Abstract
OBJECTIVE: spa-typing of methicillin-resistant Staphylococcus aureus (MRSA) has been widely used in clinical diagnostics and epidemiological studies. We aimed 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 spa gene was amplified by colony PCR using SensiMix™ HRM kit and melting temperature (Tm) and melting curves of the amplicons was analyzed in close tubes using a Rotor-Gene 6000 instrument.
RESULTS: Fifteen out of nineteen spa-types each had distinct Tm, which was sufficient to unambiguously type each of these spa-types. The remaining 4 spa-types cannot be separated by Tm alone: t008 and t2770 shared Tm (80.3°C) and t021 and t311 shared Tm (80.0°C). But, they can be separated based on the shapes of their melting curves. There are discrepancies between ours and previous studies, suggesting that standardization reminds a challenge for cross references.
CONCLUSION: HRM-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-references between labs globally.
METHODS: The polymorphic X region of spa gene was amplified by colony PCR using SensiMix™ HRM kit and melting temperature (Tm) and melting curves of the amplicons was analyzed in close tubes using a Rotor-Gene 6000 instrument.
RESULTS: Fifteen out of nineteen spa-types each had distinct Tm, which was sufficient to unambiguously type each of these spa-types. The remaining 4 spa-types cannot be separated by Tm alone: t008 and t2770 shared Tm (80.3°C) and t021 and t311 shared Tm (80.0°C). But, they can be separated based on the shapes of their melting curves. There are discrepancies between ours and previous studies, suggesting that standardization reminds a challenge for cross references.
CONCLUSION: HRM-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-references between labs globally.
| Original language | English |
|---|---|
| Article number | S1201-9712(15)00109-5 |
| Pages (from-to) | 1-18 |
| Number of pages | 18 |
| Journal | International Journal of Infectious Diseases |
| Early online date | 11 May 2015 |
| DOIs | |
| Publication status | Published - 2015 |
Keywords
- high-resolution melting (HRM)
- spa-types
- melting temperature (Tm)
- methicillin resistant Staphylococcus aureus (MRSA)