scholarly journals Regulation of mtrF Expression in Neisseria gonorrhoeae and Its Role in High-Level Antimicrobial Resistance

2005 ◽  
Vol 187 (11) ◽  
pp. 3713-3720 ◽  
Author(s):  
Jason P. Folster ◽  
William M. Shafer
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Sublethal Concentration ◽  
Human Pathogen ◽  
Mucosal Surfaces ◽  
Induction Process ◽  
Inner Membrane Protein ◽  
High Level ◽  
Triton X

ABSTRACT The obligate human pathogen Neisseria gonorrhoeae uses the MtrC-MtrD-MtrE efflux pump to resist structurally diverse hydrophobic antimicrobial agents (HAs), some of which bathe mucosal surfaces that become infected during transmission of gonococci. Constitutive high-level HA resistance occurs by the loss of a repressor (MtrR) that negatively controls transcription of the mtrCDE operon. This high-level HA resistance also requires the product of the mtrF gene, which is located downstream and transcriptionally divergent from mtrCDE. MtrF is a putative inner membrane protein, but its role in HA resistance mediated by the MtrC-MtrD-MtrE efflux pump remains to be determined. High-level HA resistance can also be mediated through an induction process that requires enhanced transcription of mtrCDE when gonococci are grown in the presence of a sublethal concentration of Triton X-100. We now report that inactivation of mtrF results in a significant reduction in the induction of HA resistance and that the expression of mtrF is enhanced when gonococci are grown under inducing conditions. However, no effect was observed on the induction of mtrCDE expression in an MtrF-negative strain. The expression of mtrF was repressed by MtrR, the major repressor of mtrCDE expression. In addition to MtrR, another repressor (MpeR) can downregulate the expression of mtrF. Repression of mtrF by MtrR and MpeR was additive, demonstrating that the repressive effects mediated by these regulators are independent processes.

scholarly journals Molecular pathways to high-level azithromycin resistance in Neisseria gonorrhoeae

2021 ◽  
Author(s):  
J G E Laumen ◽  
S S Manoharan-Basil ◽  
E Verhoeven ◽  
S Abdellati ◽  
I De Baetselier ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Ribosomal Proteins ◽  
Efflux Pump ◽  
23S Rrna ◽  
Rrna Gene ◽  
23S Rrna Gene ◽  
Evolution Of Resistance ◽  
High Level ◽  
Azithromycin Resistance ◽  
Level Resistance

Abstract Background The prevalence of azithromycin resistance in Neisseria gonorrhoeae is increasing in numerous populations worldwide. Objectives To characterize the genetic pathways leading to high-level azithromycin resistance. Methods A customized morbidostat was used to subject two N. gonorrhoeae reference strains (WHO-F and WHO-X) to dynamically sustained azithromycin pressure. We tracked stepwise evolution of resistance by whole genome sequencing. Results Within 26 days, all cultures evolved high-level azithromycin resistance. Typically, the first step towards resistance was found in transitory mutations in genes rplD, rplV and rpmH (encoding the ribosomal proteins L4, L22 and L34 respectively), followed by mutations in the MtrCDE-encoded efflux pump and the 23S rRNA gene. Low- to high-level resistance was associated with mutations in the ribosomal proteins and MtrCDE efflux pump. However, high-level resistance was consistently associated with mutations in the 23S ribosomal RNA, mainly the well-known A2059G and C2611T mutations, but also at position A2058G. Conclusions This study enabled us to track previously reported mutations and identify novel mutations in ribosomal proteins (L4, L22 and L34) that may play a role in the genesis of azithromycin resistance in N. gonorrhoeae.

scholarly journals Overexpression of the MtrC-MtrD-MtrE Efflux Pump Due to an mtrR Mutation Is Required for Chromosomally Mediated Penicillin Resistance in Neisseria gonorrhoeae

2002 ◽  
Vol 184 (20) ◽  
pp. 5619-5624 ◽  
Author(s):  
Wendy L. Veal ◽  
Robert A. Nicholas ◽  
William M. Shafer
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Base Pair ◽  
Efflux Pump ◽  
Penicillin Resistance ◽  
Single Base ◽  
Base Pair Deletion ◽  
Resistance To Penicillin ◽  
Single Base Pair ◽  
High Level ◽  
Level Resistance

