Antibiotic stress, genetic response and altered permeability of E. coli. - Related Documents




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896801.0000Antibiotic stress, genetic response and altered permeability of E. coli. BACKGROUND: Membrane permeability is the first step involved in resistance of bacteria to an antibiotic. The number and activity of efflux pumps and outer membrane proteins that constitute porins play major roles in the definition of intrinsic resistance in Gram-negative bacteria that is altered under antibiotic exposure. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe the genetic regulation of porins and efflux pumps of Escherichia coli during prolonged exposure to increasing concentrations of tetracycline and demonstrate, with the aid of quantitative real-time reverse transcriptase-polymerase chain reaction methodology and western blot detection, the sequence order of genetic expression of regulatory genes, their relationship to each other, and the ensuing increased activity of genes that code for transporter proteins of efflux pumps and down-regulation of porin expression. CONCLUSIONS/SIGNIFICANCE: This study demonstrates that, in addition to the transcriptional regulation of genes coding for membrane proteins, the post-translational regulation of proteins involved in the permeability of Gram-negative bacteria also plays a major role in the physiological adaptation to antibiotic exposure. A model is presented that summarizes events during the physiological adaptation of E. coli to tetracycline exposure.200717426813
896510.9999Resistance characterization and transcriptomic analysis of imipenem-induced drug resistance in Escherichia coli. BACKGROUND: Bacteria can develop resistance to various antibiotics under selective pressure, leading to multifaceted changes in resistance mechanisms. Transcriptomic sequencing allows for the observation of transcriptional level alterations in cells under antibiotic stress. Understanding the bacterial response to such stress is essential for deciphering their strategy against drug-resistant antibiotics and identifying potential targets for antibiotic development. METHODS: This study using wild-type (WT) Escherichia coli (E. coli) discovered that continuous in vitro induction screening for imipenem-resistant strains resulted in bacteria with enhanced biofilm-forming ability and mutations in antibiotic target sites. Transcriptomic sequencing of the resistant bacteria revealed significant changes in carbon and amino acid metabolism, nutrient assimilation, substance transport, nucleotide metabolism, protein biosynthesis, and cell wall biosynthesis. The up-regulated drug efflux genes were disrupted using gene knockout technology. Drug sensitivity tests indicated that drug efflux has a minimal effect on imipenem resistance. RESULTS: This suggests a strategy for E. coli drug resistance involving the reduction of unnecessary substance synthesis and metabolism, coupled with an increase in activities that aid in resisting foreign threats.202439624129
896920.9999Breaching the Barrier: Genome-Wide Investigation into the Role of a Primary Amine in Promoting E. coli Outer-Membrane Passage and Growth Inhibition by Ampicillin. Gram-negative bacteria are problematic for antibiotic development due to the low permeability of their cell envelopes. To rationally design new antibiotics capable of breaching this barrier, more information is required about the specific components of the cell envelope that prevent the passage of compounds with different physiochemical properties. Ampicillin and benzylpenicillin are β-lactam antibiotics with identical chemical structures except for a clever synthetic addition of a primary amine group in ampicillin, which promotes its accumulation in Gram-negatives. Previous work showed that ampicillin is better able to pass through the outer membrane porin OmpF in Escherichia coli compared to benzylpenicillin. It is not known, however, how the primary amine may affect interaction with other cell envelope components. This study applied TraDIS to identify genes that affect E. coli fitness in the presence of equivalent subinhibitory concentrations of ampicillin and benzylpenicillin, with a focus on the cell envelope. Insertions that compromised the outer membrane, particularly the lipopolysaccharide layer, were found to decrease fitness under benzylpenicillin exposure, but had less effect on fitness under ampicillin treatment. These results align with expectations if benzylpenicillin is poorly able to pass through porins. Disruption of genes encoding the AcrAB-TolC efflux system were detrimental to survival under both antibiotics, but particularly ampicillin. Indeed, insertions in these genes and regulators of acrAB-tolC expression were differentially selected under ampicillin treatment to a greater extent than insertions in ompF. These results suggest that maintaining ampicillin efflux may be more significant to E. coli survival than full inhibition of OmpF-mediated uptake. IMPORTANCE Due to the growing antibiotic resistance crisis, there is a critical need to develop new antibiotics, particularly compounds capable of targeting high-priority antibiotic-resistant Gram-negative pathogens. In order to develop new compounds capable of overcoming resistance a greater understanding of how Gram-negative bacteria are able to prevent the uptake and accumulation of many antibiotics is required. This study used a novel genome wide approach to investigate the significance of a primary amine group as a chemical feature that promotes the uptake and accumulation of compounds in the Gram-negative model organism Escherichia coli. The results support previous biochemical observations that the primary amine promotes passage through the outer membrane porin OmpF, but also highlight active efflux as a major resistance factor.202236409154
