Project description:Understanding constraints which shape antibiotic resistance is key for predicting and controlling drug resistance. Here, we performed high-throughput laboratory evolution of Escherichia coli. The transcriptome, resistance, and genomic profiles for the evolved strains in 48 environments were quantitatively analyzed. By analyzing the quantitative datasets through interpretable machine learning techniques, the emergence of low dimensional phenotypic states within the 192 strains was observed. Further analysis revealed the underlying biological processes responsible for the distinct states. We also report a novel constraint which leads to decelerated evolution. These findings bridge the genotypic, gene expression, and drug resistance space, and lead to a comprehensive understanding of constraints for antibiotic resistance.
Project description:We used RNA-seq to profile E. coli K-12 MG1655 strains subjected to adaptive laboratory evolution after chorismate synthase knockouts. Either isochorismate synthase (menF) or isochorismate synthase AND chorismate lyase (ubiC) was deleted from a strain of E. coli K-12 MG1655 that had already been previously adapted for growth on glucose minimal media. RNA-seq profiles of the original glucose-adapted strain, the 2 deletion strains, and 4 laboratory-evolved strains from each deletion are included in duplicate. ubiC catalyzes the first committed step of ubiquinone synthesis, an important molecule for the electron transport chain. Thus, these experiments allowed assessment of cellular adaptations to restore energy metabolism capability.
Project description:We conducted whole genome sequencing on eight evolved E. coli strains (S1–S8) and the parental wild-type (WT) strain to identify mutations arising from ofloxacin treatments. These strains (S1-S8), generated through fluoroquinolone-mediated adaptive laboratory evolution (ALE), exhibited varying levels of tolerance and resistance. The ALE experiment involved intermittent antibiotic treatments of eight independent cultures over 22 days. The untreated WT strain served as a baseline to pinpoint mutations in the evolved strains.
Project description:Laboratory adaptive evolution experiments were conducted using serial passage of E. coli in M9 minimal medium supplemented with either 2 g/L of lactate for 60 days or 2 g/L of glycerol for 44 days. 7 parallel evolution strains were generated for growth on lactate and 7 parallel evolution strains were generated for growth on glycerol. Affymetrix arrays were used to study the time-course change in gene expression from unevolved E. coli (day 0) to a midpoint evolved strain (day 20) and evolutionary endpoints
Project description:Escherichia coli is an important human pathogen, among others a cause of severe diarrhea diseases and urinary tract infections. The ability to distinguish different pathogenic E. coli subspecies is crucial for correct treatment of the infection. Characterization and quantification of clinical isolates proteomes can provide details of the organisms’ metabolism and specific virulence factors. We performed a systematic quantitative proteomic analysis on a representative selection of 16 pathogenic and 2 commensal E. coli strains, together with 5 pathogenic Shigella strains. The analysis yielded a dataset of more than 4 thousand proteins, with an average of 2 thousand proteins per strain and 980 proteins common to all strains. Statistical comparison of label-free quantitative levels of 750 proteins, which were quantified in all strains, revealed that levels of a majority of the shared proteins vary substantially among specific strains. Theses quantitative protein profiles clearly distinguished E. coli strains from Shigella and largely separated commensal E. coli strains from intestinal and extraintestinal E. coli isolates.
Project description:E. coli frequently encounters oxidative stress both in its natural environment or in industrial biotechnology. Elucidating the mechanisms behind tolerance to oxidative stress would be beneficial for understanding pathogenesis as well as improving production strain fitness. We make use of adaptive laboratory evolution to develop two strains of E. coli which exhibit 500% increased tolerance to paraquat stress compared to wild type. Evolved strains tolerate oxidative stress by reduction of flux through TCA, dyregulation of iron-uptake genes, and up-regulation of cell motility or iron-sulfur cluster repair genes.
Project description:Acetate is a cost-effective and sustainable carbon source that, despite its potential, remains underutilized. This study employed biosensor-assisted adaptive laboratory evolution (ALE) to enhance itaconic acid production and acetate metabolism in Escherichia coli. The evolved E. coli W strains exhibited 65% increase in itaconic acid production and 71% increase in growth rate, and 45% increase in itaconic acid yield. A common 31-kb genomic deletion was identified in the evolved strains, with two genes, ecw_m2276 and ecw_m2277, driving the observed phenotypic changes. The evolved strains exhibited an intensified stringent response, which enhanced the acetate-utilizing pathway and resulted in over a 5,000% increase in the expression of the glyoxylate shunt, thereby boosting microbial growth. Overexpression of relA further replicated these enhanced phenotypes. Our findings highlight not only significant physiological improvements but also present a novel strategy for enhancing microbial growth and bioproduction from acetate, offering valuable insights for industrial biotechnology applications.
Project description:C. glutamicum strains adapted to higher growth temperatures were obtained through an adaptive laboratory evolution experiment. To elucidate molecular basis for thermotolerance acquired by the evolved strains, we examined transcriptional responses of the evolved and parental strains to thermal stress using microarray technology.
Project description:Laboratory adaptive evolution experiments were conducted using serial passage of E. coli in M9 minimal medium supplemented with either 2 g/L of lactate for 60 days or 2 g/L of glycerol for 44 days. 7 parallel evolution strains were generated for growth on lactate and 7 parallel evolution strains were generated for growth on glycerol. Affymetrix arrays were used to study the time-course change in gene expression from unevolved E. coli (day 0) to a midpoint evolved strain (day 20) and evolutionary endpoints Biological replicate arrays were conducted for each of the time points tested for the different evolution strains