scholarly journals Assessing the role of the RND efflux pump in metronidazole resistance of Helicobacter pylori by RT-PCR assay

2010 ◽  
Vol 5 (02) ◽  
pp. 88-93 ◽  
Author(s):  
Jalil Fallah Mehrabadi ◽  
Mehrandokht Sirous ◽  
Naser Ebrahimi Daryani ◽  
Saeed Eshraghi ◽  
Bahram Akbari ◽  
...  
Keyword(s):  
Efflux Pump ◽  
Efflux Pumps ◽  
Clinical Isolates ◽  
Transcriptional Analysis ◽  
Rt Pcr ◽  
Helicobactor Pylori ◽  
Homologous Genes ◽  
Metronidazole Resistance ◽  
Rnd Efflux Pump ◽  
H Pylori

Introduction: Metronidazole is a significant antibiotic used for eradication of Helicobactor pylori infections and it is of notice that metronidazole-resistant clinical isolates have been found in high rates worldwide. While the RND family of efflux pumps plays a central role in drug resistance among Gram-negative bacteria, this is questionable for H. pylori. Methodology: To understand whether TolC homologues of RND pumps contribute to metronidazole resistance in H. pylori isolates, expression of four TolC homologous genes of five resistant clinical isolates exposed to varying concentrations of metronidazole were evaluated by RT-PCR and transcriptional analysis. Results: The results indicate that excess amounts of metronidazole are able to increase the expression level of these genes at the transcriptional stage. Conclusions: Therefore, it may be hypothesized that use of metronidazole in H. pyori infection can induce metronidazole resistance. Furthermore, the RND family of efflux pumps may contribute to metronidazole resistance in clinical isolates of H. pylori.

scholarly journals Furazolidone- and Nitrofurantoin-ResistantHelicobacter pylori: Prevalence and Role of Genes Involved in Metronidazole Resistance

2001 ◽  
Vol 45 (1) ◽  
pp. 306-308 ◽  
Author(s):  
Dong H. Kwon ◽  
Miae Lee ◽  
J. J. Kim ◽  
J. G. Kim ◽  
F. A. K. El-Zaatari ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Clinical Isolates ◽  
Metronidazole Resistance ◽  
H Pylori

ABSTRACT The prevalence of furazolidone, nitrofurantoin, and metronidazole resistance among Helicobacter pylori strains was assessed with 431 clinical isolates. Fifty-two percent were metronidazole resistant, compared to 2% (7 of 431) with resistance to furazolidone and nitrofurantoin. All seven furazolidone- and nitrofurantoin-resistant isolates were also metronidazole resistant.rdxA, frxA, and fdxB knockouts did not result in furazolidone or nitrofurantoin resistance. These data suggest that furazolidone and nitrofurantoin may be good alternatives to metronidazole for treating H. pylori infection.

scholarly journals Systematic Analysis of Efflux Pump-Mediated Antiseptic Resistance in Staphylococcus aureus Suggests a Need for Greater Antiseptic Stewardship

mSphere ◽  
2020 ◽  
Vol 5 (1) ◽  
Author(s):  
Patrick T. LaBreck ◽  
Audrey C. Bochi-Layec ◽  
Joshua Stanbro ◽  
Gina Dabbah-Krancher ◽  
Mark P. Simons ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Transcriptional Analysis ◽  
Systematic Analysis ◽  
Daily Lives ◽  
Single Strain ◽  
Content Type ◽  
Subinhibitory Concentrations ◽  
Different Strains

