Intestinal carriage of colistin-resistant Enterobacteriaceae at Saint Georges Hospital in Lebanon. - Related Documents




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91301.0000Intestinal carriage of colistin-resistant Enterobacteriaceae at Saint Georges Hospital in Lebanon. OBJECTIVES: The increase in resistance to antibiotics has led to the revival of colistin as the last option for treatment, which automatically led to an increase of colistin-resistant, Gram-negative bacteria. In this study, we report the presence of clinical colistin-resistant Enterobacteriaceae isolated from a Lebanese hospital. METHODS: From 23 rectal swabs, eight colistin-resistant clinical strains (five Escherichia coli, two Enterobacter cloacae, and one Klebsiella pneumoniae) were isolated. Antibiotic susceptibility testing was performed using the disk diffusion method and Etest. The broth microdilution method was used to determine colistin susceptibility. Reverse transcription polymerase chain reaction (RT-PCR), standard PCR and sequencing were used to investigate genes encoding for extended-spectrum β-lactamases, carbapenemases and colistin resistance. Genotyping of these isolates was conducted by multilocus sequence typing (MLST). RESULTS: Results of antibiotic susceptibility testing revealed that all isolates were resistant to colistin. They had MICs for colistin that ranged from 8 to 32 mg/L. Real-time PCR results showed that five strains harboured bla(TEM-1) and one strain harboured bla(TEM-163). Moreover, four strains were positive for bla(CTX-M-15), bla(CTX-M-103) and bla(CTX-M-189), and K. pneumoniae harboured bla(SHV-1). Observed colistin resistance was linked to amino acid substitutions into protein sequences of pmrA/B, phoP/Q, and mgrB. Interestingly, we report here a mutation in the mgrB regulator and pmrA/B, phoP/Q in colistin-resistant E. cloacae and E. coli clinical isolates for the first time in Lebanon. CONCLUSION: This study highlights the presence of colistin-resistant Gram-negative bacteria in a Lebanese hospital, which is worrisome. An urgent strategy needs to be adopted to avoid the spread of such bacteria.202031838239
91210.9999Carbapenem and colistin-resistant bacteria in North Lebanon: Coexistence of mcr-1 and NDM-4 genes in Escherichia coli. INTRODUCTION: The increasing incidence of infections caused by multidrug-resistant bacteria is considered a global health problem. This study aimed to investigate this resistance in Gram-negative bacteria isolated from patients hospitalized in North-Lebanon. METHODOLOGY: All isolates were identified using the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Antibiotic susceptibility testing was achieved using disk diffusion, E-test and Broth microdilution methods. Phenotypic detection of carbapenemase was carried out using the CarbaNP test. RT-PCR, standard-PCR and sequencing were performed to detect resistance genes and oprD gene. Conjugal transfer was carried out between our isolates and Escherichia coli J53 to detect the genetic localization of resistance genes. MLST was conducted to determine the genotype of each isolate. RESULTS: Twenty-three carbapenem-resistant Enterobacterales of which eight colistin-resistant Escherichia coli, and Twenty carbapenem-resistant Pseudomonas aeruginosa were isolated. All isolates showed an imipenem MIC greater than 32 mg/mL with MICs for colistin greater than 2 mg/L for E. coli isolates. All the Enterobacterales isolates had at least one carbapenemase-encoding gene, with E. coli isolates coharboring blaNDM-4 and mcr-1 genes. Moreover, 16/20 Pseudomonas aeruginosa harbored the blaVIM-2 gene and 18/20 had mutations in the oprD gene. MLST revealed that the isolates belonged to several clones. CONCLUSIONS: We report here the first description in the world of clinical E. coli isolates coharboring blaNDM-4 and mcr-1 genes, and K. pneumoniae isolates producing NDM-6 and OXA-48 carbapenemases. Also, we describe the emergence of NDM-1-producing E. cloacae in Lebanon. Screening for these isolates is necessary to limit the spread of resistant microorganisms in hospitals.202134343118
