Logo of jcmPermissionsJournals.ASM.orgJournalJCM ArticleJournal InfoAuthorsReviewers
J Clin Microbiol. 1990 May; 28(5): 843–850.
PMCID: PMC267821

Molecular detection and identification of enteroviruses using enzymatic amplification and nucleic acid hybridization.


Analysis of enteroviral genomes has revealed that the 5' nontranslated region is highly conserved, providing consensus sequences for the design of oligonucleotides which should anneal to most, if not all, human enteroviral RNAs. We designed and used a pair of such generic primers to enzymatically amplify cDNA from coxsackievirus group B types 1 through 6, poliovirus types 1 through 3, 4 coxsackievirus A types, and 29 echoviruses. The polymerase chain reaction (PCR) products generated with these enteroviral primers were analyzed by agarose gel electrophoresis, Southern blotting, or slot blot hybridization. A genotype-specific PCR was used to detect coxsackievirus B3, to the exclusion of other enteroviruses, by using a coxsackievirus B3 genome-specific primer pair that was derived from sequences coding for part of a capsid protein. A technique is demonstrated by which individual genotypes, for which no sequence information is known, can be identified by high-criterion hybridization analysis following amplification with generic enterovirus PCR primers.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.9M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Click on the image to see a larger version.

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Beck MA, Tracy SM. Murine cell-mediated immune response recognizes an enterovirus group-specific antigen(s). J Virol. 1989 Oct;63(10):4148–4156. [PMC free article] [PubMed]
  • Laird-Meeter K, van Domburg R, Bos E, Hugenholtz PG. Survival at 5 to 10 years after aorto-coronary bypass operations in 1041 consecutive patients. Eur Heart J. 1987 May;8(5):449–456. [PubMed]
  • Cook AF, Vuocolo E, Brakel CL. Synthesis and hybridization of a series of biotinylated oligonucleotides. Nucleic Acids Res. 1988 May 11;16(9):4077–4095. [PMC free article] [PubMed]
  • DiLella AG, Woo SL. Hybridization of genomic DNA to oligonucleotide probes in the presence of tetramethylammonium chloride. Methods Enzymol. 1987;152:447–451. [PubMed]
  • Easton AJ, Eglin RP. The detection of coxsackievirus RNA in cardiac tissue by in situ hybridization. J Gen Virol. 1988 Feb;69(Pt 2):285–291. [PubMed]
  • Feinberg AP, Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. [PubMed]
  • Hogle JM, Chow M, Filman DJ. Three-dimensional structure of poliovirus at 2.9 A resolution. Science. 1985 Sep 27;229(4720):1358–1365. [PubMed]
  • Iizuka N, Kuge S, Nomoto A. Complete nucleotide sequence of the genome of coxsackievirus B1. Virology. 1987 Jan;156(1):64–73. [PubMed]
  • Innis MA, Myambo KB, Gelfand DH, Brow MA. DNA sequencing with Thermus aquaticus DNA polymerase and direct sequencing of polymerase chain reaction-amplified DNA. Proc Natl Acad Sci U S A. 1988 Dec;85(24):9436–9440. [PMC free article] [PubMed]
  • Jamison RM. An electron microscopic study of the intracellular development of echovirus 22. Arch Gesamte Virusforsch. 1974;44(3):184–194. [PubMed]
  • Jenkins O, Booth JD, Minor PD, Almond JW. 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. [PubMed]
  • Kandolf R, Ameis D, Kirschner P, Canu A, Hofschneider PH. 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. [PMC free article] [PubMed]
  • Kandolf R, Hofschneider PH. Molecular cloning of the genome of a cardiotropic Coxsackie B3 virus: full-length reverse-transcribed recombinant cDNA generates infectious virus in mammalian cells. Proc Natl Acad Sci U S A. 1985 Jul;82(14):4818–4822. [PMC free article] [PubMed]
  • Kitamura N, Semler BL, Rothberg PG, Larsen GR, Adler CJ, Dorner AJ, Emini EA, Hanecak R, Lee JJ, van der Werf S, et al. Primary structure, gene organization and polypeptide expression of poliovirus RNA. Nature. 1981 Jun 18;291(5816):547–553. [PubMed]
  • Klein RD, Selsing E, Wells RD. A rapid microscale technique for isolation of recombinant plasmid DNA suitable for restriction enzyme analysis. Plasmid. 1980 Jan;3(1):88–91. [PubMed]
  • Kwoh DY, Davis GR, Whitfield KM, Chappelle HL, DiMichele LJ, Gingeras TR. Transcription-based amplification system and detection of amplified human immunodeficiency virus type 1 with a bead-based sandwich hybridization format. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1173–1177. [PMC free article] [PubMed]
