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Stability of severe acute respiratory syndrome coronavirus 2 RNA in placenta and fetal cells

2020; Elsevier BV; Volume: 224; Issue: 1 Linguagem: Inglês

10.1016/j.ajog.2020.09.003

1097-6868

Léo Pomar, Karin Nielsen‐Saines, David Baud,

SARS-CoV-2 detection and testing

We have read with great interest the systematic review and metaanalysis by Kotlyar and colleagues1Kotlyar A. Grechukhina O. Chen A. et al.Vertical transmission of COVID-19: a systematic review and meta-analysis.Am J Obstet Gynecol. 2020; ([Epub ahead of print])Abstract Full Text Full Text PDF Scopus (6) Google Scholar reporting a pooled proportion of 3.2% for vertical transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The authors indicate the proportion of positive SARS-CoV-2 RNA testing in neonatal blood, urine, placental samples, and amniotic fluid, without considering the stability of viral RNA as a major limitation in the diagnosis of maternal-fetal transmission. RNA is very susceptible to degradation, which occurs through hydrolysis and ribonuclease activity. Clinical samples are particularly vulnerable to RNA degradation by the action of host nucleases.2Nelson P.T. Baldwin D.A. Scearce L.M. Oberholtzer J.C. Tobias J.W. Mourelatos Z. Microarray-based, high-throughput gene expression profiling of microRNAs.Nat Methods. 2004; 1: 155-161Crossref PubMed Scopus (554) Google Scholar In the case of the diagnosis of vertical transmission of viruses, RNA is considerably less stable than DNA in the placental and fetal samples and requires more steps for detection at the laboratory level. A critical challenge for RNA preservation and detection in these samples is to prevent degradation by the nuclease during the sampling and purification processes.3Barnes M.G. Tsoras M. Thompson S.D. Martinez H. Iverson B. Nuñez R. Ambient temperature stabilization of purified RNA in GenTegra™ for use in Affymetrix Human Exon 1.0 ST arrays.BioTechniques. 2010; 48: 468-469Crossref Scopus (4) Google Scholar The storage and transportation of clinical samples are also at risk of RNA hydrolysis, which represents a limitation for healthcare settings with a decentralized laboratory.4Martinez H. Beaudry G. Veer J. et al.Ambient temperature storage of RNA for use in a RT-qPCR test for the detection of GCC mRNA.Cancer Res. 2011; 70: 819Google Scholar For example, the H5N1 RNA was undetectable if stored 24 hours at room temperature, whereas it remained detectable more than 40 days when stored in cold temperature, in RNA-safe buffer, or in dry pellet matrix, without exposure to high temperatures.5Relova D. Rios L. Acevedo A.M. Coronado L. Perera C.L. Pérez L.J. Impact of RNA degradation on viral diagnosis: an understated but essential step for the successful establishment of a diagnosis network.Vet Sci. 2018; 5: 19Crossref Scopus (26) Google Scholar The same is true for Zika virus, which rapidly degrades if not stored in RNA lather, often becoming undetectable once frozen specimens are thawed.6Corman V.M. Rasche A. Baronti C. et al.Assay optimization for molecular detection of Zika virus.Bull World Health Organ. 2016; 94: 880-892Crossref PubMed Scopus (119) Google Scholar,7Gorchakov R. Berry R.M. Patel S.M. El Sahly H.M. Ronca S.E. Murray K.O. Optimizing PCR detection of Zika virus from various body fluids.Am J Trop Med Hyg. 2019; 100: 427-433Crossref PubMed Scopus (18) Google Scholar These limitations have been well documented for reverse transcription–polymerase chain reaction (RT-PCR) assays in cases of congenital Zika virus infections.10Adebanjo T. Godfred-Cato S. Viens L. et al.Update: interim guidance for the diagnosis, evaluation, and management of infants with possible congenital Zika virus infection - United States, October 2017.MMWR Morb Mortal Wkly Rep. 2017; 66: 1089-1099Crossref PubMed Scopus (152) Google Scholar, 8Pomar L. Vouga M. Lambert V. et al.Maternal-fetal transmission and adverse perinatal outcomes in pregnant women infected with Zika virus: prospective cohort study in French Guiana.BMJ. 2018; 363: k4431Crossref PubMed Scopus (104) Google Scholar, 9Schaub B. Vouga M. Najioullah F. et al.Analysis of blood from Zika virus-infected fetuses: a prospective case series.Lancet Infect Dis. 2017; 17: 520-527Abstract Full Text Full Text PDF PubMed Scopus (74) Google Scholar Conversely, genomic content from DNA viruses, such as Cytomegalovirus, is easily purified from whole blood or any other tissue or fluid (placenta, fetal liver and brain, amniotic fluid, urine, cerebrospinal fluid) and is less subjected to deterioration, increasing the sensitivity of PCR assays for the diagnosis of vertical transmissions.11Ross S.A. Ahmed A. Palmer A.L. et al.Urine collection method for the diagnosis of congenital cytomegalovirus infection.Pediatr Infect Dis J. 2015; 34: 903-905Crossref PubMed Scopus (31) Google Scholar,12Roberts T.C. Buller R.S. Gaudreault-Keener M. et al.Effects of storage temperature and time on qualitative and quantitative detection of cytomegalovirus in blood specimens by shell vial culture and PCR.J Clin Microbiol. 1997; 35: 2224-2228Crossref PubMed Google Scholar When maternal infection occurs during the first trimester of pregnancy, DNA viruses are detectable throughout the pregnancy in fetal and placental tissues. This contrasts with RNA viruses (such as Zika virus or SARS-CoV-2), which are only transiently present and detectable.9Schaub B. Vouga M. Najioullah F. et al.Analysis of blood from Zika virus-infected fetuses: a prospective case series.Lancet Infect Dis. 2017; 17: 520-527Abstract Full Text Full Text PDF PubMed Scopus (74) Google Scholar Therefore, the absence of the detection of an RNA virus does not necessarily mean that the infection of the given tissue is absent. This issue should be mentioned in any study investigating the potential evidence of vertical transmission of SARS-CoV-2. Vertical transmission of coronavirus disease 2019: a systematic review and meta-analysisAmerican Journal of Obstetrics & GynecologyVol. 224Issue 1PreviewThis study aimed to conduct a systematic review of the current literature to determine estimates of vertical transmission of coronavirus disease 2019 based on early RNA detection of severe acute respiratory syndrome coronavirus 2 after birth from various neonatal or fetal sources and neonatal serology. Full-Text PDF ReplyAmerican Journal of Obstetrics & GynecologyVol. 224Issue 1PreviewWe thank Pomar and colleagues1 for their interest in our article and the important point they raised in their letter. We acknowledge that RNA degrades more easily than DNA and that sampling, purification processes, storage, and transport conditions can influence RNA stability, which may impact the ability to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in placental and certain fetal samples using molecular assays. However, this seems to be much less of a concern when assessing the presence of SARS-CoV-2 RNA in neonatal nasopharyngeal (NP) specimens. Full-Text PDF