91520.9999Detection of Plasmid-Mediated Mobile Colistin Resistance Gene (mcr-1) in Enterobacterales Isolates from a University Hospital. PURPOSE: Colistin represents one of the last treatment options for infections caused by multi-drug resistant (MDR) Enterobacterales. The emergence of a plasmid-mediated mobile colistin resistance-1 (mcr-1) gene has raised serious concerns about its potential dissemination among bacteria. METHODS: In this study, we evaluated the chromogenic medium, CHROMID(®) Colistin Resistance (COLR) agar, for the rapid detection of colistin-resistant Enterobacterales using broth microdilution (BMD) as a reference method. We also attempted to detect mcr-1, -2, -3, -4, and -5 genes, as well as the insertion sequence ISApl1 via polymerase chain reaction (PCR), followed by sequencing of mcr gene(s). RESULTS: Among the 100 studied Enterobacterales isolates, 53% of them were colistin-resistant, with higher rate among Klebsiella pneumoniae (75%) as compared to Escherichia coli (44.4%). The COLR agar showed 83.2% sensitivity and 97.9% specificity for the detection of colistin resistance. Among colistin-resistant isolates, mcr-1 gene was only detected in four (7.5%) E. coli isolates. The ISApl1 was not found among mcr-1 positive isolates. Sequencing of mcr-1 gene revealed nucleotide sequence homogeneity with the wild-type mcr-1 gene in BLAST. CONCLUSION: The COLR agar is a promising phenotypic method for the detection of colistin-resistant Enterobacterales. Multiplex PCR followed by sequencing can be used for mcr genes' detection and characterization.202134408450
91430.9999Investigation of colistin heteroresistance and the colistin resistance genes mcr-1 to mcr-5 in Escherichia coli and Klebsiella pneumoniae isolates in a tertiary hospital in Turkey. INTRODUCTION: Heteroresistance is not detected by traditional antimicrobial susceptibility testing methods and may lead to treatment failures. Investigating the presence of plasmid-mediated colistin resistance genes is important because of the horizontal transmission of the relevant genes between bacterial species. This study aimed to investigate the presence of colistin heteroresistance and the colistin resistance genes mcr-1 to mcr-5 in Escherichia coli and Klebsiella pneumoniae isolates. METHODOLOGY: A total of 254 isolates, including 100 E. coli and 154 K. pneumoniae strains isolated from clinical samples, were included in the study. Colistin susceptibility was evaluated using the broth microdilution method for all strains. Heteroresistance screening was performed using the gradient strip test. Eight strains were evaluated for heteroresistance by population analysis profiling (PAP). The colistin resistance genes mcr-1 to mcr-5 were investigated by multiplex polymerase chain reaction (PCR) in colistin-resistant K. pneumoniae isolates. Multilocus sequence typing (MLST) analysis was performed on two K. pneumoniae strains. RESULTS: Colistin resistance was not detected in the E. coli isolates and was detected in 16.23% (25/154) of the K. pneumoniae isolates. No heteroresistant bacteria were detected by the gradient strip test or by PAP. All colistin-resistant isolates were negative for the mcr genes. The two isolates analyzed by MLST were ST14 and ST2096. CONCLUSIONS: Periodic follow-up of colistin heteroresistance is useful for administering appropriate antibiotic therapy. In addition, the investigation of colistin resistance genes is important for infection control measures.202439693153
90940.9999First Description of Colistin and Tigecycline-Resistant Acinetobacter baumannii Producing KPC-3 Carbapenemase in Portugal. Herein, we describe a case report of carbapenem-resistant Acinetobacter baumannii and Klebsiella pneumoniae isolates that were identified from the same patient at a Tertiary University Hospital Centre in Portugal. Antimicrobial susceptibility and the molecular characterization of resistance and virulence determinants were performed. PCR screening identified the presence of the resistance genes bla(KPC-3), bla(TEM-1) and bla(SHV-1) in both isolates. The KPC-3 K. pneumoniae isolate belonged to the ST-14 high risk clone and accumulated an uncommon resistance and virulence profile additional to a horizontal dissemination capacity. In conclusion, the molecular screening led to the first identification of the A. baumannii KPC-3 producer in Portugal with a full antimicrobial resistance profile including tigecycline and colistin.201830404152
