Abstract: OBJECTIVE To explore the prevalence of cytomegalovirus infection in neonates with hearing impairment and further analyze the relationship between viral load in urine and hearing impairment so as to provide guidance for clinical study. METHODS A total of 100 neonates with hearing impairment who were treated in the hospital from Jan 2011 to Jan 2015 were recruited as the study objects, then the brainstem auditory evoked potential (BAEP) examination was conducted for all the enrolled neonates, the fasting venous blood specimens were collected to detect IgM antibody of cytomegalovirus, the cytomegalovirus DNA contents in urine were determined for the neonates with cytomegalovirus infection by using fluorescence quantitative PCR, and the relationship between the viral load in urine and the hearing impairment was observed. RESULTS Of the 100 neonates with hearing impairment, 20 (20.0%) had the cytomegalovirus infection; 60.0% of the neonates with cytomegalovirus infection showed abnormal BAEP, significantly higher than 28.8% of the neonates with non-cytomegalovirus infection (P<0.05). The viral load of the cytomegalovirus infection neonates with abnormal BAEP was significantly higher than that of the cytomegalovirus infection neonates with normal BAEP (P<0.05). As the viral load 5.1 was set as the critical point, 11 neonates had high viral load, and 9 neonates had low viral load; 54.5% of the neonates with high viral load had severe damage to hearing threshold, significantly higher than 22.2% of the neonates with low viral load (P<0.05). CONCLUSION The neonates with cytomegalovirus infections tend to have the hearing impairment; the viral load in urine is positively correlated with the degree of hearing impairment but can not indicate the type of impairment.