Congenital Cytomegalovirus Infection in the Netherlands: Birth Prevalence and Risk Factors

ArticleinJournal of Medical Virology 83(10):1777-82 · October 2011with 68 Reads
DOI: 10.1002/jmv.22181 · Source: PubMed
Abstract
Congenital cytomegalovirus (CMV) infection is the most common congenital viral infection worldwide. The sequela encountered most frequently is hearing impairment, affecting approximately one out of five infants congenitally infected. Data on the birth prevalence and risk factors of congenital CMV infection in the Netherlands are scarce. The aim of this study was to determine the birth prevalence of congenital CMV in the Netherlands. A sample of 6,500 dried blood spots (DBS) from infants born in the Netherlands was tested anonymously for CMV DNA. The sample was stratified by the number of live births in different regions of the Netherlands of the year 2007. Additionally, on a regional level, risk factors for congenital CMV were analyzed. The birth prevalence of congenital CMV in the Netherlands was 0.54% (35/6,433, 95%CI 0.36-0.72). Congenital CMV infection was significantly higher in regions with more than 15% young children (0-5 years) compared with regions with a lower proportion of young children (OR 5.9, 95%CI 1.4-25.2). Congenital CMV infection was significantly higher in regions with more than 30% immigrants compared with regions with a lower proportion of immigrants (OR 2.2, 95%CI 1.1-4.6). This association was strongest for regions with more than 30% non-Western immigrants (OR 3.3, 95%CI 1.5-7.5). Based on the knowledge of the natural history of congenital CMV infection, approximately 1,000 children are born with congenital CMV infection in the Netherlands annually, of whom eventually approximately 180 children (0.1% of all newborns) will be affected by long term sequelae, with hearing loss being the symptom encountered most frequently.
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  • Article
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    Cytomegalovirus (CMV) is a leading cause of congenital infection in neonates. Most infants with congenital CMV infection are asymptomatic at birth and not diagnosed on routine clinical examination. To identify these at-risk infants early in life, polymerase chain reaction (PCR) assays are done to screen large populations of newborn infants. We carried out a pilot study to estimate the prevalence of CMV in saliva from newborns by DNA PCR assay. This study was performed from January 2012 to March 2012 at a maternity hospital in the south of Tehran. All newborns aged between 1 to 14 days born at this hospital were enrolled. Saliva specimens from newborns were collected by swabbing the inside of the baby's mouth and stored at -70 degrees C until PCR processing for virus detection. Six-hundred and twenty infants between 1 to 14 days of age were enrolled during the study period of two months. The PCR assay was positive for CMV in 2 newborns [0.3%]. Both of these infants were asymptomatic for congenital CMV at birth and also when followed up at three months and six months of age. Our findings reveal that because of a low yield of positive results, screening for congenital CMV infection would not be cost-effective in Iranian neonates.
  • Article
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  • Article
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  • Article
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  • Article
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  • Article
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  • Article
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  • Article
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  • Article
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  • Article
    Because maternal seropositivity for CMV is associated with substantial protection against congenital CMV infection, prevention measures have focused mainly on seronegative pregnant women for decades. However, population-wide insight in the contribution of nonprimary infection (reactivation and/or re-infection with a different strain) on the most common sequela, hearing loss, is missing. A population-based prediction model was developed to estimate the proportion of congenital CMV-related hearing loss resulting from nonprimary maternal infection. Incorporated was a meta-analysis of the risk of hearing loss, calculating pooled proportions of children with hearing loss after nonprimary and primary infection. Subsequently, the model was applied for worldwide present population seroprevalences (range 30–95%). It was estimated that, for all population seroprevalences, nonprimary maternal infections are responsible for the majority of congenital CMV infections. This proportion increased with seroprevalence, ranging from 57% (95%CI 24–85%) to 96% (95% CI 88–99%) for seroprevalences of 30% to 95%. Our meta-analysis (six reports) showed that the risk of hearing loss after nonprimary infection was 11% (28/253 children, 95% CI 7–15%) versus 13% (50/385 children, 95% CI 10–16%) after primary infection. Incorporating this risk into our model, we estimated that nonprimary infections also accounted for the majority of CMV-related hearing loss. This proportion ranged from 53% (95% CI 13–86%) to 95% (95% CI 62–99%) for seroprevalences of 30% to 95%. Our data underline the worldwide contribution of nonprimary infections in causing CMV-related hearing loss. These results imply that prevention research such as vaccine and hygiene studies should not only be directed at seronegative but also seropositive pregnant women. Copyright
  • Article
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  • Article
    Only 10-15% of neonates with congenital cytomegalovirus infection have symptoms at birth. The most common symptoms are intrauterine growth retardation, microcephaly, petechiae, jaundice, hepatosplenomegaly, intracranial abnormalities, ophthalmological abnormalities and hearing loss.- Symptomatic and asymptomatic CMV infections can both have long-term effects.- CMV infection during pregnancy is diagnosed using a blood test and possible testing of the amniotic fluid for viral DNA. Infection of the fetus may be prevented by treating the mother with CMV hyperimmune globulin.- In the neonate a diagnosis can be made by viral culture or PCR in urine. PCR in saliva could be an alternative. Blood testing is of limited value.- If symptoms of CMV infection occur in the neonate, such as petechiae, microcephaly, central nervous system abnormalities, sensorineural hearing loss or chorioretinitis, antiviral treatment should be considered.- Long-term follow-up is advisable because of the possibility of delayed-onset hearing loss and chorioretinitis.
  • Article
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  • Article
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  • Article
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  • Article
    Full-text available
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  • Article
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  • Article
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  • Article
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    Intrauterine infections are a crucial pathogenic factor exerting an appreciable influence on the development of the fetus. They can provoke intrauterine death, cause multiple lesions in the organs and tissues as well as long-term complications that manifest themselves at the later stages of the growth and development of the child. One of such complications is the sensorineural loss of hearing. The importance of hearing impairment arises from the high prevalence of tis condition and frequent incapacitation it causes in the patients. The present publication is focused on various mechanisms underlying the development of hearing impairment depending on the primary infection. Special attention is given to the methods of diagnostics and treatment of intrauterine infections.
  • Article
    Purpose of review: Until recently, management options in congenital cytomegalovirus (cCMV) infection have been either conservative or termination of pregnancy. However, medical therapies aimed at reducing the risk of infection and/or its severity have recently been investigated. Recent findings: In a phase 2 open label, nonrandomized trial, valaciclovir (ValACV) was given to women carrying a CMV-infected fetus. ValACV was associated with a greater proportion of asymptomatic neonates when compared with a historical cohort (82 vs. 43%). However, the study design and the small number of treated women limit its applicability. Even though initial observational data suggested that hyperimmune globulin (HIG) therapy in pregnancy was associated with a significantly lower risk of cCMV, its efficacy has not been borne out in a subsequent phase 2 randomized, placebo controlled, double-blind study [cCMV 30% in the HIG group, 44% in the placebo group (P = 0.13)]. Furthermore, 11% of fetuses in the HIG group had transient or permanent abnormalities, compared with 16% in the placebo group. Summary: ValACV might have a promising role in the antenatal treatment of cCMV infection, but definitive recommendations require further research. The use of HIG should currently be limited to the research setting.Video abstract http://links.lww.com/COID/A18.