90750.9998Clonal Dissemination of Plasmid-Mediated Carbapenem and Colistin Resistance in Refugees Living in Overcrowded Camps in North Lebanon. Carbapenem and colistin-resistant bacteria represent a global public health problem. Refugees carrying these bacteria and living in inadequate shelters can spread these microorganisms. The aim of this study was to investigate the intestinal carriage of these bacteria in Syrian refugees in Lebanon. Between June and July 2019, 250 rectal swabs were collected from two refugee camps in North Lebanon. Swabs were cultured on different selective media. Antibiotic susceptibility testing was performed using the disk diffusion method. Carbapenemase-encoding genes and mcr genes were investigated using real-time polymerase chain reaction (RT-PCR) and standard polymerase chain reaction (PCR). Epidemiological relatedness was studied using multilocus sequence typing (MLST). From 250 rectal swabs, 16 carbapenem-resistant, 5 colistin-resistant, and 4 colistin and carbapenem-resistant Enterobacteriaceae were isolated. The isolates exhibited multidrug-resistant phenotypes. Seven Klebsiella pneumoniae isolates harboured the bla(OXA-48) gene, and in addition four K. pneumoniae had mutations in the two component systems pmrA/pmrB, phoP/phoQ and co-harboured the bla(NDM-1) gene. Moreover, the bla(NDM-1) gene was detected in six Escherichia coli and three Enterobacter cloacae isolates. The remaining five E. coli isolates harboured the mcr-1 gene. MLST results showed several sequence types, with a remarkable clonal dissemination. An urgent strategy needs to be adopted in order to avoid the spread of such resistance in highly crowded underserved communities.202134943690
150360.9998OXA-48 Carbapenemase-Encoding Transferable Plasmids of Klebsiella pneumoniae Recovered from Egyptian Patients Suffering from Complicated Urinary Tract Infections. Gram-negative bacteria are common causes of urinary tract infections (UTIs). Such pathogens can acquire genes encoding multiple mechanisms of antimicrobial resistance, including carbapenem resistance. The aim of this study was to detect the carbapenemase-producing ability of some Gram-negative bacterial isolates from urine specimens of patients suffering from complicated UTIs at two vital tertiary care hospitals in Cairo, Egypt; to determine the prevalence of carbapenemase genes among plasmid-bearing isolates; and explore the possibility of horizontal gene transfer to other bacterial species. The collected isolates were subjected to antimicrobial susceptibility testing, phenotypic analysis of carbapenemase production, and molecular detection of plasmid-borne carbapenemase genes, then the extracted plasmids were transformed into competent E. coli DH5α. A total of 256 Gram-negative bacterial clinical isolates were collected, 65 (25.4%) isolates showed carbapenem resistance of which 36 (55.4%) were carbapenemase-producers, and of these 31 (47.7%) harbored plasmids. The extracted plasmids were used as templates for PCR amplification of bla(KPC), bla(NDM), bla(VIM), bla(OXA-48,) and bla(IMP) carbapenemase genes. The bla(OXA-48) gene was detected in 24 (77.4%) of the tested isolates while bla(VIM) gene was detected in 8 (25.8%), both bla(KPC) and bla(NDM) genes were co-present in 1 (3.2%) isolate. Plasmids carrying the bla(OXA-48) gene from 4 K. pneumoniae clinical isolates were successfully transformed into competent E. coli DH5α. The transformants were carbapenemase-producers and acquired resistance to some of the tested antimicrobial agents as compared to untransformed E. coli DH5α. The study concluded that the rate of carbapenem resistance among Gram-negative bacterial uropathogens in Cairo, Egypt is relatively high and can be transferred horizontally to other bacterial host(s).202134571766
