Role of histone-like protein H-NS in multidrug resistance of Escherichia coli. - Related Documents




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64201.0000Role of histone-like protein H-NS in multidrug resistance of Escherichia coli. The histone-like protein H-NS is a major component of the bacterial nucleoid and plays a crucial role in global gene regulation of enteric bacteria. It is known that the expression of a variety of genes is repressed by H-NS, and mutations in hns result in various phenotypes, but the role of H-NS in the drug resistance of Escherichia coli has not been known. Here we present data showing that H-NS contributes to multidrug resistance by regulating the expression of multidrug exporter genes. Deletion of the hns gene from the DeltaacrAB mutant increased levels of resistance against antibiotics, antiseptics, dyes, and detergents. Decreased accumulation of ethidium bromide and rhodamine 6G in the hns mutant compared to that in the parental strain was observed, suggesting the increased expression of some drug exporter(s) in this mutant. The increased drug resistance and decreased drug accumulation caused by the hns deletion were completely suppressed by deletion of the multifunctional outer membrane channel gene tolC. At least eight drug exporter systems require TolC for their functions. Among these, increased expression of acrEF, mdtEF, and emrKY was observed in the Deltahns strain by quantitative real-time reverse transcription-PCR analysis. The Deltahns-mediated multidrug resistance pattern is quite similar to that caused by overproduction of the AcrEF exporter. Deletion of the acrEF gene greatly suppressed the level of Deltahns-mediated multidrug resistance. However, this strain still retained resistance to some compounds. The remainder of the multidrug resistance pattern was similar to that conferred by overproduction of the MdtEF exporter. Double deletion of the mdtEF and acrEF genes completely suppressed Deltahns-mediated multidrug resistance, indicating that Deltahns-mediated multidrug resistance is due to derepression of the acrEF and mdtEF drug exporter genes.200414973023
64310.9997Effect of overexpression of small non-coding DsrA RNA on multidrug efflux in Escherichia coli. OBJECTIVES: Several putative and proven drug efflux pumps are present in Escherichia coli. Because many such efflux pumps have overlapping substrate spectra, it is intriguing that bacteria, with their economically organized genomes, harbour such large sets of multidrug efflux genes. To understand how bacteria utilize these multiple efflux pumps, it is important to elucidate the process of pump expression regulation. The aim of this study was to determine a regulator of the multidrug efflux pump in this organism. METHODS: We screened a genomic library of E. coli for genes that decreased drug susceptibility in this organism. The library was developed from the chromosomal DNA of the MG1655 strain, and then the recombinant plasmids were transformed into an acrB-deleted strain. Transformants were screened for resistance to various antibiotics including oxacillin. RESULTS: We found that the multidrug susceptibilities of the acrB-deleted strain were decreased by the overexpression of small non-coding DsrA RNA as well as by the overexpression of known regulators of multidrug efflux pumps. Plasmids carrying the dsrA gene conferred resistance to oxacillin, cloxacillin, erythromycin, rhodamine 6G and novobiocin. DsrA decreased the accumulation of ethidium bromide in E. coli cells. Furthermore, expression of mdtE was significantly increased by dsrA overexpression, and the decreased multidrug susceptibilities modulated by DsrA were dependent on the MdtEF efflux pump. CONCLUSIONS: These results indicate that DsrA modulates multidrug efflux through activation of genes encoding the MdtEF pump in E. coli.201121088020
77320.9996Mutational Activation of Antibiotic-Resistant Mechanisms in the Absence of Major Drug Efflux Systems of Escherichia coli. Mutations are one of the common means by which bacteria acquire resistance to antibiotics. In an Escherichia coli mutant lacking major antibiotic efflux pumps AcrAB and AcrEF, mutations can activate alternative pathways that lead to increased antibiotic resistance. In this work, we isolated and characterized compensatory mutations of this nature mapping in four different regulatory genes, baeS, crp, hns, and rpoB. The gain-of-function mutations in baeS constitutively activated the BaeSR two-component regulatory system to increase the expression of the MdtABC efflux pump. Missense or insertion mutations in crp and hns caused derepression of an operon coding for the MdtEF efflux pump. Interestingly, despite the dependence of rpoB missense mutations