633030.9999Transcriptomic study of ciprofloxacin resistance in Streptomyces coelicolor A3(2). Soil organisms exhibit resistance to a wide range of antibiotics as they either need to protect themselves from endogenous antibiotics or from those present in their soil environment. The soil could serve as a reservoir for resistance mechanisms that have already emerged or have the potential to emerge in clinically important bacteria. Streptomyces coelicolor, a non-pathogenic soil-dwelling organism, is thus used as a model for the study of intrinsic resistance. Preliminary screening of several compounds showed that S. coelicolor had high intrinsic resistance for the fluoroquinolone group of antibiotics. We subjected the bacteria to sub-inhibitory concentrations of ciprofloxacin and studied the transcriptomic response using microarrays. The data were supported with various biochemical and phenotypic assays. Ciprofloxacin treatment leads to differential expression of many genes with enhanced mRNA expression of its target, DNA gyrase gene. High induction of DNA repair pathways was also observed and many transporters were upregulated. Ciprofloxacin was found to induce ROS formation in a dose dependent manner. Reduction of ROS via anti-oxidants increased the effective MIC of the drug in the bacteria. The regulation of antibiotic resistance in S. coelicolor was studied systematically and contribution of different mechanisms in the development of resistance was assessed. Our data suggest that multiple mechanisms work in coordination to facilitate the cell to combat the stress due to ciprofloxacin.201324100886
896740.9999Distinct transcriptomic response of S. coelicolor to ciprofloxacin in a nutrient-rich environment. With the rising threat of anti-microbial resistance (AMR), there is an urgent need to enhance efficacy of existing antibiotics. Understanding the myriad mechanisms through which bacteria evade these drugs would be of immense value to designing novel strategies against them. Streptomyces coelicolor A3(2) M145 belongs to the actinomyctes species that are responsible for more than two-thirds of antibiotics. This group of bacteria therefore encodes for various mechanisms that can resist both endogenous and non-endogenous antibiotics. In an earlier study, we had studied the transcriptomic response of these bacteria to ciprofloxacin, when cultured in a minimal media. In this work, we investigate why the minimum inhibitory concentration of the drug increases by fourfold when the bacteria are grown in a nutrient-rich media. Through transcriptomic, biochemical, and microscopic studies, we show that S. coelicolor responds to ciprofloxacin in a concentration-dependent manner. While, sub-inhibitory concentration of the drug primarily causes oxidative stress, the inhibitory concentration of ciprofloxacin evokes a more severe genome-wide response in the cell, which ranges from the familiar upregulation of the SOS response and DNA repair pathways to the widespread alterations in the central metabolism pathway to accommodate the increased needs of nucleotides and other precursors. Further, the upregulation of peptidoglycan synthesis genes, along with microscopy images, suggest alterations in the cell morphology to increase fitness of the bacteria during the antibiotic stress. The data also points to the enhanced efflux activity in cells cultured in rich media that contributes significantly towards reducing intracellular drug concentration and thus promotes survival.201830327831
892950.9999Interplay in the selection of fluoroquinolone resistance and bacterial fitness. Fluoroquinolones are antibacterial drugs that inhibit DNA Gyrase and Topoisomerase IV. These essential enzymes facilitate chromosome replication and RNA transcription by regulating chromosome supercoiling. High-level resistance to fluoroquinolones in E. coli requires the accumulation of multiple mutations, including those that alter target genes and genes regulating drug efflux. Previous studies have shown some drug-resistance mutations reduce bacterial fitness, leading to the selection of fitness-compensatory mutations. The impact of fluoroquinolone-resistance on bacterial fitness was analyzed in constructed isogenic strains carrying up to 5 resistance mutations. Some mutations significantly decreased bacterial fitness both in vitro and in vivo. We identified low-fitness triple-mutants where the acquisition of a fourth resistance mutation significantly increased fitness in vitro and in vivo while at the same time dramatically decreasing drug susceptibility. The largest effect occurred with the addition of a parC mutation (Topoisomerase IV) to a low-fitness strain carrying resistance mutations in gyrA (DNA Gyrase) and marR (drug efflux regulation). Increased fitness was accompanied by a significant change in the level of gyrA promoter activity as measured in an assay of DNA supercoiling. In selection and competition experiments made in the absence of drug, parC mutants that improved fitness and reduced susceptibility were selected. These data suggest that natural selection for improved growth in bacteria with low-level resistance to fluoroquinolones could in some cases select for further reductions in drug susceptibility. Thus, increased resistance to fluoroquinolones could be selected even in the absence of further exposure to the drug.200919662169