150270.9998Tunisian Multicenter Study on the Prevalence of Colistin Resistance in Clinical Isolates of Gram Negative Bacilli: Emergence of Escherichia coli Harbouring the mcr-1 Gene. BACKGROUND: Actually, no data on the prevalence of plasmid colistin resistance in Tunisia are available among clinical bacteria. OBJECTIVES: This study aimed to investigate the current epidemiology of colistin resistance and the spread of the mcr gene in clinical Gram-negative bacteria (GNB) isolated from six Tunisian university hospitals. METHODS: A total of 836 GNB strains were inoculated on COL-R agar plates with selective screening agar for the isolation of GNB resistant to colistin. For the selected isolates, mcr genes, beta-lactamases associated-resistance genes and molecular characterisation were screened by PCRs and sequencing. RESULTS: Colistin-resistance was detected in 5.02% (42/836) of the isolates and colistin-resistant isolates harboured an ESBL (bla(CTX-M-15)) and/or a carbapenemase (bla(OXA-48), bla(VIM)) encoding gene in 45.2% of the cases. The mcr-1 gene was detected in four E. coli isolates (0.59%) causing urinary tract infections and all these isolates also contained the bla(TEM-1) gene. The bla(CTX-M-15) gene was detected in three isolates that also carried the IncY and IncFIB replicons. The genetic environment surrounding the mcr-carrying plasmid indicated the presence of pap-2 gene upstream mcr-1 resistance marker with unusual missing of ISApl1 insertion sequence. THE CONCLUSIONS: This study reports the first description of the mcr-1 gene among clinical E. coli isolates in Tunisia and provides an incentive to conduct routine colistin susceptibility testing in GNB clinical isolates.202236290048
99180.9998Characterization of extended-spectrum beta-lactamases in Enterobacteriaceae causing nosocomial infections in a Zagreb University Hospital. The bacteria producing extended-spectrum beta-lactamases (ESBLs) are increasingly reported. production of ESBLs by Gram-negative bacteria is the major mechanism of resistance to oxymino-cephalosporins and aztreonam. the aim of the present study was to characterize ESBLs produced by Enterobacteriaceae, collected during 2003-2005 in a University Hospital in Zagreb, and to determine the risk factors associated with nosocomial infections due to them. 76 isolates of Enterobacteriaceae were included in the study. Antibiotic susceptibility testing was performed by disk-diffusion and broth microdilution method according to CLSI. beta-lactamases were characterized by PCR and sequencing of bla(ESBL )genes. plasmids were extracted by alkaline lysis method and digested with EcoRI enzyme. Most of the strains displayed CAZ phenotype meaning a higher level of resistance to ceftazidime compared to cefotaxime and ceftriaxone. 50 strains produced SHV-ESBL, 28 tem and 8 CTX-M beta-lactamase. Sequencing of bla(SHV )genes from representative strains revealed SHV-5 beta-lactamase in 6 strains whereas sequencing of bla(CTX-M )genes identified CTX-M-3 beta-lactamase in 3 and CTX-M-15 in 5 strains. Strains were assigned to groups from A to f according to plasmid fingerprinting. The spread of SHV-5-producing strains throughout the hospital units could be due to selective pressure of ceftazidime which is widely prescribed in our hospital thus favoring survival of strains possessing a mutation at the Ambler position 240 responsible for ceftazidime and aztreonam resistance.200919567348
