scholarly journals Formation in vitro of colistin resistance in carbapenem-resistant Gram-negative bacteria and its biological cost

2021 ◽  
Author(s):  
D. V. Tapalski ◽  
T. A. Petrovskaya ◽  
A. E. Kozlov
Keyword(s):  
Broth Culture ◽  
Exponential Growth Phase ◽  
Lag Phase ◽  
Gram Negative Bacteria ◽  
Colistin Resistance ◽  
Gram Negative ◽  
Broth Microdilution Method ◽  
Resistant Mutants ◽  
Biological Cost

Introduction. The spread of resistance to carbapenems among gram-negative bacteria have led to an increase in the consumption of polymyxins and the emergence of certain strains resistant to them. Polymyxin resistance is mainly associated with mutations in chromosomal genes. The development of mutational resistance to antibiotics can lead to a decrease in the viability of bacteria, which is manifested by an increase in the duration of the cell cycle, a decrease in virulence and competitive fitness. The purpose of the study was to assess in vitro the intensity of the formation of colistin resistance in carbapenemresistant clinical isolates of gram-negative bacteria, the stability of the formed emerged resistance and its biological cost.Materials and methods. For 46 strains of Klebsiella pneumoniae, 77 strains of Pseudomonas aeruginosa and 42 strains of Acinetobacter baumannii, real time polymerase chain reaction (PCR) was used to detect the genes of carbapenemases, the minimum inhibitory concentrations (MIC) of meropenem and colistin were determined by broth microdilution method. The selection of resistant subpopulations on Muller–Hinton agar with the addition of 16 mg/l colistin was carried out. For colistin-resistant mutants and their isogenic sensitive strains, the kinetic parameters of growth in broth culture were determined. Incubation and result recording were performed on an Infinite M200 microplate reader for 18.5 hours at 35°C with measurement of light scatter in the wells every 15 minutes.Results. The production of carbapenemases MBL VIM in P. aeruginosa, MBL NDM, KPC and OXA-48 in K. pneumoniae, OXA-23 and OXA-40 in A. baumannii was observed. All strains were sensitive to colistin (MIC varied from 0.062 to 2 mg/l). The colony growth on a selective medium with16 mg/l colistin was observed for 97.8% of K. pneumoniae strains, 16.9% of P. aeruginosa strains, and 61.9% of A. baumannii strains. The mutational nature of colistin resistance was confirmed for 21.7% of K. pneumoniae strains. For colistin-resistant mutants of K. pneumoniae, a significant increase in the duration of the lag phase (Tlag) was observed: 225.6 ± 7.037 min in the wild-type susceptible strains and 245.5 ± 8.726 in resistant mutants, p = 0.037. The indicators of the doubling time of the number of microbial cells in the exponential growth phase (Tdoubling) and the area under the bacterial growth curve did not differ significantly.Conclusion. A high frequency of formation of colistin resistance in vitro in carbapenemase-producing strains of K. pneumoniae was observed. The absence of significant changes in the kinetics of microbial growth in resistant strains makes it possible to predict the further spread of mutational resistance to colistin, as well as its preservation in microbial populations of K. pneumoniae even in the case of limiting the use of this antibiotic. 

scholarly journals Susceptibility trends of ceftolozane/tazobactam and comparators when tested against European Gram-negative bacterial surveillance isolates collected during 2012–18

2020 ◽  
Vol 75 (10) ◽  
pp. 2907-2913 ◽  
Author(s):  
Helio S Sader ◽  
Cecilia G Carvalhaes ◽  
Leonard R Duncan ◽  
Robert K Flamm ◽  
Dee Shortridge
Keyword(s):  
Eastern Europe ◽  
Active Compound ◽  
Western Europe ◽  
Antimicrobial Agents ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Microdilution Method ◽  
Broth Microdilution Method ◽  
Systemic Infections

