Susceptibility of Colistin-Resistant, Gram-Negative Bacteria to Antimicrobial Peptides and Ceragenins

ABSTRACT The susceptibility of colistin-resistant clinical isolates of Klebsiella pneumoniae to ceragenins and antimicrobial peptides (AMPs) suggests that there is little to no cross-resistance between colistin and ceragenins/AMPs and that lipid A modifications are found in bacteria with modest changes in susceptibility to ceragenins and with high levels of resistance to colistin. These results suggest that there are differences in the resistance mechanisms to colistin and ceragenins/AMPs.

Download Full-text

Pathogenicity Genomic Island-Associated CrpP-Like Fluoroquinolone-Modifying Enzymes among Pseudomonas aeruginosa Clinical Isolates in Europe

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00489-20 ◽

2020 ◽

Vol 64 (7) ◽

Cited By ~ 1

Author(s):

José Manuel Ortiz de la Rosa ◽

Patrice Nordmann ◽

Laurent Poirel

Keyword(s):

Pseudomonas Aeruginosa ◽

Genomic Island ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Genomic Islands ◽

Clinical Isolates ◽

Gram Negative Bacteria ◽

Gram Negative ◽

Content Type ◽

Genes Encoding

ABSTRACT Many transferable quinolone resistance mechanisms have been identified in Gram-negative bacteria. The plasmid-encoded 65-amino-acid-long ciprofloxacin-modifying enzyme CrpP was recently identified in Pseudomonas aeruginosa isolates. We analyzed a collection of 100 clonally unrelated and multidrug-resistant P. aeruginosa clinical isolates, among which 46 were positive for crpP-like genes, encoding five CrpP variants conferring variable levels of reduced susceptibility to fluoroquinolones. These crpP-like genes were chromosomally located as part of pathogenicity genomic islands.

Download Full-text

A Molecular Perspective on Colistin and Klebsiella pneumoniae: Mode of Action, Resistance Genetics, and Phenotypic Susceptibility

Diagnostics ◽

10.3390/diagnostics11071165 ◽

2021 ◽

Vol 11 (7) ◽

pp. 1165

Author(s):

Rita Elias ◽

Aida Duarte ◽

João Perdigão

Keyword(s):

Klebsiella Pneumoniae ◽

Mode Of Action ◽

Lipid A ◽

Drug Susceptibility ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Drug Susceptibility Testing ◽

Gram Negative Bacteria ◽

Colistin Resistance ◽

Gram Negative

Klebsiella pneumoniae is a rod-shaped, encapsulated, Gram-negative bacteria associated with multiple nosocomial infections. Multidrug-resistant (MDR) K. pneumoniae strains have been increasing and the therapeutic options are increasingly limited. Colistin is a long-used, polycationic, heptapeptide that has regained attention due to its activity against Gram-negative bacteria, including the MDR K. pneumoniae strains. However, this antibiotic has a complex mode of action that is still under research along with numerous side-effects. The acquisition of colistin resistance is mainly associated with alteration of lipid A net charge through the addition of cationic groups synthesized by the gene products of a multi-genic regulatory network. Besides mutations in these chromosomal genes, colistin resistance can also be achieved through the acquisition of plasmid-encoded genes. Nevertheless, the diversity of molecular markers for colistin resistance along with some adverse colistin properties compromises the reliability of colistin-resistance monitorization methods. The present review is focused on the colistin action and molecular resistance mechanisms, along with specific limitations on drug susceptibility testing for K. pneumoniae.

Download Full-text

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

10.1101/2020.05.24.112334 ◽

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.

Download Full-text

National Surveillance of Antimicrobial Susceptibility of Bacteremic Gram-Negative Bacteria with Emphasis on Community-Acquired Resistant Isolates: Report from the 2019 Surveillance of Multicenter Antimicrobial Resistance in Taiwan (SMART)

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01089-20 ◽

2020 ◽

Vol 64 (10) ◽

Author(s):

Po-Yu Liu ◽

Yu-Lin Lee ◽

Min-Chi Lu ◽

Pei-Lan Shao ◽

Po-Liang Lu ◽

...

