A Novel Tigecycline Adjuvant ML-7 Reverses the Susceptibility of Tigecycline-Resistant Klebsiella pneumoniae

The increasing incidence of tigecycline resistance undoubtedly constitutes a serious threat to global public health. The combination therapies had become the indispensable strategy against this threat. Herein, 11 clinical tigecycline-resistant Klebsiella pneumoniae which mainly has mutations in ramR, acrR, or macB were collected for tigecycline adjuvant screening. Interestingly, ML-7 hydrochloride (ML-7) dramatically potentiated tigecycline activity. We further picked up five analogs of ML-7 and evaluated their synergistic activities with tigecycline by using checkerboard assay. The results revealed that ML-7 showed certain synergy with tigecycline, while other analogs exerted attenuated synergistic effects among tigecycline-resistant isolates. Thus, ML-7 was selected for further investigation. The results from growth curves showed that ML-7 combined with tigecycline could completely inhibit the growth of bacteria, and the time-kill analysis revealed that the combination exhibited synergistic bactericidal activities for tigecycline-resistant isolates during 24 h. The ethidium bromide (EtBr) efflux assay demonstrated that ML-7 could inhibit the functions of efflux pump. Besides, ML-7 disrupted the proton motive force (PMF) via increasing ΔpH, which in turn lead to the inhibition of the functions of efflux pump, reduction of intracellular ATP levels, as well as accumulation of ROS. All of which promoted the death of bacteria. And further transcriptomic analysis revealed that genes related to the mechanism of ML-7 mainly enriched in ABC transporters. Taken together, these results revealed the potential of ML-7 as a novel tigecycline adjuvant to circumvent tigecycline-resistant Klebsiella pneumoniae.

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Synergistic Effect of Colistin Combined with PFK-158 against Colistin-Resistant Enterobacteriaceae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00271-19 ◽

2019 ◽

Vol 63 (7) ◽

Cited By ~ 3

Author(s):

Youwen Zhang ◽

Xiukun Wang ◽

Xue Li ◽

Limin Dong ◽

Xinxin Hu ◽

...

Keyword(s):

Synergistic Effect ◽

Synergistic Effects ◽

Survival Rates ◽

New Combination ◽

Resistant Bacteria ◽

Content Type ◽

Checkerboard Assay ◽

High Level ◽

Time Kill

ABSTRACT As increasing numbers of colistin-resistant bacteria emerge, new therapies are urgently needed to treat infections caused by these pathogens. The discovery of new combination therapies is one important way to solve such problems. Here, we report that the antitumor drug PFK-158 and its analogs PFK-015 and 3PO can exert synergistic effects with colistin against colistin-resistant Enterobacteriaceae, including mcr-1-positive or high-level-colistin-resistant (HLCR) isolates, as shown by a checkerboard assay. The results of a time-kill assay revealed that colistin combined with PFK-158 continuously eliminated colistin-resistant Escherichia coli 13-43, Klebsiella pneumoniae H04, and Enterobacter cloacae D01 in 24 h. Images from scanning electron microscopy (SEM) at 5 h postinoculation confirmed the killing effect of the combination. Finally, in vivo treatment showed that PFK-158 had a better synergistic effect than its analogs. Compared to the corresponding rates after colistin monotherapy, the survival rates of systemically infected mice were significantly increased 30% or 60% when the mice received an intravenous injection of colistin in combination with 15 mg/kg of body weight PFK-158. These results have important implications for repurposing PFK-158 to combat colistin resistance.

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In vitro and in vivo synergistic effects of tigecycline combined with aminoglycosides on carbapenem-resistant Klebsiella pneumoniae

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkab122 ◽

2021 ◽

Author(s):

Wentao Ni ◽

Deqing Yang ◽

Jie Guan ◽

Wen Xi ◽

Dexun Zhou ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Synergistic Effects ◽

