Prevalence and characterization of carbapenemase producing isolates of Enterobacteriaceae obtained from clinical and environmental samples: Efflux pump inhibitor study

Laribacter hongkongensis is a food-borne bacterium associated with community-acquired gastroenteritis and diarrhoea. Quinolone resistance was recently reported in bacterial isolates from aquatic products, but the molecular mechanisms for resistance were still unknown. In this study, a total of 157 L. hongkongensis strains were isolated from grass carps (n = 443) and Chinese tiger frogs (n = 171). Twenty-one ciprofloxacin-resistant strains were analysed for mutations in quinolone resistance-determining regions (QRDR), acquired quinolone resistance (AQR) genes and the role of efflux pumps in resistance. All QRDR mutations in gyrA (codons 85 and 89) and parC (codons 83 and 231) were found to be closely associated with ciprofloxacin resistance. The AQR gene aac(6′)-Ib-cr was found in 42.9 % (9/21) of the resistant strains, but qnrA, qnrB, qnrC, qnrD, qnrS and qepA were not detected. No significant change of MICs to ciprofloxacin was observed in the presence of an efflux pump inhibitor, indicating the role of efflux pump was probably absent. All 21 ciprofloxacin-resistant strains showed different electrophoretic patterns, which suggested they were not genetically related. These data highlight the importance of QRDR mutations and the AQR gene aac(6′)-Ib-cr during the development of quinolone resistance in a heterogeneous population of L. hongkongensis.

Download Full-text

EFFLUX MEDIATED MULTIDRUG RESISTANT PSEUDOMONAS AERUGINOSA ISOLATED FROM DIFFERENT ENVIRONMENTAL SOURCES

Journal of Natural Sciences Engineering and Technology ◽

10.51406/jnset.v17i1.1903 ◽

2019 ◽

Vol 17 (1) ◽

pp. 110-120

Author(s):

G. C. AGU ◽

B. T. THOMAS ◽

O. O. SALAMI ◽

O. D. POPOOLA

Keyword(s):

Pseudomonas Aeruginosa ◽

Environmental Samples ◽

Efflux Pump ◽

Opportunistic Pathogen ◽

Antibiotics Resistance ◽

Drug Resistant ◽

Efflux Pump Inhibitor ◽

Pump System ◽

Carbonyl Cyanide

Pseudomonas aeruginosa is an important opportunistic pathogen and one of the leading causes of multi-drug resistant nosocomial infections. This study was therefore carried out to determine the resistance nature, and the role of efflux pump in multidrug resistance of Pseudomonas aeruginosa isolated from different environmental sources using the efflux pump inhibitor, Carbonyl Cyanide 3-Chlorophenylhydrazone (CCCP). A total of 220 environmental samples were collected and processed following standard techniques. Susceptibility to antibiotics was performed using disc diffusion methods as described by the Clinical and Laboratory Standards Institute. Activity of the efflux pump system was carried out using the efflux pump inhibitor, CCCP. Results obtained identified 100 (45.5%) Pseudomonas aeruginosa and 72 (32.7%) other strains of Pseudomonas spp. The susceptibility testing revealed that all the identified strains of Pseudomonas aeruginosa that were subjected to susceptibility test were significantly resistant to ampicillin and cefotaxime, But the resistance profile of isolates to tetracycline, chloramphenicol, ceftriaxone, cefuroxime and perfloxacin were 93%, 72.1%, 79.1%, 58.1% and 51.2% respectively. However, imipenem was the most sensitive (100%), followed by cefepime (65%) and gentamicin (44%). Carbonyl Cyanide 3-Chlorophenylhydrazone decreased the minimum inhibitory concentration (MIC) of the isolates by 2 folds. Results obtained have shown the ubiquitous presence of multi-drug resistant P. aeruginosa from the environmental samples examined. Furthermore, it indicated the role of efflux pump in antibiotics resistance in P. aeruginosa isolates which indicate that P. aeruginosa strains from environmental sources could resist antibiotics by the efflux mechanism.