Abstract Background The Program to Assess Ceftolozane/Tazobactam Susceptibility (PACTS) monitors the in vitro activity of ceftolozane/tazobactam and numerous antimicrobial agents against Gram-negative bacteria worldwide. Objectives To evaluate the activity of ceftolozane/tazobactam and resistance trends among Pseudomonas aeruginosa and Enterobacterales isolates in Europe between 2012 and 2018. Methods P. aeruginosa (7503) and Enterobacterales (30 582) isolates were collected from 53 medical centres in 26 countries in Europe and the Mediterranean region and tested for susceptibility by reference broth microdilution method in a central laboratory. MIC results were interpreted using EUCAST criteria. Results Ceftolozane/tazobactam was the most active compound tested against P. aeruginosa isolates after colistin, with overall susceptibility rates of 94.1% in Western Europe and 80.9% in Eastern Europe. Moreover, ceftolozane/tazobactam retained activity against 75.2% and 59.2% of meropenem-non-susceptible P. aeruginosa isolates in Western and Eastern Europe, respectively. Tobramycin was the third most active compound tested against P. aeruginosa, with susceptibility rates of 88.6% and 70.9% in Western and Eastern Europe, respectively. Ceftolozane/tazobactam was active against 94.5% of all Enterobacterales and 96.1% of meropenem-susceptible isolates from Western Europe. In Eastern Europe, ceftolozane/tazobactam was active against 79.4% of Enterobacterales overall and 86.2% of meropenem-susceptible isolates. Discussion Antimicrobial susceptibility rates for agents commonly used to treat serious systemic infections varied widely among nations and geographic regions and were generally lower in Eastern Europe compared with Western Europe. Ceftolozane/tazobactam demonstrated potent activity against P. aeruginosa, including MDR strains, and retained activity against most meropenem-susceptible Enterobacterales causing infection in European medical centres.

scholarly journals Combining Colistin with Furanone C-30 Rescues Colistin Resistance of Gram-Negative Bacteria in Vitro and in Vivo

2021 ◽  
Author(s):  
Ying Zhang ◽  
Yishuai Lin ◽  
Xiaodong Zhang ◽  
Liqiong Chen ◽  
Chunyan Xu ◽  
...  
Keyword(s):  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Gram Negative Bacteria ◽  
Colistin Resistance ◽  
Gram Negative ◽  
C 30

Colistin is among the few antibiotics effective against multidrug-resistant Gram-negative bacteria (GNB) clinical isolates. However, colistin-resistant GNB strains have emerged in recent years.

scholarly journals In vitro evolution of colistin resistance in the Klebsiella pneumoniae complex follows multiple evolutionary trajectories with variable effects on fitness and virulence characteristics

2020 ◽  
Author(s):  
Axel B. Janssen ◽  
Dennis J. Doorduijn ◽  
Grant Mills ◽  
Malbert R.C. Rogers ◽  
Marc J.M. Bonten ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Lipid A ◽  
Multidrug Resistant ◽  
Sensu Stricto ◽  
Cross Resistance ◽  
Gram Negative Bacteria ◽  
Colistin Resistance ◽  
Gram Negative ◽  
Evolutionary Trajectories

