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J. Clin. Microbiol. doi:10.1128/JCM.01391-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Evaluation of different CMV DNA PCR protocols on dried blood spots from consecutive cases of congenital CMV infected neonates

Department of Microbiology, UZ Brussel, Laarbeeklaan 101, 1090 Brussel, Belgium; Department of Microbiology and Immunology, Assistance Publique-Hôpitaux de Paris, Antoine Béclère hospital, 157 av. de la porte de Trivaux 92141 Clamart, Faculty of Medecine Paris-Sud, France; Department of ENT, UZ Brussel, Laarbeeklaan 101, 1090 Brussel, Belgium

^* To whom correspondence should be addressed. Email: anne.naessens{at}uzbrussel.be.

	Abstract

Two extraction protocols and two amplification methods for CMV DNA on dried blood spots were evaluated for the retrospective diagnosis of congenital CMV infection. During the period 1996-2006 a urine screening program detected 76 congenitally infected neonates. Stored Guthrie cards from 55 cases and 12 controls were tested. From each card 2 spots of dried blood were cut and evaluated in two centers. CMV DNA was extracted from a whole single spot. Center 1 used a 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, extracted DNA from each blood spot was evaluated with the amplification method used by the collaborating centre. The sensitivity was 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 (phenol-chloroform procedure) with the most sensitive PCR (real-time PCR). The detection rate was not influenced by the duration of storage of the spots. The sensitivity was higher in congenital infected cases due to a primary maternal CMV infection, regardless the protocol used. However, the difference did reach significance only for the least sensitive protocol (p = 0.036).