Synergistic Combination of AS101 and Azidothymidine against Clinical Isolates of Carbapenem-Resistant Klebsiella pneumoniae.
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ABSTRACT:
SUBMITTER: Sung CL
PROVIDER: S-EPMC8706163 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Carbapenem-resistant Enterobacteriaceae (CRE) is listed as an urgent threat by the World Health Organization because of the limited therapeutic options, rapid evolution of resistance mechanisms, and worldwide dissemination. Colistin is a common backbone agent among the "last-resort" antibiotics for CRE; however, its emerging resistance among CRE has taken the present dilemma to the next level. Azidothymidine (AZT), a thymidine analog used to treat human immunodeficiency virus/acquired immunodeficiency syndrome, has been known to possess antibacterial effects against Enterobacteriaceae. In this study, we investigated the combined effects of AZT and colistin in 40 clinical isolates of colistin-resistant, carbapenem-resistant K. pneumoniae (CCRKP). Eleven of the 40 isolates harbored Klebsiella pneumoniae carbapenemase. The in vitro checkerboard method and in vivo nematode killing assay both revealed synergistic activity between the two agents, with fractional inhibitory concentration indexes of ≤0.5 in every strain. Additionally, a significantly lower hazard ratio was observed for the nematodes treated with combination therapy (0.288; p < 0.0001) compared with either AZT or colistin treatment. Toxicity testing indicated potentially low toxicity of the combination therapy. Thus, the AZT-colistin combination could be a potentially favorable therapeutic option for treating CCRKP.
| S-EPMC7764370 | biostudies-literature
Project description:Klebsiella pneumoniae is an opportunistic pathogen causing nosocomial and community-acquired infections. Klebsiella has developed resistance against antimicrobials including the last resort class; carbapenem. Currently, treatment options for carbapenem-resistant-Klebsiella (CRK) are very limited. This study aims to restore carbapenem effectiveness against CRK using celastrol and thymol. Clinical Klebsiella isolates were identified using biochemical and molecular methods. Antimicrobial susceptibility was determined using disk-diffusion method. Carbapenemase-production was tested phenotypically and genotypically. Celastrol and thymol-MICs were determined and the carbapenemase-inhibitory effect of sub-MICs was investigated. Among 85 clinical Klebsiella isolates, 72 were multi-drug-resistant and 43 were meropenem-resistant. Phenotypically, 39 isolates were carbapenemase-producer. Genotypically, blaNDM1 was detected in 35 isolates, blaVIM in 17 isolates, blaOXA in 18 isolates, and blaKPC was detected only in 6 isolates. Celastrol showed significant inhibitory effect against carbapenemase-hydrolytic activity. Meropenem-MIC did not decrease in presence of celastrol, only 2-fold decrease was observed with thymol, while 4-64 fold decrease was observed when meropenem was combined with both celastrol and thymol. Furthermore, thymol increased CRK cell wall-permeability. Molecular docking revealed that celastrol is superior to thymol for binding to KPC and VIM-carbapenemase. Our study showed that celastrol is a promising inhibitor of multiple carbapenemases. While meropenem-MIC were not affected by celastrol alone and decreased by only 2-folds with thymol, it decreased by 4-64 folds in presence of both celastrol and thymol. Thymol increases the permeability of CRK-envelope to celastrol. The triple combination (meropenem/celastrol/thymol) could be useful for developing more safe and effective analogues to restore the activity of meropenem and other β-lactams.
| S-EPMC9640353 | biostudies-literature
Project description:Carbapenem-resistant Enterobacteriaceae (CRE), especially carbapenem-resistant Klebsiella pneumoniae (CRKP), are among the largest pathogenic threats to humans. The available antibiotic treatment options for combating CRKP are limited. Colistin-resistant Enterobacteriaceae (CoRE) have also been reported worldwide, including in Thailand. Therefore, this study aimed (1) to determine minimum inhibitory concentrations (MICs) and synergistic activities of antibiotics of CRKP, and (2) to determine the probability target of attainment (PTA) and cumulative fraction of response (CFR) using pharmacokinetic/pharmacodynamic (PK/PD) data. Clinical CRKP isolates were obtained from Phramongkutklao Hospital (June to November 2020). Broth microdilution and checkerboard techniques were used to determine the mono- and synergistic activities of antibiotics. Carbapenemase and mcr-1 genes were also identified by polymerase chain reaction (PCR). The optimal antibiotic regimens were evaluated using Monte Carlo simulations. Forty-nine CRKP isolates were collected, 40 of which were CoRKP strains. The MIC50 and MIC90 of tigecycline, amikacin, and gentamicin were 1 and 2 µg/mL, 4 and 16 µg/mL, and 0.25 and 4 µg/mL, respectively. None of any isolates expressed the mcr-1 gene, whereas blaOXA-48 (53.1%) and blaOXA-48 plus blaNDM (42.9%) were detected. Synergistic activity was observed in 8.2% of isolates for tigecycline combined with amikacin or gentamicin. Additive activity was observed in 75.5% of isolates for tigecycline-amikacin and 69.4% for tigecycline-gentamicin, and no antagonism was observed. High-dose antibiotic regimens achieved the PTA target. The general recommended dose of combination regimens began with 200 mg tigecycline and 25 mg/kg amikacin, or 7 mg/kg gentamicin, followed by 100 mg tigecycline every 12 h and 15 mg/kg amikacin or 5 mg/kg gentamicin every 24 h. In conclusion, tigecycline plus aminoglycosides might be a potential regimen against CRKP and CoRKP. The appropriate combination regimen based on MIC-based dose adjustment can improve optimal antibiotic dosing. Further research via clinical studies will be necessary to confirm these results.