Molecular Characteristics ◽

Synergistic Activity ◽

Significant Activity ◽

Clinical Pharmacokinetics ◽

Carbapenem Resistant ◽

Time Kill

Abstract Objectives Carbapenem-resistant Klebsiella pneumoniae (CR-KP) infections represent severe threats to public health worldwide. The aim of this study was to assess potential synergistic interaction between tigecycline and aminoglycosides via in vitro and in vivo studies. Methods Antibiotic resistance profiles and molecular characteristics of 168 CR-KP clinical isolates were investigated by susceptibility testing, PCR and MLST. Chequerboard tests and time–kill assays were performed for 20 CR-KP isolates to evaluate in vitro synergistic effects of tigecycline combined with aminoglycosides. A tissue-cage infection model of rats was established to evaluate in vivo synergistic effects. Different doses of tigecycline and aminoglycosides alone or in combination were administered for 7 days via tail vein injection. Antibiotic efficacy was evaluated in tissue-cage fluid and emergence of resistance was screened. Results The chequerboard tests showed that this combination displayed synergistic or partial synergistic activity against CR-KP. The time–kill assays further demonstrated that strong synergistic effects of such a combination existed against isolates that were susceptible to both drugs but for resistant isolates no synergy was observed if clinical pharmacokinetics were taken into consideration. The in vivo study showed that the therapeutic effectiveness of combination therapies was better than that of monotherapy for susceptible isolates, suggesting in vivo synergistic effects. Furthermore, combinations of tigecycline with an aminoglycoside showed significant activity in reducing the occurrence of tigecycline-resistant mutants. Conclusions Compared with single drugs, tigecycline combined with aminoglycosides could exert synergistic effects and reduce the emergence of tigecycline resistance. Such a combination might be an effective alternative when treating CR-KP infections in clinical practice.

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Curcumin-meropenem synergy in carbapenem resistant Klebsiella pneumoniae curcumin-meropenem synergy

Iranian Journal of Microbiology ◽

10.18502/ijm.v13i3.6397 ◽

2021 ◽

Author(s):

Dumrul Gülen ◽

Birol Şafak ◽

Berna Erdal ◽

Betül Günaydın

Keyword(s):

Klebsiella Pneumoniae ◽

Bacterial Infections ◽

Efflux Pump ◽

Synergistic Effects ◽

Treatment Options ◽

Resistance Pattern ◽

Carbapenem Resistant ◽

Efflux Pump Inhibition ◽

Disk Method

Background and Objectives: The frequency of multiple resistant bacterial infections, including carbapenems, is increasing worldwide. As the decrease in treatment options causes difficulties in treatment, interest in new antimicrobials is increasing. One of the promising natural ingredients is curcumin. It is known to be effective in bacteria such as Pseudomonas aeruginosa, Escherichia coli, Burkholderia pseudomallei through efflux pump inhibition, toxin inhibition and enzymes. However, because its bioavailability is poor, it seffectiveness occurs in combination with antibiotics. In the study, the interaction of meropenem and curcumin in carbapenemase producing strains of Klebsiella pneumoniae was tested. Materials and Methods: Thirty-nine Klebsiella pneumoniae isolates, resistant to meropenem, were used in this study. From those 15 MBL, 6 KPC, 17 OXA-48 and 1 AmpC resistance pattern were detected by combination disk method. Meropenem and Curcumin MIC values were determined by liquid microdilution. Checkerboard liquid microdilution was used to determine the synergy between meropenem and curcumin. Results: Synergistic effects were observed in 4 isolates producing MBL, 3 isolates producing KPC, 4 isolates producing OXA-48, and 1 isolates producing AmpC (totally 12 isolates) according to the calculated FICI. No antagonistic effects were observed in any isolates. Conclusion: Curcumin was thought to be an alternative antimicrobial in combination therapies that would positively contribute to the treatment of bacterial infection. The effectiveness of this combination should be confirmed by other in vitro and clinical studies.

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Synergistic Antibacterial Effects of Meropenem in Combination with Aminoglycosides against Carbapenem-Resistant Escherichia coli Harboring blaNDM-1 and blaNDM-5

Antibiotics ◽

10.3390/antibiotics10081023 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1023

Author(s):

Pawarisa Terbtothakun ◽

Ozioma Forstinus Nwabor ◽

Thanyaluck Siriyong ◽

Supayang P. Voravuthikunchai ◽

Sarunyou Chusri

Keyword(s):

