scholarly journals Transcriptomics reveals how minocycline-colistin synergy overcomes antibiotic resistance in multidrug-resistant Klebsiella pneumoniae

2021 ◽  
Author(s):  
Thea Brennan-Krohn ◽  
Alexandra Grote ◽  
Shade Rodriguez ◽  
James E Kirby ◽  
Ashlee M. Earl
Keyword(s):  
Klebsiella Pneumoniae ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
High Rate ◽  
Striking Similarity ◽  
Synergistic Activity ◽  
Transcriptional Profiles ◽  
Peptidoglycan Biosynthesis ◽  
Treatment Conditions ◽  
Polymyxin E

Multidrug resistant gram-negative bacteria are a rapidly growing public health threat, and the development of novel antimicrobials has failed to keep pace with their emergence. Synergistic combinations of individually ineffective drugs present a potential solution, yet little is understood about the mechanisms of most such combinations. Here, we show that the combination of colistin (polymyxin E) and minocycline has a high rate of synergy against colistin-resistant and minocycline-intermediate or -resistant strains of Klebsiella pneumoniae. Furthermore, using RNA-Seq, we characterized the transcriptional profiles of these strains when treated with the drugs individually and in combination. We found a striking similarity between the transcriptional profiles of bacteria treated with the combination of colistin and minocycline at individually subinhibitory concentrations and those of the same isolates treated with minocycline alone. We observed a similar pattern with the combination of polymyxin B nonapeptide (a polymyxin B analogue that lacks intrinsic antimicrobial activity) and minocycline. We also found that genes involved in polymyxin resistance and peptidoglycan biosynthesis showed significant differential gene expression in the different treatment conditions, suggesting possible mechanisms for the antibacterial activity observed in the combination. These findings suggest that the synergistic activity of this combination against bacteria resistant to each drug alone involves sublethal outer membrane disruption by colistin, which permits increased intracellular accumulation of minocycline.

scholarly journals PUNICA GRANATUM RIND EXTRACT: ANTIBIOTIC POTENTIATOR AND EFFLUX PUMP INHIBITOR OF MULTIDRUG RESISTANT KLEBSIELLA PNEUMONIAE CLINICAL ISOLATES

2017 ◽  
Vol 10 (3) ◽  
pp. 191 ◽  
Author(s):  
Zumaana Rafiq ◽  
Sreevidya Narasimhan ◽  
Magesh Haridoss ◽  
Rosy Vennila ◽  
Rama Vaidyanathan
Keyword(s):  
Klebsiella Pneumoniae ◽  
Efflux Pump ◽  
Ethanol Extract ◽  
Punica Granatum ◽  
Multidrug Resistant ◽  
Synergistic Activity ◽  
Sequential Fractionation ◽  
Efflux Pump Inhibitor ◽  
Fold Reduction ◽  
Significant Area

ABSTRACTObjective: With a rise in multidrug resistant (MDR) bacterial isolates, search for antibiotics or compounds that could act synergistically with themis a significant area of research. Efflux-mediated resistance, in particular, is a great hurdle that needs to be overcome. In an effort to identify suchsynergistic compounds and potential efflux pump inhibitors (EPI), we analyzed the rind of Punica granatum (pomegranate) against MDR clinicalKlebsiella pneumoniae isolates.Methods: Sequential fractionation of P. granatum rind ethanol (PGR) extract was carried out to obtain hexane, butanol and water fractions.Antibacterial activity of the plant extracts was confirmed, and synergistic interaction with antibiotics was determined by the checkerboard assay. Gaschromatography-mass spectrometry (GC-MS) analysis was performed to identify the phytochemical constituents of the hexane extract. To study EPIactivity of the extracts, norfloxacin accumulation assay was carried out.Results: PGR ethanol extract was found to have synergistic activity with ciprofloxacin, levofloxacin, ceftazidime, cefoxitin, meropenem, and gentamicinresulting in fold decrease of minimum inhibitory concentration (MIC) ranging from 2 to 32 fold. The hexane fraction was found to have maximumsynergistic activity resulting in a 32-fold reduction of ciprofloxacin MIC followed by butanol and water fractions. The PGR ethanol extract was alsofound to have efflux inhibition activity by the norfloxacin accumulation assay. Of the sequential fractions, the butanol fraction had maximum effluxinhibition activity.Conclusion: Therefore, our study shows that PGR extract can potentiate the effect of antibiotics on MDR bacteria, and the mode of action is likely tobe due to EPI.Keywords: Punica granatum rind, Pomegranate, Synergy with antibiotics, Multidrug resistant, Klebsiella pneumoniae, Efflux pump inhibition.