Download Full-text

Characterization of Plasmid-Encoded Resistance-Nodulation-Division Efflux Pump in Klebsiella pneumoniae and Klebsiella quasipneumoniae from Patients in China

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02075-20 ◽

2020 ◽

Author(s):

Ruowen He ◽

Yongqiang Yang ◽

Yiping Wu ◽

Lan-Lan Zhong ◽

Yanxian Yang ◽

...

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Klebsiella Pneumoniae ◽

Urinary Tract Infections ◽

Efflux Pump ◽

Multidrug Resistant ◽

Efflux Pump Inhibitor ◽

Resistance Nodulation Division ◽

Tract Infections

Two multidrug-resistant (MDR) mcr-1 harboring Klebsiella pneumoniae isolates from patients with urinary tract infections, and one MDR Klebsiella quasipneumoniae isolate from a patient with bloodstream infection were identified to carry tmexCD1-toprJ1. The addition of the efflux pump inhibitor reduced the tigecycline MIC against all three isolates by 8- to 16-fold. The pKQBSI104-1 transferred from K. quasipneumoniae to Escherichia coli J53 via conjugation. The tmexCD1-toprJ1-carrying plasmids pKP15ZE495-1 (102,569 bp) and pKQBSI104-1 (121,996 bp) were completely sequenced and analyzed.

Download Full-text

New Trimethoprim-Like Molecules: Bacteriological Evaluation and Insights into Their Action

Antibiotics ◽

10.3390/antibiotics10060709 ◽

2021 ◽

Vol 10 (6) ◽

pp. 709

Author(s):

Marta Jorba ◽

Marina Pedrola ◽

Ouldouz Ghashghaei ◽

Rocío Herráez ◽

Lluis Campos-Vicens ◽

...

Keyword(s):

Inhibitory Concentration ◽

Efflux Pump ◽

Synergistic Effects ◽

Nicotinamide Adenine Dinucleotide Phosphate ◽

Detailed Characterization ◽

Low Concentrations ◽

Pump Activity ◽

Bacteriological Evaluation ◽

Kinetics Of

This work reports a detailed characterization of the antimicrobial profile of two trimethoprim-like molecules (compounds 1a and 1b) identified in previous studies. Both molecules displayed remarkable antimicrobial activity, particularly when combined with sulfamethoxazole. In disk diffusion assays on Petri dishes, compounds 1a and 1b showed synergistic effects with colistin. Specifically, in combinations with low concentrations of colistin, very large increases in the activities of compounds 1a and 1b were determined, as demonstrated by alterations in the kinetics of bacterial growth despite only slight changes in the fractional inhibitory concentration index. The effect of colistin may be to increase the rate of antibiotic entry while reducing efflux pump activity. Compounds 1a and 1b were susceptible to extrusion by efflux pumps, whereas the inhibitor phenylalanine arginyl β-naphthylamide (PAβN) exerted effects similar to those of colistin. The interactions between the target enzyme (dihydrofolate reductase), the coenzyme nicotinamide adenine dinucleotide phosphate (NADPH), and the studied molecules were explored using enzymology tools and computational chemistry. A model based on docking results is reported.

Download Full-text

The Impact of an Efflux Pump Inhibitor on the Activity of Free and Liposomal Antibiotics against Pseudomonas aeruginosa

Pharmaceutics ◽

10.3390/pharmaceutics13040577 ◽

2021 ◽

Vol 13 (4) ◽

pp. 577

Author(s):