Escherichia Coli ◽

Antimicrobial Agents ◽

Synergistic Effects ◽

Treatment Options ◽

Molecular Characteristics ◽

Carbapenem Resistant ◽

Aminoglycoside Therapy ◽

Checkerboard Assay ◽

Almost All ◽

Time Kill

Infections due to carbapenem-resistant Escherichia coli (CREC) are problematic due to limitation in treatment options. Combination therapies of existing antimicrobial agents have become a reliable strategy to control these infections. In this study, the synergistic effects of meropenem in combination with aminoglycosides were assessed by checkerboard and time-kill assays. Of the 35 isolates, 19 isolates (54.3%) were resistant to carbapenems (imipenem and meropenem) with the MIC ranges from 16 to 128 µg/mL. These isolates were resistant to almost all antibiotic classes. Molecular characteristics revealed co-harboring of carbapenemase (blaNDM-1, blaNDM-5 and blaOXA-48) and extended-spectrum β-lactamases (ESBL) genes (blaCTX-M, blaSHV and blaTEM). The checkerboard assay displayed synergistic effects of meropenem and several aminoglycosides against most CREC isolates. Time-kill assays further demonstrated strong synergistic effects of meropenem in combination with either amikacin, gentamicin, kanamycin, streptomycin, and tobramycin. The results suggested that meropenem in combination with aminoglycoside therapy might be an efficient optional treatment for infections cause by CREC.

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Comparative In Vitro Activities of First and Second-Generation Ceragenins Alone and in Combination with Antibiotics Against Multidrug-Resistant Klebsiella pneumoniae Strains

Antibiotics ◽

10.3390/antibiotics8030130 ◽

2019 ◽

Vol 8 (3) ◽

pp. 130 ◽

Cited By ~ 5

Author(s):

Berna Ozbek-Celik ◽

Damla Damar-Celik ◽

Emel Mataraci-Kara ◽

Cagla Bozkurt-Guzel ◽

Paul B. Savage

Keyword(s):

Klebsiella Pneumoniae ◽

Second Generation ◽

Synergistic Effects ◽

Multidrug Resistant ◽

Antimicrobial Activities ◽

Resistant Bacteria ◽

Microdilution Method ◽

Kill Curve ◽

Time Kill

Objectives: The ceragenins, or CSAs, were designed to mimic the activities of antimicrobial peptides and represent a new class of antimicrobial agent. The aim of this study was to comparatively investigate the antimicrobial activities of first/second generation ceragenins and various antibiotics against multidrug-resistant (MDR) Klebsiella pneumoniae, including colistin-resistant bacteria. Also, the synergistic effects of two ceragenins with colistin or meropenem were investigated with six K. pneumoniae strains presenting different resistant patterns. Methods: Minimal inhibition concentrations (MICs) were determined by the microdilution method according to the CLSI. Antibiotic combination studies were evaluated by the time–kill curve method. Results: MIC50 and MIC90 values of tested ceragenins ranged from 8 to 32 mg/L and 16 to 128 mg/L. Overall, among the ceragenins tested, CSA-131 showed the lowest MIC50 and MIC90 values against all microorganisms. The MICs of the ceragenins were similar or better than tested antibiotics, except for colistin. Synergistic activities of CSA-131 in combination with colistin was found for strains both at 1× MIC and 4× MIC. No antagonism was observed with any combination. Conclusions: First-generation ceragenins CSA-13 and CSA-44 and second-generation ceragenins CSA-131, CSA-138 and CSA-142 have significant antimicrobial effects on MDR K. pneumoniae. Mechanisms allowing resistance to clinical comparator antibiotics like colistin did not impact the activity of ceragenins. These results suggest that ceragenins may play a role in treating infections that are resistant to known antibiotics.

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Interplay among Different Fosfomycin Resistance Mechanisms in Klebsiella pneumoniae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01911-20 ◽

2020 ◽

Vol 65 (3) ◽

Author(s):