ABSTRACT The importance of the mtrCDE-encoded efflux pump in conferring chromosomally mediated penicillin resistance on certain strains of Neisseria gonorrhoeae was determined by using genetic derivatives of penicillin-sensitive strain FA19 bearing defined mutations (mtrR, penA, and penB) donated by a clinical isolate (FA6140) expressing high-level resistance to penicillin and antimicrobial hydrophobic agents (HAs). When introduced into strain FA19 by transformation, a single base pair deletion in the mtrR promoter sequence from strain FA6140 was sufficient to provide high-level resistance to HAs (e.g., erythromycin and Triton X-100) but only a twofold increase in resistance to penicillin. When subsequent mutations in penA and porIB were introduced from strain FA6140 into strain WV30 (FA19 mtrR) by transformation, resistance to penicillin increased incrementally up to a MIC of 1.0 μg/ml. Insertional inactivation of the gene (mtrD) encoding the membrane transporter component of the Mtr efflux pump in these transformant strains and in strain FA6140 decreased the MIC of penicillin by 16-fold. Genetic analyses revealed that mtrR mutations, such as the single base pair deletion in its promoter, are needed for phenotypic expression of penicillin and tetracycline resistance afforded by the penB mutation. As penB represents amino acid substitutions within the third loop of the outer membrane PorIB protein that modulate entry of penicillin and tetracycline, the results presented herein suggest that PorIB and the MtrC-MtrD-MtrE efflux pump act synergistically to confer resistance to these antibiotics.

scholarly journals Cryo-EM Structures of a Gonococcal Multidrug Efflux Pump Illuminate a Mechanism of Drug Recognition and Resistance

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
Meinan Lyu ◽  
Mitchell A. Moseng ◽  
Jennifer L. Reimche ◽  
Concerta L. Holley ◽  
Vijaya Dhulipala ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Drug Resistance ◽  
Neisseria Gonorrhoeae ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Multidrug Efflux ◽  
Multidrug Efflux Pump ◽  
Transmitted Infection ◽  
Content Type ◽  
Cryo Electron Microscopy

ABSTRACT Neisseria gonorrhoeae is an obligate human pathogen and causative agent of the sexually transmitted infection (STI) gonorrhea. The most predominant and clinically important multidrug efflux system in N. gonorrhoeae is the multiple transferrable resistance (Mtr) pump, which mediates resistance to a number of different classes of structurally diverse antimicrobial agents, including clinically used antibiotics (e.g., β-lactams and macrolides), dyes, detergents and host-derived antimicrobials (e.g., cationic antimicrobial peptides and bile salts). Recently, it has been found that gonococci bearing mosaic-like sequences within the mtrD gene can result in amino acid changes that increase the MtrD multidrug efflux pump activity, probably by influencing antimicrobial recognition and/or extrusion to elevate the level of antibiotic resistance. Here, we report drug-bound solution structures of the MtrD multidrug efflux pump carrying a mosaic-like sequence using single-particle cryo-electron microscopy, with the antibiotics bound deeply inside the periplasmic domain of the pump. Through this structural approach coupled with genetic studies, we identify critical amino acids that are important for drug resistance and propose a mechanism for proton translocation. IMPORTANCE Neisseria gonorrhoeae has become a highly antimicrobial-resistant Gram-negative pathogen. Multidrug efflux is a major mechanism that N. gonorrhoeae uses to counteract the action of multiple classes of antibiotics. It appears that gonococci bearing mosaic-like sequences within the gene mtrD, encoding the most predominant and clinically important transporter of any gonococcal multidrug efflux pump, significantly elevate drug resistance and enhance transport function. Here, we report cryo-electron microscopy (EM) structures of N. gonorrhoeae MtrD carrying a mosaic-like sequence that allow us to understand the mechanism of drug recognition. Our work will ultimately inform structure-guided drug design for inhibiting these critical multidrug efflux pumps.

