Antibiotic resistance ofKlebsiella pneumoniae through β-arrestin recruitment-induced β-lactamase signaling pathway. - Related Documents




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506301.0000Antibiotic resistance ofKlebsiella pneumoniae through β-arrestin recruitment-induced β-lactamase signaling pathway. Overuse and misuse of antibiotics leads to rapid evolution of antibiotic-resistant bacteria and antibiotic resistance genes. Klebsiella pneumoniae has become the most common pathogenic bacterium accountable for nosocomial infections due to its high virulence factor and general occurrence of resistance to most antibiotics. The β-lactamase signaling pathway has been suggested to be involved in antibiotic resistance against β-lactams in Klebsiella pneumoniae. In the present study, the molecular mechanism of the antibiotic resistance of Klebsiella pneumoniae was investigated and the results indicated involvement of the β-arrestin recruitment-induced β-lactamase signaling pathway. Antimicrobial susceptibility of Klebsiella pneumoniae was assessed using automated systems and extended-spectrum β-lactamase (ESBL) and β-arrestin expression levels in Klebsiella pneumoniae were analyzed by reverse-transcription quantitative PCR. β-lactam resistance in Klebsiella pneumoniae was determined using β-lactam agar screening plates. The results demonstrated that β-arrestin recruitment was increased in Klebsiella pneumoniae with antibiotic resistance (AR-K.P.) compared with that in the native Klebsiella pneumoniae strain (NB-K.P.). Increased production of ESBL was observed in AR-K.P. after treatment with the β-lactam penicillin. Of note, inhibition of β-arrestin recruitment significantly suppressed ESBL expression in AR-K.P. and in addition, genes encoding β-arrestin and ESBL were upregulated in Klebsiella pneumoniae. Restoration of endogenous β-arrestin markedly increased antibiotic resistance of Klebsiella pneumoniae to β-lactam. Knockdown of endogenous β-arrestin downregulated antibiotic resistance genes and promoted the inhibitory effects of β-lactam antibiotic treatment on Klebsiella pneumoniae growth. In conclusion, the present study identified that β-arrestin recruitment was associated with growth and resistance to β-lactams, which suggested that β-arrestin regulating ESBL expression may be a potential target for addressing antibiotic resistance to β-lactams in Klebsiella pneumoniae.201829563975
505310.9995Effects of different carbapenemase and siderophore production on cefiderocol susceptibility in Klebsiella pneumoniae. The resistance mechanism of Gram-negative bacteria to the siderophore antibiotic cefiderocol is primarily attributed to carbapenemase and siderophore uptake pathways; however, specific factors and their relationships remain to be fully elucidated. Here, we constructed cefiderocol-resistant Klebsiella pneumoniae (CRKP) strains carrying different carbapenemases and knocked out siderophore genes to investigate the roles of various carbapenemases and siderophores in the development of cefiderocol resistance. Antimicrobial susceptibility testing revealed that both bla(NDM) and bla(KPC) significantly increased the minimum inhibitory concentration (MIC) of Klebsiella pneumoniae (KP) to cefiderocol, while bla(OXA-48) showed a modest increase. Notably, KP expressing NDM exhibited a higher cefiderocol MIC compared to KP expressing KPC, although expression of NDM alone did not induce cefiderocol resistance. Laboratory evolutionary experiments demonstrated that combining pNDM with mutations in the siderophore uptake receptor gene cirA and pKPC with a mutation in the two-component system gene envZ led to KP reaching a high level of cefiderocol resistance. Although combining pOXA with mutations in the two-component system gene baeS did not induce cefiderocol resistance, it significantly reduced susceptibility. Moreover, siderophores could influence the development of cefiderocol resistance. Strains deficient in enterobactin exhibited increased susceptibility to cefiderocol, while deficiencies in yersiniabactin and salmochelin showed no significant alterations. In conclusion, carbapenemase gene expression facilitates cefiderocol resistance, but its presence alone is insufficient. Cefiderocol resistance in CRKP typically involves abnormal expression of certain genes and other factors, such as mutations in siderophore uptake receptor genes and two-component system genes. The enterobactin siderophore synthesis gene entB may also contribute to resistance.202439470196