M. Ortiz-Padilla ◽

I. Portillo-Calderón ◽

B. de Gregorio-Iaria ◽

J. Blázquez ◽

J. Rodríguez-Baño ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Growth Curves ◽

Resistant Mutant ◽

Bactericidal Effect ◽

Clinical Isolates ◽

Growth Monitoring ◽

Content Type ◽

Fosfomycin Resistance ◽

Time Kill

ABSTRACT The objectives of this study were to characterize the role of the uhpT, glpT, and fosA genes in fosfomycin resistance in Klebsiella pneumoniae and evaluate the use of sodium phosphonoformate (PPF) in combination with fosfomycin. Seven clinical isolates of K. pneumoniae and the reference strain (ATCC 700721) were used, and their genomes were sequenced. ΔuhpT, ΔglpT, and ΔfosA mutants were constructed from two isolates and K. pneumoniae ATCC 700721. Fosfomycin susceptibility testing was done by the gradient strip method. Synergy between fosfomycin and PPF was studied by checkerboard assay and analyzed using SynergyFinder. Spontaneous fosfomycin mutant frequencies at 64 and 512 mg/liter, in vitro activity using growth curves with fosfomycin gradient concentrations (0 to 256mg/liter), and time-kill assays at 64 and 307 mg/liter were evaluated with and without PPF (0.623 mM). The MICs of fosfomycin against the clinical isolates ranged from 16 to ≥1,024 mg/liter. The addition of 0.623 mM PPF reduced fosfomycin MIC between 2- and 8-fold. Deletion of fosA led to a 32-fold decrease. Synergistic activities were observed with the combination of fosfomycin and PPF (most synergistic area at 0.623 mM). The lowest fosfomycin-resistant mutant frequencies were found in ΔfosA mutants, with decreases in frequency from 1.69 × 10−1 to 1.60 × 10−5 for 64 mg/liter of fosfomycin. In the final growth monitoring and time-kill assays, fosfomycin showed a bactericidal effect only with the deletion of fosA and not with the addition of PPF. We conclude that fosA gene inactivation leads to a decrease in fosfomycin resistance in K. pneumoniae. The pharmacological approach using PPF did not achieve enough activity, and the effect decreased with the presence of fosfomycin-resistant mutations.

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In VitroSynergy of Colistin Combinations against Colistin-Resistant Acinetobacter baumannii, Pseudomonas aeruginosa, and Klebsiella pneumoniae Isolates

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.05996-11 ◽

2012 ◽

Vol 56 (9) ◽

pp. 4856-4861 ◽

Cited By ~ 72

Author(s):

Céline Vidaillac ◽

Lothaire Benichou ◽

Raphaël E. Duval

Keyword(s):

Pseudomonas Aeruginosa ◽

Klebsiella Pneumoniae ◽

Acinetobacter Baumannii ◽

Population Analysis ◽

Multidrug Resistant ◽

Colistin Resistance ◽

Content Type ◽

Checkerboard Assay ◽

Time Kill

ABSTRACTColistin resistance, although uncommon, is increasingly being reported among Gram-negative clinical pathogens, and an understanding of its impact on the activity of antimicrobials is now evolving. We evaluated the potential for synergy of colistin plus trimethoprim, trimethoprim-sulfamethoxazole (1/19 ratio), or vancomycin against 12 isolates ofAcinetobacter baumannii(n= 4),Pseudomonas aeruginosa(n= 4), andKlebsiella pneumoniae(n= 4). The strains included six multidrug-resistant clinical isolates,K. pneumoniaeATCC 700603,A. baumanniiATCC 19606,P. aeruginosaATCC 27853, and their colistin-resistant derivatives (KPm1, ABm1, and PAm1, respectively). Antimicrobial susceptibilities were assessed by broth microdilution and population analysis profiles. The potential for synergy of colistin combinations was evaluated using a checkerboard assay, as well as static time-kill experiments at 0.5× and 0.25× MIC. The MIC ranges of vancomycin, trimethoprim, and trimethoprim-sulfamethoxazole (1/19) were ≥128, 4 to ≥128, and 2/38 to >128/2,432 μg/ml, respectively. Colistin resistance demonstrated little impact on vancomycin, trimethoprim, or trimethoprim-sulfamethoxazole MIC values. Isolates with subpopulations heterogeneously resistant to colistin were observed to various degrees in all tested isolates. In time-kill assays, all tested combinations were synergistic against KPm1 at 0.25× MIC and 0.5× MIC and ABm1 and PAm1 at 0.5× MIC. In contrast, none of the tested combinations demonstrated synergy against any colistin-susceptibleP. aeruginosaisolates and clinical strains ofK. pneumoniaeisolates. Only colistin plus trimethoprim or trimethoprim-sulfamethoxazole was synergistic and bactericidal at 0.5× MIC againstK. pneumoniaeATCC 700603. Colistin resistance seems to promote thein vitroactivity of unconventional colistin combinations. Additional experiments are warranted to understand the clinical significance of these observations.