ABSTRACT Staphylococcus aureus-associated infections can be difficult to treat due to multidrug resistance. Thus, infection prevention is critical. Cationic antiseptics, such as chlorhexidine (CHX) and benzalkonium chloride (BKC), are liberally used in health care and community settings to prevent infection. However, increased administration of antiseptics has selected for S. aureus strains that show reduced susceptibilities to cationic antiseptics. This increased resistance has been associated with carriage of specific efflux pumps (QacA, QacC, and NorA). Since prior published studies focused on different strains and on strains carrying only a single efflux gene, the relative importance of these various systems to antiseptic resistance is difficult to ascertain. To overcome this, we engineered a collection of isogenic S. aureus strains that harbored norA, qacA, and qacC, individually or in combination. MIC assays showed that qacA was associated with increased resistance to CHX, cetrimide (CT), and BKC, qacC was associated with resistance to CT and BKC, and norA was necessary for basal-level resistance to the majority of tested antiseptics. When all three pumps were present in a single strain, an additive effect was observed in the MIC for CT. Transcriptional analysis revealed that expression of qacA and norA was significantly induced following exposure to BKC. Alarmingly, in a strain carrying qacA and norA, preexposure to BKC increased CHX tolerance. Overall, our results reveal increased antiseptic resistance in strains carrying multiple efflux pumps and indicate that preexposure to BKC, which is found in numerous daily-use products, can increase CHX tolerance. IMPORTANCE S. aureus remains a significant cause of disease within hospitals and communities. To reduce the burden of S. aureus infections, antiseptics are ubiquitously used in our daily lives. Furthermore, many antiseptic compounds are dual purpose and are found in household products. The increased abundance of antiseptic compounds has selected for S. aureus strains that carry efflux pumps that increase resistance to antiseptic compounds; however, the effect of carrying multiple pumps within S. aureus is unclear. We demonstrated that an isogenic strain carrying multiple efflux pumps had an additive resistance phenotype to cetrimide. Moreover, in a strain carrying qacA and norA, increased chlorhexidine tolerance was observed after the strain was preexposed to subinhibitory concentrations of a different common-use antiseptic. Taken together, our findings demonstrate cooperation between antiseptic resistance efflux pumps and suggest that their protective phenotype may be exacerbated by priming with subinhibitory concentrations of household antiseptics.

Tigecycline efflux in Acinetobacter baumannii is mediated by TetA in synergy with RND-type efflux transporters

2020 ◽  
Vol 75 (5) ◽  
pp. 1135-1139 ◽  
Author(s):  
Wuen Ee Foong ◽  
Jochen Wilhelm ◽  
Heng-Keat Tam ◽  
Klaas M Pos
Keyword(s):  
Acinetobacter Baumannii ◽  
Efflux Pump ◽  
Gene Knockout ◽  
Drug Susceptibility ◽  
Major Facilitator Superfamily ◽  
Rt Pcr ◽  
E Coli ◽  
Rnd Efflux Pump ◽  
Major Facilitator

Abstract Objectives To investigate the role of Major Facilitator Superfamily (MFS)-type transporters from Acinetobacter baumannii AYE in tigecycline efflux. Methods Two putative tetracycline transporter genes of A. baumannii AYE (tetA and tetG) were heterologously expressed in Escherichia coli and drug susceptibility assays were conducted with tigecycline and three other tetracycline derivatives. The importance of TetA in tigecycline transport in A. baumannii was determined by complementation of tetA in WT and Resistance Nodulation cell Division (RND) gene knockout strains of A. baumannii ATCC 19606. Gene expression of the MFS-type tetA gene and RND efflux pump genes adeB, adeG and adeJ in A. baumannii AYE in the presence of tigecycline was analysed by quantitative real-time RT–PCR. Results Overproduction of TetA or TetG conferred resistance to doxycycline, minocycline and tetracycline in E. coli. Cells expressing tetA, but not those expressing tetG, conferred resistance to tigecycline, implying that TetA is a determinant for tigecycline transport. A. baumannii WT and RND-knockout strains complemented with plasmid-encoded tetA are significantly less susceptible to tigecycline compared with non-complemented strains. Efflux pump genes tetA and adeG are up-regulated in A. baumannii AYE in the presence of subinhibitory tigecycline concentrations. Conclusions TetA plays an important role in tigecycline efflux of A. baumannii by removing the drug from cytoplasm to periplasm and, subsequently, the RND-type transporters AdeABC and AdeIJK extrude tigecycline across the outer membrane. When challenged with tigecycline, tetA is up-regulated in A. baumannii AYE. Synergy between TetA and the RND-type transporters AdeABC and/or AdeIJK appears necessary for A. baumannii to confer higher tigecycline resistance via drug efflux.

scholarly journals Involvement of the AcrAB-TolC Efflux Pump in the Resistance, Fitness, and Virulence of Enterobacter cloacae

2012 ◽  
Vol 56 (4) ◽  
pp. 2084-2090 ◽  
Author(s):  
Astrid Pérez ◽  
Margarita Poza ◽  
Ana Fernández ◽  
Maria del Carmen Fernández ◽  
Susana Mallo ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Pathogenic Bacteria ◽  
Efflux Pump ◽  
Enterobacter Cloacae ◽  
Efflux Pumps ◽  
Clinical Isolates ◽  
Wild Type ◽  
Content Type