  • Article
    Objective: Congenital cytomegalovirus infection (cCMV) can cause symptoms at birth as well as long-term impairment. This study estimates cCMV-related healthcare costs in the Netherlands in early childhood. Design, setting and patients: In a nationwide retrospective cohort study, 156 children with cCMV were identified by testing 31 484 neonatal dried blood spots for cCMV. Use of healthcare resources in the first 6 years of life by children with cCMV and a matched cCMV-negative control group were analysed. Mean costs per child were calculated by multiplying healthcare resource use by its reference prices. Exposure: Children with cCMV were compared with cCMV-negative children. Main outcome measures: The average total healthcare costs per child were based on the average costs for hospital admissions and consultations by healthcare providers. Results: Mean healthcare costs of children with cCMV (€6113, n=133) were higher than children without cCMV (€3570, n=274), although statistically not significant, with a mean difference of €2544 (95% CI €-451 to €5538). The costs of children with long-term impairment were two times higher in children with cCMV (€17 205) compared with children without cCMV (€8332). Conclusions: Children with cCMV, especially those with long-term impairment and those symptomatic at birth, accrue higher healthcare costs than cCMV-negative children in the first 6 years of life, although this is not statistically significant. This economic impact is of importance in the evaluation of preventive measures against cCMV. Trial registration number: NTR3582.
  • Article
    Full-text available
    Genotyping of cytomegalovirus (CMV) is useful to examine potential differences in the pathogenicity of strains and to demonstrate coinfection with multiple strains involved in CMV disease in adults and congenitally infected newborns. Studies on genotyping of CMV in dried blood spots (DBS) are rare and have been hampered by the small amount of dried blood available. In this study, two multiplex real-time PCR assays for rapid gB and gH genotyping of CMV in DBS were developed. Validation of the assays with 39 CMV-positive plasma samples of transplant recipients and 21 urine specimens of congenitally infected newborns was successful in genotyping 100% of the samples, with gB1 and gB3 being the most prevalent genotypes. Multiple gB and gH genotypes were detected in 36% and 33% of the plasma samples, respectively. One urine sample from a newborn with symptomatic congenital CMV was positive for gB1 and gB2. DBS of congenitally infected newborns (n = 41) were tested using 9 μl of dried blood, and genotypes were detected in 81% (gB) and 73% (gH) of the samples, with gB3 being the most prevalent genotype. No clear association of specific genotypes with clinical outcome was observed. In conclusion, the CMV gB and gH PCR assays were found to be rapid, sensitive for detecting mixed infections, and suitable for direct usage on DBS. These assays are efficient tools for genotyping of CMV in DBS of congenitally infected newborns.
  • Article
    Full-text available
    Capillary blood samples from 63 infants collected 3-7 days after birth, and thereafter stored on filter papers for 12-18 y, were tested for the presence of CMV DNA by the polymerase chain reaction (PCR) method. Of 16 infants with proven congenital CMV infection (positive virus isolation test in urine sampled within 1 week of age), 13 (81%) had a positive CMV PCR test and 3 (19%) a negative PCR test. All blood samples from 16 control infants without congenital CMV infection (negative virus isolation test in urine sampled within 1 week of age) were CMV PCR-negative. When 31 samples on filter papers stored above or below the samples of the infected infants were tested, 6 (19%) had a weak reactivity. This suggests that CMV DNA can be transferred from one filter paper to another during storage. We conclude that PCR performed on dried blood stored on filter paper is a useful method in the retrospective diagnostics of congenital CMV infection. Consideration must be given, however, to the possibility of transfer of CMV DNA from blood samples stored nearby.
  • Evaluatie van de neonatale hielprikscreening bij kinderen geboren in 2007 Congenital cytomegalovirus infection and hearing loss
    • Rijpstra A Lanting Ci
    • Breuning-Boers Jm
    • Verkerk
    • Ph
    Lanting CI, Rijpstra A, Breuning-Boers JM, Verkerk PH. Evaluatie van de neonatale hielprikscreening bij kinderen geboren in 2007. TNO-rapport KvL/P&Z 2008.119. 2010. Pass RF. 2005. Congenital cytomegalovirus infection and hearing loss. Herpes 12:50–55.
  • Evaluatie van de neonatale hielprikscreening bij kinderen geboren in 2007
    • Ci Lanting
    • A Rijpstra
    • Jm Breuning-Boers
    • Ph Verkerk
    Lanting CI, Rijpstra A, Breuning-Boers JM, Verkerk PH. Evaluatie van de neonatale hielprikscreening bij kinderen geboren in 2007. TNO-rapport KvL/P&Z 2008.119. 2010.
  • Birth prevalence and implications for disease burden of congenital CMV in the Netherlands b Symptomatic: Petechiae, jaundice, hepatosplenomegaly, thrombocytopenia, chorioretinitis, seizures, microcephaly, intracranial calcifications, or fetal hydrops
    • Figlanting
    Fig. 1. Birth prevalence and implications for disease burden of congenital CMV in the Netherlands. a [Lanting et al., 2010]. b Symptomatic: Petechiae, jaundice, hepatosplenomegaly, thrombocytopenia, chorioretinitis, seizures, microcephaly, intracranial calcifications, or fetal hydrops [Dollard et al., 2007]. c [Dollard et al., 2007]. d Sequelae: Sensorineural hearing loss (uni-and bilateral), cognitive deficit (mental retardation, neurological impairment and developmental delay), and motor deficit [Dollard et al., 2007].
  • Article
    Context Vaccine development to prevent congenital cytomegalovirus (CMV) infection has been impeded by the uncertainty over whether maternal immunity protects the fetus.Objective To determine whether the presence of maternal antibodies to CMV significantly reduces the risk of congenital CMV infection in future pregnancies.Design, Setting, and Participants Cohort study of 3461 multiparous women from a population with a high rate of congenital CMV infection who delivered newborns screened for congenital CMV infection between 1993 and 1998, and whose cord serum specimen from a previous delivery could be retrieved and tested for antibody to CMV.Main Outcome Measure Congenital CMV infection according to maternal immune status, age, race, parity, and socioeconomic status.Results Of 604 newborns born to initially seronegative mothers, congenital CMV infection occurred in 18 (3.0%). In contrast, of 2857 newborns born to immune mothers, congenital CMV infection occurred in 29 (1.0%) Two factors, preconception maternal immunity (adjusted risk ratio, 0.31; 95% confidence interval, 0.17-0.58) and maternal age of 25 years or older (adjusted risk ratio, 0.19; 95% confidence interval, 0.07-0.49), were highly protective against congenital CMV infection. No other factors were associated with a reduction in the risk of congenital CMV infection.Conclusion Naturally acquired immunity results in a 69% reduction in the risk of congenital CMV infection in future pregnancies.
  • Article
    Background: Dried blood spots (DBS) may be valuable in the diagnosis of congenital cytomegalovirus (CMV) infection. However, the 2007 European Quality Control for Molecular Diagnostics (QCMD) proficiency testing programme showed that CMV DNA detection in DBS was lacking sensitivity in a considerable number of participating laboratories. Objective: To compare DNA extraction methods for DBS for detecting CMV. Sensitivity and applicability of the methods for high-throughput usage were assessed. Study design: Guthrie cards were spotted with CMV DNA-positive whole blood (n = 15). DNA was extracted from the DBS using different extraction methods, followed by CMV amplification by means of real-time PCR. Results: Significant differences between the extraction methods with respect to the sensitivity were found. Optimal sensitivity was achieved when samples were tested in triplicate, demonstrating that the methods in general operated around their detection limits. Triplicate testing using the protocol by Barbi et al. [Barbi M, et al. Cytomegalovirus DNA detection in Guthrie cards: a powerful tool for diagnosing congenital infection. J Clin Virol 2000;17:159-65], representing the most sensitive methods, resulted in sensitivities of 100%, 86%, and 50% for DBS with CMV DNA loads of 5-4, 4-3, and 3-2 log10 copies/ml, respectively. This indicates that sensitivity limitations apply in the clinically relevant concentration range. Few methods appeared suitable for 96-well format high-throughput testing. Discussion: When considering universal neonatal screening for congenital CMV infection, an assay which is both sensitive and applicable for high-throughput testing is required. The protocol by Barbi et al. and the BioRobot Universal System appear appropriate candidates currently available for 96-well format application in neonatal screening using DBS.
