Camellia oleifera, a woody plant that produces edible oil, is indigenous to China. The devastating disease of anthracnose inflicts significant financial losses onCa. oleifera. The primary causative agent of anthracnose onCa. oleiferaisColletotrichum fructicola. Chitin, a pivotal constituent of fungal cell walls, assumes a critical function in their proliferation and maturation. To study the biological functions of chitin synthase 1(Chs1) inC. fructicola, theCfCHS1gene knockout mutants, ∆Cfchs1-1 and ∆Cfchs1-2, and their complementary strain, ∆Cfchs1/CfCHS1, ofC. fructicolawere generated. Our results showed that the colony diameters of wild-type and complement-strain ∆Cfchs1/CfCHS1, mutant ∆Cfchs1-1 and ∆Cfchs1-2 cultured on the CM and MM medium were 5.2, 5.0, 2.2 and 2.4 cm and 4.0, 4.0, 2.1 and 2.6 cm, respectively, which were significantly smaller for the mutant than for the wild type and complement strain; the inhibition rates on the CM medium supplemented with H2O2, DTT, SDS and CR were 87.0% and 88.5%, 29.6% and 27.1%, 88.0% and 89.4%, and 41.7% and 28.7%, respectively, for the mutant strains, ∆Cfchs1-1 and ∆Cfchs1-2, which were significantly higher than those for the other two strains; the rate of hyphal tips with CFW fluorescence in ∆Cfchs1-1 and ∆Cfchs1-2 was 13.3% and 15.0%, which was significantly lower than those for the other two strains; the mutant strains, ∆Cfchs1-1 and ∆Cfchs1-2, lost the ability to produce conidia; the mutant strains showed weaker pathogenicity on wounded and unwoundedCa. oleiferaleaves than the wild type and complement strain. The findings of this study suggest that CfChs1 plays a crucial role in the growth and development, stress responses, and pathogenicity ofC. fructicola. Thus, this gene could be a potential target for developing novel fungicide.