on MdtABC for their antibiotic resistance phenotype, neither the expression of the mdtABCD-baeSR operon nor that of other known antibiotic efflux pumps went up. Instead, the transcriptome sequencing (RNA-seq) data revealed a gene expression profile resembling that of a "stringent" RNA polymerase where protein and DNA biosynthesis pathways were downregulated but pathways to combat various stresses were upregulated. Some of these activated stress pathways are also controlled by the general stress sigma factor RpoS. The data presented here also show that compensatory mutations can act synergistically to further increase antibiotic resistance to a level similar to the efflux pump-proficient parental strain. Together, the findings highlight a remarkable genetic ability of bacteria to circumvent antibiotic assault, even in the absence of a major intrinsic antibiotic resistance mechanism. IMPORTANCE Antibiotic resistance among bacterial pathogens is a chronic health concern. Bacteria possess or acquire various mechanisms of antibiotic resistance, and chief among them is the ability to accumulate beneficial mutations that often alter antibiotic targets. Here, we explored E. coli's ability to amass mutations in a background devoid of a major constitutively expressed efflux pump and identified mutations in several regulatory genes that confer resistance by activating specific or pleiotropic mechanisms.202133972351
77130.9996The multiple antibiotic resistance operon of enteric bacteria controls DNA repair and outer membrane integrity. The multiple antibiotic resistance (mar) operon of Escherichia coli is a paradigm for chromosomally encoded antibiotic resistance in enteric bacteria. The locus is recognised for its ability to modulate efflux pump and porin expression via two encoded transcription factors, MarR and MarA. Here we map binding of these regulators across the E. coli genome and identify an extensive mar regulon. Most notably, MarA activates expression of genes required for DNA repair and lipid trafficking. Consequently, the mar locus reduces quinolone-induced DNA damage and the ability of tetracyclines to traverse the outer membrane. These previously unrecognised mar pathways reside within a core regulon, shared by most enteric bacteria. Hence, we provide a framework for understanding multidrug resistance, mediated by analogous systems, across the Enterobacteriaceae. Transcription factors MarR and MarA confer multidrug resistance in enteric bacteria by modulating efflux pump and porin expression. Here, Sharma et al. show that MarA also upregulates genes required for lipid trafficking and DNA repair, thus reducing antibiotic entry and quinolone-induced DNA damage.201729133912
77440.9996The 2019 Garrod Lecture: MDR efflux in Gram-negative bacteria-how understanding resistance led to a new tool for drug discovery. The AcrAB-TolC MDR efflux system confers intrinsic MDR and overproduction confers clinically relevant resistance to some antibiotics active against Gram-negative bacteria. The system is made up of three components, namely AcrA, AcrB and TolC, otherwise known as the AcrAB-TolC tripartite system. Inactivation or deletion of a gene encoding one of the constituent proteins, or substitution of a single amino acid in the efflux pump component AcrB that results in loss of efflux function, confers increased antibiotic susceptibility. Clinically relevant resistance can be mediated by a mutation in acrB that changes the way AcrB substrates are transported. However, it is more common that resistant clinical and veterinary isolates overproduce the AcrAB-TolC MDR efflux system. This is due to mutations in genes such as marR and ramR that encode repressors of transcription factors (MarA and RamA, respectively) that when produced activate expression of the acrAB and tolC genes thereby increasing efflux. The Lon protease degrades MarA and RamA to return the level of efflux to that of the WT. Furthermore, the levels of AcrAB-TolC are regulated by CsrA. Studies with fluorescent reporters that report levels of acrAB and regulatory factors allowed the development of a new tool for discovering efflux inhibitors. Screens of the Prestwick Chemical Library and a large library from a collaborating pharmaceutical company have generated a number of candidate compounds for further research.201931626705
896850.9995Antibiotic 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