889760.9999Clinically relevant mutant DNA gyrase alters supercoiling, changes the transcriptome, and confers multidrug resistance. Bacterial DNA is maintained in a supercoiled state controlled by the action of topoisomerases. Alterations in supercoiling affect fundamental cellular processes, including transcription. Here, we show that substitution at position 87 of GyrA of Salmonella influences sensitivity to antibiotics, including nonquinolone drugs, alters global supercoiling, and results in an altered transcriptome with increased expression of stress response pathways. Decreased susceptibility to multiple antibiotics seen with a GyrA Asp87Gly mutant was not a result of increased efflux activity or reduced reactive-oxygen production. These data show that a frequently observed and clinically relevant substitution within GyrA results in altered expression of numerous genes, including those important in bacterial survival of stress, suggesting that GyrA mutants may have a selective advantage under specific conditions. Our findings help contextualize the high rate of quinolone resistance in pathogenic strains of bacteria and may partly explain why such mutant strains are evolutionarily successful. IMPORTANCE: Fluoroquinolones are a powerful group of antibiotics that target bacterial enzymes involved in helping bacteria maintain the conformation of their chromosome. Mutations in the target enzymes allow bacteria to become resistant to these antibiotics, and fluoroquinolone resistance is common. We show here that these mutations also provide protection against a broad range of other antimicrobials by triggering a defensive stress response in the cell. This work suggests that fluoroquinolone resistance mutations may be beneficial under a range of conditions.201323882012
899570.9998Interaction between mutations and regulation of gene expression during development of de novo antibiotic resistance. Bacteria can become resistant not only by horizontal gene transfer or other forms of exchange of genetic information but also by de novo by adaptation at the gene expression level and through DNA mutations. The interrelationship between changes in gene expression and DNA mutations during acquisition of resistance is not well documented. In addition, it is not known whether the DNA mutations leading to resistance always occur in the same order and whether the final result is always identical. The expression of >4,000 genes in Escherichia coli was compared upon adaptation to amoxicillin, tetracycline, and enrofloxacin. During adaptation, known resistance genes were sequenced for mutations that cause resistance. The order of mutations varied within two sets of strains adapted in parallel to amoxicillin and enrofloxacin, respectively, whereas the buildup of resistance was very similar. No specific mutations were related to the rather modest increase in tetracycline resistance. Ribosome-sensed induction and efflux pump activation initially protected the cell through induction of expression and allowed it to survive low levels of antibiotics. Subsequently, mutations were promoted by the stress-induced SOS response that stimulated modulation of genetic instability, and these mutations resulted in resistance to even higher antibiotic concentrations. The initial adaptation at the expression level enabled a subsequent trial and error search for the optimal mutations. The quantitative adjustment of cellular processes at different levels accelerated the acquisition of antibiotic resistance.201424841263
896680.9998Gene expression profile of Campylobacter jejuni in response to macrolide antibiotics. Campylobacter jejuni is a foodborne pathogen that causes gastroenteritis in humans and has developed resistance to various antibiotics. The primary objective of this research was to examine the network of antibiotic resistance in C. jejuni. The study involved the wild and antibiotic-resistant strains placed in the presence and absence of antibiotics to review their gene expression profiles in response to ciprofloxacin via microarray. Differentially expressed genes (DEGs) analysis and Protein-Protein Interaction (PPI) Network studies were performed for these genes. The results showed that the resistance network of C. jejuni is modular, with different genes involved in bacterial motility, capsule synthesis, efflux, and amino acid and sugar synthesis. Antibiotic treatment resulted in the down-regulation of cluster genes related to translation, flagellum formation, and chemotaxis. In contrast, cluster genes involved in