A Plasmid-Borne System To Assess the Excision and Integration of Staphylococcal Cassette Chromosome mec Mediated by CcrA and CcrB.
A Plasmid-Borne System To Assess the Excision and Integration of Staphylococcal Cassette Chromosome mec Mediated by CcrA and CcrB.
复制标题
评估 CcrA 和 CcrB 介导的葡萄球菌盒染色体 mec 切除和整合的质粒携带系统。
DOI：
10.1128/jb.00078-15
复制
发表时间：
2015
影响因子：
3.2
通讯作者：
Archer,GordonL
中科院分区：
文献类型：
--
作者：
Wang,Lei;Ahmed,MostafaH;Safo,MartinK;Archer,GordonL
Resistance to methicillin and other β-lactam antibiotics in staphylococci is due tomecA, which is carried on a genomic island, staphylococcal cassette chromosomemec(SCCmec). The chromosomal excision and integration of SCCmecare mediated by the site-specific recombinase CcrAB or CcrC, encoded within this element. A plasmid-borne system was constructed to assess the activities of CcrA and CcrB in the excision and integration of SCCmecin Escherichia coli and Staphylococcus aureus. The excision frequency in E. coli mediated by CcrAB from methicillin-resistant S. aureus (MRSA) strain N315 was only 9.2%, while the integration frequency was 31.4%. In S. aureus the excision and integration frequencies were 11.0% and 18.7%, respectively. Truncated mutants identified the N-terminal domain of either CcrB or CcrA to be necessary for both integration and excision, while the C-terminal domain was important for recombination efficiency. Site-directed mutagenesis of the N-terminal domain identified S11 and R79 of CcrA and S16, R89, T149, and R151 of CcrB to be residues essential for catalytic activities, and the critical location of these residues was consistent with a model of the tertiary structure of the N terminus of CcrA and CcrB. Furthermore, CcrAB and CcrC, cloned from a panel of 6 methicillin-resistant S. aureus strains and 2 methicillin-resistant Staphylococcus epidermidis strains carrying SCCmectypes II, IV, and V, also catalyzed integration at rates 1.3 to 10 times higher than the rates at which they catalyzed excision, similar to the results from N315. The tendency of SCCmecintegration to be favored over excision may explain the low spontaneous excision frequency seen among MRSA strains.IMPORTANCESpontaneous excision of the genomic island (SCCmec) that encodes resistance to beta-lactam antibiotics (methicillin resistance) in staphylococci would convert a methicillin-resistant strain to a methicillin-susceptible strain, improving therapy of difficult-to-treat infections. This study characterizes a model system by which the relative frequencies of excision and integration can be compared. Using a plasmid-based model for excision and integration mediated by the recombinases CcrA and CcrB, integration occurred at a higher frequency than excision, consistent with the low baseline excision frequency seen in most strains. This model system can now be used to study conditions and drugs that may raise the SCCmecexcision frequency and generate strains that are beta-lactam susceptible.