| S-EPMC8234075 | biostudies-literature
Project description:Colistin- and carbapenem-resistant Enterobacteriaceae cases are increasing at alarming rates worldwide. Drug repurposing is receiving greater attention as an alternative approach in light of economic and technical barriers in antibiotics research. The immunomodulation agent ammonium trichloro(dioxoethylene-O,O'-)tellurate (AS101) was repurposed as an antimicrobial agent against colistin- and carbapenem-resistant Klebsiella pneumoniae (CRKP). 134 CRKP isolates were collected between 2012 and 2015 in Taiwan. The in vitro antibacterial activities of AS101 was observed through broth microdilution, time-kill assay, and electron microscopy. Pharmaceutical manipulation and RNA microarray were applied to investigate these antimicrobial mechanisms. Caenorhabditis elegans, a nematode animal model, and the Institute for Cancer Research (ICR) mouse model was employed for the evaluation of in vivo efficacy. The in vitro antibacterial results were found for AS101 against colistin- and CRKP isolates, with minimum inhibitory concentration (MIC) values ranging from <0.5 to 32 μg/mL. ROS-mediated antibacterial activity eliminated 99.9% of bacteria within 2-4 h. AS101 also extended the median survival time in a C. elegans animal model infected with a colistin-resistant CRKP isolate and rescued lethally infected animals in a separate mouse model of mono-bacterial sepsis by eliminating bacterial organ loads. These findings support the use of AS101 as an antimicrobial agent for addressing the colistin and carbapenem resistance crisis.
| S-EPMC8400984 | biostudies-literature
Project description:Co-administration of antibiotics with synergistic effects is one method to combat carbapenem-resistant organisms. Although the synergistic effects of tigecycline combined with aminoglycosides against carbapenem-resistant Klebsiella pneumoniae (CRKP) have been demonstrated in vitro and in animal models, the underlying mechanism remains elusive. Here we used proteomics analysis to assess the short-term bacterial responses to tigecycline and aminoglycosides alone or in combination. Emergence of tigecycline resistance during treatment and the susceptibility of tigecycline-resistant strains to aminoglycosides was further evaluated. The proteomic responses to tigecycline and aminoglycosides were divergent in monotherapy, with proteomic alterations to combination therapy dominated by tigecycline. Adaptive responses to tigecycline were associated with the upregulation of oxidative phosphorylation and translation-related proteins. These responses might confer CRKP hypersensitivity towards aminoglycosides by increasing the drug uptake and binding targets. Meanwhile, tigecycline might perturb adaptive responses to aminoglycosides through inhibition of heat shock response. Tigecycline-resistant strains could be isolated within 24 h exposure even in strains without heteroresistance, and the sensitivity to aminoglycosides significantly increased in resistant strains. Overall, these findings demonstrated that adaption to tigecycline in CRKP was a double-edged sword associated with the synergistic killing in tigecycline-aminoglycoside combination. Evolutionary hypersensitivity can provide novel insight into the mechanisms of antibiotic synergistic effects.
