Novel mutations in CYP51B from Penicillium digitatum involved in prochloraz resistance

Green mold caused by Penicillium digitatum is one of the most serious postharvest diseases of citrus fruit, and it is ubiquitous in all citrus growing regions in the world. Sterol 14α-demethylase (CYP51) is one of the key enzymes of sterol biosynthesis in the biological kingdom and a prime target of antifungal drugs. Mutations in CYP51s have been found to be correlated with resistance to azole fungicides in many fungal species. To investigate the mechanism of resistance to prochloraz (PRC) in P. digitatum, the PRC sensitivity was determined in vitro in this study to assess the sensitivity of 78 P. digitatum isolates collected in Hubei province. The results showed that 25 isolates were prochloraz-resistant (PRC-R), including six high-resistant (HR) strains, twelve medium-resistant (MR) and seven low-resistant (LR) strains. A sequence analysis showed no consistent point mutations of PdCYP51A in the PRC-R strains, but four substitutions of CYP51B were found, Q309H in LR strains, Y136H and Q309H in HR strains, and G459S and F506I in MR strains, which corresponded to the four sensitivity levels. Based on the sequence alignment analysis and homology modeling followed by the molecular docking of the PdCYP51B protein, the potential correlation between the mutations and PRC resistance is proposed.