  • Article
    Background: The reference method of cytomegalovirus (CMV) isolation from urine or saliva is not a feasible routine technique for all newborns, and laboratory diagnosis of this infection would be useful both for epidemiological purposes and to enable prompt institution of adequate measures to identify and correct late sequelae. Extraction and amplification of viral DNA from dried blood spots (DBS) collected from babies in the first days of life during routine screening for genetic and metabolic disorders has been proposed for the early diagnosis of viral congenital infections.
  • Article
    Congenital cytomegalovirus (CMV) infection is an important public health problem with approximately 7 in 1,000 newborns infected and consequently at risk for hearing impairment. Newborn hearing screening will fail to detect this hearing impairment in approximately half of the cases because late onset hearing loss is frequent. Hearing impairment has profound impact on cognitive and social development of children and their families, determining most of the disease burden of congenital CMV infection. The potential value of newborn screening for congenital CMV is increasingly discussed. To date, many experts acknowledge the benefit of antiviral treatment in the prevention of hearing deterioration in newborns with neurological symptoms, and the benefit of early identification of late-onset hearing impairment by means of extensive audiological follow up of infected infants. These opinions imply that the potential of newborn screening for CMV would lie in the identification of the large proportion of asymptomatic congenitally infected newborns at risk for developing late-onset hearing loss. Experience with postnatal antiviral treatment of symptomatic newborns is encouraging, but has not been studied in asymptomatic congenitally infected newborns. A large-scale study on the safety and effectiveness of combined screening and antiviral therapy for congenital CMV infection is the necessary next step to take and should not be delayed.
  • Article
    To the Editor: Dr Boppana and colleagues1 concluded that DBS real-time PCR assays are not suitable for screening newborns for congenital CMV infection due to their insufficient sensitivity. We believe that this is a premature conclusion, based on a number of considerations that were not sufficiently discussed in this article.First, the sensitivity of DBS testing is highly variable, largely depending on the nucleic acid extraction methodology used,2 so conclusions cannot be generalized. It appears that this problem can be reduced by using optimized techniques that differ from those applied in the study by Boppana et al.1 In addition, performing independent triplicate testing to increase sensitivity has been advocated,2 an approach not used in this study.
  • Article
    To the Editor: The study by Dr Boppana and colleagues1 conducted newborn screening for congenital cytomegalovirus (CMV) infection and found dried blood spots (DBS) to have a testing sensitivity of 28% to 34%. We believe that the authors' conclusion that DBS PCR assays are not suitable for screening newborns for congenital CMV infection is premature.The low sensitivity may have been due in part to the specific methods used. It may be that neither the M48 robotic system nor the column extraction method used is optimal for detection of low copy number viral DNA in DBS. Demonstration of poor sensitivity with a particular diagnostic test, or even several tests, does not preclude future development of a test with sufficient sensitivity.
  • Article
    Infection with human cytomegalovirus (HCMV) is the most common congenital virus infection, affecting about 0.5-2% of newborns. Using DBS on Guthrie cards, it is possible to discriminate congenital from postnatal HCMV-infection. However, a recent European trial revealed serious problems in detection of low HCMV-DNA levels from DBS-filter-cards (Barbi et al., 2008).(7) Evaluation of the most sensitive combination of sample size, DNA extraction method and PCR system for the detection of low copy numbers of HCMV-DNA from DBS-filter-cards. We compared three different manual extraction methods for the detection of HCMV-DNA out of DBS: the QIAmp-blood-Mini-Kit, a heat-extraction-method and traditional phenol-chloroform extraction. Additionally, we tested an automated nucleic acid extraction system (NucliSense EasyMag/Biomerieux). Different punch-sizes of DBS spiked with defined HCMV AD169-DNA copy numbers were analyzed. For detection, we used a quantitative in-house-LightCycler-PCR targeting the gB-region using the hybridisation-probe-format. We compared the sensitivity of the real-time-PCR with IE1Ex4-targeted nested-PCR. The highest sensitivity with 200 copies HCMV DNA/ml was achieved using the phenol-chloroform method in combination with the nested-PCR and 6mm, 3x3mm punches or the whole DBS. The QIAmp-blood-Mini-Kit also showed a very high sensitivity by using the whole DBS and the nested-PCR. These results may have strong implications for retrospective diagnosis of congenital HCMV (cHCMV) infection, since a defined combination of the area of punch, the extraction method, and PCR method determine the probability of detection of viral DNA from DBS according to a logistic model.
  • Article
    To determine the birth prevalence of cytomegalovirus (CMV) in a population-based sample of newborns by use of dried blood spots compared with previous studies that used established detection methods, and to evaluate risk factors and birth outcomes for congenital CMV infection. A total of 3972 newborn dried blood spots collected for the California Newborn Screening Program were tested for presence of CMV DNA. Demographic and pregnancy data were obtained from linked newborn screening and live-birth records. CMV prevalence among newborns by maternal race and ethnicity was 0.9% for blacks, 0.8% for Hispanics, 0.6% for whites, and 0.6% for Asians. Among Hispanics (n = 2053), infants who were infected had younger mothers (23 vs 26 years, P = .03), and prevalence was higher for children with no father information provided (2.6% vs 0.6%, P = .03). Overall CMV infection was associated with low birth weight (prevalence ratios [95% CI]: 3.4 [1.4-8.5]) and preterm birth (2.7 [1.4-5.1]). CMV viral loads were inversely related to birth weight and gestational age (both P = .03). CMV prevalence measured with dried blood spots was similar to reports using standard viral culture methods. Dried blood spots may be suitable for detection of CMV infection in newborns and warrant further evaluation. Congenital CMV infection may contribute to low birth weight and preterm birth.
  • Article
    A significant number of asymptomatic newborns infected with congenital cytomegalovirus (CMV) will present with permanent childhood hearing impairment (PCHI) during early childhood. To investigate the role of congenital CMV infection in causing PCHI in the Netherlands, and assess the efficacy of two different hearing screening strategies and the developmental outcome following each strategy. We included 192 children with PCHI at the age of 3-5 years, who were offered hearing screening in their first year of life. Dried blood spots from 171 children were available for CMV detection using real-time PCR. The results of eight previously tested samples were also available. Clinical baseline characteristics were collected from medical records and the Child Development Inventory was used to investigate the developmental outcome. The rate of congenital CMV among the 179 children was 8% (14/179) and 23% (9/39) among children with profound PCHI. Two of eight CMV-positive children with PCHI at the age of 3-5 years had passed the newborn hearing screening (NHS) test. Developmental outcome measures showed a significantly greater delay in language comprehension in children with both PCHI and congenital CMV infection (the largest in symptomatic children) than in the children with PCHI without congenital CMV infection. Congenital CMV infection is important in the etiology of PCHI. Universal NHS is not a guarantee of normal hearing and development in childhood for children with congenital CMV infection. This is a problem which might be solved by universal congenital CMV screening.