scholarly journals MpeR Regulates themtrEfflux Locus in Neisseria gonorrhoeae and Modulates Antimicrobial Resistance by an Iron-Responsive Mechanism

2012 ◽  
Vol 56 (3) ◽  
pp. 1491-1501 ◽  
Author(s):  
Alexandra Dubon Mercante ◽  
Lydgia Jackson ◽  
Paul J. T. Johnson ◽  
Virginia A. Stringer ◽  
David W. Dyer ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Efflux Pump ◽  
Regulatory System ◽  
Logarithmic Phase ◽  
Free Iron ◽  
Content Type ◽  
Protein Encoding ◽  
Late Logarithmic Phase ◽  
Physiologic Parameters ◽  
Inner Membrane Protein

ABSTRACTPrevious studies have shown that the MpeR transcriptional regulator produced byNeisseria gonorrhoeaerepresses the expression ofmtrF, which encodes a putative inner membrane protein (MtrF). MtrF works as an accessory protein with the Mtr efflux pump, helping gonococci to resist high levels of diverse hydrophobic antimicrobials. Regulation ofmpeRhas been reported to occur by an iron-dependent mechanism involving Fur (ferric uptake regulator). Collectively, these observations suggest the presence of an interconnected regulatory system in gonococci that modulates the expression of efflux pump protein-encoding genes in an iron-responsive manner. Herein, we describe this connection and report that levels of gonococcal resistance to a substrate of themtrCDE-encoded efflux pump can be modulated by MpeR and the availability of free iron. Using microarray analysis, we found that themtrRgene, which encodes a direct repressor (MtrR) ofmtrCDE, is an MpeR-repressed determinant in the late logarithmic phase of growth when free iron levels would be reduced due to bacterial consumption. This repression was enhanced under conditions of iron limitation and resulted in increased expression of themtrCDEefflux pump operon. Furthermore, as judged by DNA-binding analysis, MpeR-mediated repression ofmtrRwas direct. Collectively, our results indicate that both genetic and physiologic parameters (e.g., iron availability) can influence the expression of themtrefflux system and modulate levels of gonococcal susceptibility to efflux pump substrates.

scholarly journals Molecular pathways to high-level azithromycin resistance in Neisseria gonorrhoeae

2020 ◽  
Author(s):  
J.G.E. Laumen ◽  
S.S. Manoharan-Basil ◽  
E Verhoeven ◽  
S Abdellati ◽  
I De Baetselier ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Ribosomal Proteins ◽  
Efflux Pump ◽  
23S Rrna ◽  
Rrna Gene ◽  
23S Rrna Gene ◽  
Evolution Of Resistance ◽  
High Level ◽  
Azithromycin Resistance ◽  
Level Resistance

AbstractObjectivesThe prevalence of azithromycin resistance in Neisseria gonorrhoeae is increasing in numerous populations worldwide. The aim of this study was to characterize the genetic pathways leading to high-level azithromycin resistance.MethodsA customized morbidostat was used to subject two N. gonorrhoeae reference strains (WHO-F and WHO-X) to dynamically sustained azithromycin pressure. We tracked stepwise evolution of resistance by whole genome sequencing.ResultsWithin 26 days, all cultures evolved high-level azithromycin resistance. Typically, the first step towards resistance was found in transitory mutations in genes rplD, rplV and rpmH (encoding the ribosomal proteins L4, L22 and L34 respectively), followed by mutations in the MtrCDE-encoded efflux pump and the 23S rRNA gene. Low-to high-level resistance was associated with mutations in the ribosomal proteins and MtrCDE-encoded efflux pump. However, high-level resistance was consistently associated with mutations in the 23S ribosomal RNA - mainly the well-known A2059G and C2611T mutations, but also at position A2058G.ConclusionsThis study enabled us to track previously reported mutations and identify novel mutations in ribosomal proteins (L4, L22 and L34) that may play a role in the genesis of azithromycin resistance in N. gonorrhoeae.

