Skip to main content
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Eukaryotic Cell
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems
  • Log in
  • My alerts
  • My Cart

Main menu

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Archive
    • Minireviews
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JCM
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Eukaryotic Cell
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems

User menu

  • Log in
  • My alerts
  • My Cart

Search

  • Advanced search
Journal of Clinical Microbiology
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Archive
    • Minireviews
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JCM
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
Virology

Comparison of Use of Cerebrospinal Fluid, Serum, and Throat Swab Specimens in Diagnosis of Enteroviral Acute Neurological Infection by a Rapid RNA Detection PCR Assay

Laurent Andréoletti, Nathalie Blassel-Damman, Anny Dewilde, Louis Vallée, Robin Cremer, Didier Hober, Pierre Wattré
Laurent Andréoletti
Laboratoire de Virologie,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Nathalie Blassel-Damman
Laboratoire de Virologie,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Anny Dewilde
Laboratoire de Virologie,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Louis Vallée
Service des Maladies Infectieuses Neurologiques Infantiles, and
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Robin Cremer
Service de Réanimation Pédiatrique, Centre Hospitalier Universitaire de Lille, 59037 Lille cedex, France
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Didier Hober
Laboratoire de Virologie,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Pierre Wattré
Laboratoire de Virologie,
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
DOI: 10.1128/JCM.36.2.589-591.1998
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

ABSTRACT

A PCR assay for detection of enterovirus RNA in multiple specimen types from patients with neurological infections was evaluated. Combined PCR assay of cerebrospinal fluid and serum (systemic specimens) was more sensitive than assaying either specimen alone in children but not in adults. Compared with PCR in systemic specimens, detection of enterovirus RNA in throat swabs showed a sensitivity of 62.5% and a specificity of 75.6%.

Enteroviruses (EVs) of thePicornaviridae family, including the polioviruses, echoviruses, group A and B coxsackieviruses, and more recently numbered EVs (67 distinct serotypes) are important human pathogens (10, 14). Neurological infections with the nonpolio EVs are common and important causes of morbidity in both children and adults (1, 3, 6). Previous reports have clearly established the etiological role of these viruses in aseptic meningitis, encephalitis, and chronic meningoencephalitis, as well as in paralytic myelitis, cerebellar ataxia, Guillain-Barré syndrome, and transverse myelitis (10). However, attempts to isolate EVs from cerebrospinal fluid (CSF), pharyngeal, and stool samples are frequently unsuccessful because of the low viral titer in clinical specimens and because some serotypes grow poorly in cell culture (4). Therefore, PCR techniques for the detection of the enterovirus genome have been introduced (2, 9, 11).

In this report, we used a commercially available PCR assay which utilizes a single enzyme for both reverse transcription (RT) and PCR steps, incorporates uracil-N-glycosylase to prevent carryover contaminations, and detects the EV amplicons in a microwell colorimetric assay within 6 h. Using this PCR assay, we investigated the potential utility of a PCR assay of both CSF and serum specimens (systemic specimens) for rapid diagnosis of acute EV neurological disease in adult and children patients. Moreover, the EV PCR results from throat swabs were compared to those obtained from systemic specimens.

Forty-four children and 17 adults hospitalized for suspected neurological virus infections were included. CSF specimens were obtained by lumbar puncture. Whole-blood samples were obtained by peripheral veinous puncture, and the sera were sterilely discarded after centrifugation (10 min, 4°C, 3,500 × g). Throat swab specimens were obtained by using the Virocult system (Medical Wire and Equipment, Corsham, United Kingdom) (7).

CSF and throat swabs specimens were inoculated in duplicate into 24-well plates covered with monolayers of human diploid fibroblasts (MRC-5) and buffalo green monkey kidney (BGMK) cells as previously described (13).

EV RT-PCR assays were performed with the amplicor Roche EV kit (Amplicor PCR Diagnostics, Hoffmann-Laroche, Basel, Switzerland) according to the manufacturer’s instructions. Briefly, after chemical denaturation, the single-stranded PCR products were hybridized to the probe and detected with the avidin-horseradish peroxidase system. Results were scored as positive when the optical density value at 450 nm was >0.34 (7). χ2 testing with Yates correction was used to compare the results of biological assays (P values of <0.05 were considered significant).