  • Article
    Dried blood spots (DBS) may be valuable in the diagnosis of congenital cytomegalovirus (CMV) infection. However, the 2007 European Quality Control for Molecular Diagnostics (QCMD) proficiency testing programme showed that CMV DNA detection in DBS was lacking sensitivity in a considerable number of participating laboratories. To compare DNA extraction methods for DBS for detecting CMV. Sensitivity and applicability of the methods for high-throughput usage were assessed. Guthrie cards were spotted with CMV DNA-positive whole blood (n=15). DNA was extracted from the DBS using different extraction methods, followed by CMV amplification by means of real-time PCR. Significant differences between the extraction methods with respect to the sensitivity were found. Optimal sensitivity was achieved when samples were tested in triplicate, demonstrating that the methods in general operated around their detection limits. Triplicate testing using the protocol by Barbi et al. [Barbi M, et al. Cytomegalovirus DNA detection in Guthrie cards: a powerful tool for diagnosing congenital infection. J Clin Virol 2000;17:159-65], representing the most sensitive methods, resulted in sensitivities of 100%, 86%, and 50% for DBS with CMV DNA loads of 5-4, 4-3, and 3-2log(10)copies/ml, respectively. This indicates that sensitivity limitations apply in the clinically relevant concentration range. Few methods appeared suitable for 96-well format high-throughput testing. When considering universal neonatal screening for congenital CMV infection, an assay which is both sensitive and applicable for high-throughput testing is required. The protocol by Barbi et al. and the BioRobot Universal System appear appropriate candidates currently available for 96-well format application in neonatal screening using DBS.
  • Article
    Full-text available
    Reliable methods to screen newborns for congenital cytomegalovirus (CMV) infection are needed for identification of infants at increased risk of hearing loss. Since dried blood spots (DBS) are routinely collected for metabolic screening from all newborns in the United States, there has been interest in using DBS polymerase chain reaction (PCR)-based methods for newborn CMV screening. To determine the diagnostic accuracy of DBS real-time PCR assays for newborn CMV screening. Between March 2007 and May 2008, infants born at 7 US medical centers had saliva specimens tested by rapid culture for early antigen fluorescent foci. Results of saliva rapid culture were compared with a single-primer (March 2007-December 2007) and a 2-primer DBS real-time PCR (January 2008-May 2008). Infants whose specimens screened positive on rapid culture or PCR had congenital infection confirmed by the reference standard method with rapid culture testing on saliva or urine. Sensitivity, specificity, and positive and negative likelihood ratios (LRs) of single-primer and 2-primer DBS real-time PCR assays for identifying infants with confirmed congenital CMV infection. Congenital CMV infection was confirmed in 92 of 20,448 (0.45%; 95% confidence interval [CI], 0.36%-0.55%) infants. Ninety-one of 92 infants had positive results on saliva rapid culture. Of the 11,422 infants screened using the single-primer DBS PCR, 17 of 60 (28%) infants had positive results with this assay, whereas, among the 9026 infants screened using the 2-primer DBS PCR, 11 of 32 (34%) screened positive. The single-primer DBS PCR identified congenital CMV infection with a sensitivity of 28.3% (95% CI, 17.4%-41.4%), specificity of 99.9% (95% CI, 99.9%-100%), positive LR of 803.7 (95% CI, 278.7-2317.9), and negative LR of 0.7 (95% CI, 0.6-0.8). The positive and negative predictive values of the single-primer DBS PCR were 80.9% (95% CI, 58.1%-94.5%) and 99.6% (95% CI, 99.5%-99.7%), respectively. The 2-primer DBS PCR assay identified infants with congenital CMV infection with a sensitivity of 34.4% (95% CI, 18.6%-53.2%), specificity of 99.9% (95% CI, 99.9%-100.0%), positive LR of 3088.9 (95% CI, 410.8-23 226.7), and negative LR of 0.7 (95% CI, 0.5-0.8). The positive and negative predictive values of the 2-primer DBS PCR were 91.7% (95% CI, 61.5%-99.8%) and 99.8% (95% CI, 99.6%-99.9%), respectively. Among newborns, CMV testing with DBS real-time PCR compared with saliva rapid culture had low sensitivity, limiting its value as a screening test.
  • Article
    Full-text available
    Cytomegalovirus (CMV) and Parvovirus B19 infections acquired during pregnancy may result in developmental disabilities of the foetus. This study evaluates the occupational risk of these infections in female day care personnel. IgG seroprevalence was determined in 310 Dutch day care workers and 158 nursing school students. CMV seroprevalence was age-related, starting at 21% in those <20 years and reaching 65% in those >35 years. Between the ages of 20 and 24 years the CMV prevalence was higher in day care personnel than in controls, 50% versus 31% (p = 0.03). In the first 2 years of employment the risk of attracting CMV was significantly increased (OR(adj) = 3.80; p < 0.001) and the occupational risk was also increased (OR(adj) 2.19; p < 0.001). Parvovirus seropositivity (71-77%) was not related to age or working at a day care centre. In conclusion, an occupational risk was observed for CMV, but not for Parvovirus infection in female day care personnel.
  • Article
    In a prospective study of 148 children from urbanized southern Finland 3 were found to be congenitally and 48 perinatally infected with cytomegalovirus (CMV), while 6 developed "late" infection during the first year of life. During pregnancy and the first year after delivery 23 of the mothers had no CMV antibodies; none of the children of these seronegative mothers developed any type of CMV infection. Fresh blood exchange transfusions did not increase the risk of CMV infection. The data support the hypothesis that the mother is the source of perinatal CMV infection. Children with a low birthweight not due to prematurity, and first children seem to run a greater risk of acquiring perinatal CMV infection. If the child is breast fed up to the age of 2 months the risk seems to be increased. Perinatal CMV infection gave rise to no symptoms or signs and had no effect on growth or on motor and psychosocial development during the first year of life.
  • Article
    Screening of 3060 neonates for congenital cytomegalovirus (CMV) infection by virus excretion in the urine showed an overall incidence of 0.4%. The incidence was about 1% for mothers between 16 and 25 years and only 0.2% for mothers between 25 and 35. No mothers over 35 years of age gave birth to congenitally infected infants. The percentage of women in the child-bearing age susceptible to CMV infection was estimated by the absence of CMV complement-fixing antibodies in cord sera and ranged from 48% to 33% with increasing age. None of the infected infants showed obvious signs of congenital CMV infection at birth. At follow-up, two infants showed slight, but transient symptoms compatible with a foetal infection; a pair of premature twins exhibited retarded physical and psychomotor development, but this could just as well be ascribed to the prematurity itself. None of the infants had detectable CMV--IgM antibodies in cord sera, but a trend towards elevated total IgM concentration in cord sera and elevated virus excretion titres appeared in the infants with symptoms. With the very low incidence and no signs of sensomotor sequelae the preliminary conclusion is that foetal CMV infection in our population by no means has a significance to deserve screening or a vaccination programme.
  • Article
    To determine if women at risk of having babies infected with cytomegalovirus (CMV) can be identified antenatally. Prospective serological and demographic study of pregnant women and virological study of their newborn infants. Teaching hospital in London. 3315 pregnant women and 2737 of their babies. Quantitative detection of CMV IgG antibodies; qualitative detection of CMV IgM antibodies; demographic characteristics of mothers; qualitative and quantitative titration of CMV viruria in newborn. Congenital CMV infection was found in nine newborn babies (0.33%) two of whom had symptoms. Serological testing of the nine mothers showed four primary and five recurrent infections; both of the symptomatic children were born in the latter group. Testing for CMV specific IgM antibodies or quantitation of IgG antibodies in early pregnancy sera could not differentiate those women at risk of giving birth to babies infected or damaged by CMV from the rest of the population. Quantitation of viruria confirmed that those babies most at risk of CMV disease have the highest titres of CMV. (i) Since laboratory tests in pregnant women cannot reliably identify fetuses at risk of disease, screening for asymptomatic maternal infection coupled with termination of pregnancy cannot be recommended. (ii) Since 'immune' women can still give birth to babies affected by CMV, we propose that future CMV vaccines should be used to immunize children with the aim of eradicating CMV infection in preference to selective immunization of sero-susceptible females.