AbstractThe increasing prevalence of multidrug-resistant Gram-negative opportunistic pathogens, including Klebsiella pneumoniae, has led to a resurgence in the use of colistin as a last-resort drug. Colistin is a cationic lipopeptide antibiotic that selectively acts on Gram-negative bacteria through electrostatic interactions with anionic phosphate groups of the lipid A moiety of lipopolysaccharides (LPS). Colistin resistance in K. pneumoniae is mediated through loss of these phosphate groups, or modification with cationic groups (e.g. 4-amino-4-deoxy-L-arabinose (L-Ara4N), or phosphoethanolamine), but also hydroxylation of acyl-groups of lipid A. Here, we study the in vitro evolutionary trajectories towards colistin resistance in clinical K. pneumoniae complex strains (three K. pneumoniae sensu stricto strains and one K. variicola subsp. variicola strain) and their impact on fitness and virulence characteristics.Through population sequencing during the in vitro evolution experiment, we found that resistance develops through a combination of single nucleotide polymorphisms (SNPs), insertion and deletions (indels), and the integration of insertion sequence (IS) elements, affecting genes associated with LPS biosynthesis and modification, and capsule structures. The development of colistin resistance decreased the maximum growth rate of one K. pneumoniae sensu stricto strain, but not in the other three K. pneumoniae sensu lato strains. Colistin-resistant strains had lipid A modified through hydroxylation, palmitoylation, and L-Ara4N addition. Colistin-resistant K. pneumoniae sensu stricto strains exhibited cross-resistance to LL-37, in contrast to the K. variicola subsp. variicola strain that did not change in susceptibility to LL-37. Virulence, as determined in a Caenorhabditis elegans survival assay, was higher in two colistin-resistant strains.Our study suggests that nosocomial K. pneumoniae complex strains can rapidly develop colistin resistance de novo through diverse evolutionary trajectories upon exposure to colistin. This effectively shortens the lifespan of this last-resort antibiotic for the treatment of infections with multidrug-resistant Klebsiella.Author summaryBacteria that frequently cause infections in hospitalised patients are becoming increasingly resistant to antibiotics. Colistin is a positively charged antibiotic that is used for the treatment of infections with multidrug-resistant Gram-negative bacteria. Colistin acts by specifically interacting with the negatively charged LPS molecule in the outer membrane of Gram-negative bacteria. Colistin resistance is mostly mediated through modification of LPS to reduce its negative charge. Here, we use a laboratory evolution experiment to show that strains belonging to the Klebsiella pneumoniae complex, a common cause of multidrug-resistant hospital-acquired infections, can rapidly accumulate mutations that reduce the negative charge of LPS without an appreciable loss of fitness. Colistin resistance can lead to cross-resistance to an antimicrobial peptide of the human innate immune system, but can increase susceptibility to serum, and virulence in a nematode model. These findings show that extensively resistant K. pneumoniae complex strains may rapidly develop resistance to the last-resort antibiotic colistin via different evolutionary trajectories, while retaining their ability to cause infections.

Antimicrobial activity of POL7306 tested against clinical isolates of Gram-negative bacteria collected worldwide

2020 ◽  
Vol 75 (6) ◽  
pp. 1518-1524 ◽  
Author(s):  
Helio S Sader ◽  
Paul R Rhomberg ◽  
Leonard R Duncan ◽  
Hans H Locher ◽  
Glenn E Dale ◽  
...  
Keyword(s):  
Protein Targeting ◽  
Treatment Options ◽  
Asia Pacific ◽  
Surveillance Program ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Microdilution Method ◽  
Broth Microdilution Method ◽  
The Usa

Abstract Background POL7306 belongs to a new class of peptidomimetic outer-membrane-protein-targeting antibiotics with a novel mechanism of action. POL7306 is in development for the treatment of infections caused by antimicrobial-resistant Gram-negative bacteria and has demonstrated low cytotoxicity and nephrotoxicity. Methods A total of 891 isolates were collected by the SENTRY Antimicrobial Surveillance Program from 134 medical centres in Europe (n = 424; 41 centres in 18 nations), the USA (n = 411 isolates from 67 centres), the Asia-Pacific region (n = 24; 15 centres in 6 nations) and Latin America (n = 32; 11 centres in 9 nations) and included 558 Enterobacterales, 310 non-fermenters and 23 fastidious organisms. Susceptibility testing was performed using the reference broth microdilution method and the medium was supplemented with 0.002% polysorbate-80 for testing POL7306. Resistant subsets were characterized by WGS. Results POL7306 demonstrated potent in vitro activity against Enterobacterales [including carbapenem-resistant (MIC50/90, 0.06/0.25 mg/L), ESBL-producing (MIC50/90, 0.06/0.12 mg/L), KPC-producing (MIC50/90, 0.12/0.25 mg/L), MBL-producing (MIC50/90, 0.06/0.25 mg/L), colistin-non-susceptible, mcr-negative (MIC50/90, 0.5/2 mg/L) and mcr-positive (MIC50/90, 0.12/0.25 mg/L) Enterobacterales], Pseudomonas aeruginosa (MIC50/90, 0.25/0.25 mg/L), Acinetobacter baumannii (MIC50/90, 0.06/0.12 mg/L) and Stenotrophomonas maltophilia (MIC50/90, 0.06/0.25 mg/L). Conclusions POL7306 demonstrated potent activity against a large collection of Gram-negative organisms collected worldwide that included colistin-resistant, XDR and ESBL- and carbapenemase-producing isolates for which there are currently limited treatment options.