| S-EPMC9280366 | biostudies-literature
Project description:Objective:The emergence of carbapenem-resistant and hypervirulent Klebsiella pneumoniae (CR-hvKp) strains poses a significant public threat, and effective antimicrobial therapy is urgently needed. Recent studies indicated that apramycin is a potent antibiotic with good activity against a range of multi-drug resistant pathogens. In this study, we evaluated the in vitro activity of apramycin against clinical CR-hvKp along with carbapenem-resistant non-hvKp (CR-non-hvKp) isolates. Methods:Broth microdilution method was used to evaluate the in vitro activities of apramycin, gentamicin, amikacin, imipenem, meropenem, doripenem, ertapenem and other comparator "last-resort" antimicrobial agents, including ceftazidime-avibactam, colistin and tigecycline, against eighty-four CR-hvKp and forty CR-non-hvKp isolates collected from three Chinese hospitals. Multilocus Sequence typing (MLST), molecular capsule typing (wzi sequencing) and antimicrobial resistance genes were examined by PCR and Sanger sequencing. Pulsed-field gel electrophoresis and next generation sequencing were conducted on selected isolates. Results:Among the 84 CR-hvKp isolates, 97.6, 100, 97.6, and 100% were resistant to imipenem, meropenem, doripenem and ertapenem, respectively. Apramycin demonstrated an MIC50/MIC90 of 4/8 ?g/mL against the CR-hvKp isolates. In contrast, the MIC50/MIC90 for amikacin and gentamicin were >64/>64 ?g/mL. All CR-hvKp isolates were susceptible to ceftazidime-avibactam, colistin and tigecycline with the MIC50/MIC90 values of 0.5/1, 0.25/0.5, 1/1, respectively. For CR-non-hvKp, The MIC50/90 values for apramycin, gentamicin and amikacin were 2/8, >64/>64, and >64/>64 ?g/mL, respectively. There were no statistical significance in the resistance rates of antimicrobial agents between CR-hvKp and CR-non-hvKp groups (p > 0.05). Genetic analysis revealed that all CR-hvKp isolates harbored bla KPC-2, and 94% (n = 79) belong to the ST11 high-risk clone. 93.6% (44/47) of amikacin or gentamicin resistant strains carried 16S rRNA methyltransferases gene rmtB. Conclusion:Apramycin demonstrated potent in vitro activity against CR-hvKp isolates, including those were resistant to amikacin or gentamicin. Further studies are needed to evaluate the applicability of apramycin to be used as a therapeutic antibiotic against CR-hvKp infections.
| S-EPMC7083131 | biostudies-literature
Project description:The emergence and dissemination of carbapenemases, bacterial enzymes able to inactivate most β-lactam antibiotics, in Enterobacteriaceae is of increasing concern. The concurrent spread of resistance against colistin, an antibiotic of last resort, further compounds this challenge further. Whole-genome sequencing (WGS) can play a significant role in the rapid and accurate detection/characterization of existing and emergent resistance determinants, an essential aspect of public health surveillance and response activities to combat the spread of antimicrobial resistant bacteria. In the current study, WGS data was used to characterize the genomic content of antimicrobial resistance genes, including those encoding carbapenemases, in 10 multidrug-resistant Klebsiella pneumoniae isolates from Pakistan. These clinical isolates represented five sequence types: ST11 (n = 3 isolates), ST14 (n = 3), ST15 (n = 1), ST101 (n = 2), and ST307 (n = 1). Resistance profiles against 25 clinically-relevant antimicrobials were determined by broth microdilution; resistant phenotypes were observed for at least 15 of the 25 antibiotics tested in all isolates except one. Specifically, 8/10 isolates were carbapenem-resistant and 7/10 isolates were colistin-resistant. The blaNDM-1 and blaOXA-48 carbapenemase genes were present in 7/10 and 5/10 isolates, respectively; including 2 isolates carrying both genes. No plasmid-mediated determinants for colistin resistance (e.g. mcr) were detected, but disruptions and mutations in chromosomal loci (i.e. mgrB and pmrB) previously reported to confer colistin resistance were observed. A blaOXA-48-carrying IncL/M-type plasmid was found in all blaOXA-48-positive isolates. The application of WGS to molecular epidemiology and surveillance studies, as exemplified here, will provide both a more complete understanding of the global distribution of MDR isolates and a robust surveillance tool useful for detecting emerging threats to public health.