  • Article
    8278 (56%) of 14 789 pregnant women who were screened for cytomegalovirus (CMV) antibodies at their first antenatal visit were seropositive. 42 (3 per 1000) infants screened were congenitally infected. 3 (7%) have serious handicaps, 14 (33%) have minor or transient problems, and 25 (60%) have so far had no problems. In 26 of the 42 mothers with infected infants CMV antibodies were present in the first antenatal blood sample. 28 (67%) of the infected infants were born to mothers who had experienced a primary infection in pregnancy and 7 (17%) to mothers who had experienced recurrent infection. In the remaining 7 (17%) it was not possible to determine the type of maternal infection. One handicapped infant was born after a probable recurrent CMV infection during pregnancy. These preliminary findings suggest that routine screening of pregnant women to detect evidence of primary CMV infection is not helpful.
  • Article
    A rapid assay for detection of cytomegalovirus (CMV) in saliva was evaluated as a screening method for congenital infection. Samples of saliva were examined by detection of early antigen fluorescent foci (DEAFF) and standard tissue culture (TC). Results were compared with those from urine DEAFF. CMV was detected in saliva from 31 (1.7%) of 1870 newborns, 26 by DEAFF and TC, 1 by DEAFF alone, and 4 by TC alone. Urine DEAFF was positive in 28 of these 31 newborns. The sensitivities of various tests were saliva TC, 96.8%; saliva DEAFF, 87.1 %; and urine DEAFF, 90.3%. A change in transport medium for 825 saliva samples resulted in improved sensitivities: saliva TC and saliva DEAFF, 100%; urine DEAFF, 92.3%. Screening saliva of newborns for CMV appears to be at least as sensitive a method for detecting congenital infection as detection of viruria; saliva can be collected with less difficulty and expense than urine.
  • Article
    Cytomegalovirus (CMV) is the leading cause of congenital viral infection in the United States. To prevent damaging congenital CMV infections, it is necessary to have accurate population estimates of prevalence and to identify maternal factors associated with an elevated risk of congenital infection in the newborn. From 1980 through 1990, 17,163 offspring of predominately low-income nonwhite women who delivered at a public hospital and 9892 newborns ofpredominately mid- to upper-income white women who delivered at a private hospital were screened for congenital CMV infection. Women <20 years old (adjusted prevalence odds ratio [POR], 4.8; 95% confidence interval [CI], 2.6–8.9) at the public hospital and all nonwhite women (adjusted POR, 1.6; 95% CI, 1.1–2.2) had an increased risk of delivering an infected newborn. Newborns of adolescent women in both populations had the highest prevalence of clinically apparent infection. Offspring of nonwhite low-income adolescents are at greatest risk for congenital CMV infection and more damaging sequelae.
  • Screening newborns for congenital cytomegalovirus infection Implementing neonatal screening for congenital cytomega-lovirus: Addressing the deafness of policy makers
    • De Jjc
    • Vossen Actm
    • Acm
    De Vries JJC, Vossen ACTM, Kroes ACM. 2010. Screening newborns for congenital cytomegalovirus infection. JAMA 304:407. De Vries JJC, Vossen ACTM, Kroes ACM, Van der Zeijst BAM. 2011. Implementing neonatal screening for congenital cytomega-lovirus: Addressing the deafness of policy makers. Rev Med Virol
  • To time the onset of cytomegalovirus (CMV) infection in patients (n = 39) with CMV associated neonatal cholestasis by analysing CMV DNA on Guthrie cards sampled at 3 days of age. CMV infection was diagnosed by serology/urine isolation or by CMV DNA detection (polymerase chain reaction) in liver biopsy specimens. In order to time the infection dry blood filter paper discs were punched out from stored Guthrie cards. After phenol-choloroform extraction CMV DNA was detected by nested polymerase chain reaction. All cards from control children (n = 8) with congenital CMV tested positive; none of the negative controls (n = 4) did so. Two of 39 cholestatic infants were CMV DNA positive; their mothers had serological signs compatible with infection during the second half of the pregnancy. All other cholestatic infants tested negative. CMV DNA was not detected in most of the children using Guthrie cards, suggesting that infection developed at or soon after birth.
  • Article
    Full-text available
    This report summarizes knowledge accumulated in a long-term study of congenital and maternal cytomegalovirus (CMV) infection in Sweden. Some new findings are included. We considered diagnostic methods, sources of maternal infection (including occupational risks), roles of primary and secondary maternal infections, transmission to foetuses, incidence, symptoms and prognosis of established congenital infection and relative importance of such infection in infantile sensorineural deafness, microcephaly and type 1 diabetes mellitus. Virus isolation testing was done 1977-1985 on 16,474 newborns. 76 (0.5%) congenitally infected infants were found, 22/76 (29%) with transient neonatal symptoms and 11/60 (18%) with neurological symptoms by the age of 7 y. Type of maternal CMV infection was serologically determined in 62/76 cases (30 primary, 32 secondary). CNS disturbances in the infants occurred after both primary (all trimesters) and secondary maternal infections. The negative potential of secondary maternal infections might be an obstacle to preventive vaccination.
  • Article
    Although cytomegalovirus infection is the most common infection transmitted via the placenta, there are no guidelines for routine screening to detect children congenitally infected with cytomegalovirus. From 1993 to 1997, maternal serum and cord vein blood of newborns was screened for HCMV-IgM (n = 21,183). Urine was examined for HCMV-excretion during the first postnatal week to prove HCMV infection in children who expressed HCMV-IgM in cord vein blood (n = 13) or who were born to mothers positive for HCMV-IgM in the serum (n = 234), or when both cord vein blood and maternal serum were positive for HCMV-IgM (n = 6). Congenital HCMV infection was detected in 17 newborns. To determine the incidence of congenital HCMV infection, only those mother/child pairs were selected in whom serum and cord vein blood were investigated (n = 5967 mother/child pairs). In this group 13 newborns were infected. The observed incidence for congenital HCMV infection is 0.21%. It is concluded that that this screening programme will detect those children at risk for congenital HCMV infection. These children have to be examined for virus excretion in the urine. Although the observed incidence is only 0.21%, congenital HCMV infection is a problem that can no longer be neglected because of its long-term sequelae.
  • Article
    A simple and reliable diagnosis of congenital cytomegalovirus infection is necessary both for clinical and epidemiological purposes. This could be accomplished through the demonstration of cytomegalovirus (CMV) DNA in blood spots (DBS) on Guthrie cards. (1) To assess the sensitivity and specificity of the method (DBS test) in diagnosing congenital CMV infection compared with viral isolation and (2) to evaluate the applications of the test to the late diagnosis of congenital CMV. The method was tested on the cards of (1) 509 babies examined through viral isolation within their third week of life (72 positive cases) and (2) 191 children studied after 3 weeks of life (25 days to 5 years). Blood was eluted from Guthrie cards and heat extracted. The products of a nested polymerase chain reaction (PCR) amplifying one region in the CMV glycoprotein B (gB) gene were detected by agarose gel electrophoresis. DBS test was positive in all 72 congenitally infected babies and in four of the 437 negative at cytomegalovirus isolation (sensitivity 100%, specificity 99%). Infection in 16 of the 92 infants with a late viral isolation was demonstrated to be congenital by the test, which also detected congenital infection in 18 of 83 children in whom viral culture was not performed (13 with and five without symptoms). Fifty-six additional control cases tested negative. DBS test is a reliable assay for diagnosing congenital cytomegalovirus infection and could be used as an alternative to viral culture. It is able to reveal whether ascertained CMV infection is congenital or postnatal at an age when viral isolation is not able to do so. It can assess the role of risky procedures such as transfusion and it can ascertain the etiology of morbid conditions diagnosed late or of controversial origin.