In the group of children, we detected specific EV RNA sequences in 22.7% (10 of 44) of CSF specimens, whereas the rates of EV isolation by cell culture were only 2.3% (1 of 44) in these samples (Table1). At the same time, detection of EV RNA in serum was positive in 20.45% (9 of 44) of children studied (Table1). This positive EV RNAemia was associated with a positive EV PCR result for CSF specimens in three patients with aseptic meningitis and in one patient with Guillain-Barré syndrome. Interestingly, a positive EV RNAemia result allowed us to establish the etiological diagnosis of neurological virus infection in one patient with encephalitis and in three patients with aseptic meningitis (Table 1). Combination of EV PCR testing of CSF and serum specimens was more sensitive than a single PCR test of a CSF (14 of 44 versus 10 of 44;P = 0.014) or of a serum (14 of 44 versus 9 of 44;P = 0.007) specimen from infants.

View this table:
  • View inline
  • View popup
Table 1.

Enteroviral RT-PCR and cell culture isolation results for CSF, serum, and throat specimens from patients with suspected neurological EV infections

In the adult patient group, we detected EV RNA in 43.7% (7 of 16) of CSF specimens tested, whereas no EV strain was recovered from these specimens by cell culture isolation. A positive detection of EV RNA in serum was observed in one patient with aseptic meningitis and in one patient with myelitis (Table 1). The percentage of positive EV RNA detection was not significantly different between the combined EV PCR assay for CSF and serum specimens and the single PCR detection of a CSF (8 of 15 versus 7 of 16; P = 0.87) or a serum (8 of 15 versus 2 of 16; P = 0.075) specimen.

Throat specimens were positive by PCR in 31.8% of the children and in 11.8% of the adults studied (Table 1). The overall performances of the PCR test for throat swabs versus the PCR test for systemic specimens are shown in Table 2. Of the 16 throat specimens positive by PCR, only 10 were correlated to a positive EV detection in one of the two systemic specimens (sensitivity of 62.5%); of the 45 throat specimens negative by PCR, 34 were correlated to an absence of EV RNA sequences detectable by PCR in CSF and/or serum (specificity of 75.6%) (Table 2).

View this table:
  • View inline
  • View popup
Table 2.

Comparison of EV RT-PCR results obtained from a peripheral (throat) specimen and systemic (CSF and serum) specimens taken from patients

Previous reports demonstrated the advantages of the PCR assay used in this work for diagnosis of neurological EV infection over traditional tissue culture isolation from CSF (7, 9, 11). In our prospective study, more diagnoses of an enteroviral neurological syndrome were achieved by PCR-microwell hybridization of CSF than by cell culture isolation (Table 1). The low percentages of enteroviral isolation from CSF specimens could be explained by poorly cultivable enteroviral serotypes or by a small number of infectious particles in CSF samples at the time of CSF puncture (4, 15). In order to investigate the diagnostic value of EV viremia in neurological syndromes, we compared the results of the detection of EV RNA by PCR in CSF and serum specimens taken from children and adult patients (Table1). The detection of EV RNA either in CSF or in serum proved enteroviral infection, whereas a positive PCR detection in throat swabs alone was considered not significant (11). A positive EV PCR assay of serum was observed in 5 of 10 children and in only 1 of 7 adult patients with a positive EV PCR result in the CSF sample. An isolated positive EV PCR detection in serum was observed in four children and in one adult patient suffering from an acute EV infection. Moreover, the results demonstrated that a combined PCR assay of CSF and serum was significantly more sensitive than a single PCR detection of CSF or of serum specimens from children patients, suggesting that this approach improves the diagnosis of neurological enteroviral diseases in infants. Comparatively to the child group, adult patients demonstrated a lower percentage (20.45 versus 12.5%) of positive EV RNAemia (Table1). Moreover, the EV RNA sequence detection by PCR was more sensitive in CSF than in serum (7 of 16 versus 2 of 16; P = 0.056), suggesting the value of PCR analysis of CSF in adult patients. The discrepant results obtained between the adult and children patient groups could be linked to a shorter viremia phase after the onset of neurological symptoms in adults (5, 6, 8).

We evaluated the performance of PCR testing of throat swabs versus PCR testing of systemic (CSF and serum) specimens. The low percentages of sensitivity (62.5%) and specificity (75.6%) clearly demonstrated the low diagnostic value of detection of EV RNA in throat specimens from children and adult patients (Table 2). The sensitivity of PCR of throat swabs did not exceed that of tissue culture and was lower than that obtained by Rotbart et al. in a similar study (12) (Table1). These discrepancies could be explained by the presence of high levels of RNase activity or by Taq DNA polymerase inhibitors in the tested throat swabs (2).