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Modulation of the drug resistance by Platonia insignis Mart. extract, ethyl acetate fraction and Morelloflavone/Volkensiflavone (biflavonoids) in Staphylococcus aureus strains overexpressing efflux pump genes

Current Drug Metabolism ◽

10.2174/1389200221666200523155617 ◽

2020 ◽

Vol 21 ◽

Author(s):

Andressa Kelly Ferreira e Silva ◽

Antonielly Campinho dos Reis ◽

Emanuelly Elanny Andrade Pinheiroc ◽

Jonas Nascimento de Sousa ◽

Felipe Araújo de Alcântara Oliveira ◽

...

Keyword(s):

Natural Products ◽

Ethyl Acetate ◽

Native Species ◽

Efflux Pump ◽

Public Health Problem ◽

Ethyl Acetate Fraction ◽

Multidrug Resistant ◽

Global Public Health ◽

Minimal Inhibitory Concentrations ◽

Subinhibitory Concentrations

Background: Microbial resistance to antibiotics is a global public health problem, which requires urgent attention. Platonia insignis is a native species from the eastern Brazilian Amazon, used in the treatment of burns and wounds. Objectives: To evaluate the antimicrobial activity of the hydroalcoholic extract of P. insignis (PIHA), the ethyl acetate fraction (PIAE), and its subfraction containing a mixture of biflavonoids (BF). Moreover, the effect of these natural products on the antibiotic activity against S. aureus strains overexpressing efflux pump genes was also evaluated. Methods: Minimal inhibitory concentrations were determined against different species of microorganisms. To evaluate the modulatory effect on the Norfloxacin-resistance, the MIC of this antibiotic was determined in the absence and presence of the natural products at subinhibitory concentrations. Inhibition of the EtBr efflux assays were conducted in the absence or presence of the natural products. Results: PIHA showed a microbicidal effect against S. aureus and C. albicans, while PIAE was bacteriosctatic for S. aureus. PIAE and BF at subinhibitory concentrations were able to reduce the MIC of Norfloxacin acting as modulating agents. BF was able to inhibit the efflux of EtBr efflux in S. aureus strains overexpressing specific efflux pump genes. Conclusion: P. inignisis a source of efflux pump inhibitors, including volkensiflavone and morelloflavone which were able to potentiate the Norfloxacin activity by NorA inhibition, being also able to inhibit QacA/B, TetK and MsrA. Volkensiflavone and morelloflavone could be used as adjuvant in the antibiotic therapy of multidrug resistant S. aureus strains overexpressing efflux pumps.

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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.

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Molecular Epidemiology and Characterization of Carbapenem-Resistant Klebsiella pneumoniae Isolated from Urine at a Teaching Hospital in Taiwan

Microorganisms ◽

10.3390/microorganisms9020271 ◽

2021 ◽

Vol 9 (2) ◽

pp. 271

Author(s):

Yuarn-Jang Lee ◽

Chih-Hung Huang ◽

Noor Andryan Ilsan ◽

I-Hui Lee ◽

Tzu-Wen Huang

Keyword(s):

Klebsiella Pneumoniae ◽

Teaching Hospital ◽

Efflux Pump ◽

Resistance Mechanisms ◽

Pcr Analysis ◽

Carbapenem Resistant ◽

High Prevalence ◽

Antimicrobial Susceptibility Tests ◽

Susceptibility Tests ◽

Tract Infections

Urinary tract infections (UTIs) are common in clinics and hospitals and are associated with a high economic burden. Enterobacterium Klebsiella pneumoniae is a prevalent agent causing UTIs. A high prevalence of carbapenem-resistant K. pneumoniae (CRKP) has emerged recently and is continuing to increase. Seventeen urinary CRKP isolates collected at a teaching hospital in Taiwan from December 2016 to September 2017 were analyzed to elucidate their drug resistance mechanisms. Two-thirds of the isolates were obtained from outpatients. Antimicrobial susceptibility tests demonstrated multidrug resistance in all the isolates. Multilocus sequence typing analysis showed high diversity among the isolates. PCR analysis demonstrated the presence of carbapenemases in three isolates. All isolates carried at least one other extended-spectrum β-lactamase, including TEM, DHA, and CTX-M. Fifteen isolates contained mutations in one of the outer membrane porins that were assessed. The expression levels of the acrB and/or oqxB efflux pump genes, as determined by qRT-PCR, were upregulated in 11 isolates. Six isolates might have utilized other efflux pumps or antimicrobial resistance mechanisms. These analyses demonstrated a highly diverse population and the presence of complex resistance mechanisms in urinary isolates of K. pneumoniae.

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