ABSTRACTMultidrug efflux pumps have emerged as important mechanisms of antimicrobial resistance in bacterial pathogens. In order to cause infection, pathogenic bacteria require mechanisms to avoid the effects of host-produced compounds, and express efflux pumps may accomplish this task. In this study, we evaluated the effect of the inactivation of AcrAB-TolC on antimicrobial resistance, fitness, and virulence inEnterobacter cloacae, an opportunistic pathogen usually involved in nosocomial infections. Two different clinical isolates ofE. cloacaewere used, EcDC64 (multidrug resistance overexpressing the AcrAB-TolC efflux pump) and Jc194 (basal AcrAB-TolC expression). TheacrAandtolCgenes were deleted in strains EcDC64 and Jc194 to produce, respectively, EcΔacrAand EcΔtolCand JcΔacrAand JcΔtolCknockout (KO) derivatives. Antibiotic susceptibility testing was performed with all isolates, and we discovered that these mechanisms are involved in the resistance ofE. cloacaeto several antibiotics. Competition experiments were also performed with wild-type and isogenic KO strains. The competition index (CI), defined as the mutant/wild-type ratio, revealed that theacrAandtolCgenes both affect the fitness ofE. cloacae, as fitness was clearly reduced in theacrAandtolCKO strains. The median CI values obtainedin vitroandin vivowere, respectively, 0.42 and 0.3 for EcDC64/EcΔacrA, 0.24 and 0.38 for EcDC64/EcΔtolC, 0.15 and 0.11 for Jc194/JcΔacrA, and 0.38 and 0.39 for Jc194/JcΔtolC. Use of an intraperitoneal mouse model of systemic infection revealed reduced virulence in bothE. cloacaeclinical strains when either theacrAortolCgene was inactivated. In conclusion, the structural components of the AcrAB-TolC efflux pump appear to play a role in antibiotic resistance as well as environmental adaptation and host virulence in clinical isolates ofE. cloacae.

scholarly journals Characteristics and diversity of mutations in regulatory genes of resistance-nodulation-cell division efflux pumps in association with drug-resistant clinical isolates of Acinetobacter baumannii

2020 ◽  
Author(s):  
Bahare Salehi ◽  
Zohreh Ghalavand ◽  
Abbas Yadegar ◽  
Gita Eslami
Keyword(s):  
Cell Division ◽  
Acinetobacter Baumannii ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Regulatory Genes ◽  
Clinical Isolates ◽  
Drug Resistant ◽  
Broth Microdilution Method ◽  
Rnd Efflux Pumps ◽  
Multiple Antibiotics

Abstract Background: This study aimed to characterize the regulation and expression of three putative resistance-nodulation-cell division (RND)-type efflux systems and their contribution to multidrug efflux in clinical isolates of Acinetobacter baumannii. Methods: Antimicrobial susceptibility testing (AST) of 95 A. baumannii isolates was determined by Kirby-Bauer disk diffusion for 18 antibiotics and minimum inhibitory concentration (MIC) of colistin was determined by broth microdilution method. Moreover, MIC of five classes of antibiotics was assessed using E-test strips in the presence and absence of phenylalanine-arginine beta-naphthylamide (PAβN). Regulatory genes of RND efflux pumps (AdeRS, AdeL, AdeN and BaeSR) were subjected to sequencing. The relative expression of adeB. adeG and adeJ genes was determined by quantitative real-time PCR (RT-PCR).Results: Overall, majority of isolates (93%) were extensively drug-resistant (XDR). In the phenotypic assay, efflux pump activity was observed in 40% of isolates against multiple antibiotics mainly tigecycline, but not to imipenem. Several amino acid substitutions were detected in the regulatory genes; except in AdeN. Of note, G186V in AdeS were found to be associated with overexpression of their relative efflux pumps. No insertion sequences (ISs) were detected. Conclusions: Our findings outline the role of RND efflux pumps in resistance of A. baumannii against multiple antibiotics particularly tigecycline, and point out importance of a variety of single mutations in the corresponding regulatory systems. Even though it has been concluded that multidrug resistance occurs as a result of a complex sets of different resistant mechanisms.

scholarly journals Characteristics and diversity of mutations in regulatory genes of resistance-nodulation-cell division efflux pumps in association with drug-resistant clinical isolates of Acinetobacter baumannii

2020 ◽  
Author(s):  
Bahare Salehi ◽  
Zohreh Ghalavand ◽  
Abbas Yadegar ◽  
Gita Eslami
Keyword(s):  
Cell Division ◽  
Acinetobacter Baumannii ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Regulatory Genes ◽  
Clinical Isolates ◽  
Drug Resistant ◽  
Broth Microdilution Method ◽  
Rnd Efflux Pumps ◽  
Multiple Antibiotics