632260.9995A soxRS-constitutive mutation contributing to antibiotic resistance in a clinical isolate of Salmonella enterica (Serovar typhimurium). The soxRS regulon is activated by redox-cycling drugs such as paraquat and by nitric oxide. The >15 genes of this system provide resistance to both oxidants and multiple antibiotics. An association between clinical quinolone resistance and elevated expression of the soxRS regulon has been observed in Escherichia coli, but this association has not been explored for other enteropathogenic bacteria. Here we describe a soxRS-constitutive mutation in a clinical strain of Salmonella enterica (serovar Typhimurium) that arose with the development of resistance to quinolones during treatment. The elevated quinolone resistance in this strain derived from a point mutation in the soxR gene and could be suppressed in trans by multicopy wild-type soxRS. Multiple-antibiotic resistance was also transferred to a laboratory strain of S. enterica by introducing the cloned mutant soxR gene from the clinical strain. The results show that constitutive expression of soxRS can contribute to antibiotic resistance in clinically relevant S. enterica.200111120941
894370.9995Effects of indole on drug resistance and virulence of Salmonella enterica serovar Typhimurium revealed by genome-wide analyses. BACKGROUND: Many Gram-positive and Gram-negative bacteria produce large quantities of indole as an intercellular signal in microbial communities. Indole demonstrated to affect gene expression in Escherichia coli as an intra-species signaling molecule. In contrast to E. coli, Salmonella does not produce indole because it does not harbor tnaA, which encodes the enzyme responsible for tryptophan metabolism. Our previous study demonstrated that E. coli-conditioned medium and indole induce expression of the AcrAB multidrug efflux pump in Salmonella enterica serovar Typhimurium for inter-species communication; however, the global effect of indole on genes in Salmonella remains unknown. RESULTS: To understand the complete picture of genes regulated by indole, we performed DNA microarray analysis of genes in the S. enterica serovar Typhimurium strain ATCC 14028s affected by indole. Predicted Salmonella phenotypes affected by indole based on the microarray data were also examined in this study. Indole induced expression of genes related to efflux-mediated multidrug resistance, including ramA and acrAB, and repressed those related to host cell invasion encoded in the Salmonella pathogenicity island 1, and flagella production. Reduction of invasive activity and motility of Salmonella by indole was also observed phenotypically. CONCLUSION: Our results suggest that indole is an important signaling molecule for inter-species communication to control drug resistance and virulence of S. enterica.201222632036
64480.9995The MarR repressor of the multiple antibiotic resistance (mar) operon in Escherichia coli: prototypic member of a family of bacterial regulatory proteins involved in sensing phenolic compounds. BACKGROUND: The marR gene of Escherichia coli encodes a repressor of the marRAB operon, a regulatory locus controlling multiple antibiotic resistance in this organism. Inactivation of marR results in increased expression of marA, which acts at several target genes in the cell leading to reduced antibiotic accumulation. Exposure of E. coli to sodium salicylate (SAL) induces marRAB operon transcription and antibiotic resistance. The mechanism by which SAL antagonizes MarR repressor activity is unclear. MATERIALS AND METHODS: Recombinant plasmid libraries were introduced into a reporter strain designed to identify cloned genes encoding MarR repressor activity. Computer analysis of sequence databases was also used to search for proteins related to MarR. RESULTS: A second E. coli gene, MprA, that exhibits MarR repressor activity was identified. Subsequent database searching revealed a family of 10 proteins from a variety of bacteria that share significant amino acid sequence similarity to MarR and MprA. At least four of these proteins are transcriptional repressors whose activity is antagonized by SAL or by phenolic agents structurally related to SAL. CONCLUSIONS: The MarR family is identified as a group of regulatory factors whose activity is modulated in response to environmental signals in the form of phenolic compounds. Many of these agents are plant derived. Some of the MarR homologs appear more likely to control systems expressed in animal hosts, suggesting that phenolic sensing by bacteria is important in a variety of environments and in the regulation of numerous processes.19958521301