Keyword(s):

Escherichia Coli ◽

Pseudomonas Aeruginosa ◽

Antimicrobial Resistance ◽

Klebsiella Pneumoniae ◽

Gram Negative Bacteria ◽

National Surveillance ◽

Gram Negative ◽

Content Type ◽

E Coli ◽

Carbapenem Resistant

ABSTRACT A multicenter collection of bacteremic isolates of Escherichia coli (n = 423), Klebsiella pneumoniae (n = 372), Pseudomonas aeruginosa (n = 300), and Acinetobacter baumannii complex (n = 199) was analyzed for susceptibility. Xpert Carba-R assay and sequencing for mcr genes were performed for carbapenem- or colistin-resistant isolates. Nineteen (67.8%) carbapenem-resistant K. pneumoniae (n = 28) and one (20%) carbapenem-resistant E. coli (n = 5) isolate harbored blaKPC (n = 17), blaOXA-48 (n = 2), and blaVIM (n = 1) genes.

Download Full-text

Susceptibility of Multiresistant Gram-Negative Bacteria to Fosfomycin and Performance of Different Susceptibility Testing Methods

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02048-13 ◽

2013 ◽

Vol 58 (3) ◽

pp. 1763-1767 ◽

Cited By ~ 31

Author(s):

L. V. Perdigão-Neto ◽

M. S. Oliveira ◽

C. F. Rizek ◽

C. M. D. M. Carrilho ◽

S. F. Costa ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Klebsiella Pneumoniae ◽

Susceptibility Testing ◽

Gram Negative Bacteria ◽

Testing Methods ◽

Gram Negative ◽

Content Type ◽

Agar Dilution ◽

And Performance ◽

Systemic Infections

ABSTRACTFosfomycin may be a treatment option for multiresistant Gram-negative bacteria. This study compared susceptibility methods using 94 multiresistant clinical isolates. With agar dilution (AD), susceptibilities were 81%, 7%, 96%, and 100% (CLSI) and 0%, 0%, 96%, and 30% (EUCAST), respectively, forAcinetobacter baumannii,Pseudomonas aeruginosa,Klebsiella pneumoniae, andEnterobacterspp. Categorical agreement between Etest and AD forEnterobacteriaceaeandA. baumanniiwas ≥80%. Disk diffusion was adequate only forEnterobacter. CLSI criteria for urine may be adequate for systemic infections.

Download Full-text

Discovery of a Novel Class of Boron-Based Antibacterials with Activity against Gram-Negative Bacteria

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02058-12 ◽

2013 ◽

Vol 57 (3) ◽

pp. 1394-1403 ◽

Cited By ~ 102

Author(s):

Vincent Hernandez ◽

Thibaut Crépin ◽

Andrés Palencia ◽

Stephen Cusack ◽

Tsutomu Akama ◽

...

Keyword(s):

Resistance Mechanisms ◽

Trna Synthetase ◽

Gram Negative Bacteria ◽

Aminoacyl Trna Synthetase ◽

Gram Negative ◽

Content Type ◽

E Coli ◽

Drug Classes ◽

Medical Need ◽

Infection Models

ABSTRACTGram-negative bacteria cause approximately 70% of the infections in intensive care units. A growing number of bacterial isolates responsible for these infections are resistant to currently available antibiotics and to many in development. Most agents under development are modifications of existing drug classes, which only partially overcome existing resistance mechanisms. Therefore, new classes of Gram-negative antibacterials with truly novel modes of action are needed to circumvent these existing resistance mechanisms. We have previously identified a new a way to inhibit an aminoacyl-tRNA synthetase, leucyl-tRNA synthetase (LeuRS), in fungi via the oxaborole tRNA trapping (OBORT) mechanism. Herein, we show how we have modified the OBORT mechanism using a structure-guided approach to develop a new boron-based antibiotic class, the aminomethylbenzoxaboroles, which inhibit bacterial leucyl-tRNA synthetase and have activity against Gram-negative bacteria by largely evading the main efflux mechanisms inEscherichia coliandPseudomonas aeruginosa. The lead analogue, AN3365, is active against Gram-negative bacteria, includingEnterobacteriaceaebearing NDM-1 and KPC carbapenemases, as well asP. aeruginosa. This novel boron-based antibacterial, AN3365, has good mouse pharmacokinetics and was efficacious againstE. coliandP. aeruginosain murine thigh infection models, which suggest that this novel class of antibacterials has the potential to address this unmet medical need.

