Abstract:[Objective] To explore the impacts of methylene blue (MB) , a cationic dyes usually used as tracer agent for periprosthetic joint infection, on the culture results of different bacteria in vitro. [Methods] Eight standard bacterial strains were prepared in vitro, includ- ing staphylococcus aureus (SAU) , staphylococcus epidermidis (SEP) , staphylococcus hominis (SHO) , escherichia coli (ECO) , klebsiella pneumoniae (KPN) , bacterial suspensions of acinetobacter baumannii (ABA) , streptococcus pyogenes (SPY) and candida albicans (CAL) , for simulation of bacterial culture of synovial fluid. The bacterial suspensions of all the standard bacterial strains were diluted 50 times and divided into three groups with 5 samples in each group, and then treated with 0.1%MB and 0.05%MB and 0.45% sterile saline, respective- ly. The colony count was compared among the 3 groups. [Results] The colony counts of SHO and ABA strains were ranked from low to high in sequence of 0.1%MB group <0.05% MB group < saline group, with a statistically significant difference (P<0.05) . However, there were no significant differences in colony counts in terms of SEP, ECO, KPN, SPY and CAL among the 3 groups (P>0.05) . As results of Spear- man correlation analysis, the MB concentration was significantly negatively correlated with SHO and ABA colony counts (P<0.05) , whereas which had no correlation with colony counts of SAU, SEP, ECO, KPN, SPY and CAL (P>0.05). [Conclusion] The MB has dose-dependent inhibitory effect on SHO and ABA, but has no effect on the other 6 bacterial strains.