77690.9995Exploring functional interplay amongst Escherichia coli efflux pumps. Bacterial efflux pumps exhibit functional interplay that can translate to additive or multiplicative effects on resistance to antimicrobial compounds. In diderm bacteria, two different efflux pump structural types - single-component inner membrane efflux pumps and cell envelope-spanning multicomponent systems - cooperatively export antimicrobials with cytoplasmic targets from the cell. Harnessing our recently developed efflux platform, which is built upon an extensively efflux-deficient strain of Escherichia coli, here we explore interplay amongst a panel of diverse E. coli efflux pumps. Specifically, we assessed the effect of simultaneously expressing two efflux pump-encoding genes on drug resistance, including single-component inner membrane efflux pumps (MdfA, MdtK and EmrE), tripartite complexes (AcrAB, AcrAD, MdtEF and AcrEF), and the acquired TetA(C) tetracycline resistance pump. Overall, the expression of two efflux pump-encoding genes from the same structural type did not enhance resistance levels regardless of the antimicrobial compound or efflux pump under investigation. In contrast, a combination of the tripartite efflux systems with single-component pumps sharing common substrates provided multiplicative increases to antimicrobial resistance levels. In some instances, resistance was increased beyond the product of resistance provided by the two pumps individually. In summary, the developed efflux platform enables the isolation of efflux pump function, facilitating the identification of interactions between efflux pumps.202236318669
8929100.9995Interplay 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
784110.9995Regulation of the AcrAB-TolC efflux pump in Enterobacteriaceae. Bacterial multidrug efflux systems are a major mechanism of antimicrobial resistance and are fundamental to the physiology of Gram-negative bacteria. The resistance-nodulation-division (RND) family of efflux pumps is the most clinically significant, as it is associated with multidrug resistance. Expression of efflux systems is subject to multiple levels of regulation, involving local and global transcriptional regulation as well as post-transcriptional and post-translational regulation. The best-characterised RND system is AcrAB-TolC, which is present in Enterobacteriaceae. This review describes the current knowledge and new data about the regulation of the acrAB and tolC genes in Escherichia coli and Salmonella enterica.201829128373
8942120.9995Indole-Induced Activities of β-Lactamase and Efflux Pump Confer Ampicillin Resistance in Pseudomonas putida KT2440. Indole, which is widespread in microbial communities, has received attention because of its effects on bacterial physiology. Pseudomonas putida and Pseudomonas aeruginosa can acquire ampicillin (Amp) resistance during growth on indole-Amp agar. Transcriptome, mutant, and inhibitor studies have suggested that Amp resistance induced by indole can be attributed to increased gene expression of ttgAB encoding two genes of RND-type multidrug efflux operons and an ampC encoding β-lactamase. Expression, enzyme activities, and mutational analyses indicated that AmpC β-lactamase is important for acquiring Amp resistance of P. putida in the presence of indole. Here, we show, for the first time, that volatile indole increased Amp-resistant cells. Consistent with results of the volatile indole assay, a low concentration of indole in liquid culture promoted growth initially, but led to mutagenesis after indole was depleted, which could not be observed at high indole concentrations. Interestingly, ttgAB and ampC gene expression levels correlate with the concentration of indole, which might explain the low number of Amp-mutated cells in high indole concentrations. The expression levels of genes involved in mutagenesis, namely rpoS, recA, and mutS, were also modulated by indole. Our data indicates that indole reduces Amp-induced heterogeneity by promoting expression of TtgABC or MexAB-OprM efflux pumps and the indole-induced β-lactamase in P. putida and P. aeruginosa.201728352264
4705130.9995Upregulation of outer membrane porin gene ompC contributed to enhancement of azithromycin susceptibility in multidrug-resistant Escherichia coli. The outer membrane (OM) in gram-negative bacteria contains proteins that regulate the passive or active uptake of small molecules for growth and cell function, as well as mediate the emergence of antibiotic resistance. This study aims to explore the potential mechanisms for restoring bacteria to azithromycin susceptibility based on transcriptome analysis of bacterial membrane-related genes. Transcriptome sequencing was performed by treating multidrug-resistant Escherichia coli T28R with azithromycin or in combination with colistin and confirmed by reverse transcription-quantitative PCR (RT-qPCR). Azithromycin enzyme-linked immunosorbent assay (ELISA) test, ompC gene overexpression, and molecular docking were utilized to conduct the confirmatory research of the potential mechanisms. We found that colistin combined with azithromycin led to 48 differentially expressed