Download Full-text

Analysis of the Networks Controlling the Antimicrobial-Peptide-Dependent Induction of Klebsiella pneumoniae Virulence Factors

Infection and Immunity ◽

10.1128/iai.05226-11 ◽

2011 ◽

Vol 79 (9) ◽

pp. 3718-3732 ◽

Cited By ~ 66

Author(s):

Enrique Llobet ◽

Miguel A. Campos ◽

Paloma Giménez ◽

David Moranta ◽

José A. Bengoechea

Keyword(s):

Klebsiella Pneumoniae ◽

Defense Mechanisms ◽

Lipid A ◽

Multidrug Resistant ◽

Cross Resistance ◽

New Approach ◽

Content Type ◽

Signaling Systems ◽

Transcriptional Changes ◽

Initial Target

ABSTRACTAntimicrobial peptides (APs) impose a threat to the survival of pathogens, and it is reasonable to postulate that bacteria have developed strategies to counteract them. Polymyxins are becoming the last resort to treat infections caused by multidrug-resistant Gram-negative bacteria and, similar to APs, they interact with the anionic lipopolysaccharide. Given that polymyxins and APs share the initial target, it is possible that bacterial defense mechanisms against polymyxins will be also effective against host APs. We sought to determine whether exposure to polymyxin will increaseKlebsiella pneumoniaeresistance to host APs. Indeed, exposure ofK. pneumoniaeto polymyxin induces cross-resistance not only to polymyxin itself but also to APs present in the airways. Polymyxin treatment upregulates the expression of the capsule polysaccharide operon and the loci required to modify the lipid A with aminoarabinose and palmitate with a concomitant increase in capsule and lipid A species containing such modifications. Moreover, these surface changes contribute to APs resistance and also to polymyxin-induced cross-resistance to APs. Bacterial loads of lipid A mutants in trachea and lungs of intranasally infected mice were lower than those of wild-type strain. PhoPQ, PmrAB, and the Rcs system govern polymyxin-induced transcriptional changes, and there is a cross talk between PhoPQ and the Rcs system. Our findings support the notion thatKlebsiellaactivates a defense program against APs that is controlled by three signaling systems. Therapeutic strategies directed to prevent the activation of this program could be a new approach worth exploring to facilitate the clearance of the pathogen from the airways.

Download Full-text

Nonclonal Emergence of Colistin Resistance Associated with Mutations in the BasRS Two-Component System in Escherichia coli Bloodstream Isolates

mSphere ◽

10.1128/msphere.00143-20 ◽

2020 ◽

Vol 5 (2) ◽

Cited By ~ 2

Author(s):

Axel B. Janssen ◽

Toby L. Bartholomew ◽

Natalia P. Marciszewska ◽

Marc J. M. Bonten ◽

Rob J. L. Willems ◽

...

Keyword(s):

Escherichia Coli ◽

Lipid A ◽

Gram Negative Bacteria ◽

Colistin Resistance ◽

Anionic Lipid ◽

Gram Negative ◽

Content Type ◽

E Coli ◽

Two Component ◽

Resistant Strains

ABSTRACT Infections by multidrug-resistant Gram-negative bacteria are increasingly common, prompting the renewed interest in the use of colistin. Colistin specifically targets Gram-negative bacteria by interacting with the anionic lipid A moieties of lipopolysaccharides, leading to membrane destabilization and cell death. Here, we aimed to uncover the mechanisms of colistin resistance in nine colistin-resistant Escherichia coli strains and one Escherichia albertii strain. These were the only colistin-resistant strains of 1,140 bloodstream Escherichia isolates collected in a tertiary hospital over a 10-year period (2006 to 2015). Core-genome phylogenetic analysis showed that each patient was colonized by a unique strain, suggesting that colistin resistance was acquired independently in each strain. All colistin-resistant strains had lipid A that was modified with phosphoethanolamine. In addition, two E. coli strains had hepta-acylated lipid A species, containing an additional palmitate compared to the canonical hexa-acylated E. coli lipid A. One E. coli strain carried the mobile colistin resistance (mcr) gene mcr-1.1 on an IncX4-type plasmid. Through construction of chromosomal transgene integration mutants, we experimentally determined that mutations in basRS, encoding a two-component signal transduction system, contributed to colistin resistance in four strains. We confirmed these observations by reversing the mutations in basRS to the sequences found in reference strains, resulting in loss of colistin resistance. While the mcr genes have become a widely studied mechanism of colistin resistance in E. coli, sequence variation in basRS is another, potentially more prevalent but relatively underexplored, cause of colistin resistance in this important nosocomial pathogen. IMPORTANCE Multidrug resistance among Gram-negative bacteria has led to the use of colistin as a last-resort drug. The cationic colistin kills Gram-negative bacteria through electrostatic interaction with the anionic lipid A moiety of lipopolysaccharides. Due to increased use in clinical and agricultural settings, colistin resistance has recently started to emerge. In this study, we used a combination of whole-genome sequence analysis and experimental validation to characterize the mechanisms through which Escherichia coli strains from bloodstream infections can develop colistin resistance. We found no evidence of direct transfer of colistin-resistant isolates between patients. The lipid A of all isolates was modified by the addition of phosphoethanolamine. In four isolates, colistin resistance was experimentally verified to be caused by mutations in the basRS genes, encoding a two-component regulatory system. Our data show that chromosomal mutations are an important cause of colistin resistance among clinical E. coli isolates.

