Valsa canker, caused by Valsa mali, is a destructive disease of apple in China. The pathogen infects apple branches, mainly through pruning wounds, and causes branch and tree death. To determine the conditions required for V. mali infection through pruning wounds and growth within the xylem, pruning wounds on 1- to 4-year-old apple branches were inoculated with conidia in vitro under artificially controlled conditions and in vivo in the orchard. The effects of temperature, wetness duration, and wound age on conidial infection through pruning wounds as well as hyphal growth in the xylem were examined. The results showed that, after invading through pruning wounds, V. mali hyphae grew along xylem vessels, tracheids, and rays, expanding longitudinally and laterally. The hyphae could enter adjacent xylem vessels and tracheids through micropores to form a dense hyphal network. Wetness duration did not exhibit an essential effect on conidial infection from pruning wounds. Conidia spread to pruning wounds with rainwater could infect the xylem without any other extra moisture. Temperature for V. mali conidia infection through pruning wounds and hyphal extension in the xylem ranged from 5 to 35°C, with the optimum at 20°C. Pruning wounds made in late March were susceptible to V. mali infection in March, April, and May; the susceptibility was markedly deceased by June, and the pathogen could barely infect through the pruning wounds in November. The infected pruning wounds began to show symptoms from the spring of the following year. More than half of the observed Valsa canker lesions emerged in the spring of the second year, and new canker twigs were also developed from the inoculations in the spring of the third year. March, April, and May are the critical periods for protecting pruning wounds against infection by V. mali in China, and coating pruning wounds with protective film immediately after pruning is an easy and effective measure to protect the pruning wounds.