The synthesis of total cellular proteins in Escherichia coli K12 was studied in batch culture following exposure of cells to low concentrations of monochlorophenol, pentachlorophenol and cadmium chloride. Changes in protein patterns were identified after pulse-chase labelling of proteins with [35S]methionine and subsequent two-dimensional gel electrophoresis (2D-PAGE). We demonstrated that besides the induction of some stress proteins, also a transient decrease in the rate of synthesis of other proteins occurred. Two of these proteins were identified as OmpF and aspartate transcarbamoylase (ATCase). Their transient repression appeared to be a general response to stress elicited by different pollutants and may therefore be used as a general and sensitive early warning system for pollutant stress.
Plasmid-mediated dissemination of antibiotic resistance among fecal Enterobacteriaceae in natural ecosystems may contribute to the persistence of antibiotic resistance genes in anthropogenically impacted environments. Plasmid transfer frequencies measured under laboratory conditions might lead to overestimation of plasmid transfer potential in natural ecosystems. This study assessed differences in the conjugative transfer of an IncP-1 (pKJK5) plasmid to three natural Escherichia coli strains carrying extended-spectrum beta-lactamases, by filter mating. Matings were performed under optimal laboratory conditions (rich LB medium and 37°C) and environmentally relevant temperatures (25, 15 and 9°C) or nutrient regimes mimicking environmental conditions and limitations (synthetic wastewater and soil extract). Under optimal nutrient conditions and temperature, two recipients yielded high transfer frequencies (5 × 10–1) while the conjugation frequency of the third strain was 1000-fold lower. Decreasing mating temperatures to psychrophilic ranges led to lower transfer frequencies, albeit all three strains conjugated under all the tested temperatures. Low nutritive media caused significant decreases in transconjugants (−3 logs for synthetic wastewater; −6 logs for soil extract), where only one of the strains was able to produce detectable transconjugants. Collectively, this study highlights that despite less-than-optimal conditions, fecal organisms may transfer plasmids in the environment, but the transfer of pKJK5 between microorganisms is limited mainly by low nutrient conditions.
MULTIFILE
Common cloning is often associated with instability of certain classes of DNA. Here we report on IS1 transposition as possible source of such instability. During the cloning of Arabidopsis thaliana gene into commercially available vector maintained in widely used Escherichia coli host the insertion of complete IS1 element into the intron of cloned gene was found. The transposition of the IS1 element was remarkably rapid and is likely to be sequence-specific. The use of E. coli strains that lower the copy number of vector or avoiding the presence of the problematic sequence is a solution to the inadvertent transposition of IS1. The transposition of IS1 is rare but it can occur and might confound functional studies of a plant gene.
