1887

Abstract

Real-time PCR based on the capsule transfer gene () is a significant aid in the diagnosis of meningococcal infection but fails to detect a high proportion (60 %) of non-groupable strains associated with nasopharyngeal carriage. This study aimed to design a novel real-time (TaqMan) PCR that would detect more strains of meningococci and be suitable for large-scale carriage studies. Primer and probe sequences were based on the meningococcal gene and designed specifically to exclude the highly related pseudogene in . The specificity of the assay was confirmed by testing strains of known to contain the pseudogene together with commensal strains of and . None of these was detected in the assay. strains representing a wide range of serogroups together with non-groupable strains isolated from the nasopharynx were tested by assay and the novel -based TaqMan PCR. All carriage strains were detected by the -based assay including four that gave weak or no reaction with the assay. Comparison of and assays on 71 throat swabs obtained from university students showed that the assay detected meningococcal DNA in all samples that were positive plus three that were negative but culture positive. This novel -based TaqMan assay provides a highly specific method for detecting meningococcal DNA that is more sensitive than the assay for detecting meningococcal carriage and is particularly suitable for carriage studies where non-groupable strains and other are present.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/jmm.0.45847-0
2005-05-01
2019-11-18
Loading full text...

Full text loading...

/deliver/fulltext/jmm/54/5/JM540508.html?itemId=/content/journal/jmm/10.1099/jmm.0.45847-0&mimeType=html&fmt=ahah

References

  1. Altschul, S. F., Gish, W., Miller, W., Myers, E. W. & Lipman, D. J. ( 1990;). Basic local alignment search tool. J Mol Biol 215, 403–410.[CrossRef]
    [Google Scholar]
  2. Barlow, A. K., Heckels, J. E. & Clarke, I. N. ( 1989;). The class 1 outer membrane protein of Neisseria meningitidis: gene sequence and structural and immunological similarities to gonococcal porins. Mol Microbiol 3, 131–139.[CrossRef]
    [Google Scholar]
  3. Clarke, S. C., Diggle, M. A. & Edwards, G. F. S. ( 2001;). Automated non-culture-based sequence typing of meningococci from body fluids. Br J Biomed Sci 58, 230–234.
    [Google Scholar]
  4. Communicable Disease Surveillance Centre ( 2001;). The impact of conjugate group C meningococcal vaccination. Commun Dis Rep CDR Wkly (Online) 11(2). http://www.hpa.org.uk/cdr/PDFfiles/2001/cdr0201.pdf, –234.
    [Google Scholar]
  5. Corless, C. E., Guiver, M., Borrow, R., Edwards-Jones, V., Fox, A. J. & Kaczmarski, E. B. ( 2001;). Simultaneous detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae in suspected cases of meningitis and septicaemia using real-time PCR. J Clin Microbiol 39, 1553–1558.[CrossRef]
    [Google Scholar]
  6. Diaz, J. L. & Heckels, J. E. ( 1982;). Antigenic variation of outer membrane II in colonial variants of Neisseria gonorrhoeae P9. J Gen Microbiol 128, 585–591.
    [Google Scholar]
  7. Feavers, I. M. & Maiden, M. C. J. ( 1998;). A gonococcal porA pseudogene: implications for understanding the evolution and pathogenicity of Neisseria gonorrhoeae. Mol Microbiol 30, 647–656.[CrossRef]
    [Google Scholar]
  8. Jordens, J. Z., Williams, J. N., Jones, G. R. & Heckels, J. E. ( 2002;). Detection of meningococcal carriage by culture and PCR of throat swabs and mouth gargles. J Clin Microbiol 40, 75–79.[CrossRef]
    [Google Scholar]
  9. Maiden, M. C., Suker, J., McKenna, A. J., Bygraves, J. A. & Feavers, I. M. ( 1991;). Comparison of the class 1 outer membrane proteins of eight serological reference strains of Neisseria meningitidis. Mol Microbiol 5, 727–736.[CrossRef]
    [Google Scholar]
  10. McGuinness, B., Barlow, A. K., Clarke, I. N., Farley, J. E., Anilionis, A., Poolman, J. T. & Heckels, J. E. ( 1990;). Deduced amino acid sequences of class 1 protein (PorA) from three strains of Neisseria meningitidis.Synthetic peptides define the epitopes responsible for serosubtype specificity. J Exp Med 171, 1871–1882.[CrossRef]
    [Google Scholar]
  11. McGuinness, B. T., Lambden, P. R. & Heckels, J. E. ( 1993;). Class 1 outer membrane protein of Neisseria meningitidis: epitope analysis of the antigenic diversity between strains, implications for subtype definition and molecular epidemiology. Mol Microbiol 7, 505–514.[CrossRef]
    [Google Scholar]
  12. Van der Ende, A., Hopman, C. T. & Dankert, J. ( 1999;). Deletion of porA by recombination between clusters of repetitive extragenic palindromic sequences in Neisseria meningitidis. Infect Immun 67, 2928–2934.
    [Google Scholar]
  13. Van der Ley, P., Heckels, J. E., Virji, M., Hoogerhout, P. & Pollman, J. T. ( 1991;). Topology of outer membrane porins in pathogenic Neisseria spp. Infect Immun 59, 2963–2971.
    [Google Scholar]
  14. Van der Voort, E. R., Van Dijken, H., Kuipers, B., Van der Biezen, J., Van der Ley, P., Meylis, J., Claassen, I. & Poolman, J. ( 1997;). Human B- and T-cell responses after immunization with a hexavalent PorA meningococcal outer membrane vesicle vaccine. Infect Immun 65, 5184–5190.
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/jmm.0.45847-0
Loading
/content/journal/jmm/10.1099/jmm.0.45847-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error