The Cloning of Chromosomal DNA Associated with Methicillin and Other Resistances in Staphylococcus aureus

Abstract

Competitive hybridization was used to detect the deletion of chromosomal DNA accompanying the loss of resistance to methicillin (and concomitantly, to cadmium, mercury and tetracycline) from a clinical strain of methicillin-resistant Staphylococcus aureus (MRSA). The method was also used to screen a partial plasmid library of chromosomal HindIII fragments from the MRSA strain. Eight recombinant plasmid clones were identified as containing DNA included in the deletion. These clones were used as probes to screen a phage library of the total DNA of the same MRSA strain, resulting in the isolation of overlapping recombinant phage clones carrying 24 kb of the deleted DNA. Two of the cloned HindIII fragments were associated closely with methicillin resistance, as shown by probing DNA from an independent methicillin-sensitive/resistant transduced strain pair and from two MRSA strains following growth in the presence of high concentrations of methicillin. The endonuclease map of the cloned DNA indicates the presence of four copies of a direct repeat less than 1 kb in size. The map is also consistent with the presence in the chromosome of sequences for mercury resistance (mer A mer B) and for tetracycline-resistance plasmid pT181.

Tn554 inserts in methicillin-resistant Staphylococcus aureus from Australia and England: comparison with an American methicillin-resistant group

“…To explore further the possibility of a physical relationship between Tn554 inserts and the rnec complex of resistance genes, we have characterized the Tn554 elements carried by the Australian and London Mc' groups. A variant (ANS62) of the Australian Mc' strain ANS46 that had become Mcs because of deletion of part of its mec region was especially informative (Matthews et al, 1987). The Australian and London Mc' groups were found to resemble each other and to differ from the New Jersey group by harbouring three Tn5.54 inserts, all of which had distinctive transposon-chromosomal junction sequences.…”

“…The Australian and London Mcr isolates were known to resemble the New Jersey group in having chromosomal M L S determinant(s) (Townsend et al, 1983(Townsend et al, , 1987Matthews et al, 1987). To explore further the possibility of a physical relationship between Tn554 inserts and the rnec complex of resistance genes, we have characterized the Tn554 elements carried by the Australian and London Mc' groups.…”

“…Earlier studies had shown that Mcr S . aureus isolates contain chromosomal DNA, amounting to 30 kb or more, which is also absent from Mcs strains (Beck et al, 1986;Matsuhashi et al, 1986;Matthews et al, 1987). This DNA contains the Mc' determinant mec and determinants for various other resistances (Skinner el al., 1988;Skurray et al, 1988;Ubukata et al, 1989).…”

Spread of the Brazilian epidemic clone of a multiresistant MRSA in two cities in Argentina

“…Chromosomal DNA from all MRSA isolates was digested with Cla I and, after Southern blotting, the membranes were hybridised with a specific mecA probe, a 1250-bp Pst I-Xba I fragment of the mecA gene cloned into pTZ219 [15].…”

Molecular Epidemiology of Methicillin-Resistant Staphylococcus aureus in Finland

“…The PBP2a is inducible and coded by the mecA gene, which is part of an additional DNA region, staphylococcal casette chromosome mec (SCCmec), found in methicillin-resistant strains, but not in methicillin-susceptible strains (14,128,167). SCCmec is always located in the same region in the S. aureus chromosome, between spa and purA (137,209).…”