Specific phenotypic, genomic, and fitness evolutionary trajectories toward streptomycin resistance induced by pesticide co-stressors in Escherichia coli

AbstractTo explore how co-occurring non-antibiotic environmental stressors affect evolutionary trajectories toward antibiotic resistance, we exposed susceptible Escherichia coli K-12 populations to environmentally relevant levels of pesticides and streptomycin for 500 generations. The coexposure substantially changed the phenotypic, genotypic, and fitness evolutionary trajectories, resulting in much stronger streptomycin resistance (>15-fold increase) of the populations. Antibiotic target modification mutations in rpsL and rsmG, which emerged and dominated at late stages of evolution, conferred the strong resistance even with less than 1% abundance, while the off-target mutations in nuoG, nuoL, glnE, and yaiW dominated at early stages only led to mild resistance (2.5–6-fold increase). Moreover, the strongly resistant mutants exhibited lower fitness costs even without the selective pressure and had lower minimal selection concentrations than the mildly resistant ones. Removal of the selective pressure did not reverse the strong resistance of coexposed populations at a later evolutionary stage. The findings suggest higher risks of the selection and propagation of strong antibiotic resistance in environments potentially impacted by antibiotics and pesticides.

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Specific phenotypic, genomic, and fitness evolutionary trajectories toward antibiotic resistance induced by pesticide co-stressors in Escherichia coli

10.1101/2021.07.13.452255 ◽

2021 ◽

Author(s):

Yue Xing ◽

Xiaoxi Kang ◽

Siwei Zhang ◽

Yujie Men

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Selective Pressure ◽

Fold Increase ◽

Evolutionary Stage ◽

Fitness Costs ◽

Evolutionary Trajectories ◽

Resistant Mutants ◽

K 12 ◽

Late Stages

To explore how co-occurring non-antibiotic environmental stressors affect evolutionary trajectories toward antibiotic resistance, we exposed susceptible Escherichia coli K-12 populations to environmentally relevant levels of pesticides and streptomycin for 500 generations. The coexposure substantially changed the phenotypic, genotypic, and fitness evolution trajectories, resulting in much stronger streptomycin resistance (>15-fold increase) of the populations. Antibiotic target modification mutations in rpsL and rsmG, which emerged and dominated at late stages of evolution, conferred the strong resistance even with less than 1% abundance, while the off-target mutations in nuoG, nuoL, glnE, and yaiW dominated at early stages only led to mild resistance (2.5 ~ 6-fold increase). Moreover, the strongly resistant mutants exhibited lower fitness costs even without the selective pressure and had lower minimal selection concentrations than the mildly resistant ones. Removal of the selective pressure did not reverse the strong resistance of coexposed populations at a later evolutionary stage. The findings suggest higher risks of the selection and propagation of strong antibiotic resistance in environments potentially impacted by antibiotics and pesticides.

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Potentiation by L-cysteine of the bactericidal effect of hydrogen peroxide in Escherichia coli

Journal of Bacteriology ◽

10.1128/jb.152.1.81-88.1982 ◽

1982 ◽

Vol 152 (1) ◽

pp. 81-88

Author(s):

E H Berglin ◽

M B Edlund ◽

G K Nyberg ◽

J Carlsson

Keyword(s):

Escherichia Coli ◽

Hydrogen Peroxide ◽

Metal Ion ◽

Chelating Agent ◽

Fold Increase ◽

Bactericidal Effect ◽

Anaerobic Conditions ◽

Dna Breakage ◽

E Coli ◽

K 12

Under anaerobic conditions an exponentially growing culture of Escherichia coli K-12 was exposed to hydrogen peroxide in the presence of various compounds. Hydrogen peroxide (0.1 mM) together with 0.1 mM L-cysteine or L-cystine killed the organisms more rapidly than 10 mM hydrogen peroxide alone. The exposure of E. coli to hydrogen peroxide in the presence of L-cysteine inhibited some of the catalase. This inhibition, however, could not fully explain the 100-fold increase in hydrogen peroxide sensitivity of the organism in the presence of L-cysteine. Of other compounds tested only some thiols potentiated the bactericidal effect of hydrogen peroxide. These thiols were effective, however, only at concentrations significantly higher than 0.1 mM. The effect of L-cysteine and L-cystine could be annihilated by the metal ion chelating agent 2,2'-bipyridyl. DNA breakage in E. coli K-12 was demonstrated under conditions where the organisms were killed by hydrogen peroxide.