Abstract Background: This study aimed to characterize the regulation and expression of three putative resistance-nodulation-cell division (RND)-type efflux systems and their contribution to multidrug efflux in clinical isolates of Acinetobacter baumannii.Methods: Antimicrobial susceptibility testing (AST) of 95 A. baumannii isolates was determined by Kirby-Bauer disk diffusion for 18 antibiotics and minimum inhibitory concentration (MIC) of colistin was determined by broth microdilution method. Moreover, MIC of five classes of antibiotics was assessed using E-test strips in the presence and absence of phenylalanine-arginine beta-naphthylamide (PAβN). Regulatory genes of RND efflux pumps (AdeRS, AdeL, AdeN and BaeSR) were subjected to sequencing. The relative expression of adeB. adeG and adeJ genes was determined by quantitative real-time PCR (RT-PCR).Results: Overall, majority of isolates (93%) were extensively drug-resistant (XDR). In the phenotypic assay, efflux pump activity was observed in 40% of isolates against multiple antibiotics mainly tigecycline, but not to imipenem. Several amino acid substitutions were detected in the regulatory genes; except in AdeN. Of note, G186V in AdeS were found to be associated with overexpression of their relative efflux pumps. No insertion sequences (ISs) were detected.Conclusions: Our findings outline the role of RND efflux pumps in resistance of A. baumannii against multiple antibiotics particularly tigecycline, and point out importance of a variety of single mutations in the corresponding regulatory systems. Even though it has been concluded that multidrug resistance occurs as a result of a complex sets of different resistant mechanisms.

scholarly journals Alkylaminoquinolines inhibit the bacterial antibiotic efflux pump in multidrug-resistant clinical isolates

2003 ◽  
Vol 376 (3) ◽  
pp. 801-805 ◽  
Author(s):  
Monique MALLÉA ◽  
Abdallah MAHAMOUD ◽  
Jacqueline CHEVALIER ◽  
Sandrine ALIBERT-FRANCO ◽  
Pierre BROUANT ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Microbial Pathogens ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
High Level ◽  
Antibiotic Efflux

Over the last decade, MDR (multidrug resistance) has increased worldwide in microbial pathogens by efflux mechanisms, leading to treatment failures in human infections. Several Gram-negative bacteria efflux pumps have been described. These proteinaceous channels are capable of expelling structurally different drugs across the envelope and conferring antibiotic resistance in various bacterial pathogens. Combating antibiotic resistance is an urgency and the blocking of efflux pumps is an attractive response to the emergence of MDR phenotypes in infectious bacteria. In the present study, various alkylaminoquinolines were tested as potential inhibitors of drug transporters. We showed that alkylaminoquinolines are capable of restoring susceptibilities to structurally unrelated antibiotics in clinical isolates of MDR Gram-negative bacteria. Antibiotic efflux studies indicated that 7-nitro-8-methyl-4-[2´-(piperidino)ethyl]aminoquinoline acts as an inhibitor of the AcrAB–TolC efflux pump and restores a high level of intracellular drug concentration. Inhibitory activity of this alkylaminoquinoline is observed on clinical isolates showing different resistance phenotypes.

scholarly journals Bifunctional enzyme SpoT is involved in biofilm formation of Helicobacter pylori with multidrug resistance by upregulating efflux pump Hp1174 (gluP)

2018 ◽  
Author(s):  
Xiaoran Ge ◽  
Yuying Cai ◽  
Zhenghong Chen ◽  
Sizhe Gao ◽  
Xiwen Geng ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Helicobacter Pylori ◽  
Multidrug Resistance ◽  
Biofilm Formation ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Multi Drug Resistance ◽  
Bifunctional Enzyme ◽  
Average Expression Level ◽  
H Pylori

ABSTRACTThe drug resistance of Helicobacter pylori (H. pylori) is gradually becoming a serious problem. Biofilm formation is an important factor that leads to multidrug resistance in bacteria. The ability of H. pylori to form biofilms on the gastric mucosa has been known. However, there are few studies on the regulation mechanisms of H. pylori biofilm formation and multidrug resistance. Guanosine 3’-diphosphate 5’-triphosphate and guanosine 3’,5’-bispyrophosphate [(p)ppGpp] are global regulatory factors and are synthesized in H. pylori by the bifunctional enzyme SpoT. It has been reported that (p)ppGpp is involved in the biofilm formation and multidrug resistance of various bacteria. In this study, we found that SpoT also plays an important role in H. pylori biofilm formation and multidrug resistance. Therefore, it is necessary to carry out some further studies regarding its regulatory mechanism. Considering that efflux pumps are of great importance in the biofilm formation and multidrug resistance of bacteria, we tried to find if efflux pumps controlled by SpoT participate in these activities. Then, we found that Hp1174 (glucose/galactose transporter, gluP), an efflux pump of the MFS family, is highly expressed in biofilm-forming and multi-drug resistance (MDR) H. pylori and is upregulated by SpoT. Through further research, we determined that gluP involved in H. pylori biofilm formation and multidrug resistance. Furthermore, the average expression level of gluP in clinical MDR strains was considerably higher than that in clinical drug-sensitive strains. Taken together, our results revealed a novel molecular mechanism of H. pylori tolerance to multidrug.