In summary, the results of the present study suggest the diagnostic value of a rapid combined EV RNA detection method for CSF and serum from children with acute neurological EV infections and suggest that this approach could become a screening method for diagnosis of neurological EV diseases.

ACKNOWLEDGMENTS

We thank F. Eberlé and B. Aulong of Produits Roche for supplying the Amplicor Enterovirus kits in this study. We are grateful to Sylvie Ledru for skillful assistance in statistical analysis.

This work was supported in part by grant no. 93-11 from the Centre Hospitalier et Universitaire de Lille and from the Conseil regional Nord-Pas de Calais (France).

FOOTNOTES

    • Received 4 June 1997.
    • Returned for modification 2 September 1997.
    • Accepted 4 November 1997.
  • Copyright © 1998 American Society for Microbiology

REFERENCES

  1. 1.↵
    1. Abzug M. J.,
    2. Loeffelholz M.,
    3. Rotbart H. A.
    Diagnosis of neonatal enterovirus infection by polymerase chain reaction.J. Pediatr. 126 1995 447 450
    OpenUrlCrossRefPubMedWeb of Science
  2. 2.↵
    1. Andréoletti L.,
    2. Hober D.,
    3. Belaich S.,
    4. Lobert P. E.,
    5. Dewilde A.,
    6. Wattré P.
    Rapid detection of enterovirus in clinical specimens using PCR and microwell capture hybridization assay.J. Virol. Methods 62 1996 1 10
    OpenUrlCrossRefPubMedWeb of Science
  3. 3.↵
    1. Berlin L. E.,
    2. Rorabaugh M. L.,
    3. Heldrich F.,
    4. Roberts K.,
    5. Doran T.,
    6. Modlin J. F.
    Aseptic meningitis in infants <2 years of age: diagnosis and etiology.J. Infect. Dis. 168 1993 888 892
    OpenUrlCrossRefPubMedWeb of Science
  4. 4.↵
    1. Chonmaitree T.,
    2. Ford C.,
    3. Sanders C.,
    4. Lucia H.
    Comparison of cell cultures for rapid isolation of enteroviruses.J. Clin. Microbiol. 26 1988 2576 2580
    OpenUrlAbstract/FREE Full Text
  5. 5.↵
    1. Dagan R.,
    2. Jenista J. A.,
    3. Prather S. L.,
    4. Powell K. R.,
    5. Menegus M. A.
    Viremia in hospitalized children with enterovirus infections.J. Pediatr. 104 1984 397 401
    OpenUrl
  6. 6.↵
    1. Dagan R.,
    2. Menegus M. A.
    Nonpolio enteroviruses and the febrile infant Human enterovirus infections. Rotbart H. A. 1995 239 254 American Society for Microbiology Washington, D.C
  7. 7.↵
    1. Lina B.,
    2. Pozzeto B.,
    3. Andréoletti L.,
    4. et al.
    Multicenter evaluation of a commercially available PCR assay for diagnosing enterovirus infection in a panel of cerebrospinal fluid specimens.J. Clin. Microbiol. 34 1996 3002 3006
    OpenUrlAbstract/FREE Full Text
  8. 8.↵
    1. Prather S. L.,
    2. Dagan R.,
    3. Jenista A. J.,
    4. Menegus M. A.
    The isolation of enterovirus from blood: a comparison of four processing methods.J. Med. Virol. 14 1984 221 227
    OpenUrlPubMedWeb of Science
  9. 9.↵
    1. Rotbart H. A.,
    2. Sawyer M. H.,
    3. Fast S.,
    4. Lewinsk C.,
    5. Murphy N.,
    6. Keyser E.,
    7. Spadoro J.,
    8. Kao S.,
    9. Loeffelholz S.
    Diagnosis of enteroviral meningitis by using PCR with a colorimetric microwell detection assay.J. Clin. Microbiol. 32 1994 2590 2592
    OpenUrlAbstract/FREE Full Text
  10. 10.↵
    1. Rotbart H. A.
    Enteroviral infections of the central nervous system.Clin. Infect. Dis. 20 1995 971 981
    OpenUrlCrossRefPubMedWeb of Science
  11. 11.↵
    1. Rotbart H. A.,
    2. Romero J.
    Laboratory diagnosis of enteroviral infections Human enterovirus infections. Rotbart H. A. 1995 401 418 American Society for Microbiology Washington, D.C
  12. 12.↵
    1. Rotbart H. A.,
    2. Ahmed A.,
    3. Hickey S.,
    4. Dagan R.,
    5. McCracken J. H.,
    6. Whitley R. J.,
    7. Modlin J. F.,
    8. Cascino M.,
    9. O’Connell J. F.,
    10. Menegus M. A.,
    11. Blum D.
    Diagnosis of enterovirus infection by polymerase chain reaction of multiple specimen types.Pediatr. Infect. Dis. J. 16 1997 409 411
    OpenUrlCrossRefPubMedWeb of Science
  13. 13.↵
    1. Trabelsi A.,
    2. Grattard F.,
    3. Nejmeddine M.,
    4. Aoumi M.,
    5. Bourlet T.,
    6. Pozzeto B.
    Evaluation of an enterovirus group-specific anti-VP1 monoclonal antibody in comparison with neutralization and PCR for rapid identification of enterovirus in cell culture.J. Clin. Microbiol. 33 1995 2454 2457
    OpenUrlAbstract/FREE Full Text
  14. 14.↵
    1. Woodruff J. F.
    Viral myocarditis.Am. J. Pathol. 101 1980 427 479
    OpenUrl
  15. 15.↵
    1. Yerly S.,
    2. Gervaix A.,
    3. Simonet V.,
    4. Callisch M.,
    5. Perrin L.,
    6. Wunderli W.
    Rapid and sensitive detection of enteroviruses in specimens from patients with aseptic meningitis.J. Clin. Microbiol. 34 1996 199 201
    OpenUrlAbstract/FREE Full Text
View Abstract
PreviousNext
Back to top
Download PDF
Citation Tools
Comparison of Use of Cerebrospinal Fluid, Serum, and Throat Swab Specimens in Diagnosis of Enteroviral Acute Neurological Infection by a Rapid RNA Detection PCR Assay
Laurent Andréoletti, Nathalie Blassel-Damman, Anny Dewilde, Louis Vallée, Robin Cremer, Didier Hober, Pierre Wattré
Journal of Clinical Microbiology Feb 1998, 36 (2) 589-591; DOI: 10.1128/JCM.36.2.589-591.1998