504820.9994Alliance of Efflux Pumps with β-Lactamases in Multidrug-Resistant Klebsiella pneumoniae Isolates. Nosocomial infections caused by Klebsiella pneumoniae are primarily characterized by a high prevalence of extended-spectrum β-lactamases (ESBL's) and a soaring pace of carbapenemase dissemination. Availability of limited antimicrobial agents as a therapeutic option for multidrug-resistant bacteria raises an alarming concern. This study aimed at the molecular characterization of multidrug-resistant K. pneumoniae clinical isolates and studied the role of efflux pumps in β-lactam resistance. Thirty-three isolates confirmed as ESBL-positive K. pneumoniae that harbored resistance genes to major classes of antibiotics. The results showed that CTX-M15 was the preeminent β-lactamase along with carbapenemases in ESBL-positive isolates. However, the efficacy of different antibiotics varied in the presence of lactamase inhibitors and efflux pump inhibitors (EPIs). Those showing increased efficacy of antibiotics with EPI were further explored for the expression of efflux pump genes and expressed a significantly different level of efflux pumps. We found that an isolate had higher expression of kpnF (SMR family) and kdeA (MATE family) pump genes relative to RND family pump genes. No mutations were observed in the genes for porins. Together, the findings suggest that β-lactamases are not the only single factor responsible for providing resistance against the existing β-lactam drugs. Resistance may increase many folds by simultaneous expression of RND family (the most prominent family in Gram-negative bacteria) and other efflux pump family.201931613200
157930.9994Inverse Association between the Existence of CRISPR/Cas Systems with Antibiotic Resistance, Extended Spectrum β-Lactamase and Carbapenemase Production in Multidrug, Extensive Drug and Pandrug-Resistant Klebsiella pneumoniae. Antimicrobial resistance, with the production of extended-spectrum β-lactamases (ESBL) and carbapenemases, is common in the opportunistic pathogen, Klebsiella pneumoniae. This organism has a genome that can contain clustered regularly interspaced short palindromic repeats (CRISPRs), which operate as a defense mechanism against external invaders such as plasmids and viruses. This study aims to determine the association of the CRISPR/Cas systems with antibiotic resistance in K. pneumoniae isolates from Iraqi patients. A total of 100 K. pneumoniae isolates were collected and characterized according to their susceptibility to different antimicrobial agents. The CRISPR/Cas systems were detected via PCR. The phenotypic detection of ESBLs and carbapenemases was performed. The production of ESBL was detected in 71% of the isolates. Carbapenem-resistance was detected in 15% of the isolates, while only 14% were susceptible to all antimicrobial agents. Furthermore, the bacteria were classified into multidrug (77%), extensively drug-resistant (11.0%) and pandrug-resistant (4.0%). There was an inverse association between the presence of the CRISPR/Cas systems and antibiotic resistance, as resistance was higher in the absence of the CRISPR/Cas system. Multidrug resistance in ESBL-producing and carbapenem-resistant K. pneumoniae occurred more frequently in strains negative for the CRISPR/Cas system. Thus, we conclude that genes for exogenous antibiotic resistance can be acquired in the absence of the CRISPR/Cas modules that can protect the bacteria against acquiring foreign DNA.202337370299
569840.9994Evolutionary Trajectories toward High-Level β-Lactam/β-Lactamase Inhibitor Resistance in the Presence of Multiple β-Lactamases. β-Lactam antibiotics are the first choice for the treatment of most bacterial infections. However, the increased prevalence of β-lactamases, in particular extended-spectrum β-lactamases, in pathogenic bacteria has severely limited the possibility of using β-lactam treatments. Combining β-lactam antibiotics with β-lactamase inhibitors can restore treatment efficacy by negating the effect of the β-lactamase and has become increasingly important against infections caused by β-lactamase-producing strains. Not surprisingly, bacteria with resistance to even these combinations have been found in patients. Studies on the development of bacterial