scholarly journals Inducible, but Not Constitutive, Resistance of Gonococci to Hydrophobic Agents Due to the MtrC-MtrD-MtrE Efflux Pump Requires TonB-ExbB-ExbD Proteins

2002 ◽  
Vol 46 (2) ◽  
pp. 561-565 ◽  
Author(s):  
Corinne Rouquette-Loughlin ◽  
Igor Stojiljkovic ◽  
Tara Hrobowski ◽  
Jacqueline T. Balthazar ◽  
William M. Shafer
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antimicrobial Resistance ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Energy Requirement ◽  
Constitutive Resistance ◽  
Absolute Requirement ◽  
Pump Activity ◽  
Tonb Protein ◽  
Triton X

ABSTRACT The MtrC-MtrD-MtrE efflux pump possessed by Neisseria gonorrhoeae is very similar to the MexA-MexB-OprM efflux pump of Pseudomonas aeruginosa. Because the antimicrobial resistance property afforded by the MexA-MexB-OprM efflux pump also requires the TonB protein, we asked whether a similar requirement exists for the gonococcal efflux pump. Unlike earlier studies with P. aeruginosa, we found that constitutive levels of gonococcal resistance to hydrophobic antimicrobial agents (i.e., Triton X-100 [TX-100]) did not require the TonB, ExbB, or ExbD protein. However, inducible levels of TX-100 resistance in gonococci had an absolute requirement for the TonB-ExbB-ExbD system, suggesting that such resistance in gonococci has an energy requirement above and beyond that required for constitutive pump activity.

scholarly journals Overproduction of the MtrCDE Efflux Pump in Neisseria gonorrhoeae Produces Unexpected Changes in Cellular Transcription Patterns

2014 ◽  
Vol 59 (1) ◽  
pp. 724-726 ◽  
Author(s):  
Elizabeth A. Ohneck ◽  
Maira Goytia ◽  
Corinne E. Rouquette-Loughlin ◽  
Sandeep J. Joseph ◽  
Timothy D. Read ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Efflux Pump ◽  
Transcriptional Profiling ◽  
Regulatory Proteins ◽  
Drug Efflux ◽  
Content Type ◽  
Genes Encoding ◽  
High Level ◽  
Cellular Transcription ◽  
Global Consequence

ABSTRACTThe global consequence of drug efflux gene overexpression in bacteria has not been specifically analyzed because strains showing high-level expression typically have mutations in genes encoding regulatory proteins that control other genes. Results from a transcriptional profiling study performed with a strain ofNeisseria gonorrhoeaethat is capable of high-level transcription of themtrCDEefflux pump operon independently of control by cognate regulatory proteins revealed that its overexpression has ramifications for systems other than drug efflux.

scholarly journals Induction of Multidrug Resistance Mechanism in Escherichia coli Biofilms by Interplay between Tetracycline and Ampicillin Resistance Genes

2009 ◽  
Vol 53 (11) ◽  
pp. 4628-4639 ◽  
Author(s):  
Thithiwat May ◽  
Akinobu Ito ◽  
Satoshi Okabe
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Marker Genes ◽  
Ampicillin Resistance ◽  
High Level ◽  
Biofilm Development ◽  
Level Resistance

ABSTRACT Biofilms gain resistance to various antimicrobial agents, and the presence of antibiotic resistance genes is thought to contribute to a biofilm-mediated antibiotic resistance. Here we showed the interplay between the tetracycline resistance efflux pump TetA(C) and the ampicillin resistance gene (bla TEM-1) in biofilms of Escherichia coli harboring pBR322 in the presence of the mixture of ampicillin and tetracycline. E. coli in the biofilms could obtain the high-level resistance to ampicillin, tetracycline, penicillin, erythromycin, and chloramphenicol during biofilm development and maturation as a result of the interplay between the marker genes on the plasmids, the increase of plasmid copy number, and consequently the induction of the efflux systems on the bacterial chromosome, especially the EmrY/K and EvgA/S pumps. In addition, we characterized the overexpression of the TetA(C) pump that contributed to osmotic stress response and was involved in the induction of capsular colanic acid production, promoting formation of mature biofilms. However, this investigated phenomenon was highly dependent on the addition of the subinhibitory concentrations of antibiotic mixture, and the biofilm resistance behavior was limited to aminoglycoside antibiotics. Thus, marker genes on plasmids played an important role in both resistance of biofilm cells to antibiotics and in formation of mature biofilms, as they could trigger specific chromosomal resistance mechanisms to confer a high-level resistance during biofilm formation.

