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Urinary tract infections (UTI) are the most common serious bacterial infection in febrile infants. Urinalysis (UA) is a screening test for preliminary diagnosis of UTI. UA can be performed manually or using automated techniques. We sought to compare manual versus automated UA for urine specimens obtained via catheterization in the pediatric emergency department.In this prospective study, we processed catheterized urine samples from infants with suspected UTI by both the manual method (enhanced UA) and the automated method. We defined a positive enhanced UA as ≥10 white blood cells per cubic millimeter and presence of any bacteria per 10 oil immersion fields on a Gram-stained smear. We defined a positive automated UA as ≥2 white blood cells per high-powered field and presence of any bacteria using the IRIS iQ200 ELITE. We defined a positive urine culture as growth of ≥50,000 colony-forming units per milliliter of a single uropathogen. We analyzed data using SPSS software.A total of 703 specimens were analyzed. Prevalence of UTI was 7%. For pyuria, the sensitivity and positive predictive value (PPV) of the enhanced UA in predicting positive urine culture were 83.6% and 52.5%, respectively; corresponding values for the automated UA were 79.5% and 37.5%, respectively. For bacteriuria, the sensitivity and PPV of a Gram-stained smear (enhanced UA) were 83.6% and 59.4%, respectively; corresponding values for the automated UA were 73.4%, and 26.2%, respectively. Using criteria of both pyuria and bacteriuria for the enhanced UA resulted in a sensitivity of 77.5% and a PPV of 84.4%; corresponding values for the automated UA were 63.2% and 51.6%, respectively. Combining automated pyuria (≥2 white blood cells/high-powered microscopic field) with a Gram-stained smear resulted in a sensitivity of 75.5% and a PPV of 84%.Automated UA is comparable with manual UA for detection of pyuria in young children with suspected UTI. Bacteriuria detected by automated UA is less sensitive and specific for UTI when compared with a Gram-stained smear. We recommend using either manual or automated measurement of pyuria in combination with Gram-stained smear as the preferred technique for UA of catheterized specimens obtained from children in an acute care setting.