  • Article
    A rapid test for the diagnosis of congenital CMV infection is still needed. This study evaluated the usefulness of dried blood and urine samples collected on filter paper for detecting cytomegalovirus (CMV) by the polymerase chain reaction (PCR) assay compared with the use of liquid urine. Samples were obtained from 332 infants aged 1-7 days. Liquid urine samples were collected into bags, cultured in human fibroblasts, and processed using a multiplex PCR technique. Dried urine samples were obtained by placing a piece of filter paper in contact with the infant's genitals. The heels of neonates were punctured and capillary blood was blotted onto filter paper and dried. Dried blood and urine specimens were analyzed by multiplex PCR and nested-PCR assays. A diagnosis of congenital CMV infection was established by isolating the virus, and by detecting viral DNA in the liquid urine. Of the 332 liquid urine samples collected from 332 neonates, seven (2.1%) were positive for CMV and 325 were negative, by both cell culture and PCR assay. In dried samples, CMV DNA was detectable only with a nested PCR assay. Compared with known CMV infection status, 5/7 (71.4%) neonates were positive for congenital CMV infection using dried blood samples. All 325 uninfected neonates were negative. In the dried urine samples, 4/4 CMV-infected infants gave positive tests, and all 262 uninfected infants were negative. Although further improvements in sample collection and/or processing are still needed, PCR testing on dried urine or blood collected on filter paper is a promising approach in the diagnosis of neonatal CMV infection.
  • Article
    Congenital human cytomegalovirus (hCMV) infection is the most common intrauterine viral disease in western countries. Little is known about hCMV virus load in various body fluids of congenitally infected children. To determine virus load in various body fluids. To assess the impact of hCMV virus load to predict the outcome of congenitally infected newborns and efficacy of antiviral therapy. Cord vein blood, urine, and cerebrospinal fluid (CSF) of congenitally hCMV-infected children were investigated and hCMV load was determined by quantitative polymerase chain reaction (PCR). Fourteen of 30 children had clinical symptoms and/or pathological laboratory results and 16 had none of them at birth. Ganciclovir was given to 21 children (10 of them with symptoms, 11 of them without symptoms). Viral load before and after therapy was measured. There was a significant difference between median virus load in cord vein blood (2.3 x 10(3) copies per ml) and in urine (4.2 x 10(5) copies per ml; P<0.001) at diagnosis of congenital hCMV infection. At that time, no significant difference of virus load was found between the various groups (symptomatic vs. asymptomatic; with therapy vs. without therapy), neither in serum nor in urine. Comparing median virus load in urine before (3.0 x 10(5) copies per ml) and after therapy (2.0 x 10(3) copies per ml), a significant decrease was observed (P<0.001). Virus load in CSF was always found to be less than 400 copies per ml, and only those children with symptoms showed a positive result. At birth, virus load in urine seems to be superior to that in cord vein blood to reflect the situation in the organs precisely. As predicting factor for the risk of developing symptoms, only hCMV detection in the CSF appears to be promising. The significant decrease of virus load in children with therapy may reflect the efficacy of therapy. Studies including a greater number of children are needed.
  • Article
    Diagnostic problems in identifying congenital infection cases in infancy have thus far impaired the assessment of the role of congenital cytomegalovirus (CMV) infection in the etiology of sensorineural hearing loss (SNHL). To estimate the impact of congenital infection in children with SNHL by detection of CMV DNA in stored samples of neonatal dried blood (dried blood spots test). The Guthrie cards of 130 children with hearing loss >40 dB hearing loss were retrieved from the regional screening center. CMV DNA was extracted by thermal shock and amplified by PCR. The percentage of SNHL cases attributable to congenital CMV infection was 10% (9 of 87) in infants whose SNHL had been diagnosed in their first 2 months of life and 34.2% (13 of 38) in children with deafness of unidentified cause that was diagnosed in early childhood. In the latter group 42.7% (12 of 28) of the children with a hearing loss of >70 dB were CMV-positive. The results suggest that congenital CMV infection has a more relevant role in the etiology of SNHL than previously reported. The data obtained in both groups suggest that 20 to 30% of all deafness cases are caused by CMV. The percent of congenital CMV cases alone appears to account for all the cases previously attributed to all congenital infections. More than 40% of deafness cases with an unknown cause, needing rehabilitation, are caused by congenital CMV.
  • Article
    Vaccine development to prevent congenital cytomegalovirus (CMV) infection has been impeded by the uncertainty over whether maternal immunity protects the fetus. To determine whether the presence of maternal antibodies to CMV significantly reduces the risk of congenital CMV infection in future pregnancies. Cohort study of 3461 multiparous women from a population with a high rate of congenital CMV infection who delivered newborns screened for congenital CMV infection between 1993 and 1998, and whose cord serum specimen from a previous delivery could be retrieved and tested for antibody to CMV. Congenital CMV infection according to maternal immune status, age, race, parity, and socioeconomic status. Of 604 newborns born to initially seronegative mothers, congenital CMV infection occurred in 18 (3.0%). In contrast, of 2857 newborns born to immune mothers, congenital CMV infection occurred in 29 (1.0%) Two factors, preconception maternal immunity (adjusted risk ratio, 0.31; 95% confidence interval, 0.17-0.58) and maternal age of 25 years or older (adjusted risk ratio, 0.19; 95% confidence interval, 0.07-0.49), were highly protective against congenital CMV infection. No other factors were associated with a reduction in the risk of congenital CMV infection. Naturally acquired immunity results in a 69% reduction in the risk of congenital CMV infection in future pregnancies.
  • Article
    Full-text available
    Successful preemptive cytomegalovirus (CMV) therapy in transplant patients depends on the availability of sensitive, specific, and timely diagnostic tests for CMV infections. The pp65 antigenemia assay has been used for this purpose with considerable success. Quantification of CMV DNA is currently regarded to be an alternative diagnostic approach. The precise relationship between these two methods has still to be defined, but is essential to compare diagnostic results. This study compared the results of both assays with a large series of transplant recipients in different categories. An internally controlled quantitative real-time CMV DNA PCR was used to test 409 plasma samples from solid organ transplant (SOT) and stem cell transplant (SCT) patients. Levels of CMV DNA in plasma correlated well with classified outcomes of the pp65 antigenemia test. Despite this correlation, the quantitative CMV PCR values in a class of antigen test results were within a wide range, and the definition of an optimal cutoff value for initiating treatment required further analysis by a receiver-operating characteristic curve analysis. This is essential for reactivating infections in particular. For the SCT patients the optimal cutoff value of CMV DNA load defining relevant viral reactivation (in this assay, 10,000 copies/ml) was slightly higher than that for the SOT patients (6,300 copies/ml). Based on a comparison with the established pp65 antigenemia assay, quantification of CMV DNA in plasma appeared to be capable of guiding the clinical management of transplant recipients. This approach may have important advantages, which include a superior reproducibility and sensitivity, allowing the inclusion of kinetic criteria in clinical guidelines.
