Is Polymerase Chain Reaction in Neonatal Dried Blood Spots Reliable for the Diagnosis of Congenital Cytomegalovirus Infection?

Vives-Oñós, Isabel; Codina-Grau, María Gema; Noguera-Julián, Antoni; Blázquez‐Gamero, Daniel; Fortuny, Clàudia; Baquero-Artigao, Fernando; Frick, Marie Antoinette; Saavedra-Lozano, Jesús; Goycochea-Valdivia, Walter Alfredo; Rives-Ferreiro, María Teresa; Montesdeoca-Melián, Abián; Calavia-Garsaball, Olga; Ferreras-Antolín, Laura; Marín-Soria, José Luís; Dulín-Iñiguez, Elena; Soler‐Palacín, Pere

doi:10.1097/inf.0000000000002144

Cited by 9 publications

(5 citation statements)

References 27 publications

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“…In particular, the use of saliva samples improved the inclusion rate in screening programs since many newborns had to be excluded due to unsuccessful sampling of urine [6]. Viral load levels in EDTA blood were more than 10 4 -fold lower than in buccal swabs (Fig 3, S3 Table), consistent with the reduced sensitivity reported for assays based on dried blood spots [22,[25][26][27][28]. A recent study on symptomatic cCMV infections even revealed that CMV DNA levels were undetectable in 11% of whole blood samples of clinically symptomatic patients further questioning congenital CMV screening approaches based on blood samples [29].…”

Section: Discussionsupporting

confidence: 66%

Characterization of a universal screening approach for congenital CMV infection based on a highly-sensitive, quantitative, multiplex real-time PCR assay

et al. 2020

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The majority of congenital cytomegalovirus (cCMV) infections are asymptomatic at birth and therefore not diagnosed. Approximately 10-15% of these infants develop late-onset hearing loss and other developmental disorders. Implementation of a universal screening approach at birth may allow early initiation of symptomatic interventions due to a closer follow-up of infants at risk and offers the opportunity to consider treatment of late-onset disease. Realtime PCR assays for the detection of CMV DNA in buccal swab samples demonstrated feasibility and good clinical sensitivity in comparison to a rapid culture screening assay. Because most cCMV infections remain asymptomatic, a universal screening assay that stratifies CMV infected infants according to low and high risk of late-onset cCMV disease could limit the parental anxiety and reduce follow-up costs. We therefore developed and characterized a screening algorithm based on a highly-sensitive quantitative real-time PCR assay that is compatible with centralized testing of samples from universal screening and allows to determine CMV DNA load of saliva samples either as International Units (IU)/ml saliva or IU/10 5 cell equivalents. 18 of 34 saliva samples of newborns that tested positively by the screening algorithm were confirmed by detection of CMV DNA in blood and/or urine samples obtained during the first weeks of life. All screening samples that could not be confirmed had viral loads of <2.3x10 5 IU/ml saliva (median: 6.8x10 3) or 1.3x10 5 IU/10 5 cell equivalents (median: 4.0x10 2). The viral load of screening samples with confirmed cCMV infection ranged from 7.5x10 2 to 8.2x10 9 IU/ml saliva (median: 9.3x10 7) or 1.5x10 2 to 5.6x10 10 IU/10 5 cell equivalents (median: 3.5x10 6). Clinical follow-up of these newborns with confirmed cCMV infection should reveal whether the risk of late-onset cCMV disease correlates with CMV DNA load in early life saliva samples and whether a cutoff can

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Section: Discussionsupporting

confidence: 66%

Characterization of a universal screening approach for congenital CMV infection based on a highly-sensitive, quantitative, multiplex real-time PCR assay

et al. 2020

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“…High extraction rates are of no benefit if the PCR is not sensitive. Table 3 shows that the sensitivities of the PCRs for the four CMV target genes in this study are comparable to those reported by other groups, including the commercial quantitative R-gene CMV kit (also used in this study) and RealStar CMV kit [28]. This was surprising as we had designed our PCRs for genotyping of highly variable areas of the CMV genome, while the commercial kits focus purely the on detection and quantification of highly conserved CMV genes (normally UL83).…”