resistance to β-lactam/β-lactamase inhibitor combinations have focused mainly on the effects of single, chromosomal or plasmid-borne, β-lactamases. However, clinical isolates often carry more than one β-lactamase in addition to multiple other resistance genes. Here, we investigate how the evolutionary trajectories of the development of resistance to three commonly used β-lactam/β-lactamase inhibitor combinations, ampicillin-sulbactam, piperacillin-tazobactam, and ceftazidime-avibactam, were affected by the presence of three common β-lactamases, TEM-1, CTX-M-15, and OXA-1. First-step resistance was due mainly to extensive gene amplifications of one or several of the β-lactamase genes where the amplification pattern directly depended on the respective drug combination. Amplifications also served as a stepping-stone for high-level resistance in combination with additional mutations that reduced drug influx or mutations in the β-lactamase gene bla(CTX-M-15). This illustrates that the evolutionary trajectories of resistance to β-lactam/β-lactamase inhibitor combinations are strongly influenced by the frequent and transient nature of gene amplifications and how the presence of multiple β-lactamases shapes the evolution to higher-level resistance.202235652643
155050.9994Newly Detected Transmission of bla(KPC-2) by Outer Membrane Vesicles in Klebsiella Pneumoniae. OBJECTIVE: The prevalence of carbapenem-resistant Klebsiella pneumoniae (CR-KP) is a global public health problem. It is mainly caused by the plasmid-carried carbapenemase gene. Outer membrane vesicles (OMVs) contain toxins and other factors involved in various biological processes, including β-lactamase and antibiotic-resistance genes. This study aimed to reveal the transmission mechanism of OMV-mediated drug resistance of Klebsiella (K.) pneumoniae. METHODS: We selected CR-KP producing K. pneumoniae carbapenemase-2 (KPC-2) to study whether they can transfer resistance genes through OMVs. The OMVs of CR-KP were obtained by ultracentrifugation, and incubated with carbapenem-sensitive K. pneumoniae for 4 h. Finally, the carbapenem-sensitive K. pneumoniae was tested for the presence of bla(KPC-2) resistance gene and its sensitivity to carbapenem antibiotics. RESULTS: The existence of OMVs was observed by the electron microscopy. The extracted OMVs had bla(KPC-2) resistance gene. After incubation with OMVs, bla(KPC-2) resistance gene was detected in sensitive K. pneumoniae, and it became resistant to imipenem and meropenem. CONCLUSION: This study demonstrated that OMVs isolated from KPC-2-producing CR-KP could deliver bla(KPC-2) to sensitive K. pneumoniae, allowing the bacteria to produce carbapenemase, which may provide a novel target for innovative therapies in combination with conventional antibiotics for treating carbapenem-resistant Enterobacteriaceae.202336602673
250260.9994Rapid detection of colistin resistance in Acinetobacter baumannii using MALDI-TOF-based lipidomics on intact bacteria. With the dissemination of extremely drug resistant bacteria, colistin is now considered as the last-resort therapy for the treatment of infection caused by Gram-negative bacilli (including carbapenemase producers). Unfortunately, the increase use of colistin has resulted in the emergence of resistance as well. In A. baumannii, colistin resistance is mostly caused by the addition of phosphoethanolamine to the lipid A through the action of a phosphoethanolamine transferase chromosomally-encoded by the pmrC gene, which is regulated by the two-component system PmrA/PmrB. In A. baumannii clinical isolate the main resistance mechanism to colistin involves mutations in pmrA, pmrB or pmrC genes leading to the overexpression of pmrC. Although, rapid detection of resistance is one of the key issues to improve the treatment of infected patient, detection of colistin resistance in A. baumannii still relies on MIC determination through microdilution, which is time-consuming (16-24 h). Here, we evaluated the performance of a recently described MALDI-TOF-based assay, the MALDIxin test, which allows the rapid detection of colistin resistance-related modifications to lipid A (i.e phosphoethanolamine addition). This test accurately detected all colistin-resistant A. baumannii isolates in less than 15 minutes, directly on intact bacteria with a very limited sample preparation prior MALDI-TOF analysis.201830442963
