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Introduction Spinal dysraphism (SD) is a general term used to refer to developmental abnormalities of the spine that involves many clinical conditions including myelomeningocele (MMC). In these patients, neurogenic bladder (NB) is a common and predisposing factor for renal damage; the most frequently used approach to manage this situation is based on clean intermittent catheterization (CIC) and anticholinergic drugs. Urinary tract infections (UTIs) are a significant concern for these patients, and antibiotic prophylaxis is frequently used even if it is still a debated topic of literature. The purpose of this paper is to investigate the role and the real effectiveness of antibiotic prophylaxis in the reduction of incidence of UTIs in patients with spina bifida performing CIC. Methods We collected data of all patients performing CIC, who did their last follow-up visit in the period between January 2019 and January 2021, followed at the children multidisciplinary Spina Bifida Center of A. Gemelli Hospital in Rome. Data collected included age at referral, gender, type of SD lesion, serum creatinine and cystatin C levels, the use of anticholinergic medications, antibiotic prophylaxis and type of prophylaxis (oral/endovesical), age of starting prophylaxis with its duration/adherence, number of CIC/day and its duration, episodes of UTIs in the 2 years prior to the last follow-up, and presence and grade of vesical-ureteric reflux (VUR) on cystourethrogram. Results A total of 121 patients with SD performing CIC was included in the study; 66 (54%) presented ≥ 1 episode of UTIs in the last two years and 55 (46%) none. During the study period, 85 (70%) patients received antibiotic prophylaxis (ABP group) and 36 (30%) did not (NABP group): no statistically significative difference in terms of UTI development was observed between the two groups (p = 0.17). We also evaluated compliance to the therapy; 71 patients (59%) took antibiotic prophylaxis constantly (CABP group) and 50 (41%) did not do antibiotic prophylaxis constantly or did not do antibiotic prophylaxis at all (NCABP group): we observed a statistically significative difference in terms of UTIs with a 2.2 times higher risk of development at least one episode of UTIs in NCABP group. Conclusion In conclusion, antibiotic prophylaxis performed constantly, without interruption, is associated with a lower risk of developing urinary tract infections and consequently to develop renal failure in adulthood.