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YdgG (TqsA) Controls Biofilm Formation in Escherichia coli K-12 through Autoinducer 2 Transport

Journal of Bacteriology ◽

10.1128/jb.188.2.587-598.2006 ◽

2006 ◽

Vol 188 (2) ◽

pp. 587-598 ◽

Cited By ~ 128

Author(s):

Moshe Herzberg ◽

Ian K. Kaye ◽

Wolfgang Peti ◽

Thomas K. Wood

Keyword(s):

Escherichia Coli ◽

Biofilm Formation ◽

Acid Production ◽

Polysialic Acid ◽

Fold Increase ◽

Uncharacterized Protein ◽

Biofilm Thickness ◽

Autoinducer 2 ◽

Membrane Spanning ◽

K 12

ABSTRACT YdgG is an uncharacterized protein that is induced in Escherichia coli biofilms. Here it is shown that deletion of ydgG decreased extracellular and increased intracellular concentrations of autoinducer 2 (AI-2); hence, YdgG enhances transport of AI-2. Consistent with this hypothesis, deletion of ydgG resulted in a 7,000-fold increase in biofilm thickness and 574-fold increase in biomass in flow cells. Also consistent with the hypothesis, deletion of ydgG increased cell motility by increasing transcription of flagellar genes (genes induced by AI-2). By expressing ydgG in trans, the wild-type phenotypes for extracellular AI-2 activity, motility, and biofilm formation were restored. YdgG is also predicted to be a membrane-spanning protein that is conserved in many bacteria, and it influences resistance to several antimicrobials, including crystal violet and streptomycin (this phenotype could also be complemented). Deletion of ydgG also caused 31% of the bacterial chromosome to be differentially expressed in biofilms, as expected, since AI-2 controls hundreds of genes. YdgG was found to negatively modulate expression of flagellum- and motility-related genes, as well as other known products essential for biofilm formation, including operons for type 1 fimbriae, autotransporter protein Ag43, curli production, colanic acid production, and production of polysaccharide adhesin. Eighty genes not previously related to biofilm formation were also identified, including those that encode transport proteins (yihN and yihP), polysialic acid production (gutM and gutQ), CP4-57 prophage functions (yfjR and alpA), methionine biosynthesis (metR), biotin and thiamine biosynthesis (bioF and thiDFH), anaerobic metabolism (focB, hyfACDR, ttdA, and fumB), and proteins with unknown function (ybfG, yceO, yjhQ, and yjbE); 10 of these genes were verified through mutation to decrease biofilm formation by 40% or more (yfjR, bioF, yccW, yjbE, yceO, ttdA, fumB, yjiP, gutQ, and yihR). Hence, it appears YdgG controls the transport of the quorum-sensing signal AI-2, and so we suggest the gene name tqsA.

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Increased Survival of Antibiotic-Resistant Escherichia coli inside Macrophages

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01632-12 ◽

2012 ◽

Vol 57 (1) ◽

pp. 189-195 ◽

Cited By ~ 33

Author(s):

