Valsa sordida and Botryosphaeria dothidea are two crucial necrotrophic fungal pathogens that damage many plant hosts, particularly species in the genus Populus. These two fungal pathogens occur mainly in poplar branches, stems, and twigs, causing classic symptoms such as canker lesions, canopy dieback, and wilting. Pathogen inoculation is the most efficient pathway to study the mechanism of plant disease. Besides the canker lesions around the inoculation sites on the stems, a novel developmental phenomenon, copious adventitious roots (ARs) with bright red color, were observed in poplar species after stem canker pathogen inoculations. In this study, we described the method for inducing ARs using fungal pathogens in poplar trees. The crucial step of this method is the pathogen inoculation after (phloem or epidermis) girdling manipulation. The second crucial step is the application of the moisturizing material. Compared to the moisturizing manipulation with Parafilm, wrapping the inoculated sites with household polyethylene (PE) plastic wrap can produce colorful, numerous, and robust ARs in 20 days after girdling-inoculation. Finally, white ARs sprouted from the inoculated rings in the poplar stems after shading treatment (wrapping the stems with aluminum foil). This method introduces a novel experimental system for studying root development and morphogenesis, which is crucial for understanding the biology of root development, morphogenesis, and response under disease stress. Furthermore, when combined with shading treatment, this study can provide a convenient experimental system for investigating light response-related processes, for example, the biosynthesis of flavonoids, anthocyanins, or other related metabolites, and genes or transcription factors involved in these processes.