genes, compared to azithromycin alone, such as downregulation of tolA, eptB, lpxP, and opgE and upregulation of ompC gene. Interestingly, the addition of colistin to azithromycin differentially downregulated the mph(A) gene mediating azithromycin resistance, facilitating the intracellular accumulation of azithromycin. Also, overexpression of the ompC elevated azithromycin susceptibility, and colistin contributed to further suppression of the Mph(A) activity in the presence of azithromycin. These findings suggested that colistin firstly enhanced the permeability of bacterial OM, causing intracellular drug accumulation, and then had a repressive effect on the Mph(A) activity along with azithromycin. Our study provides a novel perspective that the improvement of azithromycin susceptibility is related not only to the downregulation of the mph(A) gene and conformational remodeling of the Mph(A) protein but also the upregulation of the membrane porin gene ompC.IMPORTANCEUsually, active efflux via efflux pumps is an important mechanism of antimicrobial resistance, such as the AcrAB-TolC complex and MdtEF. Also, bacterial porins exhibited a substantial fraction of the total number of outer membrane proteins in Enterobacteriaceae, which are involved in mediating the development of the resistance. We found that the upregulation or overexpression of the ompC gene contributed to the enhancement of resistant bacteria to azithromycin susceptibility, probably due to the augment of drug uptakes caused and the opportunity of Mph(A) function suppressed by azithromycin with colistin. Under the combination of colistin and azithromycin treatment, OmpC exhibited an increased selectivity for cationic molecules and played a key role in the restoral of the antibiotic susceptibility. Investigations on the regulation of porin expression that mediated drug resistance would be important in clinical isolates treated with antibiotics.202438441474
4707140.9995Comparative transcriptome analyses of magainin I-susceptible and -resistant Escherichia coli strains. Antimicrobial peptides (AMPs) have attracted considerable attention because of their multiple and complex mechanisms of action toward resistant bacteria. However, reports have increasingly highlighted how bacteria can escape AMP administration. Here, the molecular mechanisms involved in Escherichia coli resistance to magainin I were investigated through comparative transcriptomics. Sub-inhibitory concentrations of magainin I were used to generate four experimental groups, including magainin I-susceptible E. coli, in the absence (C) and presence of magainin I (CM); and magainin I-resistant E. coli in the absence (R) and presence of magainin I (RM). The total RNA from each sample was extracted; cDNA libraries were constructed and further submitted for Illumina MiSeq sequencing. After RNA-seq data pre-processing and functional annotation, a total of 103 differentially expressed genes (DEGs) were identified, mainly related to bacterial metabolism. Moreover, down-regulation of cell motility and chaperone-related genes was observed in CM and RM, whereas cell communication, acid tolerance and multidrug efflux pump genes (ABC transporter, major facilitator and resistance-nodulation cell division superfamilies) were up-regulated in these same groups. DEGs from the C and R groups are related to basal levels of expression of homeostasis-related genes compared to CM and RM, suggesting that the presence of magainin I is required to change the transcriptomics panel in both C and R E. coli strains. These findings show the complexity of E. coli resistance to magainin I through the rearrangement of several metabolic pathways involved in bacterial physiology and drug response, also providing information on the development of novel antimicrobial strategies targeting resistance-related transcripts and proteins herein described.201830277857
8897150.9995Clinically 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
645160.9995Activation of cryptic aminoglycoside resistance in Salmonella enterica. Aminoglycoside resistance in bacteria can be acquired by several mechanisms, including drug modification, target alteration, reduced uptake and increased efflux. Here we demonstrate that increased resistance to the aminoglycosides streptomycin and spectinomycin in Salmonella enterica can be conferred by increased expression of an aminoglycoside adenyl transferase encoded by the cryptic, chromosomally located aadA gene. During growth in rich medium the wild-type strain was susceptible but mutations that impaired electron transport and conferred a small colony variant (SCV) phenotype or growth in glucose/glycerol minimal media resulted in activation of the aadA gene and aminoglycoside