Migla Miskinyte ◽

Isabel Gordo

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Bacterial Infections ◽

Resistance Mutation ◽

Fitness Cost ◽

Resistance Mutations ◽

Content Type ◽

E Coli ◽

Fitness Effects ◽

K 12

ABSTRACTMutations causing antibiotic resistance usually incur a fitness cost in the absence of antibiotics. The magnitude of such costs is known to vary with the environment. Little is known about the fitness effects of antibiotic resistance mutations when bacteria confront the host's immune system. Here, we study the fitness effects of mutations in therpoB,rpsL, andgyrAgenes, which confer resistance to rifampin, streptomycin, and nalidixic acid, respectively. These antibiotics are frequently used in the treatment of bacterial infections. We measured two important fitness traits—growth rate and survival ability—of 12Escherichia coliK-12 strains, each carrying a single resistance mutation, in the presence of macrophages. Strikingly, we found that 67% of the mutants survived better than the susceptible bacteria in the intracellular niche of the phagocytic cells. In particular, allE. colistreptomycin-resistant mutants exhibited an intracellular advantage. On the other hand, 42% of the mutants incurred a high fitness cost when the bacteria were allowed to divide outside of macrophages. This study shows that single nonsynonymous changes affecting fundamental processes in the cell can contribute to prolonged survival ofE. coliin the context of an infection.

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Aromatic Acid Metabolites of Escherichia coli K-12 Can Induce the marRAB Operon

Journal of Bacteriology ◽

10.1128/jb.00371-10 ◽

2010 ◽

Vol 192 (18) ◽

pp. 4786-4789 ◽

Cited By ~ 29

Author(s):

Lon M. Chubiz ◽

Christopher V. Rao

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Stress Tolerance ◽

Aromatic Acid ◽

Tryptophan Biosynthesis ◽

Metabolic Intermediates ◽

Acid Metabolites ◽

K 12 ◽

Solvent Stress

ABSTRACT MarR is a key regulator of the marRAB operon involved in antibiotic resistance and solvent stress tolerance in Escherichia coli. We show that two metabolic intermediates, 2,3-dihydroxybenzoate and anthranilate, involved in enterobactin and tryptophan biosynthesis, respectively, can activate marRAB transcription. We also found that a third intermediate involved in ubiquinone biosynthesis, 4-hydroxybenzoate, activates marRAB transcription in the absence of TolC. Of the three, however, only 2,3-dihydroxybenzoate directly binds MarR and affects its activity.

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Replication of Colonic Crohn's Disease Mucosal Escherichia coli Isolates within Macrophages and Their Susceptibility to Antibiotics

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00375-07 ◽

2007 ◽

Vol 52 (2) ◽

pp. 427-434 ◽

Cited By ~ 61

Author(s):

Sreedhar Subramanian ◽

Carol L. Roberts ◽

C. Anthony Hart ◽

Helen M. Martin ◽

Steve W. Edwards ◽

...

Keyword(s):

Escherichia Coli ◽

Crohn’S Disease ◽

Crohn's Disease ◽

Colonic Mucosa ◽

Human Monocyte ◽

Fold Increase ◽

Serum Concentrations ◽

Similar Extent ◽

E Coli ◽

K 12

ABSTRACT There is increasing evidence that Escherichia coli organisms are important in Crohn's disease (CD) pathogenesis. In CD tissue they are found within macrophages, and the adherent-invasive CD ileal E. coli isolate LF82 can replicate inside macrophage phagolysosomes. This study investigates replication and antibiotic susceptibility of CD colonic E. coli isolates inside macrophages. Replication of CD colonic E. coli within J774-A1 murine macrophages and human monocyte-derived macrophages (HMDM) was assessed by culture and lysis after gentamicin killing of noninternalized bacteria and verified by electron microscopy (EM). All seven CD colonic isolates tested replicated within J774-A1 macrophages by 3 h (6.36-fold ± 0.7-fold increase; n = 7 isolates) to a similar extent to CD ileal E. coli LF82 (6.8-fold ± 0.8-fold) but significantly more than control patient isolates (5.2-fold ± 0.25-fold; n = 6; P = 0.006) and E. coli K-12 (1.0-fold ± 0.1-fold; P < 0.0001). Replication of CD E. coli HM605 within HMDM (3.9-fold ± 0.7-fold) exceeded that for K-12 (1.4-fold ± 0.2-fold; P = 0.03). EM showed replicating E. coli within macrophage vacuoles. Killing of HM605 within J774-A1 macrophages following a 3-h incubation with antibiotics at published peak serum concentrations (C max) was as follows: for ciprofloxacin, 99.5% ± 0.2%; rifampin, 85.1% ± 6.6%; tetracycline, 62.8% ± 6.1%; clarithromycin, 62.1% ± 5.6% (all P < 0.0001); sulfamethoxazole, 61.3% ± 7.0% (P = 0.0007); trimethoprim, 56.3% ± 3.4% (P < 0.0001); and azithromycin, 41.0% ± 10.5% (P = 0.03). Ampicillin was not effective against intracellular E. coli. Triple antibiotic combinations were assessed at 10% C max, with ciprofloxacin, tetracycline, and trimethoprim causing 97% ± 0.0% killing versus 86% ± 2.0% for ciprofloxacin alone. Colonic mucosa-associated E. coli, particularly CD isolates, replicate within macrophages. Clinical trials are indicated to assess the efficacy of a combination antibiotic therapy targeting intramacrophage E. coli.