171570.9994Transcriptome analysis of beta-lactamase genes in diarrheagenic Escherichia coli. Beta (β)-lactamases are the most important agents that confer drug resistance among gram-negative bacteria. Continuous mutations in β-lactamases make them remarkably diverse. We carried out the transcriptome analysis of 10 β-lactamase genes of Extended-Spectrum β-lactamases (ESBL), Metallo β-lactamases (MBL), and AmpC β-lactamases (ABL) in drug-resistant and sensitive diarrheagenic E. coli (DEC) isolates obtained from children up to 5 years of age. Out of the 10 β-lactamase genes, four belonged to ESBL (TEM, SHV, CTX, and OXA); three to MBL (NDM-1, IMP, and VIM); and three to ABL (ACT, DHA and CMY) class of genes. The different categories of DEC were estimated for β-lactamases production using a set of conventional phenotypic tests, followed by detection of their messenger RNA (mRNA) expression. The study revealed a direct correlation between mRNA expression of these genes and the presence of antibiotic resistance; also corroborated by mutation analysis of the AmpC promoter region. All the 10 β-lactamase genes showed a significant increase in their expression levels in resistant isolates, compared to those of the sensitive isolates, indicating their possible role in the disease pathogenesis. Increase in mRNA expression of β-lactamase genes, and thereby virulence, may be due to multifactorial parameters causing phenotypic as well as genotypic changes. Our study highlights the necessity of instantaneous detection of β-lactamase gene expression to curb the overwhelming threat posed by emergence of drug resistance amongst the commensal E. coli strains in children from developing countries for larger public health interest.201930842518
576080.9994Downregulation of Klebsiella pneumoniae RND efflux pump genes following indole signal produced by Escherichia coli. BACKGROUND: More than a century has passed since it was discovered that many bacteria produce indole, but research into the actual biological roles of this molecule is just now beginning. The influence of indole on bacterial virulence was extensively investigated in indole-producing bacteria like Escherichia coli. To gain a deeper comprehension of its functional role, this study investigated how indole at concentrations of 0.5-1.0 mM found in the supernatant of Escherichia coli stationary phase culture was able to alter the virulence of non-indole-producing bacteria, such as Pseudomonas aeruginosa, Proteus mirabilis, and Klebsiella pneumoniae, which are naturally exposed to indole in mixed infections with Escherichia coli. RESULTS: Biofilm formation, antimicrobial susceptibility, and efflux pump activity were the three phenotypic tests that were assessed. Indole was found to influence antibiotic susceptibly of Pseudomonas aeruginosa, Proteus mirabilis and Klebsiella pneumoniae to ciprofloxacin, imipenem, ceftriaxone, ceftazidime, and amikacin through significant reduction in MIC with fold change ranged from 4 to 16. Biofilm production was partially abrogated in both 32/45 Pseudomonas aeruginosa and all eight Proteus mirabilis, while induced biofilm production was observed in 30/40 Klebsiella pneumoniae. Moreover, acrAB and oqxAB, which encode four genes responsible for resistance-nodulation-division multidrug efflux pumps in five isolates of Klebsiella pneumoniae were investigated genotypically using quantitative real-time (qRT)-PCR. This revealed that all four genes exhibited reduced expression indicated by 2^-ΔΔCT < 1 in indole-treated isolates compared to control group. CONCLUSION: The outcomes of qRT-PCR investigation of efflux pump expression have established a novel clear correlation of the molecular mechanism that lies beneath the influence of indole on bacterial antibiotic tolerance. This research provides novel perspectives on the various mechanisms and diverse biological functions of indole signaling and how it impacts the pathogenicity of non-indole-producing bacteria.202439182027
