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Background. In preterm infants cytomegalovirus (CMV) infection acquired through maternal milk has been related to morbidity. However, infants who may receive higher titers of protective antibodies from highly seropositive mothers have not been studied in detail.Methods. A cohort of 188 ≤30-week-old infants was monitored from admission to discharge. CMV-DNA, hematology, liver enzymes, neutralizing antibodies, and CMV-DNA-lactia were tested periodically.Results. Mothers of 157 infants (83.5%) were CMV-seropositive. A total of 24/157 (15.3%) infants became infected (95% confidence interval [CI], 9.8–22.6), particularly those of lower gestational age (GA; relative risk [RR], 2.32; 95% CI, 1.01–5.34 for 23–26 weeks). Low (<1:64) neutralizing antibody titers were similarly detected in CMV-infected and uninfected infants. Mean DNA-lactia in mothers was higher in CMV-infected than in uninfected infants (5.34 log vs 4.60 log). Clinical findings suggestive of CMV disease were similar in CMV-infected (50.0%) and CMV-uninfected (51.1%) infants. Although transitory, >2 laboratory test abnormalities occurred more frequently among CMV-infected (39.1%) than CMV-uninfected (2.1%) infants. More severe stages of retinopathy of prematurity (ROP) were found among CMV-infected infants (adjusted RR, 2.51; 95% CI, 1.07–5.91). Although deaths were more frequent among infected infants, none of the deaths could be directly attributed to CMV.Conclusions. Postnatal CMV infection acquired by exposure to raw maternal milk is very frequent among extremely premature newborns, being facilitated by high DNA-lactia and lower GA, regardless of maternally acquired neutralizing antibody levels. The association with advanced stages of ROP is a concern and needs to be further explored in larger studies.