88690.9998Detection of Plasmid-Mediated Resistance against Colistin in Multi-Drug-Resistant Gram-Negative Bacilli Isolated from a Tertiary Hospital. The aim of this study was to determine the prevalence of plasmid-mediated colistin resistance mcr-1 to mcr-5 genes among colistin and multi-drug-resistant Gram-negative bacilli strains isolated from patients in a tertiary hospital in Toluca, Mexico. The presence of mcr genes among the 241 strains collected was assessed by PCR. In the case of mcr-carrying E. coli, further PCR tests were performed to determine the presence of bla(CTX-M) and whether the strains belonged to the O25b-ST131 clone. Conjugation experiments were also carried out to assess the horizontal transmission of colistin resistance. A total of twelve strains (5.0%), of which four were E. coli; four were P. aeruginosa; three were K. pneumoniae, and one E. cloacae, were found to be resistant to colistin. Of these strains, two E. coli isolates were found to carry mcr-1, and Southern blot hybridization demonstrated its presence on an approximately 60 kb plasmid. Both mcr-1-carrying E. coli strains were found to co-express bla(CTX-M), belong to the O25b-ST131 clone, and horizontally transmit their colistin resistance. The results of this study confirm the presence of plasmid-mediated colistin resistance in hospitalized patients in Mexico and demonstrated that the multi-drug-resistant O25b-ST131 E. coli clone can acquire mcr genes and transmit such resistance traits to other bacteria.202337630556
1501100.9998High-level and novel mechanisms of carbapenem resistance in Gram-negative bacteria from tertiary hospitals in Nigeria. To determine the occurrence and molecular basis of carbapenem resistance in Gram-negative bacteria from tertiary hospitals in Nigeria, 182 non-duplicate Gram-negative bacterial isolates were investigated for antimicrobial susceptibility, presence of carbapenemases (tested phenotypically and genotypically), random amplified polymorphic DNA (RAPD) typing, plasmid sizing and replicon typing. Minimum inhibitory concentrations of carbapenems showed a high degree of resistance, with 67 isolates (36.8%) being resistant to all carbapenems, of which 40 (59.7%) produced enzymes able to hydrolyse imipenem. PCR and sequencing identified only 10 isolates (5.5%) carrying known carbapenemase genes, including bla(NDM), bla(VIM) and bla(GES). The majority of phenotypically carbapenem-resistant and carbapenemase-producing isolates did not carry a known carbapenemase gene. Transconjugant or transformant plasmid sizes were estimated to be 115 kb for bla(NDM)- and 93 kb for bla(VIM)-carrying plasmids. These plasmids were untypeable for replicon/incompatibility and transferred various other genes including plasmid-mediated quinolone resistance (PMQR) genes and bla(CTX-M-15). Typing showed that the isolates in this study were not clonally related. There is a high level of carbapenem resistance in Nigeria. As well as the globally relevant carbapenemases (bla(NDM), bla(VIM) and bla(GES)), there are other unknown gene(s) or variant(s) in circulation able to hydrolyse carbapenems and confer high-level resistance.201424613608
860110.9998Investigation of Plasmid-Mediated Colistin Resistance Genes (mcr-1-8) in Enterobacterales Isolates. Background The escalating global rise in multidrug-resistant gram-negative bacteria presents an increasingly substantial threat to patient safety. Over the past decade, carbapenem-resistant Enterobacterales (CRE) have emerged as one of the most critical pathogens in hospital-acquired infections, notably within intensive care units. Colistin has become one of the last-resort antimicrobial agents utilized to combat infections caused by CRE. However, the use of colistin has been accompanied by a notable increase in the prevalence of colistin-resistant bacteria. This study aimed to investigate plasmid-mediated colistin resistance genes ranging from mcr-1 to mcr-8 among members of the Enterobacterales order. Materials and methods This prospective study was conducted in the microbiology laboratory of Afyonkarahisar Health Sciences University Health Research and Practice Center between May 1, 2021 and July 31, 2022. A total of 2,646 Enterobacterales isolates were obtained from all culture-positive clinical samples sent from various clinics. Of these, 79 isolates exhibiting resistance to carbapenem antibiotics were included in the study. Among the 79 isolates, the presence of mcr-1 to mcr-8 genes was investigated in 27 isolates that