| S-EPMC5993281 | biostudies-literature
Project description:Bacterial infections remain a leading killer worldwide, which is worsened by the continuous emergence of antibiotic resistance. In particular, antibiotic-resistant Enterobacteriaceae are prevalent and extremely difficult to treat. Repurposing existing drugs and improving the therapeutic potential of existing antibiotics represent an attractive novel strategy. Azidothymidine (AZT) is an antiretroviral drug which is used in combination with other antivirals to prevent and to treat HIV/AIDS. AZT is also active against Gram-negative bacteria but has not been developed for that purpose. Here, we investigated the in vitro and in vivo efficacy of AZT in combination with colistin against antibiotic-resistant Enterobacteriaceae, including strains producing extended-spectrum beta-lactamases (ESBLs) or New Delhi metallo-beta-lactamase 1 (NDM) or carrying mobilized colistin resistance (mcr-1). The MIC was determined using the broth microdilution method. The combined effect of AZT and colistin was examined using the checkerboard method and time-kill analysis. A murine peritoneal infection model was used to test the therapeutic effect of the combination of AZT and colistin. The fractional inhibitory concentration index from the checkerboard assay demonstrated that AZT synergized with colistin against 61% and 87% of ESBL-producing Escherichia coli and Klebsiella pneumoniae strains, respectively, 100% of NDM-1-producing strains, and 92% of mcr-1-producing E. coli strains. Time-kill analysis demonstrated significant synergistic activities when AZT was combined with colistin. In a murine peritoneal infection model, AZT in combination with colistin showed augmented activities of both drugs in the treatment of NDM-1 K. pneumoniae and mcr-1 E. coli infections. The AZT and colistin combination possesses a potential to be used coherently to treat antibiotic-resistant Enterobacteriaceae infections.
| S-EPMC6325217 | biostudies-literature
Project description:The increasing incidence of carbapenemase-producing K. pneumoniae strains (CP-Kps) in the last decade has become a serious global healthcare problem. Therapeutic options for the treatment of emerging hospital clones have drastically narrowed and therefore novel approaches must be considered. Here we have isolated and characterized a lytic bacteriophage, named vB_KpnS_Kp13, that was effective against all Verona integron-encoded metallo-β-lactamase (VIM) producing K. pneumoniae isolates originating from hospital samples (urine, blood, sputum and faeces), belonging to the ST15 clonal lineage and expressing the K24 capsule. Morphological characterization of vB_KpnS_Kp13 showed that the newly identified phage belonged to the Siphoviridae family, and phylogenetic analysis showed that it is part of a distinct clade of the Tunavirinae subfamily. Functional analysis revealed that vB_KpnS_Kp13 had relatively short latent period times (18 minutes) compared to other K. pneumoniae bacteriophages and could degrade biofilm by more than 50% and 70% in 24 and 48 hours respectively. Complete in vivo rescue potential of the new phage was revealed in an intraperitoneal mouse model where phages were administered intraperitoneally 10 minutes after bacterial challenge. Our findings could potentially be used to develop specific anti-CP-Kps bacteriophage-based therapeutic strategies against major clonal lineages and serotypes.
| S-EPMC7125228 | biostudies-literature
Project description:Carbapenem-resistant Enterobacteriaceae (CRE) are resistant to most antibiotics, making CRE infections extremely difficult to treat with available agents. Klebsiella pneumoniae carbapenemases (KPC-2 and KPC-3) are predominant carbapenemases in CRE in the United States. Nacubactam is a bridged diazabicyclooctane (DBO) β-lactamase inhibitor that inactivates class A and C β-lactamases and exhibits intrinsic antibiotic and β-lactam "enhancer" activity against Enterobacteriaceae In this study, we examined a collection of meropenem-resistant K. pneumoniae isolates carrying blaKPC-2 or blaKPC-3; meropenem-nacubactam restored susceptibility. Upon testing isogenic Escherichia coli strains producing KPC-2 variants with single-residue substitutions at important Ambler class A positions (K73, S130, R164, E166, N170, D179, K234, E276, etc.), the K234R variant increased the meropenem-nacubactam MIC compared to that for the strain producing KPC-2, without increasing the meropenem MIC. Correspondingly, nacubactam inhibited KPC-2 (apparent Ki [Ki app] = 31 ± 3 μM) more efficiently than the K234R variant (Ki app = 270 ± 27 μM) and displayed a faster acylation rate (k2/K), which was 5,815 ± 582 M-1 s-1 for KPC-2 versus 247 ± 25 M-1 s-1 for the K234R variant. Unlike avibactam, timed mass spectrometry revealed an intact sulfate on nacubactam and a novel peak (+337 Da) with the K234R variant. Molecular modeling of the K234R variant showed significant catalytic residue (i.e., S70, K73, and S130) rearrangements that likely interfere with nacubactam binding and acylation. Nacubactam's aminoethoxy tail formed unproductive interactions with the K234R variant's active site. Molecular modeling and docking observations were consistent with the results of biochemical analyses. Overall, the meropenem-nacubactam combination is effective against carbapenem-resistant K. pneumoniae Moreover, our data suggest that β-lactamase inhibition by nacubactam proceeds through an alternative mechanism compared to that for avibactam.
| S-EPMC6658744 | biostudies-literature