scholarly journals Evaluation of resazurin-based assay for rapid detection of polymyxin-resistant gram-negative bacteria

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Huaiyu Jia ◽  
Renchi Fang ◽  
Jie Lin ◽  
Xuebin Tian ◽  
Yajie Zhao ◽  
...  
Keyword(s):  
Rapid Detection ◽  
Visual Detection ◽  
Detection Method ◽  
Gram Negative Bacteria ◽  
Broth Microdilution ◽  
Colistin Resistance ◽  
Acinetobacter Baumanii ◽  
Gram Negative ◽  
Microdilution Method ◽  
Broth Microdilution Method

Abstract Background Colistin resistance is considered a serious problem due to a lack of alternative antibiotics. The Rapid ResaPolymyxin Acinetobacter/Pseudomonas NP test is a resazurin reduction-based technique that relies on the visual detection of bacterial growth in the presence of a defined concentration of colistin. The aim of this study was to evaluate the performance of the Rapid ResaPolymyxin Acinetobacter/Pseudomonas NP test in the detection of colistin susceptibility in common clinical Gram-negative bacteria. Results A total of 253 clinical isolates from a teaching hospital, including Acinetobacter baumanii (n = 58, 8 colistin-resistant), Pseudomonas aeruginosa (n = 61, 11 colistin-resistant), Klebsiella pneumoniae (n = 70, 20 colistin-resistant) and Escherichia coli (n = 64, 14 colistin-resistant) were tested in this study. The sensitivity and specificity of the Rapid ResaPolymyxin Acinetobacter/Pseudomonas NP test compared to Broth microdilution method was 100 and 99%, respectively. Conclusions Our results suggest that Rapid ResaPolymyxin Acinetobacter/Pseudomonas NP test could be used as an accurate detection method for colistin resistance.

scholarly journals Synergistic activity of colistin combined with PFK-158 against colistin-resistant and colistin-susceptible Gram-negative bacteria

2020 ◽  
Author(s):  
Liqiong Chen ◽  
Kaihang Yu ◽  
Lijiang Chen ◽  
Xiangkuo Zheng ◽  
Na Huang ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Antimicrobial Agents ◽  
Gram Negative Bacteria ◽  
Synergistic Activity ◽  
Gram Negative ◽  
Broth Microdilution Method ◽  
Fermenting Bacteria ◽  
Checkerboard Assay ◽  
Experimental Strains

