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. 1992 Jan;30(1):160–165. doi: 10.1128/jcm.30.1.160-165.1992

General primer-mediated polymerase chain reaction for detection of enteroviruses: application for diagnostic routine and persistent infections.

G J Zoll 1, W J Melchers 1, H Kopecka 1, G Jambroes 1, H J van der Poel 1, J M Galama 1
PMCID: PMC265013  PMID: 1370845

Abstract

The aim of this study was to determine the applicability of the polymerase chain reaction (PCR) for routine diagnostic use and for the detection of persistent enteroviral infections. To this end, general primers were selected in the highly conserved part of the 5'-noncoding region of the enteroviral genome. They were tested on 66 different enterovirus serotypes. A specific fragment was amplified from 60 of 66 serotypes. An amplification product was not observed from coxsackievirus types A11, A17, and A24 and echovirus types 16, 22, and 23. Enteroviral RNA was detected by the PCR in routinely collected throat swabs and stool specimens that were found to be positive for enterovirus by isolation in tissue culture. Enteroviral RNA was detected in one of five myocardial biopsy specimens from patients with dilated cardiomyopathy, implicating virus persistence. No amplification product was obtained from eight control samples. Our results demonstrate the significance of the PCR for the detection of enteroviral RNA and, in particular, for the demonstration of persistent enteroviral infections.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Auvinen P., Hyypiä T. Echoviruses include genetically distinct serotypes. J Gen Virol. 1990 Sep;71(Pt 9):2133–2139. doi: 10.1099/0022-1317-71-9-2133. [DOI] [PubMed] [Google Scholar]
  2. Barrett-Connor E. Is insulin-dependent diabetes mellitus caused by coxsackievirus B infection? A review of the epidemiologic evidence. Rev Infect Dis. 1985 Mar-Apr;7(2):207–215. doi: 10.1093/clinids/7.2.207. [DOI] [PubMed] [Google Scholar]
  3. Boom R., Sol C. J., Salimans M. M., Jansen C. L., Wertheim-van Dillen P. M., van der Noordaa J. Rapid and simple method for purification of nucleic acids. J Clin Microbiol. 1990 Mar;28(3):495–503. doi: 10.1128/jcm.28.3.495-503.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bowles N. E., Dubowitz V., Sewry C. A., Archard L. C. Dermatomyositis, polymyositis, and Coxsackie-B-virus infection. Lancet. 1987 May 2;1(8540):1004–1007. doi: 10.1016/s0140-6736(87)92271-9. [DOI] [PubMed] [Google Scholar]
  5. Bowles N. E., Rose M. L., Taylor P., Banner N. R., Morgan-Capner P., Cunningham L., Archard L. C., Yacoub M. H. End-stage dilated cardiomyopathy. Persistence of enterovirus RNA in myocardium at cardiac transplantation and lack of immune response. Circulation. 1989 Nov;80(5):1128–1136. doi: 10.1161/01.cir.80.5.1128. [DOI] [PubMed] [Google Scholar]
  6. Bruce C., al-Nakib W., Forsyth M., Stanway G., Almond J. W. Detection of enteroviruses using cDNA and synthetic oligonucleotide probes. J Virol Methods. 1989 Aug;25(2):233–240. doi: 10.1016/0166-0934(89)90035-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chang K. H., Auvinen P., Hyypiä T., Stanway G. The nucleotide sequence of coxsackievirus A9; implications for receptor binding and enterovirus classification. J Gen Virol. 1989 Dec;70(Pt 12):3269–3280. doi: 10.1099/0022-1317-70-12-3269. [DOI] [PubMed] [Google Scholar]
