Objective To describe the prevalence of microcephaly in infants admitted to the neonatal intensive care units (NICUs) in eight private hospitals in south-eastern and midwestern Brazil, from 2011 to 2015.

Design Observational, cross-sectional study in a cohort of neonates.

Setting Eight private NICUs situated in the cities of Rio de Janeiro (RJ), São Paulo (SP) and Federal District of Brasilia (FDB).

Patients Neonates up to 7 days of age and >23 weeks of gestational age.

Main outcome measures Primary outcomes were the annual prevalence of microcephaly and severe microcephaly, both overall and in subgroups according to gestational age, type of microcephaly (proportional or disproportional) and the NICU's location in RJ, SP or FDB.

Results In this cohort of 8275 neonates admitted to eight private NICUs, the overall prevalence of microcephaly was 5.6% (95% CI 5.1% to 6.1%) and severe microcephaly, 1.5% (95% CI 1.2% to 1.7%). There was no annual variation in these overall estimates (p=0.48 and p=0.99, respectively), nor in the studied subgroups. Microcephaly prevalence was higher in extremely premature and term newborns (7.7% and 7.2%; p<0.001, respectively). Disproportional microcephaly was much more common than proportional microcephaly (prevalence 5.0% vs 0.6%; p<0.001).

Conclusions The prevalence of microcephaly showed little variation from 2011 to 2015 in these south-eastern and midwestern Brazilian NICUs, suggesting that the impact of the Zika epidemic in neonates that occurred mainly in the north-eastern region in 2015 did not yet affect the infants of those regions during the studied period. These prevalence results can serve as a reference for comparisons with any future Zika outbreaks in the same or similar populations from these regions.