Main moulds were isolated, identified from the fermented rice in the process of traditational Zhejiang rosy vinegar production, and their application test in the vinegar fermentation was conducted. In the total 43 isolates, 15 isolates were belonged to Aspergillus, 8 isolates to Penicillium, 3 isolates to Mucor, 2 isolates to Rhizopus, 2 isolates to Monascus, the other 13 isolates has not been identified. The results suggested that there was an implicated microbial flora in the Fahua phase of Meigui vinegar production. In the comparison of enzymes test, isolated M1 had a high glucoamylase activity(6288U), while M17 had a high protease activity (1620.1U). By the Biolog application, M1 and M17 were assigned to Rhizopus oryzae and Aspergillus oryzae respectively. Then Koji (moulded rice) of M1 and M17 were prepared and inoculated into 500kg rice to test their application of Fahua fermetnation. The obtained results suggested that the inoculation of M1 and M17 shorten the process of Fahua phase in traditional Zhejiang rosy vinegar production.