Open AccessCloning and expression of the penicillinase from a borderline methicillin‐susceptible Staphylococcus aureus strain in Escherichia coli
Author(s) -
Massidda Orietta,
Montanari Maria Pia,
Mingoia Marina,
Varaldo Pietro Emanuele
Publication year - 1994
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1994.tb06899.x
Subject(s) - ampicillin , microbiology and biotechnology , amp resistance , plasmid , beta lactamase , escherichia coli , chloramphenicol , biology , staphylococcus aureus , penicillin , chemistry , gene , antibiotics , bacteria , genetics
Abstract The blaZ gene contained in a single 17.2‐kb β‐lactamase plasmid from a borderline methicillin‐susceptible Staphylococcus aureus strain (a53) has been cloned in Escherichia coli . A Bluescript II derivative in which the ampicillin resistance gene has been replaced with the chloramphenicol resistance gene was used as a multi‐copy vector. One ampicillin‐resistant colony was detected among 31 chloramphenicol‐resistant transformants selected. This E. coli clone harbored a recombinant plasmid (pAH12) containing two different staphylococcal Hind III inserts (7.0 and 5.3 kb), of which only the former hybridized with a blaZ probe. The clone showed an ampicillin MIC of > 1024 μg ml −1 , independently of the inoculum size used, and produced large amounts of β‐lactamase, which hydrolyzed nitrocefin and penicillin G but not methicillin of the β‐lactamase substrate, padac. In contrast, S. aureus a53 hydrolyzed all four substrates. The fact that high levels of staphylococcal penicillinase are unable to cause methicillin hydrolysis confirms that penicillinase hyperproduction is unlikely to be the true mechanism responsible for the borderline phenotype. These results also suggest that the two different β‐lactamases (penicillinase and methicillinase) associated with borderline S. aureus strains have a different genetic origin.