Protein O-mannosyltransferases (PMTs) initiate O-mannosylation of secretory proteins, which are of fundamental importance in eukaryotes. The human fungal pathogen Aspergillus fumigatus possesses three genes encoding for PMTs, namely, Afpmt1, Afpmt2 and Afpmt4. We have previously shown that lack of AfPmt1 leads to a temperature-sensitive phenotype featured with severe defects in hyphal growth, conidiation, cell wall integrity and morphology at elevated temperatures. In this study, a conditional mutant P2 was constructed by replacing the native promoter of the Afpmt2 with the Aspergillus nidulans alcA promoter. Reduced expression of the Afpmt2 gene led to a lagged germination, retarded hyphal growth, reduced conidiation and defect in cell wall integrity; however, no temperature-sensitive growth was observed. Further analysis revealed that reduced expression of the Afpmt2 caused a failure of the actin re-arrangement. Our results suggest that Afpmt2 gene was required for growth and played a role distinct from that of the Afpmt1 in A. fumigatus.