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In resource-rich settings, universal adoption of a 4- rather than 6-week neonatal antiretroviral (ARV) prophylaxis regimen could reduce toxicity and results in cost savings, provided prevention of mother-to-child transmission program effectiveness is not compromised.Between January 1999 and December 2008, a 10-year study of the observational database of the Irish prevention of mother-to-child transmission program that uses a 4- rather than 6-week neonatal ARV prophylaxis regimen was undertaken. Maternal and infant data were analyzed to determine the vertical transmission rate (VTR) and infant outcome. Infants were categorized as uninfected if, off ARVs, they had 2 negative human immunodeficiency virus (HIV) polymerase chain reaction (PCR) tests, the second at 3 months of age or older.Between January 1999 and December 2008, there were 964 HIV-exposed live births. Excluding 7 early neonatal deaths, 4 weeks of ARV prophylaxis was prescribed for 957 infants: 61% received mono, 32% triple, and 7% dual therapy. Of 957 infants, 906 were uninfected, 10 infected, and 41 of indeterminate status. Twenty-four of the indeterminate status infants had at least one negative HIV PCR test at ≥6 weeks and 17 were lost to follow-up before 6 weeks of age. On the basis of 916 infants of known outcome, the VTR was 1.09% (95% confidence interval, 1.07–1.11). If restricted to 910 infants whose mothers received at least 4 weeks of antiretroviral therapy (ART), the VTR was 0.4%.This study provides evidence to support the current clinical practice toward use of a 4-week neonatal ARV prophylaxis regimen.