Abstract: OBJECTIVE To establish a microbiological database for a pharmaceutical water system in the high-risk injection manufacturer, so as to provide guidance for effective control and traceability investigation of microorganisms polluted by the pharmaceutical water system. METHODS Microorganisms were monitored and collected in the purified water system and water for injection system of large-capacity injection manufacturer for four consecutive quarters, 16S rRNA and ITS rDNA sequencing methods were used to identify contaminated microorganisms. Statistical analysis was performed by combining the information of microbial species and source. A microorganisms library of pharmaceutical water system was established. RESULTS It was shown that 1 499 strains of bacteria and 20 strains of mold were isolated from pharmaceutical water system. Microorganisms in the water for injection and purified water systems accounted for 9.0% and 91.0%, respectively. Sphingomonas and Methylobacteria were the most in purified water system, accounting for 28.9% and 25.2%, respectively. Staphylococcus and Micrococcus were most in the water for injection system, accounting for 24.9% and 18.2%, respectively. The contaminated microorganisms in the purified water system were mainly Gram-negative bacteria, accounting for 67.0%. The contaminated microorganisms in the water for injection system were mainly Gram-positive bacteria, accounting for 60.3%. The more common fungi in pharmaceutical water system were Aspergillus versicolor and Penicillium chrysogenum. In addition, the analysis found that the number of microorganisms in the purified water system accounted for more in the second and third quarters, accounting for 67.8% of the total microorganisms in the year, and the number of microorganisms in the first and fourth quarters decreased. The number of microorganisms in the water for injection system was higher in the second quarter, and the numbers were similar in the remaining three seasons. CONCLUSION Establishing a microbial library for a pharmaceutical water system can guide the operation and maintenance of pharmaceutical water system and disinfection, and also can provide a source of traceability investigation for microbial contamination incidents in high-risk pharmaceutical manufacturer.