Abstract: In recent years, the problem of water shortage has gradually become prominent, and to solve the contradiction between the supply and demand of drinking water resources, the development, and utilization of unconventional water source is especially important. The secondary effluent of the sewage treatment plant can be used as an unconventional water source for drinking water reuse through an advanced treatment. The removal methods of pathogenic microorganisms in the secondary effluent include disinfection, advanced oxidation, media filtration, and membrane filtration. Slow filtration is employed as an advanced treatment way to examine its efficacy in removing opportunistic pathogens（OPs） and dissolved organic carbon（DOC） from the secondary effluent under various operating circumstances（C/N and pH of influent）. Additionally, the OPs removal mechanism of slow filtration and the community structure of microorganisms on its surface are studied.The results demonstrate that slow filtration with biofilm is superior to slow filtration without biofilm in eliminating OPs and DOC under different operating conditions. Under different influent C/N conditions, the increase in C/N helps to improve the removal effect of biofilm slow filtration on OPs, while too high C/N would inhibit the removal of OPs, and the removal effect is greater than that of without biofilm slow filtration, and the optimal C/N is 10, and the removal rates of Pseudomonas aeruginosa, Legionella and Mycobacterium avium are 91.2%, 94.8%, and 93.1%, respectively. Under different influent pH conditions, the removal effect of biofilm slow filtration on OPs in water is greater than that of biofilm slow filtration effluent, and when the influent water is neutral（pH=7）, biofilm slow filtration has the best removal effect on OPs, and the removal rates of OPs under the above conditions are 87.0%, 91.3%, and 84.1%, respectively. Under different influent C/N conditions, the removal effect of biofilm slow filtration on DOC is better than that of no biofilm slow filtration, and the removal first increases and then decreases, and the C/N is the best when C/N is 10, and the removal rate is 53.8%. Under different influent pH conditions, the removal effect of biofilm slow filtration on DOC in water is better than that of no biofilm slow filtration, and when it is neutral（pH=7）, biofilm slow filtration has the best removal effect on DOC, with a removal rate of 31.5%. Under the two optimal conditions, Pseudomonas aeruginosa, Legionella, and Mycobacterium avium are positively correlated with DOC in biofilm slow filtration effluent. When the influent C/N is 10, the biofilm diversity is the highest and the species composition is the most uniform. When the pH value is neutral, the species diversity is higher, while when the pH value is acidic and alkaline, the biomass decreases and the biofilm diversity decreases.Compared with the secondary effluent, the dominant population structure and number of biofilm microorganisms on the slow filtration surface changes to different degrees under the conditions of C/N 10 and pH 7. When C/N is 10, Planctomycetes and Rhizobium are dominant bacteria. At pH 7, the dominant genera are Nitrospirae and Novosphingobium.