Objective To explore the indwelling time of needle-free infusion connectors of peripherally inserted central venous catheter (PICC). Methods Tumor patients with chemotherapy who underwent PICC maintenance in a tertiary first-class hospital from April to October 2019 were selected and divided into group A and B according to the types of PICC needle-free infusion connectors. Group A used opaque metal spring mechanical valve needle-free infusion connectors (infusion connector A), group B used transparent silica gel mechanical valve needle-free infusion connectors (infusion connector B), needle-free infusion connectors were collected for qualitative and quantitative bacterial culture, positive culture rates of infusion connectors were compared between two groups when PICC was retained for 4-, 8-, 15-, and >21 days. Results A total of 174 patients were included in the study, quantitative bacterial culture of 11 patients with PICC needle-free infusion connectors was positive, positive rate was 6.32%, positive rates of group A and group B were 8.70% (8/92) and 3.66% (3/82) respectively. There was significant difference in the positive rate of quantitative culture of needle-free infusion connectors with different indwelling time in group A (P < 0.05), but there was no significant difference in group B (P>0.05); positive rates of qualitative culture of needle-free infusion connectors with different indwelling time were not significantly different in both group A and group B (P>0.05). There was no significant difference in the positive rates of both quantitative and qualitative culture of needle-free infusion connectors with the same indwelling time between group A and group B (both P>0.05). Positive rate of culture was 0 for 8-14 indwelling days in group A and for 15-21 indwelling days in group B. 11 strains of 5 species of bacteria were isolated, Bacillus cereus (4 strains) was most common specie. Conclusion Quantitative bacterial culture of infusion connectors in group A with different indwelling time are diffe-rent, there is no difference in bacterial culture positive rate between transfusion connectors A and B with the same indwelling time.