Douweh Leyla Gbian ◽

Abdelwahab Omri

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Efflux Pump ◽

Antimicrobial Activities ◽

Dilution Method ◽

Efflux Pump Inhibitor ◽

Signal Production ◽

Lung Infections ◽

The Impact ◽

Microbroth Dilution

The eradication of Pseudomonas aeruginosa in cystic fibrosis patients has become continuously difficult due to its increased resistance to treatments. This study assessed the efficacy of free and liposomal gentamicin and erythromycin, combined with Phenylalanine arginine beta-naphthylamide (PABN), a broad-spectrum efflux pump inhibitor, against P. aeruginosa isolates. Liposomes were prepared and characterized for their sizes and encapsulation efficiencies. The antimicrobial activities of formulations were determined by the microbroth dilution method. Their activity on P. aeruginosa biofilms was assessed, and the effect of sub-inhibitory concentrations on bacterial virulence factors, quorum sensing (QS) signals and bacterial motility was also evaluated. The average diameters of liposomes were 562.67 ± 33.74 nm for gentamicin and 3086.35 ± 553.95 nm for erythromycin, with encapsulation efficiencies of 13.89 ± 1.54% and 51.58 ± 2.84%, respectively. Liposomes and PABN combinations potentiated antibiotics by reducing minimum inhibitory and bactericidal concentrations by 4–32 fold overall. The formulations significantly inhibited biofilm formation and differentially attenuated virulence factor production as well as motility. Unexpectedly, QS signal production was not affected by treatments. Taken together, the results indicate that PABN shows potential as an adjuvant of liposomal macrolides and aminoglycosides in the management of lung infections in cystic fibrosis patients.

Download Full-text

Characterization of resistance mechanisms of Enterobacter cloacae Complex co-resistant to carbapenem and colistin

BMC Microbiology ◽

10.1186/s12866-021-02250-x ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Shixing Liu ◽

Renchi Fang ◽

Ying Zhang ◽

Lijiang Chen ◽

Na Huang ◽

...

Keyword(s):

Lipid A ◽

Efflux Pump ◽

Resistance Mechanism ◽

Carbapenem Resistance ◽

Enterobacter Cloacae ◽

Resistance Mechanisms ◽

Colistin Resistance ◽

Carbapenem Resistant ◽

Horizontal Spread

Abstract Background The emergence of carbapenem-resistant and colistin-resistant ECC pose a huge challenge to infection control. The purpose of this study was to clarify the mechanism of the carbapenems and colistin co-resistance in Enterobacter cloacae Complex (ECC) strains. Results This study showed that the mechanisms of carbapenem resistance in this study are: 1. Generating carbapenemase (7 of 19); 2. The production of AmpC or ESBLs combined with decreased expression of out membrane protein (12 of 19). hsp60 sequence analysis suggested 10 of 19 the strains belong to colistin hetero-resistant clusters and the mechanism of colistin resistance is increasing expression of acrA in the efflux pump AcrAB-TolC alone (18 of 19) or accompanied by a decrease of affinity between colistin and outer membrane caused by the modification of lipid A (14 of 19). Moreover, an ECC strain co-harboring plasmid-mediated mcr-4.3 and blaNDM-1 has been found. Conclusions This study suggested that there is no overlap between the resistance mechanism of co-resistant ECC strains to carbapenem and colistin. However, the emergence of strain co-harboring plasmid-mediated resistance genes indicated that ECC is a potential carrier for the horizontal spread of carbapenems and colistin resistance.

Download Full-text

Characterization of RarA, a Novel AraC Family Multidrug Resistance Regulator in Klebsiella pneumoniae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00456-12 ◽

2012 ◽

Vol 56 (8) ◽

pp. 4450-4458 ◽

Cited By ~ 59

Author(s):

Mark Veleba ◽

Paul G. Higgins ◽

Gerardo Gonzalez ◽

Harald Seifert ◽

Thamarai Schneiders

Keyword(s):