homeostasis, capsule formation, and cation efflux were up-regulated. The study also found that macrolide antibiotics inhibit the progression of C. jejuni infection by inactivating topoisomerase enzymes and increasing the activity of epimerase enzymes, trying to compensate for the effect of DNA twisting. Then, the bacterium limits the movement to conserve energy. Identifying the antibiotic resistance network in C. jejuni can aid in developing drugs to combat these bacteria. Genes involved in cell division, capsule formation, and substance transport may be potential targets for inhibitory drugs. Future research must be directed toward comprehending the underlying mechanisms contributing to the modularity of antibiotic resistance and developing strategies to disrupt and mitigate the growing threat of antibiotic resistance effectively.202438393387
896490.9998Analysis of the Oxidative Stress Regulon Identifies soxS as a Genetic Target for Resistance Reversal in Multidrug-Resistant Klebsiella pneumoniae. In bacteria, the defense system deployed to counter oxidative stress is orchestrated by three transcriptional factors, SoxS, SoxR, and OxyR. Although the regulon that these factors control is known in many bacteria, similar data are not available for Klebsiella pneumoniae. To address this data gap, oxidative stress was artificially induced in K. pneumoniae MGH78578 using paraquat and the corresponding oxidative stress regulon recorded using transcriptome sequencing (RNA-seq). The soxS gene was significantly induced during oxidative stress, and a knockout mutant was constructed to explore its functionality. The wild type and mutant were grown in the presence of paraquat and subjected to RNA-seq to elucidate the soxS regulon in K. pneumoniae MGH78578. Genes that are commonly regulated both in the oxidative stress and soxS regulons were identified and denoted as the oxidative SoxS regulon; these included a group of genes specifically regulated by SoxS. Efflux pump-encoding genes and global regulators were identified as part of this regulon. Consequently, the isogenic soxS mutant was found to exhibit a reduction in the minimum bactericidal concentration against tetracycline compared to that of the wild type. Impaired efflux activity, allowing tetracycline to be accumulated in the cytoplasm to bactericidal levels, was further evaluated using a tetraphenylphosphonium (TPP(+)) accumulation assay. The soxS mutant was also susceptible to tetracycline in vivo in a zebrafish embryo model. We conclude that the soxS gene could be considered a genetic target against which an inhibitor could be developed and used in combinatorial therapy to combat infections associated with multidrug-resistant K. pneumoniae. IMPORTANCE Antimicrobial resistance is a global health challenge. Few new antibiotics have been developed for use over the years, and preserving the efficacy of existing compounds is an important step to protect public health. This paper describes a study that examines the effects of exogenously induced oxidative stress on K. pneumoniae and uncovers a target that could be useful to harness as a strategy to mitigate resistance.202134098732
6326100.9998Identification of novel metronidazole-inducible genes in Mycobacterium smegmatis using a customized amplification library. The incidence of antibiotic resistance in pathogenic bacteria is rising. Bacterial resistance may be a natural defense of organisms, or it may result from spontaneous mutations or the acquisition of exogenous resistance genes. We grew spontaneous metronidazole-resistant Mycobacterium smegmatis mutants on solid medium cultures and employed differential expression using a customized amplification library to analyze the global gene profiles of metronidazole-resistant mutants under hypoxic conditions. In total, 66 genes involved in metronidazole resistance were identified and functionally characterized using the gene role category of M. smegmatis. Overall, genes associated with cell wall synthesis, such as methyltransferase and glycosyltransferase, and genes encoding drug transporters were highly expressed. The genes may be involved in the natural drug resistance of mycobacteria by increasing mycobacterial cell wall permeability and the efflux pumps of active drugs. In addition, the genes may play a role in dormancy. The genes identified in this study may lead to a better understanding of the mechanisms of metronidazole resistance during dormancy.200818373646
6333110.9998Outer Membrane Proteins form Specific Patterns in Antibiotic-Resistant Edwardsiella tarda. Outer membrane proteins of Gram-negative bacteria play key roles in antibiotic resistance. However, it is unknown whether outer membrane proteins that respond to antibiotics behave in a specific manner. The present study specifically investigated the differentially expressed outer membrane proteins of an antibiotic-resistant bacterium, Edwardsiella tarda, a Gram-negative pathogen that can lead to unnecessary mass medication of antimicrobials and consequently resistance development in aquaculture and a spectrum of intestinal and extraintestinal diseases in humans. The comparison of a clinically isolated strain to the laboratory derived kanamycin-, tetracycline-, or chloramphenicol-resistant