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Print

Alerts
Sign In to Email Alerts with your Email Address
Email

Thank you for sharing this Journal of Clinical Microbiology article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
Comparison of Use of Cerebrospinal Fluid, Serum, and Throat Swab Specimens in Diagnosis of Enteroviral Acute Neurological Infection by a Rapid RNA Detection PCR Assay
(Your Name) has forwarded a page to you from Journal of Clinical Microbiology
(Your Name) thought you would be interested in this article in Journal of Clinical Microbiology.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Comparison of Use of Cerebrospinal Fluid, Serum, and Throat Swab Specimens in Diagnosis of Enteroviral Acute Neurological Infection by a Rapid RNA Detection PCR Assay
Laurent Andréoletti, Nathalie Blassel-Damman, Anny Dewilde, Louis Vallée, Robin Cremer, Didier Hober, Pierre Wattré
Journal of Clinical Microbiology Feb 1998, 36 (2) 589-591; DOI: 10.1128/JCM.36.2.589-591.1998
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

KEYWORDS

central nervous system infections
enterovirus
Enterovirus Infections
pharynx
polymerase chain reaction
RNA, Viral

Related Articles

Cited By...

About

  • About JCM
  • Editor in Chief
  • Board of Editors
  • Editor Conflicts of Interest
  • For Reviewers
  • For the Media
  • For Librarians
  • For Advertisers
  • Alerts
  • RSS
  • FAQ
  • Permissions
  • Journal Announcements

Authors

  • ASM Author Center
  • Submit a Manuscript
  • Article Types
  • Resources for Clinical Microbiologists
  • Ethics
  • Contact Us

Follow #JClinMicro

@ASMicrobiology

       

ASM Journals

ASM journals are the most prominent publications in the field, delivering up-to-date and authoritative coverage of both basic and clinical microbiology.

About ASM | Contact Us | Press Room

 

ASM is a member of

Scientific Society Publisher Alliance

 

American Society for Microbiology
1752 N St. NW
Washington, DC 20036
Phone: (202) 737-3600

 

Copyright © 2021 American Society for Microbiology | Privacy Policy | Website feedback

Print ISSN: 0095-1137; Online ISSN: 1098-660X