scholarly journals Efficacy of Antibiotic Combinations against Multidrug-Resistant Pseudomonas aeruginosa in Automated Time-Lapse Microscopy and Static Time-Kill Experiments

2020 ◽  
Vol 64 (6) ◽  
Author(s):  
Anna Olsson ◽  
Pikkei Wistrand-Yuen ◽  
Elisabet I. Nielsen ◽  
Lena E. Friberg ◽  
Linus Sandegren ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Time Lapse ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Positive Interactions ◽  
Synergistic Activity ◽  
Content Type ◽  
Time Lapse Microscopy ◽  
Time Kill ◽  
Antibiotic Combination

ABSTRACT Antibiotic combination therapy is used for severe infections caused by multidrug-resistant (MDR) Gram-negative bacteria, yet data regarding which combinations are most effective are lacking. This study aimed to evaluate the in vitro efficacy of polymyxin B in combination with 13 other antibiotics against four clinical strains of MDR Pseudomonas aeruginosa. We evaluated the interactions of polymyxin B in combination with amikacin, aztreonam, cefepime, chloramphenicol, ciprofloxacin, fosfomycin, linezolid, meropenem, minocycline, rifampin, temocillin, thiamphenicol, or trimethoprim by automated time-lapse microscopy using predefined cutoff values indicating inhibition of growth (≤106 CFU/ml) at 24 h. Promising combinations were subsequently evaluated in static time-kill experiments. All strains were intermediate or resistant to polymyxin B, antipseudomonal β-lactams, ciprofloxacin, and amikacin. Genes encoding β-lactamases (e.g., blaPAO and blaOXA-50) and mutations associated with permeability and efflux were detected in all strains. In the time-lapse microscopy experiments, positive interactions were found with 39 of 52 antibiotic combination/bacterial strain setups. Enhanced activity was found against all four strains with polymyxin B used in combination with aztreonam, cefepime, fosfomycin, minocycline, thiamphenicol, and trimethoprim. Time-kill experiments showed additive or synergistic activity with 27 of the 39 tested polymyxin B combinations, most frequently with aztreonam, cefepime, and meropenem. Positive interactions were frequently found with the tested combinations, against strains that harbored several resistance mechanisms to the single drugs, and with antibiotics that are normally not active against P. aeruginosa. Further study is needed to explore the clinical utility of these combinations.

scholarly journals Elucidating the Regulon of Multidrug Resistance Regulator RarA in Klebsiella pneumoniae

2013 ◽  
Vol 57 (4) ◽  
pp. 1603-1609 ◽  
Author(s):  
Shyamasree De Majumdar ◽  
Mark Veleba ◽  
Sarah Finn ◽  
Séamus Fanning ◽  
Thamarai Schneiders
Keyword(s):  
Klebsiella Pneumoniae ◽  
Cell Envelope ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Protein Synthesis Inhibitors ◽  
Content Type ◽  
Phenotypic Microarray ◽  
Genes Encoding ◽  
Clusters Of Orthologous Groups ◽  
Microarray Analyses

