Congenital Cytomegalovirus Conference - Congenital CMV ...

Objectives: 1)Determine the prevalence of congenital CMV infection in of 3-5 year old children with
PCHI in the Netherlands, 2) quantify the effect of the screening strategy on the age at PCHI detection in
children with congenital CMV 3) describe the developmental outcome of these children.
Methods: Children born in the Netherlands between 1-1-2003 and 31-12-2005 with PCHI (Definition:
hearing deficit of ≥40dB in the better ear) registered at Audiological Centres are eligible for participation
in the DECIBEL-study providing they have been offered a Dutch hearing screening strategy in the first
year of life. Assessments: CMV DNA detection on neonatally acquired dried blood samples. The Child
Development Inventory-NL to investigate general development and additional developmental- and
audiological parameters from medical records.
Results: 482 children (born 1-1-2003 to 31-12-2005) were registered with PCHI at Audiological Centres
in the Netherlands. Preliminary results of CMV-DNA testing in children in the DECIBEL-study show
approximately 10-20% CMV-DNA positivity. Of the 3-5 year olds with PCHI and congenital CMV
infection, more children had passed the neonatal hearing screening strategy than children in the (later)
distraction screening hearing strategy. Results of the CDI-NL show a small overall delay in children with
PCHI and congenital CMV.
Conclusions: Early hearing screening strategies produce some negative screening results in children
with congenital CMV infection, presumably because of progressive and delayed onset hearing loss.
Developmental delay often accompanies PCHI in these children.
O-40 Evaluation of DNA extraction methods using dried blood spots for diagnosing congenital CMV
infection.
Jutte J.C. de Vries, Eric C.J. Claas, Aloys C.M. Kroes, Ann C.T.M. Vossen. Leiden University Medical
Center, Leiden, Netherlands.
Background: Cytomegalovirus (CMV) is the most common cause of congenital infection worldwide and
occurs when CMV is transmitted from mother to fetus. The most frequently encountered symptom of
congenital CMV infection is sensorineural hearing loss (SNHL). The only reliable method to diagnose
congenital CMV in older children is CMV detection in dried blood spots (DBS) sampled within one week
after birth. Since the first publication on the usage of DBS for diagnosing congenital CMV infection,
several DNA extraction methods for DBS followed by CMV PCR have become available. However, a
recent quality assessment programme by Quality Control for Molecular Diagnostics (QCMD) studying
CMV DNA detection in DBS among European and South African laboratories showed that the assays as
employed had a worrisome lack of sensitivity as well as specificity.
Objectives: To compare DNA extraction methods for DBS for CMV DNA detection. Sensitivity, specificity
and suitability of the methods for high-throughput usage were assessed.
Methods: Guthrie cards were spotted with CMV-positive whole blood from transplant recipients with a
broad range of CMV plasma loads. DNA was extracted from DBS using a panel of six non-commercial
and commercial DNA extraction methods, followed by CMV real-time PCR amplification. Samples were
tested in triplicate with negative control punches between each sample.
Results: DBS with high CMV loads (plasma loads ≥ 4 log10 c/ml) were positive in all triplicates of all
DNA extraction methods tested. However, using DBS with lower CMV loads, the ability to detect CMV
DNA decreased markedly, depending on the method used. Furthermore, CMV loads extracted from
the DBS varied widely. Specificity of the methods tested was high. Some extraction methods appeared
suitable for medium-throughput applications, only few methods appeared suitable for automated
applications using a 96-wells format.
Conclusions: The differences in sensitivity and capacity of the DNA extraction methods tested limit the
possible applications. Both highly sensitive and high-throughput methods are required when considering
newborn screening for congenital CMV infection.
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