Differentiation of Austrian Salmonella serotypes using high-resolution melting curve analysis
Abstract number: O102
Zeinzinger J., Stoeger A., Kornschober C., Mach R., Allerberger F., Ruppitsch W., Pietzka A.
Objectives: Effective epidemiological surveillance requires the accurate subtyping of strains. The standard method for differentiation of Salmonella strains is serotyping. Despite the utility of serotyping, testing with a complete set of sera is time consuming and requires a well trained technician; problems associated with antiserum production and occurrence of strains for which a serotype antigen cannot be detected prompted reference laboratories to pursue molecular approaches for serotyping.
The aim of this study was to assess the usefulness of high-resolution melting (HRM) curve analysis on the LightCycler 480 PCR system as a tool for accurate and fast molecular typing of Salmonella enterica subsp. enterica strains.
Methods: A collection of Salmonella isolates representing diverse serotypes was recently analysed using Amplified Fragment Length Polymorphism (AFLP) (1). A phylogenetic cluster comprising the serotypes Blockley, Virchow, Braenderup, Manhattan and Muenchen was chosen to evaluate the potential of HRM analysis for molecular typing. A 171 bp fragment of the gyrase B gene (gyrB) was amplified for HRM curve analysis on a LightCycler 480 instrument (Roche Diagnostics, Penzberg, Germany).
Results: HRM curve analysis of a 171 bp amplicon of the gyrB gene resulted in five different melting curves (Figure 1), and thus allowed the rapid and accurate discrimination of the investigated serotypes.
Figure 1. Melting curve profiles of the tested Salmonella serotypes.
Conclusion: HRM analysis is a new single-step closed-tube screening method for fast mutation detection and can therefore be used to discriminate even genetically closely related samples. HRM analysis has the potential to complement the classical serotyping of Salmonella isolates due to its discriminatory power and simplicity.
1. Pietzka A., Stöger A., Kornschober C., Zeinzinger J., Ruppitsch W., Franz Allerberger F. (2008) Amplified fragment length polymorphism of diverse Salmonella enterica serovars for serotype differentiation and identification of serotype specific genetic markers. Infection. 36(s1): 7273.
|Session name:||Abstracts 20th European Congress of Clinical Microbiology and Infectious Diseases|
|Location:||Vienna, Austria, 10 - 13 April 2010|
|Back to top|