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Background: Cytomegalovirus (CMV) is the most common cause of congenital infections in newborns which can lead to long-term complications in more than half of the cases with symptomatic infection at birth time. Unfortunately, neonates with congenital CMV infection will mostly remain undiagnosed because the golden time for detection is limited to the first 3 weeks of infants' life. This study aimed to determine the prevalence of congenital CMV infection in newborns admitted to intensive care units of hospitals in Tehran, Iran and assess related risk factors associated with the infection. Methods: In this cross-sectional study from April to October 2017, newborns within the first three weeks of life who were admitted to the neonatal intensive care units (NICUs) of university-affiliated hospitals in Tehran, Iran, were eligible for enrollment. CMV infection in neonates was diagnosed through testing infants' Guthrie cards and detection of viral DNA via an in-house nested-PCR assay. Congenital CMV infection in neonates with positive results was confirmed by testing urine specimens as a sensitive and gold standard sample. Related data (demographic and maternal factors) were collected by questionnaires and analyzed. Results: Congenital cytomegalovirus infection was diagnosed in 8 of 63 newborns (12.7%). Hearing loss was seen in 2 infected infants. The mean of head circumferences among infected neonates was significantly lower than that observed in uninfected cases. Infants with CMV related symptoms had statistically more chance to have infection (P=0.02). We also found Guthrie cards as a reliable sample with high sensitivity for CMV detection assays. Conclusion: The current study showed a high rate of symptomatic congenital CMV infection among neonates attending on NICU sections of hospitals in Tehran, Iran. It is of crucial importance to note that based on evidence, diagnosis of infants with congenital CMV infection at early stages could help to decrease the burden of long)term diseases if associated with prompt interventions and reduce the costs of late)ineffective treatment. Therefore, routine screening of newborns for congenital CMV infection via Guthrie cards is suggested. [ABSTRACT FROM AUTHOR]