scholarly journals Decreased Azithromycin Susceptibility ofNeisseria gonorrhoeae Due to mtrRMutations

1999 ◽  
Vol 43 (10) ◽  
pp. 2468-2472 ◽  
Author(s):  
Leticia Zarantonelli ◽  
Graciela Borthagaray ◽  
Eun-Hee Lee ◽  
William M. Shafer
Keyword(s):  
Antimicrobial Agents ◽  
Efflux Pump ◽  
Amino Acid Replacement ◽  
Cross Resistance ◽  
Binding Motif ◽  
Coding Region ◽  
Pump System ◽  
Insertional Inactivation ◽  
Ofneisseria Gonorrhoeae ◽  
Triton X

ABSTRACT Single-dose azithromycin therapy has recently been used in Uruguay for the treatment of uncomplicated gonococcal infections. As part of an active surveillance study to monitor the emergence of antibiotic resistance in gonococcal isolates, we examined the levels of azithromycin susceptibility in 51 consecutive isolates obtained from males with uncomplicated gonococcal urethritis. Isolates with decreased susceptibility to azithromycin (MICs, 0.25 to 0.5 μg/ml) were common, and these isolates often displayed cross-resistance to hydrophobic antimicrobial agents (erythromycin and Triton X-100). Resistance to erythromycin and Triton X-100 is frequently due to overexpression of the mtrCDE-encoded efflux pump mediated by mutations in themtrR gene, which encodes a transcriptional repressor that modulates expression of the mtrCDE operon. Accordingly, we questioned whether clinical isolates that express decreased azithromycin susceptibility harbor mtrR mutations. Promoter mutations that would decrease the level of expression ofmtrR as well as a missense mutation at codon 45 in themtrR-coding region that would result in a radical amino acid replacement within the DNA-binding motif of MtrR were found in these strains. When these mutations were transferred into azithromycin-susceptible strain FA19 by transformation, the susceptibility of gonococci to azithromycin was decreased by nearly 10-fold. The mtrCDE-encoded efflux pump system was responsible for this property since insertional inactivation of themtrC gene resulted in enhanced susceptibility of gonococci to azithromycin. We conclude that the mtrCDE-encoded efflux pump can recognize azithromycin and that the emergence of gonococcal strains with decreased susceptibility to azithromycin can, in part, be explained by mtrR mutations.

scholarly journals In Vitro Susceptibilities of 400 Spanish Isolates ofNeisseria gonorrhoeae to Gemifloxacin and 11 Other Antimicrobial Agents

2000 ◽  
Vol 44 (9) ◽  
pp. 2543-2544 ◽  
Author(s):  
S. Berrón ◽  
J. A. Vázquez ◽  
M. J. Giménez ◽  
L. de la Fuente ◽  
L. Aguilar
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Antimicrobial Agents ◽  
Vitro Activity ◽  
Penicillin Resistance ◽  
In Vitro Activity ◽  
Ofneisseria Gonorrhoeae ◽  
High Level

ABSTRACT The in vitro activity of gemifloxacin versus those of 11 other antimicrobial agents against 400 strains of Neisseria gonorrhoeae was determined by microdilution with supplemented GC agar. A total of 37.5% of the strains were β-lactamase positive. A total of 70 and 6.4% of the β-lactamase-negative strains exhibited intermediate and high-level penicillin resistance, respectively. Ceftriaxone and gemifloxacin were the most active drugs (MICs at which 90% of isolates are inhibited, 0.01 versus 0.007 μg/ml, respectively), with 100% of strains inhibited by 0.12 μg/ml.

Sign in / Sign up

Export Citation Format

Share Document