Open AccessExploitation of a β-lactamase reporter gene fusion in the carbapenem antibiotic production operon to study adaptive evolution in Erwinia carotovora
Author(s) -
Steven D. Bowden,
George P. C. Salmond
Publication year - 2006
Publication title -
microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.019
H-Index - 179
eISSN - 1465-2080
pISSN - 1350-0872
DOI - 10.1099/mic.0.28575-0
Subject(s) - erwinia , operon , microbiology and biotechnology , biology , carbapenem , gene , antibiotics , fusion gene , enterobacteriaceae , bacteria , genetics , escherichia coli
Erwinia carotovora subsp. carotovora strain ATTn10 produces the β -lactam antibiotic 1-carbapen-2-em-3-carboxylic acid (carbapenem) by expressing the carABCDEFGH operon. Mutants exhibiting increased carbapenem gene transcription were positively selected using an engineered strain with a functional β -lactamase translational fusion in carH , the last gene of the operon. However, spontaneous ampicillin-resistant mutants were isolated even when transcription of carH : : blaM was blocked by a strongly polar mutation in carE . The mechanism of resistance was shown to be due to cryptic IS 10 elements transposing upstream of carH : : blaM , thereby providing new promoters enabling carH : : blaM transcription. Southern blots showed that IS 10 was present in multicopy in ATTn10. In addition, a Tn 10 genetic remnant was discovered. The results offer insights into the genetic archaeology of strain ATTn10 and highlight the powerful impacts of cryptic IS elements in bacterial adaptive evolution.