Objective: To explore the practical application value of urine bacterial culture combined with routine urine testing in patients with urinary tract infection, compare the diagnostic performance of single detection method and combined detection, and provide a reference basis for clinical rapid identification, reasonable diagnosis and treatment of urinary tract infection. Methods: 82 patients with suspected urinary tract infection admitted to our hospital from January 2025 to January 2026 were selected as the subjects of this study. All subjects completed urine routine and urine bacterial culture examinations simultaneously. Urine bacterial culture results were used as the gold standard for clinical diagnosis, and the sensitivity, specificity, and diagnostic accuracy of single urine routine testing and the combined testing of the two methods were calculated, as well as the distribution of major pathogenic bacteria. Results: Among 82 suspected patients, 51 cases were confirmed to have urinary tract infection by urine bacterial culture. The sensitivity of the single urine routine test was 74.51%, the specificity was 70.97%, and the diagnostic accuracy was 73.17%. After the two methods were combined for testing, the sensitivity increased to 94.12%, the specificity was 90.32%, and the diagnostic accuracy was 92.68%. The diagnostic indicators of the combined testing were significantly better than those of urine routine testing alone, and the difference in data between groups was statistically significant (P<0.05). Among the pathogenic bacteria isolated, Escherichia coli accounted for the highest proportion. Conclusion: Routine urinalysis can complete the rapid initial screening of urinary tract infection, but when used alone, there are certain cases of missed diagnosis and misdiagnosis. Combining routine urinalysis and urine bacterial culture for testing can effectively improve the overall diagnostic level. It can not only achieve rapid screening, but also clarify the pathogenic bacteria and drug susceptibility, which has very practical significance in guiding the rational selection of antibacterial drugs in clinical practice.