  • Article
    Detection of viral DNA in dried blood spots using the Guthrie card (DBS test) is a reliable and practical method of diagnosing congenital cytomegalovirus (CMV) infection. The test lends itself to epidemiological studies to establish the prevalence of the infection, but also to neonatal screening for secondary prevention of sequelae. These applications would be facilitated if it were possible to use smaller samples and do the test on pools of individual cases. To ascertain whether doing the test on smaller, pooled samples still accurately identifies neonates with congenital CMV infection. We tested DBS from: (A) 39 laboratory reference cases; (B) 156 neonates suspected of having congenital CMV infection; (C) 119 children examined for the retrospective diagnosis of congenital CMV; (D) mock specimens prepared with known amounts of viral DNA. The test using only one third of the usual amount of dried blood was 100% sensitive and specific compared to the standard DBS test (A) and to viral isolation (A and B). Pools of three single cases gave the same results as viral isolation (B) and the small-sample test (B and C). All the versions of the test gave a detection limit of 400 copies/ml. The modified procedure can accurately diagnose congenital CMV infection. It achieves savings in both the patient material and the costs of testing.
  • Article
    To determine if protective behavior prevents child-to-mother transmission of cytomegalovirus (CMV) during pregnancy. We studied 166 seronegative mothers (94% white women; mean age, 33 years) with a child <36 months of age attending a day care facility. Mothers, either pregnant or attempting pregnancy, were randomly assigned by day care center to either a control or intervention group. Mothers in the intervention group received instructions for hand washing, glove use, and for avoiding types of intimate contact with their child. The control group received no instructions or information about their serologic status or whether their child was shedding CMV. In the intervention group, 7.8% of women (9 of 115) seroconverted, as did 7.8% of women (4 of 51) in the control group. Two independent predictors of maternal infection were (1) a child shedding and (2) a mother attempting pregnancy at enrollment. For 41 women attempting pregnancy at enrollment with a child shedding CMV, 10 of 24 became infected compared with only 1 of 17 women who were already pregnant at enrollment ( P = .008). For seronegative women who already know they are pregnant, intervention may be highly effective for preventing CMV acquisition.
  • Article
    Cytomegalovirus (CMV) is one of the most common causes of congenital infection without an effective treatment or an effective vaccine available to date. The emphasis has to be on preventive strategies, which rely on the epidemiological situation. The incidence of congenital CMV infections, however, is not known for The Netherlands. Therefore, a prospective virological study was carried out in a population of 7,524 pregnant women and 7,793 newborns. CMV-specific IgG antibodies were determined in cord blood by ELISA. When CMV antibodies were present, a CMV specific PCR was performed on the throat swab. A positive PCR was confirmed by urine culture. In addition, the seroepidemiology for CMV was investigated in the metropolitan region (Amsterdam and Rotterdam) which has a different ethnic composition. Congenital CMV infection was found in 7 infants (0.9 per 1,000). None had symptoms at birth or during 24 month follow-up. Carriage or CMV was 41%, with a variation between 35% and 100% depending on ethnicity. The ethnic composition in the south-eastern region was different from that in large cities, but similar to that in the rest of the country. The incidence of congenital CMV infections in The Netherlands is the lowest described to date, which does not justify special preventive policies.
  • Article
    Sensorineural hearing loss is the most frequent sequela of congenital cytomegalovirus (CMV) infection, and epidemiological evidence also suggests that congenital CMV infection is responsible for a substantial proportion of sensorineural hearing loss in children. Hearing loss due to congenital CMV infection can be present at birth or can appear later, usually during the first year of life; it usually worsens as the infant or child ages. Follow-up of children with congenital CMV infection should include repeated audiological testing. Based on the benefits of early detection of hearing loss, one could propose screening all infants for congenital CMV infection so that those with hearing impairment can be identified as early as possible by appropriate audiological follow-up. Antiviral treatment that could improve hearing outcome, with a safety profile suitable for use in minimally ill infants, would clearly increase the benefit of universal screening for congenital CMV infection.
  • Article
    Congenital human cytomegalovirus (HCMV) infection affects 1% of children and is the most common infectious cause of sensorineural hearing loss. Due to the difficulty of diagnosing deafness and other neurological disorders in infants, affected individuals may not be recognized until much later when active infection has resolved and culture is no longer informative. To overcome this problem, congenital HCMV infection was diagnosed retrospectively by testing residual blood samples collected from newborns and dried on perinatal cards as part of the North Carolina Newborn Screening Program. We modified the Qiagen method for purifying DNA from dried blood spots to increase the sample size and recovery of the lysate. A multiplex, real-time TaqMan polymerase chain reaction assay on an ABI 7900 instrument measured a highly conserved segment of the HCMV polymerase gene and the APOB human control gene. HCMV DNA was detected in blood dried on perinatal cards from all seven infants with culture-proven congenital infection, and all 24 negative control cases lacked detectable HCMV DNA. Our findings suggest that it is possible to diagnose congenital HCMV infection using dried blood collected up to 20 months earlier. Further studies are warranted on patients with hearing loss or other neurological deficits to determine the percentage that is attributable to congenital HCMV infection.
  • Article
    There are no studies on the detection of cytomegalovirus (CMV) DNA by molecular methods in the saliva of newborn infants in large scale screening programs. To evaluate the usefulness of saliva as a sample for the neonatal screening of congenital CMV infection as compared to urine when processed by a PCR. Saliva and urine samples were obtained during the first week of life. Both samples were attempted to be obtained from the first 2816 neonates. Subsequently, only saliva was obtained from other 1623 infants. Urine and saliva were processed by DNA-PCR. Confirmation of positive results was done by PCR and virus isolation by 3 weeks after birth. A urine sample was not obtainable from 893/2816 (31.7%) infants. Both saliva and urine samples were obtained from the remaining 1923 infants. Of these, 28 (1.45%) were CMV-infected. There was 99.7% agreement between the results with both samples. CMV excretion was similar when PCR was applied to urine (1.3%) or to saliva (1.2%) samples. Among the subsequent 1623 infants for whom only a saliva sample was planned for screening, 16 (0.98%) were CMV-infected. Saliva samples are as useful as urine for the identification of CMV-DNA in large use for screening programs.
  • Article
    Two recognized sources of maternal cytomegalovirus infection are young children and sexual activity. Previous studies evaluated either maternal exposures to young children or sexual activity, but these studies did not evaluate whether both of these maternal cytomegalovirus sources contribute to increases in congenital cytomegalovirus infections within populations with a high prevalence of infection among women of childbearing age. Our objective with this study was to investigate whether maternal cytomegalovirus exposure through young children and by sexual activity increases the risk for congenital cytomegalovirus infection in their offspring. A case-control study of 519 women from a delivery population in Birmingham, AL, between December 1992 and July 1998 was undertaken to measure the association between maternal cytomegalovirus exposures and an increased risk for congenital cytomegalovirus infection in their infants. Routine newborn cytomegalovirus screening at the hospital identified infants with congenital cytomegalovirus infection. The cases (n = 150) were women who delivered an infant with congenital cytomegalovirus infection, and the control subjects (n = 369) were randomly selected from the delivery population of women whose newborns were uninfected. Investigation of exposures included using a standardized maternal interview, prenatal and medical chart abstraction, and laboratory confirmation of cytomegalovirus infection. Significant associations between congenital cytomegalovirus infection and caring for preschool children in the year before delivery, onset of sexual activity < 2 years before delivery, sexually transmitted diseases during pregnancy, household size > 3 people, and maternal age < 25 years were identified. Women who cared for preschool children in the year before delivery and also became sexually active within the 2 years before delivery were at greatest risk for delivering an infant with congenital cytomegalovirus infection. Caring for young children and recent onset of sexual activity contribute to an increased risk for congenital cytomegalovirus infection in the offspring of young women.
