment was ineffective for the catalase-positive Agrobacterium strain. When 100 ppm H2O2 was applied both the catalase positive and catalase negative strain were affected. These results clearly indicate that the efficacy of the H2O2 treatment is strongly dependent on the particular Agrobacterium strain present in the greenhouse. Moreover, the presence of a catalase-positive Agrobacterium strain is able to protect other bacteria against H2O2 treatment as well. Altogether, our data suggest that the treatment against hairy roots disease should be attuned to the specific Agrobacterium strain present in the greenhouse.