ABSTRACTRarA is an AraC-type regulator inKlebsiella pneumoniae, which, when overexpressed, confers a low-level multidrug-resistant (MDR) phenotype linked to the upregulation of both theacrABandoqxABefflux genes. IncreasedrarAexpression has also been shown to be integral in the development of tigecycline resistance in the absence oframAinK. pneumoniae. Given its phenotypic role in MDR, microarray analyses were performed to determine the RarA regulon. Transcriptome analysis was undertaken using strains Ecl8ΔrarA/pACrarA-2 (rarA-expressing construct) and Ecl8ΔrarA/pACYC184 (vector-only control) using bespoke microarray slides consisting of probes derived from the genomic sequences ofK. pneumoniaeMGH 78578 (NC_009648.1) and Kp342 (NC_011283.1). Our results show thatrarAoverexpression resulted in the differential expression of 66 genes (42 upregulated and 24 downregulated). Under the COG (clusters of orthologous groups) functional classification, the majority of affected genes belonged to the category of cell envelope biogenesis and posttranslational modification, along with genes encoding the previously uncharacterized transport proteins (e.g., KPN_03141,sdaCB, andleuE) and the porin OmpF. However, genes associated with energy production and conversion and amino acid transport/metabolism (e.g.,nuoA,narJ, andproWX) were found to be downregulated. Biolog phenotype analyses demonstrated thatrarAoverexpression confers enhanced growth of the overexpresser in the presence of several antibiotic classes (i.e., beta-lactams and fluoroquinolones), the antifungal/antiprotozoal compound clioquinol, disinfectants (8-hydroxyquinoline), protein synthesis inhibitors (i.e., minocycline and puromycin), membrane biogenesis agents (polymyxin B and amitriptyline), DNA synthesis (furaltadone), and the cytokinesis inhibitor (sanguinarine). Both our transcriptome and phenotypic microarray data support and extend the role of RarA in the MDR phenotype ofK. pneumoniae.

scholarly journals Polymyxin Triple Combinations against Polymyxin-Resistant, Multidrug-Resistant, KPC-Producing Klebsiella pneumoniae

2020 ◽  
Vol 64 (8) ◽  
Author(s):  
Su Mon Aye ◽  
Irene Galani ◽  
Heidi Yu ◽  
Jiping Wang ◽  
Ke Chen ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Triple Therapy ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Infection Model ◽  
Bacterial Killing ◽  
Standard Dosage ◽  
Time Kill

ABSTRACT Resistance to polymyxin antibiotics is increasing. Without new antibiotic classes, combination therapy is often required. We systematically investigated bacterial killing with polymyxin-based combinations against multidrug-resistant (including polymyxin-resistant), carbapenemase-producing Klebsiella pneumoniae. Monotherapies and double- and triple-combination therapies were compared to identify the most efficacious treatment using static time-kill studies (24 h, six isolates), an in vitro pharmacokinetic/pharmacodynamic model (IVM; 48 h, two isolates), and the mouse thigh infection model (24 h, six isolates). In static time-kill studies, all monotherapies (polymyxin B, rifampin, amikacin, meropenem, or minocycline) were ineffective. Initial bacterial killing was enhanced with various polymyxin B-containing double combinations; however, substantial regrowth occurred in most cases by 24 h. Most polymyxin B-containing triple combinations provided greater and more sustained killing than double combinations. Standard dosage regimens of polymyxin B (2.5 mg/kg of body weight/day), rifampin (600 mg every 12 h), and amikacin (7.5 mg/kg every 12 h) were simulated in the IVM. Against isolate ATH 16, no viable bacteria were detected across 5 to 25 h with triple therapy, with regrowth to ∼2-log10 CFU/ml occurring at 48 h. Against isolate BD 32, rapid initial killing of ∼3.5-log10 CFU/ml at 5 h was followed by a slow decline to ∼2-log10 CFU/ml at 48 h. In infected mice, polymyxin B monotherapy (60 mg/kg/day) generally was ineffective. With triple therapy (polymyxin B at 60 mg/kg/day, rifampin at 120 mg/kg/day, and amikacin at 300 mg/kg/day), at 24 h there was an ∼1.7-log10 CFU/thigh reduction compared to the starting inoculum for all six isolates. Our results demonstrate that the polymyxin B-rifampin-amikacin combination significantly enhanced in vitro and in vivo bacterial killing, providing important information for the optimization of polymyxin-based combinations in patients.