strains identified their respective outer membrane proteins expression patterns, which are distinct to each other. Similarly, the same approach was utilized to profile the patterns in double antibiotic-resistant bacteria. Surprisingly, one pattern is always dominant over the other as to these three antibiotics; the pattern of chloramphenicol is over tetracycline, which is over kanamycin. This type of pattern was also confirmed in clinically relevant multidrug-resistant bacteria. In addition, the presence of plasmid encoding antibiotic-resistant genes also alters the outer membrane protein profile in a similar manner. Our results demonstrate that bacteria adapt the antibiotic stress through the regulation of outer membrane proteins expression. And more importantly, different outer membrane protein profiles were required to cope with different antibiotics. This type of specific pattern provides the rationale for the development of novel strategy to design outer membrane protein arrays to identify diverse multidrug resistance profiles as biomarkers for clinical medication.201728210241
8894120.9998Genome Recombination-Mediated tRNA Up-Regulation Conducts General Antibiotic Resistance of Bacteria at Early Stage. Bacterial antibiotic resistance sets a great challenge to human health. It seems that the bacteria can spontaneously evolve resistance against any antibiotic within a short time without the horizontal transfer of heterologous genes and before accumulating drug-resistant mutations. We have shown that the tRNA-mediated translational regulation counteracts the reactive oxygen species (ROS) in bacteria. In this study, we demonstrated that isolated and subcultured Escherichia coli elevated its tRNAs under antibiotic stress to rapidly provide antibiotic resistance, especially at the early stage, before upregulating the efflux pump and evolving resistance mutations. The DNA recombination system repaired the antibiotic-induced DNA breakage in the genome, causing numerous structural variations. These structural variations are overrepresented near the tRNA genes, which indicated the cause of tRNA up-regulation. Knocking out the recombination system abolished the up-regulation of tRNAs, and coincidently, they could hardly evolve antibiotic resistance in multiple antibiotics, respectively. With these results, we proposed a multi-stage model of bacterial antibiotic resistance in an isolated scenario: the early stage (recombination-tRNA up-regulation-translational regulation); the medium stage (up-regulation of efflux pump); the late stage (resistant mutations). These results also indicated that the bacterial DNA recombination system and tRNA could be targeted to retard the bacterial spontaneous drug resistance.202135126332
772130.9998A Transcriptomic Approach to Identify Novel Drug Efflux Pumps in Bacteria. The core genomes of most bacterial species include a large number of genes encoding putative efflux pumps. The functional roles of most of these pumps are unknown, however, they are often under tight regulatory control and expressed in response to their substrates. Therefore, one way to identify pumps that function in antimicrobial resistance is to examine the transcriptional responses of efflux pump genes to antimicrobial shock. By conducting complete transcriptomic experiments following antimicrobial shock treatments, it may be possible to identify novel drug efflux pumps encoded in bacterial genomes. In this chapter we describe a complete workflow for conducting transcriptomic analyses by RNA sequencing, to determine transcriptional changes in bacteria responding to antimicrobials.201829177833
6328140.9998Inactivation of MarR gene homologs increases susceptibility to antimicrobials in Bacteroides fragilis. Bacteroides fragilis is the strict anaerobic bacteria most commonly found in human infections, and has a high mortality rate. Among other virulence factors, the remarkable ability to acquire resistance to a variety of antimicrobial agents and to tolerate nanomolar concentrations of oxygen explains in part their success in causing infection and colonizing the mucosa. Much attention has been given to genes related to multiple drug resistance derived from plasmids, integrons or transposon, but such genes are also detected in chromosomal systems, like the mar (multiple antibiotic resistance) locus, that confer resistance to a range of drugs. Regulators like MarR, that control expression of the locus mar, also regulate resistance to organic solvents, disinfectants and oxygen reactive species are important players in these events. Strains derived from the parental strain 638R, with mutations in the genes hereby known as marRI (BF638R_3159) and marRII (BF638R_3706) were constructed by gene disruption using a suicide plasmid. Phenotypic response of the mutant strains to hydrogen peroxide, cell survival assay against exposure to oxygen, biofilm formation, resistance to bile salts and resistance to antibiotics was evaluated. The results showed that the mutant strains exhibit statistically significant differences in their response to oxygen stress, but no changes were observed in survival when exposed to bile salts. Biofilm formation was not affected by either gene disruption. Both mutant strains however, became more sensitive to multiple antimicrobial drugs tested. This indicates that as observed in other bacterial species, MarR are an important resistance mechanism in B. fragilis.201828847541