Multidrug Resistance ◽

Klebsiella Pneumoniae ◽

Efflux Pump ◽

Content Type ◽

Resistance Phenotype ◽

Serratia Proteamaculans ◽

Virulence Potential ◽

Article Type ◽

Acrab Efflux Pump

ABSTRACTTranscriptional regulators, such as SoxS, RamA, MarA, and Rob, which upregulate the AcrAB efflux pump, have been shown to be associated with multidrug resistance in clinically relevant Gram-negative bacteria. In addition to the multidrug resistance phenotype, these regulators have also been shown to play a role in the cellular metabolism and possibly the virulence potential of microbial cells. As such, the increased expression of these proteins is likely to cause pleiotropic phenotypes.Klebsiella pneumoniaeis a major nosocomial pathogen which can express the SoxS, MarA, Rob, and RamA proteins, and the accompanying paper shows that the increased transcription oframAis associated with tigecycline resistance (M. Veleba and T. Schneiders, Antimicrob. Agents Chemother. 56:4466–4467, 2012). Bioinformatic analyses of the availableKlebsiellagenome sequences show that an additional AraC-type regulator is encoded chromosomally. In this work, we characterize this novel AraC-type regulator, hereby called RarA (Regulator of antibiotic resistance A), which is encoded inK. pneumoniae,Enterobactersp. 638,Serratia proteamaculans568, andEnterobacter cloacae. We show that the overexpression ofrarAresults in a multidrug resistance phenotype which requires a functional AcrAB efflux pump but is independent of the other AraC regulators. Quantitative real-time PCR experiments show thatrarA(MGH 78578 KPN_02968) and its neighboring efflux pump operonoqxAB(KPN_02969_02970) are consistently upregulated in clinical isolates collected from various geographical locations (Chile, Turkey, and Germany). Our results suggest thatrarAoverexpression upregulates theoqxABefflux pump. Additionally, it appears thatoqxR, encoding a GntR-type regulator adjacent to theoqxABoperon, is able to downregulate the expression of theoqxABefflux pump, where OqxR complementation resulted in reductions to olaquindox MICs.

Download Full-text

Enhanced efficacy of putative efflux pump inhibitor/antibiotic combination treatments versus MDR strains ofPseudomonas aeruginosain aGalleria mellonella in vivoinfection model

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkv111 ◽

2015 ◽

Vol 70 (8) ◽

pp. 2271-2278 ◽

Cited By ~ 27

Author(s):

Dougal H. Adamson ◽

Vasare Krikstopaityte ◽

Peter J. Coote

Keyword(s):

Efflux Pump ◽

Efflux Pump Inhibitor ◽

Combination Treatments ◽

Antibiotic Combination

Download Full-text

Staphylococcus aureus Mutants Isolated via Exposure to Nonfluorinated Quinolones: Detection of Known and Unique Mutations

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.45.12.3422-3426.2001 ◽

2001 ◽

Vol 45 (12) ◽

pp. 3422-3426 ◽

Cited By ~ 10

Author(s):

Siddhartha Roychoudhury ◽

Tracy L. Twinem ◽

Kelly M. Makin ◽

Mark A. Nienaber ◽

Chuiying Li ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Sequence Analysis ◽

Efflux Pump ◽

Serial Passage ◽

Efflux Pump Inhibitor ◽

In Vitro Development ◽

Development Of Resistance ◽

High Level ◽

Vitro Development

ABSTRACT The in vitro development of resistance to the new nonfluorinated quinolones (NFQs; PGE 9262932, PGE 4175997, and PGE 9509924) was investigated in Staphylococcus aureus. At concentrations two times the MIC, step 1 mutants were isolated more frequently with ciprofloxacin and trovafloxacin (9.1 × 10−8 and 5.7 × 10−9, respectively) than with the NFQs, gatifloxacin, or clinafloxacin (<5.7 × 10−10). Step 2 and step 3 mutants were selected via exposure of a step 1 mutant (selected with trovafloxacin) to four times the MICs of trovafloxacin and PGE 9262932. The step 1 mutant contained the known Ser80-Phe mutation in GrlA, and the step 2 and step 3 mutants contained the known Ser80-Phe and Ser84-Leu mutations in GrlA and GyrA, respectively. Compared to ciprofloxacin, the NFQs were 8-fold more potent against the parent and 16- to 128-fold more potent against the step 3 mutants. Mutants with high-level NFQ resistance (MIC, 32 μg/ml) were isolated by the spiral plater-based serial passage technique. DNA sequence analysis of three such mutants revealed the following mutations: (i) Ser84-Leu in GyrA and Glu84-Lys and His103-Tyr in GrlA; (ii) Ser-84Leu in GyrA, Ser52-Arg in GrlA, and Glu472-Val in GrlB; and (iii) Ser84-Leu in GyrA, Glu477-Val in GyrB, and Glu84-Lys and His103-Tyr in GrlA. Addition of the efflux pump inhibitor reserpine (10 μg/ml) resulted in 4- to 16-fold increases in the potencies of the NFQs against these mutants, whereas it resulted in 2-fold increases in the potencies of the NFQs against the parent.

Download Full-text