  8. Chapman N. M., Tracy S., Gauntt C. J., Fortmueller U. Molecular detection and identification of enteroviruses using enzymatic amplification and nucleic acid hybridization. J Clin Microbiol. 1990 May;28(5):843–850. doi: 10.1128/jcm.28.5.843-850.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Coller B. A., Chapman N. M., Beck M. A., Pallansch M. A., Gauntt C. J., Tracy S. M. Echovirus 22 is an atypical enterovirus. J Virol. 1990 Jun;64(6):2692–2701. doi: 10.1128/jvi.64.6.2692-2701.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cornelissen M. T., Smits H. L., Briët M. A., van den Tweel J. G., Struyk A. P., van der Noordaa J., ter Schegget J. Uniformity of the splicing pattern of the E6/E7 transcripts in human papillomavirus type 16-transformed human fibroblasts, human cervical premalignant lesions and carcinomas. J Gen Virol. 1990 May;71(Pt 5):1243–1246. doi: 10.1099/0022-1317-71-5-1243. [DOI] [PubMed] [Google Scholar]
  11. Hughes P. J., North C., Jellis C. H., Minor P. D., Stanway G. The nucleotide sequence of human rhinovirus 1B: molecular relationships within the rhinovirus genus. J Gen Virol. 1988 Jan;69(Pt 1):49–58. doi: 10.1099/0022-1317-69-1-49. [DOI] [PubMed] [Google Scholar]
  12. Hughes P. J., North C., Minor P. D., Stanway G. The complete nucleotide sequence of coxsackievirus A21. J Gen Virol. 1989 Nov;70(Pt 11):2943–2952. doi: 10.1099/0022-1317-70-11-2943. [DOI] [PubMed] [Google Scholar]
  13. Hyypiä T., Auvinen P., Maaronen M. Polymerase chain reaction for human picornaviruses. J Gen Virol. 1989 Dec;70(Pt 12):3261–3268. doi: 10.1099/0022-1317-70-12-3261. [DOI] [PubMed] [Google Scholar]
  14. Iizuka N., Kuge S., Nomoto A. Complete nucleotide sequence of the genome of coxsackievirus B1. Virology. 1987 Jan;156(1):64–73. doi: 10.1016/0042-6822(87)90436-3. [DOI] [PubMed] [Google Scholar]
  15. Jenkins O., Booth J. D., Minor P. D., Almond J. W. The complete nucleotide sequence of coxsackievirus B4 and its comparison to other members of the Picornaviridae. J Gen Virol. 1987 Jul;68(Pt 7):1835–1848. doi: 10.1099/0022-1317-68-7-1835. [DOI] [PubMed] [Google Scholar]
  16. Jin O., Sole M. J., Butany J. W., Chia W. K., McLaughlin P. R., Liu P., Liew C. C. Detection of enterovirus RNA in myocardial biopsies from patients with myocarditis and cardiomyopathy using gene amplification by polymerase chain reaction. Circulation. 1990 Jul;82(1):8–16. doi: 10.1161/01.cir.82.1.8. [DOI] [PubMed] [Google Scholar]
  17. Kandolf R., Ameis D., Kirschner P., Canu A., Hofschneider P. H. In situ detection of enteroviral genomes in myocardial cells by nucleic acid hybridization: an approach to the diagnosis of viral heart disease. Proc Natl Acad Sci U S A. 1987 Sep;84(17):6272–6276. doi: 10.1073/pnas.84.17.6272. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kwok S., Higuchi R. Avoiding false positives with PCR. Nature. 1989 May 18;339(6221):237–238. doi: 10.1038/339237a0. [DOI] [PubMed] [Google Scholar]
  19. Lindberg A. M., Stålhandske P. O., Pettersson U. Genome of coxsackievirus B3. Virology. 1987 Jan;156(1):50–63. doi: 10.1016/0042-6822(87)90435-1. [DOI] [PubMed] [Google Scholar]
  20. Melchers W. J., Schift R., Stolz E., Lindeman J., Quint W. G. Human papillomavirus detection in urine samples from male patients by the polymerase chain reaction. J Clin Microbiol. 1989 Aug;27(8):1711–1714. doi: 10.1128/jcm.27.8.1711-1714.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Najarian R., Caput D., Gee W., Potter S. J., Renard A., Merryweather J., Van Nest G., Dina D. Primary structure and gene organization of human hepatitis A virus. Proc Natl Acad Sci U S A. 1985 May;82(9):2627–2631. doi: 10.1073/pnas.82.9.2627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Olive D. M., Al-Mufti S., Al-Mulla W., Khan M. A., Pasca A., Stanway G., Al-Nakib W. Detection and differentiation of picornaviruses in clinical samples following genomic amplification. J Gen Virol. 1990 Sep;71(Pt 9):2141–2147. doi: 10.1099/0022-1317-71-9-2141. [DOI] [PubMed] [Google Scholar]