250890.9993Genetics of Acquired Antibiotic Resistance Genes in Proteus spp. Proteus spp. are commensal Enterobacterales of the human digestive tract. At the same time, P. mirabilis is commonly involved in urinary tract infections (UTI). P. mirabilis is naturally resistant to several antibiotics including colistin and shows reduced susceptibility to imipenem. However higher levels of resistance to imipenem commonly occur in P. mirabilis isolates consecutively to the loss of porins, reduced expression of penicillin binding proteins (PBPs) PBP1a, PBP2, or acquisition of several antibiotic resistance genes, including carbapenemase genes. In addition, resistance to non-β-lactams is also frequently reported including molecules used for treating UTI infections (e.g., fluoroquinolones, nitrofurans). Emergence and spread of multidrug resistant P. mirabilis isolates, including those producing ESBLs, AmpC cephalosporinases and carbapenemases, are being more and more frequently reported. This review covers Proteus spp. with a focus on the different genetic mechanisms involved in the acquisition of resistance genes to multiple antibiotic classes turning P. mirabilis into a dreadful pandrug resistant bacteria and resulting in difficult to treat infections.202032153540
5772100.9993Molecular evaluation of colistin-resistant gene expression changes in Acinetobacter baumannii with real-time polymerase chain reaction. BACKGROUND: Acinetobacter baumannii is an important human pathogen which has recently gained increased attention due to the occurrence of drug-resistant nosocomial infections in patients suffering from immune system disorders, and those in hospital intensive care units. The aim of this research was to identify and isolate A. baumannii strains resistant to colistin, determine antibiotic resistance pattern of this bacteria, investigate the presence of colistin-resistant genes, and finally assess the effect of expression changes in pmrA and pmrB genes resistant to A. baumannii against colistin via real-time polymerase chain reaction. METHODS: The samples were initially purified and isolated using biochemical tests and Micro-gen kit. Later, the resistance pattern evaluation of validated samples to different antibiotics and colistin was carried out using two methods viz., disc diffusion and E-test. This was followed by the assessment of genes resistant to colistin via polymerase chain reaction besides gene expression changes via real-time polymerase chain reaction. RESULTS: The results of this study indicated that eleven strains of A. baumannii isolated from Shahid Rajaee Trauma Hospital were resistant to colistin. However, in the resistance pattern evaluation of A. baumannii isolated from Ali Asghar Hospital, all the strains were sensitive to colistin. In the evaluation of genes resistant to pmrA and pmrB, most of the strains resistant to colistin were carriers of these genes. Besides, in the expression assessment of these genes, it was demonstrated that expression of pmrA in the strains resistant to colistin significantly increased in relation to sensitive strains, but the expression of pmrB increased at a lower rate in the strains resistant to colistin as compared to the sensitive strains. CONCLUSION: Thus, it can be safely mentioned that increased expression of pmrA was due to the resistance of A. baumannii to colistin.201729225477
9768110.9993Inosine monophosphate overcomes the coexisting resistance of mcr-1 and bla(NDM-1) in Escherichia coli. INTRODUCTION: The rise of antibiotic-resistant bacteria, particularly those harboring mcr-1 and bla(NDM-1), threatens public health by reducing the efficacy of colistin and carbapenems. Recently, the co-spread of mcr-1 and bla(NDM-1) has been reported, and the emergence of dual-resistant Enterobacteriaceae severely exacerbates antimicrobial resistance. OBJECTIVES: This study aims to investigate the impact of mcr-1 and bla(NDM-1) expression on metabolism in Escherichia coli and to identify potential antimicrobial agents capable of overcoming the resistance conferred by these genes. METHODS: We employed non-targeted metabolomics to profile the metabolic perturbations of E. coli strains harboring mcr-1 and bla(NDM-1). The bactericidal effects of the differential metabolite, inosine monophosphate (IMP), were assessed both in vitro using time-killing assays and in vivo using a mouse infection model. The antimicrobial mechanism of IMP was elucidated through transcriptomic analysis and biochemical approaches. RESULTS: Metabolic profiling revealed significant alterations in the purine pathway, with IMP demonstrating potent bactericidal activity against E. coli strains carrying both resistance genes. IMP increased membrane permeability, disrupted proton motive force, reduced ATP levels, induced oxidative damage by promoting reactive oxygen species and inhibiting bacterial antioxidant defenses, and improved the survival rate of infected mice. CONCLUSION: Our findings suggest that IMP could be a promising candidate for combating mcr-1 and bla(NDM-1)-mediated resistance and provide a novel approach for discovering antimicrobial agents against colistin- and carbapenem-resistant bacteria.202540139526
