The integrase of the Macrococcus caseolyticus resistance island mecD (McRI_mecD ) inserts DNA site-specifically into...

Area of Science:

• Microbiology
• Genetics
• Molecular Biology

Background:

• The methicillin resistance gene mecD is found on chromosomal islands in Macrococcus caseolyticus (McRI_mecD).
• McRI_mecD contains an integrase (int) and regulatory genes (intR, xis), integrating at the rpsI gene locus in Gram-positive bacteria.
• Related integrases near the rpsI gene in other species suggest potential transfer of McRI_mecD.

Purpose of the Study:

• To investigate the site-specific recombination function of McRI_mecD.
• To determine if McRI_mecD can integrate into the rpsI locus of other bacterial species.
• To understand the roles of int, xis, and intR in McRI_mecD integration and regulation.

Main Methods:

• Used circular model elements with 5' fragments of McRI_mecD-1 for in vivo integration studies.
• Tested integration into the rpsI locus of Staphylococcus aureus, Staphylococcus pseudintermedius, and Bacillus thuringiensis.
• Analyzed the effects of int, xis, and intR on recombination and gene regulation.

Main Results:

• The integrase (int) and attachment (att) site of McRI_mecD were sufficient for site-specific integration into the rpsI locus of S. aureus, S. pseudintermedius, and B. thuringiensis.
• The xis gene enhanced both integration and excision, influencing att site selection by the Int enzyme.
• The intR gene acted as a negative regulator for int and xis gene expression.

Conclusions:

• The int-xis genes of McRI_mecD-1 provide a site-specific recombination system.
• This system facilitates the acquisition of McRI_mecD in new bacterial hosts.
• Potential for broad-spectrum β-lactam resistance dissemination across genus barriers is highlighted.