Section: Discussionsupporting

confidence: 85%

Development of highly efficient protocols for extraction and amplification of cytomegalovirus DNA from dried blood spots for detection and genotyping of polymorphic immunomodulatory genes

et al. 2019

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Congenital cytomegalovirus (CMV) infection is a major cause of birth defects ranging from developmental disorders to stillbirth. Most newborns affected by CMV do not present with symptoms at birth but are at risk of sequelae at later stages of their childhood. Stored dried blood spots (DBS) taken at birth can be used for retrospective diagnosis of hereditary diseases, but detection of pathogens is challenged by potentially low pathogen concentrations in the small blood volume available in a DBS. Here we test four different extraction methods for optimal recovery of CMV DNA from DBS at low to high CMV titers. The recovery efficiencies varied widely between the different extractions (from 3% to 100%) with the most efficient method extracting up to 113-fold more CMV DNA than the least efficient and 8-fold more than the reference protocol. Furthermore, we amplified four immunomodulatory CMV genes from the extracted DNA: the UL40 and UL111A genes which occur as functional knockouts in some circulating CMV strains, and the highly variable UL146 and US28 genes. The PCRs specifically amplified the CMV genes at all tested titers with sufficient quality for sequencing and genotyping. In summary, we here report an extraction method for optimal recovery of CMV DNA from DBSs that can be used for both detection of CMV and for genotyping of polymorphic CMV genes in congenital CMV infection.

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“…Infants are not necessarily viremic at birth. Therefore, a negative CMV PCR test for the whole blood or plasma or newborn dried blood spots (DBS) does not exclude CMV [18,19]. We concluded that intrauterine transmission had occurred for all four patients presented here.…”

Section: Discussionmentioning

confidence: 89%

Congenital Cytomegalovirus in Algiers, Algeria, a Descriptive Case Series Study

AGGOUNE¹

2022

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Cytomegalovirus (CMV) is an endemic ubiquitous herpes virus transmitted through saliva, urine, genital secretions, mononuclear blood cells, and transplanted tissue. It is a common infection with mild symptoms or asymptomatic in the immunocompetent individuals but can be severe in the immunocompromised individuals, e.g., HIV-infected individuals, transplanted and cancer patients, and fetuses. CMV is the leading cause of congenital viral infection and the leading non-hereditary cause of sensorineural hearing loss and mental retardation in early childhood. Here, we describe the clinical and laboratory monitoring of four congenital CMV (cCMV) cases referred to Pediatric Belfort Establishment in Algiers in 2019, 2020, and 2021. All the patients had developed signs and symptoms of postnatal CMV infection with intrauterine growth retardation. The clinical manifestations differed; some presented cytopenia with or without hepatosplenomegaly and others a clinical and biological cholestasis syndrome. All our patients had intrauterine growth retardation. A CMV PCR of a urine sample was positive. Treatment for six weeks based on ganciclovir, with a relay by valganciclovir.

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Is Polymerase Chain Reaction in Neonatal Dried Blood Spots Reliable for the Diagnosis of Congenital Cytomegalovirus Infection?

Abstract: The sensitivity of CMV rt-PCR in DBS in our series was low and correlated with the bVL. Thus, a negative DBS result would not rule out cCMV infection, especially in patients with a low viremia level at birth.

Cited by 9 publications

References 27 publications

Characterization of a universal screening approach for congenital CMV infection based on a highly-sensitive, quantitative, multiplex real-time PCR assay

Characterization of a universal screening approach for congenital CMV infection based on a highly-sensitive, quantitative, multiplex real-time PCR assay

Development of highly efficient protocols for extraction and amplification of cytomegalovirus DNA from dried blood spots for detection and genotyping of polymorphic immunomodulatory genes

Congenital Cytomegalovirus in Algiers, Algeria, a Descriptive Case Series Study

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