Abstract Background: The emergence of colistin resistance among Gram-negative bacteria poses a serious public health threat and warrants immediate action. Therefore, it is necessary to enhance the antibacterial activity of colistin through the combination with other drugs. In this study, we demonstrate the synergistic activity of colistin combined with PFK-158 against colistin-susceptible but more importantly against colistin-resistant Gram-negative bacteria, including non-fermenting bacteria (P. aeruginosa, A. baumannii) and Enterobacteriaceae (E. coli and K. pneumoniae).Methods: 18 colistin-resistant and 12 colistin-susceptible Gram-negative bacteria were collected as the experimental strains, and the minimum inhibitory concentrations (MICs) of colistin and PFK-158 against all strains were determined by the broth microdilution method. The MICs of routine antimicrobial agents including aztreonam (ATM), ceftazidime (CAZ), cefepime (FEP), imipenem (IMP), ciprofloxacin (CIP), levofloxacin (LVX), gentamicin (GEN), tobramycin (TOB) for all 30 experimental strains were determined by bioMerieux VITEK-2 (BioMérieux, Marcy-l’Étoile, France). The synergistic activity of colistin combined with PFK-158 in vitro was assessed using the checkerboard assay and the time-kill assays.Results: The results of the checkerboard assay showed that when colistin was used in combination with PFK-158, synergistic activity was observed against the 18 colistin-resistant and the 8 colistin-susceptible Gram-negative bacteria, and the remaining 4 colistin-susceptible strains showed additive activity. No irrelevant activity and antagonistic activity was observed for all strains. The results of the time-killing assays presented that the killing activity against the colistin-resistant Gram-negative bacterium were evident for the combination of colistin and PFK-158, compared with the groups adding colistin or PFK-158 alone. Conclusions: In conclusion, our results strongly exhibited that the combination of colistin and PFK-158 displayed the significant synergistic activity against all tested colistin-resistant and most colistin-susceptible Gram-negative strains. PFK-158 was found to potentiate the antibacterial activity of colistin against a wide panel of colistin-resistant and colistin-susceptible Gram-negative strains no matter what species (including non-fermenting bacteria and Enterobacteriaceae). It may be a potential new choice for the treatment of infections caused by the clinical Gram-negative strains.

scholarly journals Adjunctive transferrin to reduce the emergence of antibiotic resistance in Gram-negative bacteria

2019 ◽  
Vol 74 (9) ◽  
pp. 2631-2639 ◽  
Author(s):  
Brian M Luna ◽  
Ksenia Ershova ◽  
Jun Yan ◽  
Amber Ulhaq ◽  
Travis B Nielsen ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Bacterial Pathogens ◽  
Microbial Pathogens ◽  
Gram Negative Bacteria ◽  
Antibiotic Resistant ◽  
Gram Negative ◽  
Resistant Mutants ◽  
Time Kill ◽  
Emergence Of Resistance

AbstractBackgroundNew strategies are needed to slow the emergence of antibiotic resistance among bacterial pathogens. In particular, society is experiencing a crisis of antibiotic-resistant infections caused by Gram-negative bacterial pathogens and novel therapeutics are desperately needed to combat such diseases. Acquisition of iron from the host is a nearly universal requirement for microbial pathogens—including Gram-negative bacteria—to cause infection. We have previously reported that apo-transferrin (lacking iron) can inhibit the growth of Staphylococcus aureus in culture and diminish emergence of resistance to rifampicin.ObjectivesTo define the potential of apo-transferrin to inhibit in vitro growth of Klebsiella pneumoniae and Acinetobacter baumannii, key Gram-negative pathogens, and to reduce emergence of resistance to antibiotics.MethodsThe efficacy of apo-transferrin alone or in combination with meropenem or ciprofloxacin against K. pneumoniae and A. baumannii clinical isolates was tested by MIC assay, time–kill assay and assays for the selection of resistant mutants.ResultsWe confirmed that apo-transferrin had detectable MICs for all strains tested of both pathogens. Apo-transferrin mediated an additive antimicrobial effect for both antibiotics against multiple strains in time–kill assays. Finally, adding apo-transferrin to ciprofloxacin or meropenem reduced the emergence of resistant mutants during 20 day serial passaging of both species.ConclusionsThese results suggest that apo-transferrin may have promise to suppress the emergence of antibiotic-resistant mutants when treating infections caused by Gram-negative bacteria.

scholarly journals Preservation of Acquired Colistin Resistance in Gram-Negative Bacteria

