Retrospective Diagnostics of Congenital Cytomegalovirus Infection Performed by Polymerase Chain Reaction in Blood Stored on Filter Paper

Abstract

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.

Evaluation of Cytomegalovirus (CMV) DNA Quantification in Dried Blood Spots: Retrospective Study of CMV Congenital Infection

“…This difference can be explained as follows: (i) only 50 l of blood is extracted from a whole DBS compared to the 200 to 500 l extracted from fresh blood, and (ii) whatever the DNA extraction protocol used, it is less efficient when performed with DBS than with fresh blood. However, even if the in vitro sensitivity of CMV PCR in DBS is relatively low, its clinical sensitivity was 100% in our study and ranges from 80 to 100% in the literature (1,2,9,14). CMV DNA quantification with DBS has not been reported so far.…”

“…Policies for screening during pregnancy and at birth have not been implemented in European countries or in the United States, essentially because there is no well-established treatment for pregnant women or for newborns with CMV infection (7). Retrospective diagnosis of congenital infection has been achieved by PCR detection of the CMV DNA in dried blood spots (DBS) stored on perinatal Guthrie cards (2,5,6,8,9,14,16,17). Only one protocol (heat DNA extraction, followed by nested PCR) has been extensively evaluated in a clinical setting, with excellent sensitivity and specificity compared to that of viral isolation in the urine (1,2).…”

“…However, lower sensitivities (63 and 71%) were reported using the same protocol (5,16). Alternative methods based on either phenol-chloroform or silica extraction protocols have been proposed, but their sensitivities (81% and 100%, respectively) were studied only with small numbers of patients (9,14).…”

Adenovirus DNA is detected at increased frequency in Guthrie cards from children who develop acute lymphoblastic leukaemia

“…And, as is the case with children infected post-natally, adenoviral DNA is retained in latently infected T lymphocytes for years following primary infection (Garnett et al, 2002). Thus, the finding of species C adenovirus in neonatal blood spots is evidence of prenatal infection, as is the case for other prenatal viral infections (Barbi et al, 1996;Johansson et al, 1997;Fischler et al, 1999).…”

Hearing Loss and Congenital CMV Infection: A Systematic Review

“…Cross-contamination of adjacent stored cards has been reported. 54,[59][60][61] To understand the importance of cCMV as a cause of childhood hearing loss, we reviewed studies that conducted retrospective testing in a group of hearingimpaired children. Requirements were testing by real-time PCR for quantitative analysis of CMV DNA on DBS or on dried umbilical cords.…”