The aims of this study were to describe the adherence to CDC guidelines for intrapartum antibiotic prophylaxis (IAP) and to identify possible factors influencing noncompliance with guidelines. We conducted a retrospective study in Italy. Our cohort included women in whom antenatal Group B Streptococcus (GBS) screening was not performed, was performed, but results were not available at the time of labor or delivery and women who were positive for GBS colonization. The indications for complete execution of IAP according to revised CDC guidelines was evaluated. It was considered adequate when performed with a recommended antibiotic at least four hours prior to delivery. The cohort included 902 women. Among those who had performed rectal and vaginal swabs (or recto-vaginal swabs), results were available in 86.9% of vaginal swabs and in 87.1% of rectal swabs and GBS was detected in 59.8% of vaginal swabs and in 71% of rectal swabs. 49.2% women had indication for GBS prophylaxis. Among these, 91.1% received an antibiotic during labor. Totally appropriate IAP was performed in 36.3% deliveries, an inappropriate antibiotic was administered in 10.4% women, the remaining 45.3% women received partially appropriate IAP; of these, 15.5% had received antibiotics through an inappropriate route of administration, 18.2% an inappropriate dosage regimen. Overall, 27.5% women received intrapartum ampicillin with inappropriate timing. Multivariate analysis showed that totally appropriate prophylaxis was significantly more likely in women who had no previous live birth, who had vaginal delivery, and a positive result at antenatal GBS screening. Despite satisfactory GBS screening implementation, there is still a substantial gap between optimal and actual IAP. We hypothesize that the complexity of the CDC guidelines may partially explain this shortcoming. Future efforts will include initiatives focused at enabling and reinforcing adherence to evidence-based prevention practices.