were shown to be resistant to colistin. The identification of bacteria at the species level and antibiotic susceptibility tests were conducted using the VITEK 2 automated system (bioMérieux, USA). Colistin resistance among Enterobacterales strains exhibiting carbapenem resistance was evaluated using the broth microdilution technique (ComASP™ Colistin, Liofilchem, Italy), in accordance with the manufacturer's instructions. Results In our in vitro investigations, the minimum inhibitory concentration (MIC) values for meropenem were determined to be >8 µg/ml, whereas for colistin, the MIC50 value was >16 µg/ml and the MIC90 value was 8 µg/ml. A total of 27 colistin-resistant strains were identified among the 79 carbapenem-resistant Enterobacterales strains analyzed. The most prevalent agent among colistin-resistant strains was Klebsiella pneumoniae (K. pneumoniae), representing 66.7% of the isolates. This was followed by Proteus mirabilis (P. mirabilis) with 29.6% and Escherichia coli (E. coli) with 3.7%. The colistin resistance rate among carbapenem-resistant strains was found to be 34.2%, with colistin MIC values in strains tested by the broth microdilution method ranging from 4 to >16 µg/ml concentrations. In polymerase chain reaction (PCR) studies, the mcr-1 gene region was successfully detected by real-time PCR in the positive control isolate. Nevertheless, none of the gene regions from mcr-1 to mcr-8 were identified in our study investigating the presence of plasmid-mediated genes using a multiplex PCR kit. Conclusion Although our study demonstrated the presence of increased colistin resistance rates in carbapenem-resistant Enterobacterales isolates, it resulted in the failure to detect genes from mcr-1 to mcr-8 by the multiplex PCR method. Therefore, it is concluded that the colistin resistance observed in Enterobacteriaceae isolates in our region is not due to the mcr genes screened, but to different resistance development mechanisms.202438957246
2125120.9998Emergence of Carbapenem-Resistant Gram-Negative Isolates in Hospital Settings in Djibouti. Introduction: The antimicrobial resistance (AMR) of bacteria is increasing rapidly against all classes of antibiotics, with the increasing detection of carbapenem-resistant isolates. However, while growing prevalence has been reported around the world, data on the prevalence of carbapenem resistance in developing countries are fairly limited. In this study, we investigated and determined the resistance rate to carbapenems among multidrug-resistant Gram-negative bacteria (MDR-GNB) isolated in Djibouti and characterized their resistance mechanisms. Results: Of the 256 isolates, 235 (91.8%) were identified as Gram-negative bacteria (GNB). Of these GNBs, 225 (95.7%) isolates exhibited a multidrug resistance phenotype, and 20 (8.5%) isolates were resistant to carbapenems, including 13 Escherichia coli, 4 Acinetobacter baumannii, 2 Klebsiella pneumoniae and 1 Proteus mirabilis. The most predominant GNB in this hospital setting were E. coli and K. pneumoniae species. Carbapenemase genes such as bla(OXA-48) and bla(NDM-5) were identified, respectively, in six and four E. coli isolates, whereas the carbapenemase bla(NDM-1) was identified in three E. coli, two K. pneumoniae, one P. mirabilis and one A. baumannii. Moreover, three A. baumannii isolates co-hosted bla(OXA-23) and bla(NDM-1). Materials and Methods: A total of 256 clinical strains collected between 2019 and 2020 were identified using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF). Antibiotic susceptibility testing was performed using disk diffusion and E-test methods. Real-time polymerase chain reaction (RT-PCR), standard PCR and sequencing were used to investigate genes encoding for extended-spectrum-β-lactamases, carbapenemases and colistin resistance genes. Conclusions: We report, for the first time, the presence of MDR-GNB clinical isolates and the emergence of carbapenem-resistant isolates in Djibouti. In addition to performing antimicrobial susceptibility testing, we recommend phenotypic and molecular screening to track the spread of carbapenemase genes among clinical GNB isolates.202337508230
