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SCCmec transformation requires living donor cells in mixed biofilms
Mais Maree1, Yuri Ushijima1, Pedro B Fernandes1
1Institute of Medicine, University of Tsukuba, Japan.
Abstract:
Methicillin-resistant Staphylococcus aureus (MRSA) is an important human pathogen that has emerged through the horizontal acquisition of the staphylococcal cassette chromosome mec (SCCmec). Previously, we showed that SCCmec from heat-killed donors can be transferred via natural transformation in biofilms at frequencies of 10-8-10-7. Here, we show an improved transformation assay of SCCmec with frequencies up to 10-2 using co-cultured biofilms with living donor cells. The Ccr-attB system played an important role in SCCmec transfer, and the deletion of ccrAB recombinase genes reduced the frequency ∼30-fold. SCCmec could be transferred from either MRSA or methicillin-resistant coagulase-negative staphylococci to some methicillin-sensitive S. aureus recipients. In addition, the transformation of other plasmid or chromosomal genes is enhanced by using living donor cells. This study emphasizes the role of natural transformation as an evolutionary ability of S. aureus and in MRSA emergence.
Insights
Living donor cells significantly improve the transfer of staphylococcal cassette chromosome mec (SCCmec) via natural transformation in biofilms. This enhanced method aids understanding of MRSA emergence and gene transfer in Staphylococcus aureus.
Area of Science:
- Microbiology
- Genetics
- Molecular Biology
Background:
- Methicillin-resistant Staphylococcus aureus (MRSA) emergence is linked to horizontal gene transfer of the staphylococcal cassette chromosome mec (SCCmec).
- Previous studies demonstrated low-frequency SCCmec transfer via natural transformation using heat-killed donors in biofilms.
Purpose of the Study:
- To improve the efficiency of SCCmec natural transformation in Staphylococcus aureus biofilms.
- To investigate the role of the Ccr-attB system in SCCmec transfer.
- To assess the impact of using living donor cells on gene transfer efficiency.
Main Methods:
- Co-culturing living MRSA or methicillin-resistant coagulase-negative staphylococci donor cells with methicillin-sensitive Staphylococcus aureus recipient cells in biofilms.
- Developing an improved natural transformation assay.
- Analyzing the effect of deleting ccrAB recombinase genes on SCCmec transfer frequency.
Main Results:
- Achieved SCCmec transformation frequencies up to 10^-2 using living donor cells, a significant improvement over previous methods.
- Demonstrated that the Ccr-attB system is crucial for SCCmec transfer, with ccrAB deletion reducing frequency approximately 30-fold.
- Showed successful SCCmec transfer from resistant strains to sensitive Staphylococcus aureus recipients.
- Observed enhanced transformation of other plasmid and chromosomal genes when using living donor cells.
Conclusions:
- Natural transformation is a key evolutionary mechanism for Staphylococcus aureus, contributing to MRSA emergence.
- Utilizing living donor cells in co-cultured biofilms substantially enhances SCCmec transfer efficiency.
- The Ccr-attB system plays a vital role in the natural transformation process of SCCmec.
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