To evaluate the relationship between renal pelvis pressure and infection after ureteroscopy, using a live swine model. In anaesthetised pigs, a 1-h ureteroscopy was performed using a pressure-sensing guidewire, with renal pelvis pressure maintained at either 37 mmHg or 75 mmHg for the entire procedure and infusion with saline alone or with a standardised concentration of uropathogenic Escherichia coli strain CFT073 (1.5 × 107 colony-forming units [CFU]/mL). Venous blood sampling was performed during and after the procedure. Vital signs, inflammatory biomarkers, and renal tissue and blood cultures were assessed. In 21 pig kidneys, study groups were: 37 mmHg with saline irrigation (n = 3); 75 mmHg with saline irrigation (n = 4); 37 mmHg with saline irrigation with 1.5 × 107 CFU/mL E. coli (n = 7); and 75 mmHg with saline irrigation with 1.5 × 107 CFU/mL E. coli (n = 7). Statistically significant changes in inflammatory biomarkers were most pronounced in the group with 75 mmHg saline irrigation + E. coli and were significantly elevated compared with the control group and the group receiving E. coli irrigation at 37 mmHg. Positive blood cultures were noted in 5/7 animals treated with E. coli at 75 mmHg; no others developed bacteraemia. In this swine model of ureteroscopy, irrigation with saline + E. coli at a renal pelvis pressure of 75 mmHg resulted in bacteraemia and inflammatory biomarker elevations significantly greater than both E. coli irrigation with renal pelvis pressure maintained at 37 mmHg and the control.