The disinfection efficiency of performic acid (PFA) against various microbial contaminants has been studied in municipal secondary effluent. The study demonstrated that PFA provides rapid, efficient and safe disinfection, degrading both bacteria and viruses even at low doses. The resistance order starting from the most resistant microorganism is as follows: MS2-coliphages > DNA-coliphages > enterococci and Escherichia coli. PFA is also efficient in the elimination of Salmonella spp., Clostridium perfringens spores and Giardia cysts. The results showed that a PFA dose as low as 0.5–1 mg L–1 with contact time of 10 min was efficient in achieving and maintaining for 72 h the disinfection level required for unrestricted agricultural water reuse (≤3 log units for faecal coliforms). However, the optimal dose will depend on the quality of wastewater. Regarding the formation of by-products during disinfection with PFA, very low amounts of hydrogen peroxide and organic per-acids were observed; active oxygen was not detected. The amounts of adsorbable organically bound halogens (AOX) compounds formed were significantly lower compared to the amounts generated during chlorine disinfection. This chlorine-free solution enables compliance with microbiological criteria for various water reuse applications and is already on the market for advanced disinfection.