996130.9998Rapid Detection of New Delhi Metallo-β-Lactamase Gene Using Recombinase-Aided Amplification Directly on Clinical Samples From Children. New Delhi metallo-β-lactamase, a metallo-β-lactamase carbapenemase type, mediates resistance to most β-lactam antibiotics including penicillins, cephalosporins, and carbapenems. Therefore, it is important to detect bla (NDM) genes in children's clinical samples as quickly as possible and analyze their characteristics. Here, a recombinase-aided amplification (RAA) assay, which operates in a single one-step reaction tube at 39°C in 5-15 min, was established to target bla (NDM) genes in children's clinical samples. The analytical sensitivity of the RAA assay was 20 copies, and the various bacterial types without bla (NDM) genes did not amplify. This method was used to detect bla (NDM) genes in 112 children's stool samples, 10 of which were tested positive by both RAA and standard PCR. To further investigate the characteristics of carbapenem-resistant bacteria carrying bla (NDM) in children, 15 carbapenem-resistant bacteria (Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, Citrobacter freundii, Klebsiella oxytoca, Acinetobacter junii, and Proteus mirabilis) were isolated from the 10 samples. Notably, more than one bacterial type was isolated from three samples. Most of these isolates were resistant to cephalosporins, cefoperazone-sulbactam, piperacillin-tazobactam, ticarcillin-clavulanic acid, aztreonam, co-trimoxazole, and carbapenems. bla (NDM) (-) (1) and bla (NDM) (-) (5) were the two main types in these samples. These data show that the RAA assay has potential to be a sensitive and rapid bla (NDM) gene screening test for clinical samples. The common existence of bla (NDM) and multi-drug resistance genes presents major challenges for pediatric treatment.202134367092
910140.9998Analysis of Antimicrobial Resistance Genes (ARGs) in Enterobacterales and A. baumannii Clinical Strains Colonizing a Single Italian Patient. The dramatic increase in infections caused by critically multidrug-resistant bacteria is a global health concern. In this study, we characterized the antimicrobial resistance genes (ARGs) of K. pneumoniae, P. mirabilis, E. cloacae and A. baumannii isolated from both surgical wound and rectal swab of a single Italian patient. Bacterial identification was performed by MALDI-TOF MS and the antimicrobial susceptibility was carried out by Vitek 2 system. The characterization of ARGs was performed using next-generation sequencing (NGS) methodology (MiSeq Illumina apparatus). K. pneumoniae, P. mirabilis and E. cloacae were resistant to most β-lactams and β-lactam/β-lactamases inhibitor combinations. A. baumannii strain was susceptible only to colistin. The presence of plasmids (IncN, IncR, IncFIB, ColRNAI and Col (MGD2)) was detected in all Enterobacterales but not in A. baumannii strain. The IncN plasmid and bla(NDM-1) gene were found in K. pneumoniae, P. mirabilis and E. cloacae, suggesting a possible transfer of this gene among the three clinical species. Conjugation experiments were performed using K. pneumoniae (1 isolate), P. mirabilis (2 isolates) and E. cloacae (2 isolates) as donors and E. coli J53 as a recipient. The bla(NDM-1) gene was identified by PCR analysis in all transconjugants obtained. The presence of four different bacterial species harboring resistance genes to different classes of antibiotics in a single patient substantially reduced the therapeutic options.202336978306