  • Article
    Cytomegalovirus (CMV) infection is the most frequent congenital infection in humans and can cause permanent damage--particularly neurological--in about 20% of those infected, with or without symptoms at birth. Laboratory diagnosis is essential on account of the relatively non-specific clinical manifestations in symptomatic newborns but also because of the high frequency of asymptomatic cases that are nevertheless at risk of lesions later in life. However, these tests need samples taken within 3 weeks of birth to distinguish congenital infection from the more common, but clinically benign, perinatal infection. Tests for viral DNA have proved a valid means of diagnosing congenital CMV infection in neonatal blood dried on paper (DBS) widely used in screening for metabolic and genetic diseases, as an alternative to the conventional urine culture method. The DBS test is simpler, faster and less costly than viral isolation; in addition the samples can be safely stored for long periods, so diagnosis can be made even after several years. The sensitivity and specificity of the DBS test, compared to the reference method, have been reported to range between 71 and 100% and 99 and 100%, respectively, depending on the different studies and diagnostic criteria applied. The most interesting applications reported so far involve retrospective determination of the impact of congenital CMV in sensorineural deafness, abnormalities of cortical development, neonatal cholestasis and surveys of the prevalence of this infection in various populations. The test might be useful in the future for neonatal screening with a view to treating neonates and so avoiding the damage this disease can cause.
  • Article
    Congenital CMV is a major cause of neurological and sensory impairment in children. Reliable estimates of the prevalence of permanent sequelae and mortality associated with congenital CMV are needed to guide development of education and prevention programmes and to gauge the financial costs associated with this disease. To calculate such estimates, this review used data solely from studies in which children with congenital CMV were identified through universal screening. Based on 15 studies with a total of 117 986 infants screened, the overall CMV birth prevalence estimate was 0.7%. The percentage of infected children with CMV-specific symptoms at birth was 12.7%. The percentage of symptomatic children with permanent sequelae was 40-58%. The percentage of children without symptoms at birth who developed permanent sequelae was estimated to be 13.5%. The true burden of congenital CMV infection is unclear because data on important outcomes, such as visual impairment, are lacking and follow-up of infected children has been too short to fully identify late-onset sequelae. Therefore, the estimates of permanent sequelae associated with congenital CMV presented here are likely underestimates. Future studies should extend follow-up of CMV-infected children identified through universal screening and include the evaluation of visual impairment.
  • Article
    We reviewed studies that reported results of systematic cytomegalovirus (CMV) screening on fetuses and/or live-born infants. The overall birth prevalence of congenital CMV infection was 0.64%, but varied considerably among different study populations. About 11% of live-born infants with congenital CMV infection were symptomatic, but the inter-study differences in definitions of symptomatic cases limit the interpretation of these data. Non-white race, low socioeconomic status (SES), premature birth, and neonatal intensive care unit admittance were risk factors for congenital CMV infection. Birth prevalence increased with maternal CMV seroprevalence. Maternal seroprevalence accounted for 29% of the variance in birth prevalence between study populations. Maternal seroprevalence and birth prevalence were both higher in study populations that were ascertained at birth rather than in the prenatal period. Thus, timing of ascertainment should be considered when interpreting birth prevalence estimates. Birth prevalence was inversely correlated with mean maternal age, but this relationship was not significant when controlling for maternal seroprevalence. The rate of transmission to infants born to mothers who had a primary infection or a recurrent infection during pregnancy was 32% and 1.4%, respectively. Possible maternal primary infections (i.e. seropositive mother with CMV IgM) resulted in congenital infections about 20% of the time, but are likely to represent a mixture of primary and recurrent infections. In summary, CMV is a common congenital infection worldwide that can lead to permanent disabilities. There is an urgent need for interventions that can reduce the substantial burden of this often overlooked disease.
  • Article
    Full-text available
    We compared two protocols for extracting DNA from dried blood spots for cytomegalovirus (CMV) DNA detection and quantification by real-time PCR. Both extraction methods were reliable for the retrospective diagnosis of CMV congenital infection. Quantification of CMV DNA was valuable after normalization of viral loads with albumin gene PCR amplification results.
  • Article
    Congenital cytomegalovirus (CMV) infection is a cause of sensorineural hearing loss (SNHL) in children, but the magnitude of its contribution is uncertain. Quantifying the impact of congenital CMV infection requires an evidence-based assessment using a standard case definition of hearing loss. To determine the frequency of bilateral moderate to profound SNHL in children with congenital CMV infection and to estimate the CMV-attributable fraction of bilateral moderate to profound SNHL. A systematic review of studies of children with congenital CMV infection ascertained in an unbiased manner through universal newborn screening for CMV using viral culture in urine or saliva specimens in combination with a review of the literature on congenital CMV infection and hearing loss, including articles of all types. Approximately, 14% of children with congenital CMV infection develop SNHL of some type, and 3-5% develop bilateral moderate to profound SNHL. Among all children with bilateral moderate to profound SNHL, we estimate that 15-20% of cases are attributable to congenital CMV infection. Congenital CMV infection is one of the most important causes of hearing loss in young children, second only to genetic mutations, and is potentially preventable.
  • Article
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    Testing for viral DNA in neonatal blood dried on paper (DBS) has proved a valid means of diagnosing congenital CMV infection with both clinical and epidemiological relevance. To assess the quality of the detection of CMV-DNA on DBS in laboratories performing this test a proficiency panel consisting of nine samples with two blood spots on each filter paper was produced and distributed. Six samples were derived from whole blood, negative for CMV DNA and antibody, and spiked with cell-grown CMV Towne in various concentrations (7.3 x 102 - 9.6 x 105 copies/ml), one was a CMV positive clinical specimen (3.9 x 106 copies/ml), and two samples were CMV-negative whole blood. The 27 responding laboratories from 14 countries submitted 33 datasets obtained by means of conventional PCR (n = 5) or real-time PCR (n = 28) technologies. A correct positive result was reported in at least 91% of datasets in samples with a viral load of 8.8 x 104 copies/ml or higher. However only 59% and 12% identified the 9.4 x 103 and 7.3 x 102 copies/ml samples, respectively, correctly as positive. False positive results were reported by 9% of laboratories and in 11% of datasets. These results indicate a clear need for improvement of methods as sensitivity and false-positivity still appear to be a major problem in a considerable number of laboratories.
  • Article
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    Two protocols for the extraction of cytomegalovirus (CMV) DNA and two methods for the amplification of CMV DNA in dried blood spots were evaluated for the retrospective diagnosis of congenital CMV infection. During the period from 1996 to 2006, a urine screening program detected 76 congenitally infected neonates. Stored Guthrie cards with blood from 55 cases and 12 controls were tested. Two spots of dried blood were cut from each card and evaluated in two centers. CMV DNA was extracted from a whole single spot. Center 1 used phenol-chloroform extraction and ethanol precipitation followed by a conventional PCR. Center 2 used the NucliSens easyMAG automated DNA/RNA extraction platform (bioMérieux) followed by a real-time PCR. For evaluation of the extraction method, DNA extracted from each blood spot was evaluated by the amplification method used by the collaborating center. The sensitivities were 66% for center 1 and 73% for center 2. None of the controls were positive. A sensitivity as high as 82% could be obtained by combining the most sensitive extraction method (the phenol-chloroform procedure) with the most sensitive PCR method (real-time PCR). The detection rate was not influenced by the duration of storage of the spots. The sensitivity was higher with blood from congenitally infected cases due to a primary maternal CMV infection, regardless of the protocol used. However, the difference reached significance only for the least-sensitive protocol (P = 0.036).