Preliminary validation of a novel high-resolution melt-based typing method based on the multilocus sequence typing scheme of Streptococcus pyogenes

Richardson, L. J., Tong, S. Y. C., Towers, R. J., Huygens, F., McGregor, K, Fagan, P. K., Currie, B. J., Carapetis, J. R. and Giffard, P. M. (2011) Preliminary validation of a novel high-resolution melt-based typing method based on the multilocus sequence typing scheme of Streptococcus pyogenes. Clinical Microbiology and Infection, 17 9: 1426-1434. doi:10.1111/j.1469-0691.2010.03433.x


Author Richardson, L. J.
Tong, S. Y. C.
Towers, R. J.
Huygens, F.
McGregor, K
Fagan, P. K.
Currie, B. J.
Carapetis, J. R.
Giffard, P. M.
Title Preliminary validation of a novel high-resolution melt-based typing method based on the multilocus sequence typing scheme of Streptococcus pyogenes
Journal name Clinical Microbiology and Infection   Check publisher's open access policy
ISSN 1198-743X
1469-0691
Publication date 2011-09
Year available 2010
Sub-type Article (original research)
DOI 10.1111/j.1469-0691.2010.03433.x
Volume 17
Issue 9
Start page 1426
End page 1434
Total pages 9
Place of publication Oxford, England, U. K.
Publisher Wiley-Blackwell Publishing
Collection year 2012
Language eng
Formatted abstract
The major limitation of current typing methods for Streptococcus pyogenes, such as emm sequence typing and T typing, is that these are based on regions subject to considerable selective pressure. Multilocus sequence typing (MLST) is a better indicator of the genetic backbone of a strain but is not widely used due to high costs. The objective of this study was to develop a robust and cost-effective alternative
to S. pyogenes MLST. A 10-member single nucleotide polymorphism (SNP) set that provides a Simpson’s Index of Diversity (D) of 0.99 with respect to the S. pyogenes MLST database was derived. A typing format involving high-resolution melting (HRM) analysis of small fragments nucleated by each of the resolution-optimized SNPs was developed. The fragments were 59–119 bp in size and, based on differences in G+C content, were predicted to generate three to six resolvable HRM curves. The combination of curves across each of the 10 fragments can be used to generate a melt type (MelT) for each sequence type (ST). The 525 STs currently in the S. pyogenes MLST database are predicted to resolve into 298 distinct MelTs and the method is calculated to provide a D of 0.996 against the MLST database. The MelTs are concordant with the S. pyogenes population structure. To validate the method we examined clinical isolates of S. pyogenes of 70 STs. Curves were generated as predicted by G+C content discriminating the 70 STs into 65 distinct MelTs.
Keyword High-resolution melting
Multilocus sequence typing
Simpsons index of diversity
Single Nucleotide Polymorphism (SNP
Streptococcus pyogenes
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status Non-UQ
Additional Notes Article published online: 22 November 2010

Document type: Journal Article
Sub-type: Article (original research)
Collections: UQ Centre for Clinical Research Publications
Non HERDC
 
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