1578120.9993Association of CRISPR/Cas System with the Drug Resistance in Klebsiella pneumoniae. BACKGROUND: Klebsiella pneumoniae is a common opportunistic pathogen and its production of extended-spectrum β-lactamases (ESBL) and carbapenemases leads to drug resistance. Clustered regularly interspaced short palindromic repeats (CRISPRs) and their associated genes (Cas) are widespread in the genome of many bacteria and are a defense mechanism against foreign invaders such as plasmids and viruses. PURPOSE: To investigate the prevalence of the CRISPR/Cas system in wild type strains of K. pneumoniae in the hospital and its association with drug resistance. MATERIALS AND METHODS: A total of 136 strains were collected and characterized their susceptibility to antimicrobial agents. The prevalence of CRISPR/Cas system was detected by PCR and DNA sequencing was analyzed by CRISPRFinder. The statistical analysis of the results was performed by SPSS. RESULTS: We found that 50/136 (37%) isolates produced ESBL and 30/136 (22%) isolates were resistant to carbapenems. These isolates were liable to be multidrug resistant against β-lactams, quinolones, and aminoglycosides. Among the carbapenem-resistant isolates, blaKPC was the main drug resistance-associated gene and different types of ESBL and AmpC genes were present. Resistance to β-lactams, quinolones, aminoglycosides, tetracyclines, and β-lactams/enzyme inhibitor were higher in absence of the CRISPR/Cas system. Eighteen spacers within the CRISPR arrays matched with the genomes of plasmids or phages, some of which carried drug resistance genes. CONCLUSION: ESBL-producing and carbapenem-resistant K. pneumoniae are more likely to develop multidrug resistance and show an inverse correlation between drug resistance and CRISPR/Cas system. Absence of CRISPR/Cas modules allow for the acquisition of external drug resistance genes.202032606841
5838130.9993Alteration in the Morphological and Transcriptomic Profiles of Acinetobacter baumannii after Exposure to Colistin. Acinetobacter baumannii is often highly resistant to multiple antimicrobials, posing a risk of treatment failure, and colistin is a "last resort" for treatment of the bacterial infection. However, colistin resistance is easily developed when the bacteria are exposed to the drug, and a comprehensive analysis of colistin-mediated changes in colistin-susceptible and -resistant A. baumannii is needed. In this study, using an isogenic pair of colistin-susceptible and -resistant A. baumannii isolates, alterations in morphologic and transcriptomic characteristics associated with colistin resistance were revealed. Whole-genome sequencing showed that the resistant isolate harbored a PmrB(L208F) mutation conferring colistin resistance, and all other single-nucleotide alterations were located in intergenic regions. Using scanning electron microscopy, it was determined that the colistin-resistant mutant had a shorter cell length than the parental isolate, and filamented cells were found when both isolates were exposed to the inhibitory concentration of colistin. When the isolates were treated with inhibitory concentrations of colistin, more than 80% of the genes were upregulated, including genes associated with antioxidative stress response pathways. The results elucidate the morphological difference between the colistin-susceptible and -resistant isolates and different colistin-mediated responses in A. baumannii isolates depending on their susceptibility to this drug.202439203486
5699140.9993Presence of β-Lactamase Encoding Genes in Burkholderia cepacia Complex Isolated from Soil. Burkholderia cepacia complex has emerged as an important opportunistic bacteria group for immunocompromised patients, and it has a high level of intrinsic resistance for different antibiotic classes. Hydrolysis of β-lactam antibiotics by β-lactamases is the most common resistance mechanism in Gram-negative bacteria, and the presence of such enzymes complicates the selection of appropriate therapy. This study aimed at investigating the antimicrobial resistance profile and the presence of β-lactamase encoding genes in B. cepacia complex isolated from Brazilian soils. High-level ceftazidime resistance and several β-lactamase encoding genes were found, including the first report of bla(KPC) genes in bacteria isolated from soil.201828915359