Biofilm Formation Restrained by Subinhibitory Concentrations of Tigecyclin in Acinetobacter baumannii Is Associated with Downregulation of Efflux Pumps

Chemotherapy ◽  
2016 ◽  
Vol 62 (2) ◽  
pp. 128-133 ◽  
Author(s):  
Huale Chen ◽  
Jianming Cao ◽  
Cui Zhou ◽  
Haiyang Liu ◽  
Xiaoxiao Zhang ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Real Time ◽  
Biofilm Formation ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Extracellular Dna ◽  
Proteinase K ◽  
Quantitative Detection ◽  
Rt Pcr ◽  
Subinhibitory Concentrations

Background: Tigecycline, one of the few therapeutic options against multidrug-resistant Acinetobacter baumannii, reaches subinhibitory serum concentrations only with cautious clinical dosing and pharmacokinetics. Subinhibitory concentrations of tigecycline might induce an A. baumannii biofilm. Methods: Biofilm formation was assessed via the crystal violet staining method. We further analyzed the main biofilm components with NaIO4, proteinase K, and DNase. Real-time RT-PCR was applied for quantitative detection of biofilm potential-associated genes. Results: In this study, A. baumannii proved to be a strong biofilm producer, and we found that proteins and extracellular DNA are crucial components of the A. baumannii biofilm. Quantitative real-time RT-PCR revealed positive correlations between biofilm formation restrained by subinhibitory concentrations of tigecycline and the expression of biofilm potential-associated genes, especially the AdeFGH efflux pump gene. Conclusion: Our results suggest that downregulation of efflux pumps, especially the AdeFGH efflux pump, is probably responsible for the decline in biofilm formation in A. baumannii treated with subinhibitory concentrations of tigecyclin.

scholarly journals Characteristics and diversity of mutations in regulatory genes of resistance-nodulation-cell division efflux pumps in association with drug-resistant clinical isolates of Acinetobacter baumannii

2021 ◽  
Vol 10 (1) ◽  
Author(s):  
Bahare Salehi ◽  
Zohreh Ghalavand ◽  
Abbas Yadegar ◽  
Gita Eslami
Keyword(s):  
Cell Division ◽  
Acinetobacter Baumannii ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Regulatory Genes ◽  
Clinical Isolates ◽  
Drug Resistant ◽  
Rnd Efflux Pumps ◽  
Multiple Antibiotics

Abstract Background This study was aimed to characterize the genetic diversity and expression of three putative resistance-nodulation-cell division (RND)-type efflux systems and their contribution to multidrug efflux in clinical isolates of Acinetobacter baumannii. Methods Antimicrobial susceptibility testing of 95 A. baumannii isolates was determined by Kirby-Bauer disk diffusion for 18 antibiotics and minimum inhibitory concentration (MIC) of colistin was determined by the broth microdilution method. Moreover, the MIC of five classes of antibiotics was assessed using E-test strips in the presence and absence of phenylalanine-arginine beta-naphthylamide (PAβN). Regulatory genes of the RND efflux pumps (adeRS, adeL, adeN and baeSR) were subjected to sequencing. The relative expression of adeB, adeG and adeJ genes was determined by quantitative real-time PCR (qRT-PCR). Results Overall, the majority of isolates (94%) were extensively drug-resistant (XDR). In the phenotypic assay, efflux pump activity was observed in 40% of the isolates against multiple antibiotics mainly tigecycline. However, we found no efflux activity against imipenem. Several amino acid substitutions were detected in the products of regulatory genes; except in AdeN. Of note, G186V mutation in AdeS was found to be associated with overexpression of its efflux pump. No insertion sequences were detected. Conclusions Our findings outlined the role of RND efflux pumps in resistance of A. baumannii to multiple antibiotics particularly tigecycline, and pointed out the importance of a variety of single mutations in the corresponding regulatory systems. Further studies are required to decipher the precise role of RND efflux pumps in multidrug-resistant clinical isolates of A. baumannii.

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