Talaromyces rugulosus O1 is an ochratoxin A (OTA)-producing pathogen from grapes. The mechanisms involved in the infection of postharvest grapes by T. rugulosus O1 are not yet clear. In this study, the infection of grapes by T. rugulosus O1 and OTA production in grapes were investigated, and the role of cell wall degrading enzymes of T. rugulosus O1 and OTA in the infection of grapes by T. rugulosus O1 was explored. The results indicated that T. rugulosus O1 showed significant infection effect on grapes at 20 °C, and accumulated OTA in the wound of grapes in the infection progress. In comparison with the control, the activities of polygalacturonase (PG), pectin methylgalacturonase (PMG), exo-1,4-β-glucanase (C1) and β-glucosidase (β-G) secreted by T. rugulosus O1 in grapes were all increased in the infection process. Furthermore, the pectinase (PG and PMG) activities were higher than cellulase (C1 and β-G) activities. The expression levels of genes encoding PG and β-G in T. rugulosus O1 infecting grapes increased as the time prolong. Inoculation of T. rugulosus O1 in combination with addition of OTA in the wound of grapes resulted in higher decay incidence and lesion diameter of treated grapes compared with the treatment with T. rugulosus O1 or OTA alone. This indicated that OTA can promote the infection of grapes by T. rugulosus O1. This study provides useful information about the role of cell wall degrading enzymes and OTA in the infection of grapes by T. rugulosus O1.