8895150.9998Loss of DNA mismatch repair genes leads to acquisition of antibiotic resistance independent of secondary mutations. Antibiotic resistant bacteria have been a rising clinical concern for decades. Beyond acquisition of alleles conferring resistance, bacteria under stress (e.g., from changing environmental conditions or mutations) can have higher intrinsic resistance to antibiotics than unstressed cells. This concern is expanded for gram-negative bacteria which have a protective outer membrane serving as an additional barrier against harmful molecules such as antibiotics. Here, we report a pathway which increases antibiotic resistance (i.e., minimum inhibitory concentration) in response to inactivation of the DNA Mismatch Repair pathway (MMR). This pathway led to increased intrinsic resistance and was independent of secondary mutations. Specifically, deletion of the DNA mismatch repair genes mutL or mutS caused resistance to various antibiotics spanning different classes, molecular sizes, and mechanisms of action in several different E. coli K-12 MG1655 strains, and in Salmonella enterica serovar Typhimurium LT2. This pathway was independent of the SOS response (severe DNA damage response). However, the patterns of resistance correlated with previously reported increases in MMR mutants in rates of homoeologous recombination, homologous recombination between non-identical DNA strands. Mutations expected to lower rates of recombination in MMR mutants also decreased the resistance to most antibiotics. Finally, we found lysis occurs in MMR mutants and may contribute to resistance to other antibiotics. Our results have demonstrated a novel mechanism that increases antibiotic resistance in direct response to loss of MMR genes, and we propose this resistance involves increased rates of homoeologous recombination and cell lysis. The increased antibiotic resistance of MMR mutants provides a path for these cells to survive in antibiotics long enough to develop more specific resistance mutations and so may contribute to the development of new clinical resistance alleles.202540667202
6334160.9998Epigenetic inheritance based evolution of antibiotic resistance in bacteria. BACKGROUND: The evolution of antibiotic resistance in bacteria is a topic of major medical importance. Evolution is the result of natural selection acting on variant phenotypes. Both the rigid base sequence of DNA and the more plastic expression patterns of the genes present define phenotype. RESULTS: We investigated the evolution of resistant E. coli when exposed to low concentrations of antibiotic. We show that within an isogenic population there are heritable variations in gene expression patterns, providing phenotypic diversity for antibiotic selection to act on. We studied resistance to three different antibiotics, ampicillin, tetracycline and nalidixic acid, which act by inhibiting cell wall synthesis, protein synthesis and DNA synthesis, respectively. In each case survival rates were too high to be accounted for by spontaneous DNA mutation. In addition, resistance levels could be ramped higher by successive exposures to increasing antibiotic concentrations. Furthermore, reversion rates to antibiotic sensitivity were extremely high, generally over 50%, consistent with an epigenetic inheritance mode of resistance. The gene expression patterns of the antibiotic resistant E. coli were characterized with microarrays. Candidate genes, whose altered expression might confer survival, were tested by driving constitutive overexpression and determining antibiotic resistance. Three categories of resistance genes were identified. The endogenous beta-lactamase gene represented a cryptic gene, normally inactive, but when by chance expressed capable of providing potent ampicillin resistance. The glutamate decarboxylase gene, in contrast, is normally expressed, but when overexpressed has the incidental capacity to give an increase in ampicillin resistance. And the DAM methylase gene is capable of regulating the expression of other genes, including multidrug efflux pumps. CONCLUSION: In this report we describe the evolution of antibiotic resistance in bacteria mediated by the epigenetic inheritance of variant gene expression patterns. This provides proof in principle that epigenetic inheritance, as well as DNA mutation, can drive evolution.200818282299