  23. Petitjean J., Quibriac M., Freymuth F., Fuchs F., Laconche N., Aymard M., Kopecka H. Specific detection of enteroviruses in clinical samples by molecular hybridization using poliovirus subgenomic riboprobes. J Clin Microbiol. 1990 Feb;28(2):307–311. doi: 10.1128/jcm.28.2.307-311.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rivera V. M., Welsh J. D., Maizel J. V., Jr Comparative sequence analysis of the 5' noncoding region of the enteroviruses and rhinoviruses. Virology. 1988 Jul;165(1):42–50. doi: 10.1016/0042-6822(88)90656-3. [DOI] [PubMed] [Google Scholar]
  25. Rotbart H. A., Abzug M. J., Murray R. S., Murphy N. L., Levin M. J. Intracellular detection of sense and antisense enteroviral RNA by in situ hybridization. J Virol Methods. 1988 Dec;22(2-3):295–301. doi: 10.1016/0166-0934(88)90111-5. [DOI] [PubMed] [Google Scholar]
  26. Rotbart H. A. Enzymatic RNA amplification of the enteroviruses. J Clin Microbiol. 1990 Mar;28(3):438–442. doi: 10.1128/jcm.28.3.438-442.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Rotbart H. A., Kinsella J. P., Wasserman R. L. Persistent enterovirus infection in culture-negative meningoencephalitis: demonstration by enzymatic RNA amplification. J Infect Dis. 1990 Apr;161(4):787–791. doi: 10.1093/infdis/161.4.787. [DOI] [PubMed] [Google Scholar]
  28. Ryan M. D., Jenkins O., Hughes P. J., Brown A., Knowles N. J., Booth D., Minor P. D., Almond J. W. The complete nucleotide sequence of enterovirus type 70: relationships with other members of the picornaviridae. J Gen Virol. 1990 Oct;71(Pt 10):2291–2299. doi: 10.1099/0022-1317-71-10-2291. [DOI] [PubMed] [Google Scholar]
  29. Skinner M. A., Racaniello V. R., Dunn G., Cooper J., Minor P. D., Almond J. W. New model for the secondary structure of the 5' non-coding RNA of poliovirus is supported by biochemical and genetic data that also show that RNA secondary structure is important in neurovirulence. J Mol Biol. 1989 May 20;207(2):379–392. doi: 10.1016/0022-2836(89)90261-1. [DOI] [PubMed] [Google Scholar]
  30. Toyoda H., Kohara M., Kataoka Y., Suganuma T., Omata T., Imura N., Nomoto A. Complete nucleotide sequences of all three poliovirus serotype genomes. Implication for genetic relationship, gene function and antigenic determinants. J Mol Biol. 1984 Apr 25;174(4):561–585. doi: 10.1016/0022-2836(84)90084-6. [DOI] [PubMed] [Google Scholar]
  31. Yousef G. E., Bell E. J., Mann G. F., Murugesan V., Smith D. G., McCartney R. A., Mowbray J. F. Chronic enterovirus infection in patients with postviral fatigue syndrome. Lancet. 1988 Jan 23;1(8578):146–150. doi: 10.1016/s0140-6736(88)92722-5. [DOI] [PubMed] [Google Scholar]
  32. Zhang H. Y., Yousef G. E., Bowles N. E., Archard L. C., Mann G. F., Mowbray J. F. Detection of enterovirus RNA in experimentally infected mice by molecular hybridisation: specificity of subgenomic probes in quantitative slot blot and in situ hybridisation. J Med Virol. 1988 Dec;26(4):375–386. doi: 10.1002/jmv.1890260405. [DOI] [PubMed] [Google Scholar]

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