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Core oligosaccharide portion of lipopolysaccharide plays important roles on multiple antibiotic resistance in Escherichia coli

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00341-21 ◽

2021 ◽

Author(s):

Jianli Wang ◽

Wenjian Ma ◽

Yu Fang ◽

Hao Liang ◽

Huiting Yang ◽

...

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Cell Envelope ◽

Gene Clusters ◽

Gram Negative Bacteria ◽

Multiple Antibiotic Resistance ◽

Core Oligosaccharide ◽

Gram Negative ◽

The Core ◽

K 12

Gram-negative bacteria are intrinsically resistant to antibiotics due to the presence of the cell envelope, but mechanisms are still not fully understood. In this study, a series of mutants that lack one or more major components associated with the cell envelope were constructed from Escherichia coli K-12 W3110. WJW02 can only synthesize Kdo 2 -lipid A which lacks the core oligosaccharide portion of lipopolysaccharide. WJW04, WJW07 and WJW08 were constructed from WJW02 by deleting the gene clusters relevant to the biosynthesis of exopolysaccharide, flagella and fimbria, respectively. WJW09, WJW010 and WJW011 cells cannot synthesize exopolysaccharide, flagella and fimbria, respectively. Comparing to the wild type W3110, mutants WJW02, WJW04, WJW07 and WJW08 cells showed decreased resistance to more than 10 different antibacterial drugs, but not the mutants WJW09, WJW010 and WJW011. This indicates that the core oligosaccharide portion of lipopolysaccharide plays important roles on multiple antibiotic resistance in E. coli and the 1 st heptose in core oligosaccharide portion is critical. Furthermore, the removal of the core oligosaccharide of LPS leads to influences on cell wall morphology, cell phenotypes, porins, efflux systems, and the respond behaviors to antibiotic stimulation. The results demonstrated the important role of lipopolysaccharide on the antibiotic resistance of Gram-negative bacteria.

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The Conjugation Window in an Escherichia coli K-12 Strain with an IncFII Plasmid

Applied and Environmental Microbiology ◽

10.1128/aem.00948-20 ◽

2020 ◽

Vol 86 (17) ◽

Cited By ~ 1

Author(s):

Brendan Headd ◽

Scott A. Bradford

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Antibiotic Resistance Genes ◽