scholarly journals The polymyxin B-induced transcriptomic response of a clinical, multidrug-resistant Klebsiella pneumoniae involves multiple regulatory elements and intracellular targets

BMC Genomics ◽  
2016 ◽  
Vol 17 (S8) ◽  
Author(s):  
Pablo Ivan Pereira Ramos ◽  
Márlon Grégori Flores Custódio ◽  
Guadalupe del Rosario Quispe Saji ◽  
Thiago Cardoso ◽  
Gisele Lucchetti da Silva ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Polymyxin B ◽  
Regulatory Elements ◽  
Multidrug Resistant ◽  
Transcriptomic Response ◽  
Intracellular Targets

scholarly journals 2151. Biofilm and Metallo-β-Lactamase (MBL) Production Among Imipenem-Resistant Pseudomonas aeruginosa and Acinetobacter spp.

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S729-S730
Author(s):  
Yang Metok ◽  
Supram Hosuru Subramanya ◽  
Upendra Thapa Shrestha ◽  
Leandro Reus Rodrigues Perez ◽  
Nabaraj Adhikari ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Hospital Setting ◽  
Polymyxin B ◽  
Microtiter Plate ◽  
Multidrug Resistant ◽  
High Rate ◽  
Bacterial Strains ◽  
Microbiological Techniques ◽  
Acinetobacter Spp ◽  
Imipenem Resistance

Abstract Background Pseudomonas aeruginosa and Acinetobacter spp. head the list of hospital-acquired infections. Resistance to carbapenem as reserve drug is under threat with the emergence of Metallo-β-lactamase (MBL) and biofilm producing bacterial strains. This study was thus undertaken to determine the rate of MBL and biofilm production among imipenem-resistant P. aeruginosa (IRPA) and imipenem-resistant Acinetobacter spp. (IRAS) isolates. Methods A total of 79 P. aeruginosa and 117 Acinetobacter spp. were isolated from different clinical specimens of patients visiting Manipal Teaching Hospital, Pokhara Nepal from July 2016 to January 2017. Isolation, identification and antibiotic susceptibility testing of the isolates were performed by standard microbiological techniques. Combined disc test and Epsilometer test (E-test) were employed to detect MBL in IRPA and IRAS isolates. Microtiter plate using crystal violet method was employed for detection of biofilm in imipenem-resistant isolates. Results 9 (11.4%) of P. aeruginosa and 49 (41.9%) of Acinetobacter spp. were Multidrug Resistant (MDR). Similarly, 22 (27.8%) of P. aeruginosa and 23 (19.7%) of Acinetobacter spp. were Extensively Drug Resistant (XDR). Imipenem resistance was detected among 15 (19%) P. aeruginosa and 57 (48.7%) Acinetobacter spp. isolates. 8 (53.3%) of IRPA and 22 (38.6%) of IRAS isolates were MBL producers while all (100%) of IRPA and 47 (82.5%) of IRAS were biofilm producers. All the biofilm producer IRPA isolates were XDR and 62.5% of XDR IRAS strains were moderate biofilm producers. However, 80% of IRPA, 49.1% of IRAS and 63% of both MBL producer isolates were weak biofilm formers. Polymyxin B and ampicillin-sulbactam showed a better degree of susceptibility against MBL cum biofilm producer IRPA and IRAS isolates respectively. Conclusion The study showed high propensity of IRPA and IRAS to form biofilm, which is strongly associated with higher drug resistance. Such high rate of MBL and biofilm producing P. aeruginosa and Acinetobacter spp. alarms the rapid spread of such strains in our hospital setting. Disclosures All authors: No reported disclosures.

scholarly journals Antimicrobial Susceptibility Testing for Polymyxins: Challenges, Issues, and Recommendations

