Urinary tract infection is a bacterial infection caused by bacteria from the digestive tract entering the urinary tract. It is more prevalent in women and remains the most common bacterial infection in humans. The study aimed to assess the prevalence of urinary tract infections (UTIs) in pregnant women, identify and diagnose uropathogenic bacteria, and examine the antibiotic susceptibility of the isolated bacteria. A total of 283 urine specimens were collected from pregnant and non-pregnant females of different ages who visited Amedi hospital between October 2020 and January 2021. The samples were cultured and incubated on Blood agar and MacConkey agar plates for 24 hours at 37°C. Bacterial isolates were identified, and their antibiotic sensitivity was assessed using the Vitek-2 method. The study found a total UTI rate of 42.4% (120/283), with higher rates in pregnant patients (58.3%) compared to non-pregnant patients (41.7%). Among 120 positive urine cultures, bacterial isolates were identified in 92.5% (111/120), with Gram-positive bacteria being more prevalent (51.4%) than Gram-negative (48.6%). The most common Gram-negative bacteria was Escherichia coli (29.7%), while Gram-positive bacteria included Enterococcus faecalis, Staphylococcus haemolyticus, and Streptococcus agalactiae (each 12.6%; 14/111). The rate of fungal positive cultures (Candida spp.) was 7.5%. Gram-negative isolates showed high sensitivity to tigecycline (99.0%) and ertapenem (80.6%), but low sensitivity to ampicillin (2.7%) and cefuroxime (16.1%). Gram-positive isolates were highly sensitive to linezolid (98.7%) and tigecycline (98.5%). The findings of this study are valuable for understanding the nature of urinary tract infections (UTIs) and guiding appropriate treatment, leading to a reduction in the misuse of antibiotics.