6329170.9998Autoinducer-2 influences tetracycline resistance in Streptococcus suis by regulating the tet(M) gene via transposon Tn916. The concern over increasing resistance to tetracyclines (TCs), such as tetracycline and chlortetracycline, necessitates exploration of new approaches to combating infection in antimicrobial therapy. Given that bacteria use the chemical language of autoinducer 2 (AI-2) signaling molecules in order to communicate and regulate group behaviors, we asked whether the AI-2 signaling influence the tetracyclines antibiotics susceptibility in S. suis. Our present work demonstrated that MIC increased when exogenous AI-2 was added, when compared to the wild type strain. When grown in the presence of sub-MIC of antibiotics, it has been shown that exogenous AI-2 increases growth rate and biofilm formation. These results suggest that the TCs resistance in S. suis could involve a signaling mechanism. Base on the above observations, transcriptomic analyses showed significant differences in the expression of tet(M) of tetracyclines resistance genes, as well as differences in Tn916 transposon related genes transcription, as judged by RT-PCR. Our results provide strong evidence that AI-2 signaling molecules is may involve in TCs antibiotic resistance in S. suis by regulating tet(M) gene via Tn916 transposon. This study may suggest that targeting AI-2 signaling in bacteria could represent an alternative approach in antimicrobial therapy.202031837515
6294180.9998Comparison of Gene Expression Profiles of Uropathogenic Escherichia Coli CFT073 after Prolonged Exposure to Subinhibitory Concentrations of Different Biocides. Biocides are chemical compounds widely used for sterilization and disinfection. The aim of this study was to examine whether exposure to subinhibitory biocide concentrations influenced transcriptional expression of genes that could improve a pathogen's drug resistance or fitness. We used DNA microarrays to investigate the transcriptome of the uropathogenic Escherichia coli strain CFT073 in response to prolonged exposure to subinhibitory concentrations of four biocides: benzalkonium chloride, chlorhexidine, hydrogen peroxide and triclosan. Transcription of a gene involved in polymyxin resistance, arnT, was increased after treatment with benzalkonium chloride. However, pretreatment of the bacteria with this biocide did not result in cross-resistance to polymyxin in vitro. Genes encoding products related to transport formed the functional group that was most affected by biocides, as 110 out of 884 genes in this category displayed altered transcription. Transcripts of genes involved in cysteine uptake, sulfate assimilation, dipeptide transport, as well as cryptic phage genes were also more abundant in response to several biocides. Additionally, we identified groups of genes with transcription changes unique to single biocides that might include potential targets for the biocides. The biocides did not increase the resistance potential of the pathogen to other antimicrobials.201931569631
8957190.9998Transcriptome Profiling Reveals Interplay of Multifaceted Stress Response in Escherichia coli on Exposure to Glutathione and Ciprofloxacin. We have previously reported that supplementation of exogenous glutathione (GSH) promotes ciprofloxacin resistance in Escherichia coli by neutralizing antibiotic-induced oxidative stress and by enhancing the efflux of antibiotic. In the present study, we used a whole-genome microarray as a tool to analyze the system-level transcriptomic changes of E. coli on exposure to GSH and/or ciprofloxacin. The microarray data revealed that GSH supplementation affects redox function, transport, acid shock, and virulence genes of E. coli. The data further highlighted the interplay of multiple underlying stress response pathways (including those associated with the genes mentioned above and DNA damage repair genes) at the core of GSH, offsetting the effect of ciprofloxacin in E. coli. The results of a large-scale validation of the transcriptomic data using reverse transcription-quantitative PCR (RT-qPCR) analysis for 40 different genes were mostly in agreement with the microarray results. The altered growth profiles of 12 different E. coli strains carrying deletions in the specific genes mentioned above with GSH and/or ciprofloxacin supplementation implicate these genes in the GSH-mediated phenotype not only at the molecular level but also at the functional level. We further associated GSH supplementation with increased acid shock survival of E. coli on the basis of our transcriptomic data. Taking the data together, it can be concluded that GSH supplementation influences the expression of genes of multiple stress response pathways apart from its effect(s) at the physiological level to counter the action of ciprofloxacin in E. coli. IMPORTANCE The emergence and spread of multidrug-resistant bacterial strains have serious medical and clinical consequences. In addition, the rate of discovery of new therapeutic antibiotics has been inadequate in last few decades. Fluoroquinolone antibiotics such as ciprofloxacin represent a precious therapeutic resource in the fight against bacterial pathogens. However, these antibiotics have been gradually losing their appeal due to the emergence and buildup of resistance to them. In this report, we shed light on the genome-level expression changes in bacteria with respect to glutathione (GSH) exposure which act as a trigger for fluoroquinolone antibiotic resistance. The knowledge about different bacterial stress response pathways under conditions of exposure to the conditions described above and potential points of cross talk between them could help us in understanding and formulating the conditions under which buildup and spread of antibiotic resistance could be minimized. Our findings are also relevant because GSH-induced genome-level expression changes have not been reported previously for E. coli.201829468195