  • La Monica N, Meriam C, Racaniello VR. Mapping of sequences required for mouse neurovirulence of poliovirus type 2 Lansing. J Virol. 1986 Feb;57(2):515–525. [PMC free article] [PubMed]
  • Li HH, Gyllensten UB, Cui XF, Saiki RK, Erlich HA, Arnheim N. Amplification and analysis of DNA sequences in single human sperm and diploid cells. Nature. 1988 Sep 29;335(6189):414–417. [PubMed]
  • Lindberg AM, Stålhandske PO, Pettersson U. Genome of coxsackievirus B3. Virology. 1987 Jan;156(1):50–63. [PubMed]
  • Murakawa GJ, Zaia JA, Spallone PA, Stephens DA, Kaplan BE, Wallace RB, Rossi JJ. Direct detection of HIV-1 RNA from AIDS and ARC patient samples. DNA. 1988 May;7(4):287–295. [PubMed]
  • Racaniello VR, Baltimore D. Molecular cloning of poliovirus cDNA and determination of the complete nucleotide sequence of the viral genome. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4887–4891. [PMC free article] [PubMed]
  • Rossmann MG, Arnold E, Erickson JW, Frankenberger EA, Griffith JP, Hecht HJ, Johnson JE, Kamer G, Luo M, Mosser AG, et al. Structure of a human common cold virus and functional relationship to other picornaviruses. Nature. 1985 Sep 12;317(6033):145–153. [PubMed]
  • Rotbart HA, Eastman PS, Ruth JL, Hirata KK, Levin MJ. Nonisotopic oligomeric probes for the human enteroviruses. J Clin Microbiol. 1988 Dec;26(12):2669–2671. [PMC free article] [PubMed]
  • Rotbart HA, Levin MJ, Villarreal LP, Tracy SM, Semler BL, Wimmer E. Factors affecting the detection of enteroviruses in cerebrospinal fluid with coxsackievirus B3 and poliovirus 1 cDNA probes. J Clin Microbiol. 1985 Aug;22(2):220–224. [PMC free article] [PubMed]
  • Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. [PubMed]
  • Saiki RK, Scharf S, Faloona F, Mullis KB, Horn GT, Erlich HA, Arnheim N. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science. 1985 Dec 20;230(4732):1350–1354. [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Seal LA, Jamison RM. Evidence for secondary structure within the virion RNA of echovirus 22. J Virol. 1984 May;50(2):641–644. [PMC free article] [PubMed]
  • Southern EM. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. [PubMed]
  • Stanway G, Hughes PJ, Mountford RC, Reeve P, Minor PD, Schild GC, Almond JW. Comparison of the complete nucleotide sequences of the genomes of the neurovirulent poliovirus P3/Leon/37 and its attenuated Sabin vaccine derivative P3/Leon 12a1b. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1539–1543. [PMC free article] [PubMed]
  • Studencki AB, Wallace RB. Allele-specific hybridization using oligonucleotide probes of very high specific activity: discrimination of the human beta A- and beta S-globin genes. DNA. 1984;3(1):7–15. [PubMed]
  • Tabor S, Richardson CC. DNA sequence analysis with a modified bacteriophage T7 DNA polymerase. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4767–4771. [PMC free article] [PubMed]
  • TAMM I, EGGERS HJ. Differences in the selective virus inhibitory action of 2-(alpha-hydroxybenzyl)-benzimidazole and guanidine HCl. Virology. 1962 Nov;18:439–447. [PubMed]
  • 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. [PubMed]
  • Tracy S. A comparison of genomic homologies among the coxsackievirus B group: use of fragments of the cloned coxsackievirus B3 genome as probes. J Gen Virol. 1984 Dec;65(Pt 12):2167–2172. [PubMed]
  • Tracy S. Comparison of genomic homologies in the coxsackievirus B group by use of cDNA:RNA dot-blot hybridization. J Clin Microbiol. 1985 Mar;21(3):371–374. [PMC free article] [PubMed]
  • Tracy S, Liu HL, Chapman NM. Coxsackievirus B3: primary structure of the 5' non-coding and capsid protein-coding regions of the genome. Virus Res. 1985 Oct;3(3):263–270. [PubMed]
  • Tracy S, Smith RA. A comparison of the genomes of polioviruses by cDNA:RNA hybridization. J Gen Virol. 1981 Jul;55(Pt 1):193–199. [PubMed]
  • Zhang HY, Yousef GE, Bowles NE, Archard LC, Mann GF, Mowbray JF. 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. [PubMed]

Articles from Journal of Clinical Microbiology are provided here courtesy of American Society for Microbiology (ASM)


Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...


  • Compound
    PubChem Compound links
  • MedGen
    Related information in MedGen
  • PubMed
    PubMed citations for these articles
  • Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...