2015 ◽  
Vol 60 (1) ◽  
pp. 609-612 ◽  
Author(s):  
Ji-Young Lee ◽  
Myung-Jin Choi ◽  
Hyeon Jin Choi ◽  
Kwan Soo Ko
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Gram Negative Bacteria ◽  
Free Medium ◽  
Colistin Resistance ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Resistant Mutants ◽  
The Stability

ABSTRACTColistin-resistant mutants were obtained from 17 colistin-susceptible strains ofAcinetobacter baumannii,Pseudomonas aeruginosa,Klebsiella pneumoniae, andEscherichia coli. The stability of colistin resistance in these mutants was investigated. Three of four colistin-resistantP. aeruginosamutants recovered colistin susceptibility in colistin-free medium; however, colistin-susceptible revertants were obtained from only one strain each ofA. baumanniiandE. coli. No susceptible revertants were obtained fromK. pneumoniaemutants.

Synthesis, In Vitro and In Silico Studies of Indolequinone Derivatives against Clinically Relevant Bacterial Pathogens

2020 ◽  
Vol 20 (3) ◽  
pp. 192-208 ◽  
Author(s):  
Talita Odriane Custodio Leite ◽  
Juliana Silva Novais ◽  
Beatriz Lima Cosenza de Carvalho ◽  
Vitor Francisco Ferreira ◽  
Leonardo Alves Miceli ◽  
...  
Keyword(s):  
In Silico ◽  
Bacterial Infections ◽  
Antimicrobial Agents ◽  
Mass Spectrometry Analysis ◽  
World Health ◽  
Gram Negative Bacteria ◽  
Gram Negative ◽  
Dione Derivative ◽  
In Silico Studies

Background: According to the World Health Organization, antimicrobial resistance is one of the most important public health threats of the 21st century. Therefore, there is an urgent need for the development of antimicrobial agents with new mechanism of action, especially those capable of evading known resistance mechanisms. Objective: We described the synthesis, in vitro antimicrobial evaluation, and in silico analysis of a series of 1H-indole-4,7-dione derivatives. Methods: The new series of 1H-indole-4,7-diones was prepared with good yield by using a copper(II)- mediated reaction between bromoquinone and β-enamino ketones bearing alkyl or phenyl groups attached to the nitrogen atom. The antimicrobial potential of indole derivatives was assessed. Molecular docking studies were also performed using AutoDock 4.2 for Windows. Characterization of all compounds was confirmed by one- and two-dimensional NMR techniques 1H and 13C NMR spectra [1H, 13C – APT, 1H x 1H – COSY, HSQC and HMBC], IR and mass spectrometry analysis. Results: Several indolequinone compounds showed effective antimicrobial profile against Grampositive (MIC = 16 µg.mL-1) and Gram-negative bacteria (MIC = 8 µg.mL-1) similar to antimicrobials current on the market. The 3-acetyl-1-(2,5-dimethylphenyl)-1H-indole-4,7-dione derivative exhibited an important effect against different biofilm stages formed by a serious hospital life-threatening resistant strain of Methicillin-Resistant Staphylococcus aureus (MRSA). A hemocompatibility profile analysis based on in vitro hemolysis assays revealed the low toxicity effects of this new series. Indeed, in silico studies showed a good pharmacokinetics and toxicological profiles for all indolequinone derivatives, reinforcing their feasibility to display a promising oral bioavailability. An elucidation of the promising indolequinone derivatives binding mode was achieved, showing interactions with important sites to biological activity of S. aureus DNA gyrase. These results highlighted 3-acetyl-1-(2-hydroxyethyl)-1Hindole- 4,7-dione derivative as broad-spectrum antimicrobial prototype to be further explored for treating bacterial infections. Conclusion: The highly substituted indolequinones were obtained in moderate to good yields. The pharmacological study indicated that these compounds should be exploited in the search for a leading substance in a project aimed at obtaining new antimicrobials effective against Gram-negative bacteria.

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