1760150.9993Proteomic analysis of clinical isolate of Stenotrophomonas maltophilia with blaNDM-1, blaL1 and blaL2 β-lactamase genes under imipenem treatment. The co-occurrence of L1 and AmpR-L2 with bla(NDM-1) gene with an upstream 250-bp promoter was detected in a clinical isolate of Stenotrophomonas maltophilia DCPS-01, which was resistant to all β-lactams and sensitive only to colistin and fluoroquinolones. To investigate expression of resistance genes and the molecular mechanisms of bacteria resistance to carbapenems, proteomic profiles of the isolate was passaged with and without the drug by using 2D-PAGE. The results showed that 33 genes exhibiting a ≥3-fold change were identified as candidates that may help S. maltophilia survive drug selection. Strikingly, L1 was expressed more highly in cells grown with imipenem, and the abundant NDM-1 further increased, while very little L2 was detected even following induction. Specific activities for β-lactamase revealed that L2 remained at constitutive low levels (10.6 U/mg), while L1 and NDM-1 showed clear activity (69.8 U/mg). Our data support that imipenem could specifically and reversibly induce L1 and NDM-1, which together played key roles in drug resistance in DCPS-01. Although NDM-1 mediated resistance to carbapenems has been found in very few cases, to our knowledge, this is the first proteomics research of S. maltophilia with NDM-1, giving very broad-spectrum antibiotic resistance profiles.201222702735
2455160.9993Molecular Mechanisms of Colistin Resistance Among Klebsiella Pneumoniae Strains. BACKGROUND: The increasing rate of infections caused by multiple drug resistant gram-negative bacteria has led to resuscitation of colistin. As a result, colistin resistance, mainly among Klebsiella pneumoniae strains has also been increased. The aim of this study was to investigate molecular mechanisms behind colistin resistance. METHODS: Twenty colistin-resistant K. pneumoniae strains isolated from clinical samples of different patients were involved in this study. VITEK2 automated ID/AST system (Biomeriux, France) was used for the identification and also the susceptibility testing for antibiotics other than colistin. Colistin susceptibility was determined by broth microdilution method. To identify the mechanisms of resistance, mutations on mgrB genes, expression levels of pmrA, pmrB, pmrC, pmrD, pmrE, pmrK, phoQ, and phoP genes, and the presence of plasmid mediated colistin resistance genes, mcr-1 and mcr-2 were investigated. RESULTS: As a result of the study, increased expression levels of the pmrA, pmrB, pmrD, pmrK, phoP, and phoQ genes were observed. All colistin resistant strains were found wild type for the mgrB gene which is thought to be esponsible for colistin resistance. Also, no mcr-1 or mcr-2 genes which are the causes of plasmid mediated colistin resistance have been detected in any of the strains. CONCLUSIONS: Among the colistin resistant K. pneumoniae strains included in our study, increased expression Levels of the genes responsible for cell membrane modifications related with colistin resistance were the most common mechanisms.201931307167
5054170.9993In vitro resistance development gives insights into molecular resistance mechanisms against cefiderocol. Cefiderocol, a novel siderophore cephalosporin, demonstrates promising in vitro activity against multidrug-resistant Gram-negative bacteria, including carbapenemase-producing strains. Nonetheless, only a few reports are available regarding the acquisition of resistance in clinical settings, primarily due to its recent usage. This study aimed to investigate cefiderocol resistance using an in vitro resistance development model to gain insights into the underlying molecular resistance mechanisms. Cefiderocol susceptible reference strains (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa) and a clinical Acinetobacter baumannii complex isolate were exposed to increasing cefiderocol concentrations using a high-throughput resistance development model. Cefiderocol susceptibility testing was performed