2018 ◽  
Vol 57 (4) ◽  
Author(s):  
Fereshteh Ezadi ◽  
Abdollah Ardebili ◽  
Reza Mirnejad
Keyword(s):  
Bacterial Infections ◽  
Susceptibility Testing ◽  
Agar Medium ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Professional Organizations ◽  
Gram Negative ◽  
Microtiter Plates ◽  
Critical Issues ◽  
Polymyxin E

ABSTRACTPolymyxins, including polymyxin B and polymyxin E (colistin), are now increasingly being used worldwide to treat patients with multidrug-resistant (MDR) Gram-negative bacterial infections. This necessitates that laboratories employ an accurate and reliable method for the routine performance of polymyxin susceptibility testing. A number of reasons have accounted for the difficulties with susceptibility testing for the polymyxins, including their multicomponent composition, poor diffusion in the agar medium, adsorption to microtiter plates, the lack of a reliable susceptibility test, the lack of a specific breakpoint from professional organizations, the synergistic effect of polysorbate 80, and the development of heteroresistance. This minireview discusses such problems that impact the results of currently available susceptibility testing methods. We also provide emerging concepts on mechanisms of polymyxin resistance, including chromosomally and plasmid-mediatedmcr-related resistance. Broad-range investigations on such critical issues in relation to polymyxins can be beneficial for the implementation of effective treatment against MDR Gram-negative bacterial infections.

scholarly journals mgrBMutations Mediating Polymyxin B Resistance in Klebsiella pneumoniae Isolates from Rectal Surveillance Swabs in Brazil

2016 ◽  
Vol 60 (11) ◽  
pp. 6969-6972 ◽  
Author(s):  
Caio Augusto Martins Aires ◽  
Polyana Silva Pereira ◽  
Marise Dutra Asensi ◽  
Ana Paula D'Alincourt Carvalho-Assef
Keyword(s):  
Klebsiella Pneumoniae ◽  
Molecular Mechanisms ◽  
Clonal Complex ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Resistant Bacteria ◽  
Insertion Sequences ◽  
Missense Mutations ◽  
Content Type ◽  
Rectal Swabs

ABSTRACTWe aimed to investigate polymyxin B (PMB) resistance and its molecular mechanisms in 126Klebsiella pneumoniaeisolates from rectal swabs in Brazil. Ten isolates exhibited PMB resistance with interruption ofmgrBgene by insertion sequences or missense mutations. Most of the PMB-resistant isolates harboredblaKPC-2(n= 8) and belonged to clonal complex 258 (CC258) (n= 7). These results highlight the importance of monitoring the spread of polymyxin-resistant bacteria in hospitals, since few options remain to treat multidrug-resistant isolates.

scholarly journals In VitroActivity of Polymyxin B plus Imipenem, Meropenem, or Tigecycline against KPC-2-Producing Enterobacteriaceae with High MICs for These Antimicrobials

2015 ◽  
Vol 59 (6) ◽  
pp. 3596-3597 ◽  
Author(s):  
Natália Barth ◽  
Vanessa B. Ribeiro ◽  
Alexandre P. Zavascki
Keyword(s):  
Klebsiella Pneumoniae ◽  
Serratia Marcescens ◽  
Antimicrobial Agents ◽  
Enterobacter Cloacae ◽  
Polymyxin B ◽  
Synergistic Activity ◽  
Content Type ◽  
Time Kill ◽  
High Mics

ABSTRACTWe evaluated thein vitroactivity of polymyxin B plus imipenem, meropenem, or tigecycline against six KPC-2-producingEnterobacteriaceaestrains with high MICs for these antimicrobial agents. Polymyxin B with carbapenems, especially meropenem, were the most active combinations forKlebsiella pneumoniaeandEnterobacter cloacaeregardless of the polymyxin B concentration used in the time-kill assay. This combination was also synergistic against twoSerratia marcescensstrains that are intrinsically resistant to polymyxins. Polymyxin B and tigecycline also presented synergistic activity in most experiments.

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