Strain-specific quantitative detection of two putative biocontrol strains for suppression of ash dieback

•Workflow for a genome-based strain-specific detection by qPCR.•Detection systems were successfully established for two strains and are easily adaptable for other biocontrol agents.•The strains C2P003 and D4P002 could be traced on inoculated as well as on freshly grown parts of ash seedlings. Plant diseases caused by microorganisms are a major threat to agricultural and forest ecosystems and production. The application of microorganisms as biocontrol agents can be a more sustainable way to suppress pathogens and/or strengthen plant health. In this study, we wanted to demonstrate an efficient way to monitor the inoculation success of promising bacterial strains on plants by qPCR, using the example of common ash and the bacterial strains Luteimonas fraxinea D4P002 and Aureimonas altamirensis C2P003. For the design of the strain-specific qPCR systems, we used a workflow mainly based on the comparison of a target genome with a reference genome database by the software Neptune. We were able to find specific sequences for both strains D4P002 and C2P003, and the designed qPCR systems were found to be highly sensitive, strain specific and the applied approach showed a high recovery rate from plant material. Strain-specific qPCR assays were applied to monitor the colonization and persistence of the two bacterial strains on ash seedlings eight weeks after inoculation. Both inoculated strains were found on different organs of the ash seedlings, with the highest abundance on the older inoculated leaves. On freshly grown plant parts, the inoculated strains could also be identified. The results of this study gave us the opportunity to establish an efficient way to trace inoculation success by specific quantification of the inoculated bacterial strains. Based on the described workflow, strain-specific detection systems can be easily designed for other biocontrol agents to observe their persistence after inoculation. The suggested procedure might be helpful in the evaluation of the biocontrol of plant diseases in horticulture, forestry or agriculture.