title: Evaluation of spa-typing of methicillin-resistant Staphylococcus aureus using high-resolution melting analysis.
abstract: OBJECTIVE
  spa-typing of methicillin-resistant Staphylococcus aureus (MRSA) has been used widely in clinical diagnostics and epidemiological studies. The aim of this study was to evaluate high-resolution melting (HRM) as a rapid and cost-effective method, to replace DNA-sequencing, for spa-typing in a global collection of 50 MRSA isolates.
  
  
  METHODS
  The polymorphic X region of the spa gene was amplified by colony PCR using the SensiMix HRM kit, and the melting temperature (Tm) and melting curves of the amplicons were analyzed in close tubes using a Rotor-Gene 6000 instrument.
  
  
  RESULTS
  Fifteen out of 19 spa-types each had a distinct Tm, which was sufficient to unambiguously type each of these spa-types. The remaining four spa-types could not be separated by Tm alone: t008 and t2770 shared a Tm (80.3°C) and t021 and t311 shared a Tm (80.0°C). However, they could be separated based on the shapes of their melting curves. There are discrepancies between the findings of the present study and those of previous studies, suggesting that standardization remains a challenge for cross-referencing.
  
  
  CONCLUSIONS
  HRM-based spa-typing is reproducible, simple, rapid, and cost-effective. t037 is prevalent in Brazil and Sudan, while diverse spa-types are found in Scotland and Saudi Arabia. Standardization is required for cross-referencing between laboratories globally.
authors.affiliation: Department of Laboratory Medicine, Division of Clinical Microbiology, Karolinska Institutet, Karolinska University Hospital, Huddinge, SE-141 86 Stockholm, Sweden; Infection Prevention and Control Department, King Abdul Aziz Specialist Hospital, Taif, Saudi Arabia. Electronic address: waleedmazi@ki.se.
authors.forename: Waleed
authors.initials: W
authors.lastname: Mazi
authors.affiliation: Faculty of Health and Life Sciences, Northumbria University, Newcastle upon Tyne, UK.
authors.forename: Vartul
authors.initials: V
authors.lastname: Sangal
authors.affiliation: Department of Laboratory Medicine, Division of Clinical Microbiology, Karolinska Institutet, Karolinska University Hospital, Huddinge, SE-141 86 Stockholm, Sweden.
authors.forename: Gunnar
authors.initials: G
authors.lastname: Sandstrom
authors.affiliation: Department of Laboratory Medicine, Division of Clinical Microbiology, Karolinska Institutet, Karolinska University Hospital, Huddinge, SE-141 86 Stockholm, Sweden; University of Medical Sciences and Technology, Department of Microbiology, Khartoum, Sudan.
authors.forename: Amir
authors.initials: A
authors.lastname: Saeed
authors.affiliation: Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, UK.
authors.forename: Jun
authors.initials: J
authors.lastname: Yu
mesh_terms: D001938:Brazil; D006801:Humans; D055624:Methicillin-Resistant Staphylococcus aureus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D058889:Molecular Typing; D009691:Nucleic Acid Denaturation; D016133:Polymerase Chain Reaction; D012529:Saudi Arabia; D012606:Scotland; D017422:Sequence Analysis, DNA; D013205:Staphylococcal Protein A / Q000235:genetics*; D013397:Sudan
chemical_list: D013205:Staphylococcal Protein A
keywords: High-resolution melting (HRM); Melting temperature (Tm); Methicillin-resistant Staphylococcus aureus (MRSA); spa-types