Detection of congenital cytomegalovirus infection by real-time polymerase chain reaction analysis of saliva or urine specimens.

Viral culture of urine or saliva has been the gold standard technique for the diagnosis of congenital cytomegalovirus (CMV) infection. Results of rapid culture and polymerase chain reaction (PCR) analysis of urine and saliva specimens from 80 children were compared to determine the clinical utility of a real-time PCR assay for diagnosis of congenital CMV infection. Results of urine PCR were positive in 98.8% of specimens. Three PCR-positive urine samples were culture negative. Results of saliva PCR and culture were concordant in 78 specimens (97.5%). Two PCR-positive saliva samples were culture negative. These findings demonstrate that PCR performs as well as rapid culture of urine or saliva specimens for diagnosing congenital CMV infection and saliva specimens are easier to collect. Because PCR also offers more rapid turnaround, is unlikely to be affected by storage and transport conditions, has lower cost, and may be adapted to high-throughput situations, it is well suited for targeted testing and large-scale screening for CMV.

Genotypic diversity and mixed infection in newborn disease and hearing loss in congenital cytomegalovirus infection.

BACKGROUND: Congenital cytomegalovirus (cCMV) is a common congenital infection and a leading nongenetic cause of sensorineural hearing loss (SNHL). CMV exhibits extensive genetic variability, and infection with multiple CMV strains (mixed infection) was shown to be common in congenital CMV. The role of mixed infections in disease and outcome remains to be defined. METHODS: Genotyping of envelope glycoproteins, UL55 (gB), UL73 (gN) and UL75 (gH), was performed on saliva specimens of 79 infants from the ongoing CMV and Hearing Multicenter Screening (CHIMES) Study and on blood and urine specimens of 52 infants who participated in natural history studies at the University of Alabama at Birmingham. Genotyping of UL144 and US28 was also performed in the CHIMES cohort. The association of individual genotypes and mixed infection with clinical findings at birth and SNHL was examined. RESULTS: Thirty-seven of 131 infants (28%) were symptomatic at birth and 26 (20%) had SNHL at birth. All known genotypes of UL55, UL75, UL73 and US28 were represented, and no particular genotype was associated with symptomatic infection or SNHL. UL144 subtype C was more common in symptomatic infants but not associated with SNHL. Mixed infection was observed in 59 infants (45%) and not associated with symptoms (P = 0.43) or SNHL at birth (P = 0.82). In the cohort of 52 infants with long-term hearing outcome, mixed infection at birth was not predictive of SNHL. CONCLUSIONS: Mixed infection is common in infants with congenital CMV but is neither associated with symptomatic infection nor associated with SNHL.

Diagnostic consequences of cytomegalovirus glycoprotein B polymorphisms.

Failure of a cytomegalovirus (CMV) real-time PCR assay targeting glycoprotein B (gB) was investigated. A multiplex assay targeting gB and immediate-early 2 (IE2) genes showed discordant results (gB negative and IE positive or a >10-fold-higher viral load with IE primers) in saliva from 14.6% of CMV-infected newborns. Sequencing revealed 3 patterns of gB variations.

Saliva polymerase-chain-reaction assay for cytomegalovirus screening in newborns.

BACKGROUND: Congenital cytomegalovirus (CMV) infection is an important cause of hearing loss, and most infants at risk for CMV-associated hearing loss are not identified early in life because of failure to test for the infection. The standard assay for newborn CMV screening is rapid culture performed on saliva specimens obtained at birth, but this assay cannot be automated. Two alternatives--real-time polymerase-chain-reaction (PCR)-based testing of a liquid-saliva or dried-saliva specimen obtained at birth--have been developed. METHODS: In our prospective, multicenter screening study of newborns, we compared real-time PCR assays of liquid-saliva and dried-saliva specimens with rapid culture of saliva specimens obtained at birth. RESULTS: A total of 177 of 34,989 infants (0.5%; 95% confidence interval [CI], 0.4 to 0.6) were positive for CMV, according to at least one of the three methods. Of 17,662 newborns screened with the use of the liquid-saliva PCR assay, 17,569 were negative for CMV, and the remaining 85 infants (0.5%; 95% CI, 0.4 to 0.6) had positive results on both culture and PCR assay. The sensitivity and specificity of the liquid-saliva PCR assay were 100% (95% CI, 95.8 to 100) and 99.9% (95% CI, 99.9 to 100), respectively, and the positive and negative predictive values were 91.4% (95% CI, 83.8 to 96.2) and 100% (95% CI, 99.9 to 100), respectively. Of 17,327 newborns screened by means of the dried-saliva PCR assay, 74 were positive for CMV, whereas 76 (0.4%; 95% CI, 0.3 to 0.5) were found to be CMV-positive on rapid culture. Sensitivity and specificity of the dried-saliva PCR assay were 97.4% (95% CI, 90.8 to 99.7) and 99.9% (95% CI, 99.9 to 100), respectively. The positive and negative predictive values were 90.2% (95% CI, 81.7 to 95.7) and 99.9% (95% CI, 99.9 to 100), respectively. CONCLUSIONS: Real-time PCR assays of both liquid- and dried-saliva specimens showed high sensitivity and specificity for detecting CMV infection and should be considered potential screening tools for CMV in newborns. (Funded by the National Institute on Deafness and Other Communication Disorders.).

Simplified dosing of gentamicin for treatment of sepsis in Bangladeshi neonates.

Extended-interval dosing of gentamicin has several advantages over conventional multiple-daily dosing for the treatment of sepsis. The study was conducted to evaluate the pharmacokinetics of gentamicin for the treatment of neonatal sepsis in predetermined doses at 24- or 48-hour intervals, according to weight category, and to develop a simplified protocol for use in peripheral healthcare settings in developing countries. This prospective observational study was conducted among 59 neonates admitted to the Special Care Nursery at Dhaka Shishu Hospital, Bangladesh, with suspected sepsis and treated with antibiotics, including gentamicin. Intravenous dosing of gentamicin according to weight category was: 10 mg every 48 hours if the infant weighed < 2,000 g (n = 23), 10 mg every 24 hours if the infant weighed 2,000-2,249 g (n = 12), or 13.5 mg every 24 hours if the infant weighed 2,500-3,000 g (n = 24). Peak and trough concentrations of gentamicin and the presence of signs of nephrotoxicity and ototoxicity were determined. The mean +/- standard deviation peak concentration of gentamicin was 12.3 +/- 3.7 microg/mL in infants weighing < 2,000 g, 9.6 +/- 3.1 microg/mL in infants 2,000-2,249 g, and 10.0 +/- 3.4 microg/mL in infants 2,500-3,000 g. Initial peak concentration of gentamicin was > 12 microg/mL in 28.8% and initial trough concentration was > 2 microg/mL in 6.8% of the subjects. No signs of nephrotoxicity or ototoxicity were detected. Favourable pharmacokinetic parameters found with the simplified dosing regimen suggest that it is safe for the treatment of neonatal sepsis.