using broth microdilution. Whole-genome sequencing was employed to identify newly acquired resistance mutations. Our in vitro resistance development model led to several clones of strains exhibiting cefiderocol resistance, with MIC values 8-fold to 512-fold higher than initial levels. In total, we found 42 different mutations in 26 genes, of which 35 could be described for the first time. Putative loss-of-function mutations were detected in the envZ, tonB, and cirA genes in 13 out of 17 isolates, leading to a decrease in cefiderocol influx. Other potential resistance mechanisms included multidrug efflux pumps (baeS, czcS, nalC), antibiotic-inactivating enzymes (ampR, dacB), and target mutations in penicillin-binding-protein genes (mrcB). This study reveals new insights into underlying molecular resistance mechanisms against cefiderocol. While mutations leading to reduced influx via iron transporters was the most frequent resistance mechanism, we also detected several other novel resistance mutations causing cefiderocol resistance.202439080477
1543180.9993AmpC beta-lactamases. AmpC beta-lactamases are clinically important cephalosporinases encoded on the chromosomes of many of the Enterobacteriaceae and a few other organisms, where they mediate resistance to cephalothin, cefazolin, cefoxitin, most penicillins, and beta-lactamase inhibitor-beta-lactam combinations. In many bacteria, AmpC enzymes are inducible and can be expressed at high levels by mutation. Overexpression confers resistance to broad-spectrum cephalosporins including cefotaxime, ceftazidime, and ceftriaxone and is a problem especially in infections due to Enterobacter aerogenes and Enterobacter cloacae, where an isolate initially susceptible to these agents may become resistant upon therapy. Transmissible plasmids have acquired genes for AmpC enzymes, which consequently can now appear in bacteria lacking or poorly expressing a chromosomal bla(AmpC) gene, such as Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis. Resistance due to plasmid-mediated AmpC enzymes is less common than extended-spectrum beta-lactamase production in most parts of the world but may be both harder to detect and broader in spectrum. AmpC enzymes encoded by both chromosomal and plasmid genes are also evolving to hydrolyze broad-spectrum cephalosporins more efficiently. Techniques to identify AmpC beta-lactamase-producing isolates are available but are still evolving and are not yet optimized for the clinical laboratory, which probably now underestimates this resistance mechanism. Carbapenems can usually be used to treat infections due to AmpC-producing bacteria, but carbapenem resistance can arise in some organisms by mutations that reduce influx (outer membrane porin loss) or enhance efflux (efflux pump activation).200919136439
1577190.9993Clonal Clusters, Molecular Resistance Mechanisms and Virulence Factors of Gram-Negative Bacteria Isolated from Chronic Wounds in Ghana. Wound infections are common medical problems in sub-Saharan Africa but data on the molecular epidemiology are rare. Within this study we assessed the clonal lineages, resistance genes and virulence factors of Gram-negative bacteria isolated from Ghanaian patients with chronic wounds. From a previous study, 49 Pseudomonas aeruginosa, 21 Klebsiellapneumoniae complex members and 12 Escherichia coli were subjected to whole genome sequencing. Sequence analysis indicated high clonal diversity with only nine P. aeruginosa clusters comprising two strains each and one E. coli cluster comprising three strains with high phylogenetic relationship suggesting nosocomial transmission. Acquired beta-lactamase genes were observed in some isolates next to a broad spectrum of additional genetic resistance determinants. Phenotypical expression of extended-spectrum beta-lactamase activity in the Enterobacterales was associated with bla(CTX-M-15) genes, which are frequent in Ghana. Frequently recorded virulence genes comprised genes related to invasion and iron-uptake in E. coli, genes related to adherence, iron-uptake, secretion systems and antiphagocytosis in P. aeruginosa and genes related to adherence, biofilm formation, immune evasion, iron-uptake and secretion systems in K. pneumonia complex. In summary, the study provides a piece in the puzzle of the molecular epidemiology of Gram-negative bacteria in chronic wounds in rural Ghana.202133810142