Time Frame ◽

Environmental Aspect ◽

Content Type ◽

E Coli ◽

Donor Cells ◽

K 12 ◽

Regulatory Controls

ABSTRACT Many studies have examined the role that conjugation plays in disseminating antibiotic resistance genes in bacteria. However, relatively little research has quantitively examined and modeled the dynamics of conjugation under growing and nongrowing conditions beyond a couple of hours. We therefore examined growing and nongrowing cultures of Escherichia coli over a 24-h period to understand the dynamics of bacterial conjugation in the presence and absence of antibiotics with pUUH239.2, an IncFII plasmid containing multiantibiotic- and metal-resistant genes. Our data indicate that conjugation occurs after E. coli cells divide and before they have transitioned to a nongrowing phase. The result is that there is only a small window of opportunity for E. coli to conjugate with pUUH239.2 under both growing and nongrowing conditions. Only a very small percentage of the donor cells likely are capable of even undergoing conjugation, and not all transconjugants can become donor cells due to molecular regulatory controls and not being in the correct growth phase. Once a growing culture enters stationary phase, the number of capable donor cells decreases rapidly and conjugation slows to produce a plateau. Published models did not provide accurate descriptions of conjugation under nongrowing conditions. We present here a modified modeling approach that accurately describes observed conjugation behavior under growing and nongrowing conditions. IMPORTANCE There has been growing interest in horizontal gene transfer of antibiotic resistance plasmids as the antibiotic resistance crisis has worsened over the years. Most studies examining conjugation of bacterial plasmids focus on growing cultures of bacteria for short periods, but in the environment, most bacteria grow episodically and at much lower rates than in the laboratory. We examined conjugation of an IncFII antibiotic resistance plasmid in E. coli under growing and nongrowing conditions to understand the dynamics of conjugation under which the plasmid is transferred. We found that conjugation occurs in a narrow time frame when E. coli is transitioning from a growing to nongrowing phase and that the conjugation plateau develops because of a lack of capable donor cells in growing cultures. From an environmental aspect, our results suggest that episodic growth in nutrient-depleted environments could result in more conjugation than sustained growth in a nutrient rich environment.

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Limited and strain-specific transcriptional and growth responses to acquisition of a multidrug resistance plasmid in genetically diverse Escherichia coli lineages

10.1101/2020.10.23.351718 ◽

2020 ◽

Author(s):

Steven J. Dunn ◽

Laura Carrilero ◽

Michael Brockhurst ◽

Alan McNally

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Multidrug Resistance ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Growth Responses ◽

Fitness Costs ◽

Transcriptional Responses ◽

E Coli ◽

Resistance Plasmid

AbstractMulti-drug resistant (MDR) Escherichia coli are a major global threat to human health, wherein multi-drug resistance is primarily spread by MDR plasmid acquisition. MDR plasmids are not widely distributed across the entire E. coli species, but instead are concentrated in a small number of clones. Here, we test if diverse E. coli strains vary in their ability to acquire and maintain MDR plasmids, and if this relates to their transcriptional response following plasmid acquisition. We used strains from across the diversity of E. coli, including the common MDR lineage ST131, and the IncF plasmid, pLL35, encoding multiple antibiotic resistance genes. Strains varied in their ability to acquire pLL35 by conjugation, but all were able to stably maintain the plasmid. The effects of pLL35 acquisition on cefotaxime resistance and growth also varied among strains, with growth responses ranging from a small decrease to a small increase in growth of the plasmid-carrier relative to the parental strain. Transcriptional responses to pLL35 acquisition were limited in scale and highly strain specific. We observed significant transcriptional responses at the operon or regulon level, possibly due to stress responses or interactions with resident MGEs. Subtle transcriptional responses consistent across all strains were observed affecting functions, such as anaerobic metabolism, previously shown to be under negative frequency dependent selection in MDR E. coli. Overall there was no correlation between the magnitude of the transcriptional and growth responses across strains. Together these data suggest that fitness costs arising from transcriptional disruption are unlikely to act as a barrier to MDR plasmid dissemination in E. coli.ImportancePlasmids play a key role in bacterial evolution by transferring niche adaptive functions between lineages, including driving the spread of antibiotic resistance genes. Fitness costs of plasmid acquisition arising from the disruption of cellular processes could limit the spread of multidrug resistance plasmids. However, the impacts of plasmid acquisition are typically measured in lab-adapted strains rather than in more ecologically relevant natural isolates. Using a clinical multidrug resistance plasmid and a diverse collection of E. coli strains isolated from clinical infections and natural environments, we show that plasmid acquisition had only limited and highly strain-specific effects on bacterial growth and transcription. These findings suggest that fitness costs arising from transcriptional disruption are unlikely to act as a barrier to plasmid transmission in natural populations of E. coli.

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