resistance. Expression of the aadA gene was positively regulated by the stringent response regulator guanosine penta/tetraphosphate ((p)ppGpp). SCV mutants carrying stop codon mutations in the hemA and ubiA genes showed a streptomycin pseudo-dependent phenotype, where growth was stimulated by streptomycin. Our data suggest that this phenotype is due to streptomycin-induced readthrough of the stop codons, a resulting increase in HemA/UbiA levels and improved electron transport and growth. Our results demonstrate that environmental and mutational activation of a cryptic resistance gene can confer clinically significant resistance and that a streptomycin-pseudo-dependent phenotype can be generated via a novel mechanism that does not involve the classical rpsL mutations.201121507083
6218170.9995An allele of an ancestral transcription factor dependent on a horizontally acquired gene product. Changes in gene regulatory circuits often give rise to phenotypic differences among closely related organisms. In bacteria, these changes can result from alterations in the ancestral genome and/or be brought about by genes acquired by horizontal transfer. Here, we identify an allele of the ancestral transcription factor PmrA that requires the horizontally acquired pmrD gene product to promote gene expression. We determined that a single amino acid difference between the PmrA proteins from the human adapted Salmonella enterica serovar Paratyphi B and the broad host range S. enterica serovar Typhimurium rendered transcription of PmrA-activated genes dependent on the PmrD protein in the former but not the latter serovar. Bacteria harboring the serovar Typhimurium allele exhibited polymyxin B resistance under PmrA- or under PmrA- and PmrD-inducing conditions. By contrast, isogenic strains with the serovar Paratyphi B allele displayed PmrA-regulated polymyxin B resistance only when experiencing activating conditions for both PmrA and PmrD. We establish that the two PmrA orthologs display quantitative differences in several biochemical properties. Strains harboring the serovar Paratyphi B allele showed enhanced biofilm formation, a property that might promote serovar Paratyphi B's chronic infection of the gallbladder. Our findings illustrate how subtle differences in ancestral genes can impact the ability of horizontally acquired genes to confer new properties.201223300460
8887180.9995RpoE is a Putative Antibiotic Resistance Regulator of Salmonella enteric Serovar Typhi. Bacterial antimicrobial resistance has been associated with the up regulation of genes encoding efflux pumps and the down regulation of genes encoding outer membrane proteins (OMPs). Gene expression in bacteria is primarily initiated by sigma factors (σ factors) such as RpoE, which plays an important role in responding to many environmental stresses. Here, we report the first observation that RpoE serves as an antibiotic resistance regulator in Salmonella enteric serovar Typhi (S. Typhi). In this study, we found that the rpoE mutant (ΔrpoE) of S. Typhi GIFU10007 has elevated resistance to several antimicrobial agents, including β-lactams, quinolones, and aminoglycosides. Genomic DNA microarray analysis was used to investigate the differential gene expression profiles between a wild type and rpoE mutant in response to ampicillin. The results showed that a total of 57 genes displayed differential expression (two-fold increase or decrease) in ΔrpoE versus the wild-type strain. The expressions of two outer membrane protein genes, ompF and ompC, were significantly down-regulated in ΔrpoE (six and seven-fold lower in comparison to wild-type strain) and RamA, a member of the efflux pump AraC/XylS family, was up-regulated about four-fold in the ΔrpoE. Our results suggest RpoE is a potential antimicrobial regulator in S. Typhi, controlling both the down regulation of the OMP genes and up-regulating the efflux system.201626742769
6324190.9995Genetic and biochemical basis of tetracycline resistance. Properties of several, well characterized, tetracycline resistance determinants were compared. The determinants in Tn1721 and Tn10 (both from Gram-negative bacteria) each contain two genes; one encodes a repressor that regulates both its own transcription and that of a membrane protein that confers resistance by promoting efflux of the drug. Determinants from Gram-positive bacteria also encode efflux proteins, but expression of resistance is probably regulated by translational attenuation. The likely tetracycline binding site (a common dipeptide) in each efflux protein was predicted. The presence of the common binding site is consistent with the ability of an efflux protein originating in Bacillus species to be expressed in Escherichia coli.19863542941