Download Full-text

Comparative Evaluation of Four Phenotypic Tests for Detection of Carbapenemase-Producing Gram-Negative Bacteria

Journal of Clinical Microbiology ◽

10.1128/jcm.01853-16 ◽

2016 ◽

Vol 55 (2) ◽

pp. 510-518 ◽

Cited By ~ 30

Author(s):

Audrey Noël ◽

Te-Din Huang ◽

Catherine Berhin ◽

Martin Hoebeke ◽

Warda Bouchahrouf ◽

...

Keyword(s):

Comparative Evaluation ◽

Resistance Mechanisms ◽

Gram Negative Bacteria ◽

Bacterial Isolates ◽

Excellent Performance ◽

Electrochemical Assay ◽

Gram Negative ◽

Content Type ◽

Screening Assays ◽

Phenotypic Tests

ABSTRACTFour screening assays aimed for rapid detection of carbapenemase production from Gram-negative bacterial isolates, i.e., the Neo-Rapid Carb kit (Rosco Diagnostica A/S), the Rapidec Carba NP test (bioMérieux SA), the β Carba test (Bio-Rad Laboratories N.V.), and a homemade electrochemical assay (BYG Carba test) were evaluated against a panel comprising 328 clinical isolates (Enterobacteriaceae[n= 198] and nonfermentative Gram-negative bacilli [n= 130]) with previously characterized resistance mechanisms to carbapenems. AmongEnterobacteriaceaeisolates, the BYG Carba test and the β Carba test showed excellent sensitivities (respectively, 100% and 97.3%) and specificities (respectively, 98.9% and 97.7%). The two other assays yielded poorer performances with sensitivity and specificity of 91.9% and 83.9% for the Rapidec Carba NP test and of 89.2% and 89.7% for the Neo-Rapid Carb kit, respectively. AmongPseudomonasspp., sensitivities and specificities ranged, respectively, from 87.3% to 92.7% and from 88.2% to 94.1%. Finally, all tests performed poorly againstAcinetobacterspp., with sensitivities and specificities, respectively, ranging from 27.3% to 75.8% and from 75 to 100%. Among commercially available assays, the β Carba test appeared to be the most convenient for routine use and showed the best overall performances, especially against OXA-48-like producers. The excellent performance of the BYG Carba test againstEnterobacteriaceaewas confirmed (100% sensitivity and 98.9% specificity).

Download Full-text

In VitroActivity of Ceftazidime-Avibactam against 338 Molecularly Characterized Gentamicin-Nonsusceptible Gram-Negative Clinical Isolates Obtained from Patients in Canadian Hospitals

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00364-15 ◽

2015 ◽

Vol 59 (6) ◽

pp. 3623-3626 ◽

Cited By ~ 6

Author(s):

Andrew J. Denisuik ◽

James A. Karlowsky ◽

Tyler Denisuik ◽

Wright W. Nichols ◽

Thomas A. Keating ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

16S Rrna ◽

Klebsiella Pneumoniae ◽

Gram Negative Bacteria ◽

Aminoglycoside Resistance ◽

Single Strain ◽

Gram Negative ◽

Content Type ◽

16S Rrna Methylase

ABSTRACTThe mechanism of aminoglycoside resistance among 338 gentamicin-nonsusceptible Gram-negative bacteria (207Enterobacteriaceaeand 131Pseudomonas aeruginosa) was assessed, and thein vitroactivity of ceftazidime-avibactam against these isolates was determined. Aminoglycoside-modifying enzymes were detected in 91.8% ofEnterobacteriaceaeand 13.7% ofP. aeruginosaisolates. A single strain ofKlebsiella pneumoniaeharbored a 16S rRNA methylase (ArmA). The ceftazidime-avibactam MIC90values were 0.5 μg/ml (MIC, ≤8 μg/ml for 100% of isolates) and 16 μg/ml (MIC, ≤8 μg/ml for 87.8% of isolates) against gentamicin-nonsusceptibleEnterobacteriaceaeandP. aeruginosaisolates, respectively.

Download Full-text