2159150.9998Involvement of the AcrAB Efflux Pump in Ciprofloxacin Resistance in Clinical Klebsiella Pneumoniae Isolates. BACKGROUND: Increasing prevalence of multiple antibiotic resistance in Klebsiella pneumoniae strains confines the therapeutic options used to treat bacterial infections. OBJECTIVE: We aimed in this study to investigate the role of AcrAB and qepA efflux pumps and AAC(6')-Ib-cr enzyme in ciprofloxacin resistance and to detect the RAPD-PCR fingerprint of K. pneumoniae isolates. METHODS: A total of , 117 K. pneumoniae isolates were collected from hospitalized patients in three hospitals in Tehran, Iran, from August 2013 to March 2014. Antimicrobial susceptibility tests were performed by the disk diffusion method. Molecular identification and expression level of encoding quinolone resistance genes, acrA, acrB, qepA, and aac(6')-Ib-cr, were performed by PCR and real-- time PCR assays, respectively. All the K. pneumoniae isolates containing the mentioned genes were used simultaneously for RAPD-PCR typing. RESULTS: Colistin and carbapenems were the most efficient antibiotics against the clinical isolates of K. pneumoniae. PCR assay demonstrated that among the 117 isolates, 110 (94%) and 102 (87%) were positive for acrA and acrB gene and 5 (4%) and 100 (85%) isolates showed to have qepA and aac(6')-Ib-cr genes, respectively. Determination for AcrAB pump expression in 21% of strains demonstrated an increased expression, and the mean increase expression for acrB genes was 0.5-81. The results of RAPD-PCR reflected that in 95% CI, all isolates belonged to a clone. CONCLUSION: A high prevalence of genes encoding quinolone resistance in K. pneumoniae was detected in clinical samples. Therefore, the control of infection and prevention of drug-resistant bacteria spread need careful management of medication and identification of resistant isolates.202132888276
1624160.9998Detection of chromosomal and plasmid-mediated mechanisms of colistin resistance in Escherichia coli and Klebsiella pneumoniae from Indian food samples. OBJECTIVES: Numerous previous publications on the detection of bacterial isolates harbouring the mcr-1 gene from animals and humans strongly suggest an underlying route of transmission of colistin resistance via the food chain. The aim of this study was to investigate the presence of colistin-resistant (Col-R) bacteria in Indian food samples and to identify the underlying mechanisms conferring colistin resistance. METHODS: Raw food material, including poultry meat, mutton meat, fish, fruit and vegetables, collected from food outlets in Chennai, India, were processed to identify Col-R bacteria using eosin methylene blue agar supplemented with colistin. Colistin minimum inhibitory concentrations (MICs) were determined by the broth microdilution method. PCR for the mcr-1 and mcr-3 genes was performed on Col-R Escherichia coli and Klebsiella pneumoniae isolates. Mutations in the mgrB gene were analysed in K. pneumoniae isolates. One representative mcr-1-positive E. coli was subjected to whole-genome sequencing. RESULTS: Of 110 food samples tested, 51 (46.4%) were positive for non-intrinsic Col-R Gram-negative bacteria. Three E. coli isolates were found to harbour mcr-1, whereas none were positive for mcr-3. Ten K. pneumoniae isolates had alterations in mgrB, with mutations in four and insertional inactivation in six. CONCLUSION: The presence of Col-R bacteria and the mcr-1 gene in raw food samples further complicates the antimicrobial resistance scenario in India. To the best of our knowledge, this is the first report in the global literature on mgrB mutation and its insertional inactivation conferring Col-R in K. pneumoniae from food samples.201930244040
2458170.9998Mutation in mgrB is the major colistin resistance mechanism in Klebsiella pneumoniae clinical isolates in Tehran, Iran. Colistin is considered as one of a last resort antimicrobial agent against multidrug-resistant Gram-negative bacteria including Escherichia coli and Klebsiella pneumoniae. However, the recent emergence of colistin resistance (ColR) worldwide that severely restricts therapeutic options is a serious threat to global public health. In this study we have investigated the molecular determinants in ColR K. pneumoniae isolates collected from clinical specimens. A total of 98 E. coli and 195 K. pneumoniae clinical isolates were collected from two hospitals from August 2018 to December 2019 in Tehran, Iran. Colistin susceptibility and minimum inhibitory concentrations (MIC) were determined according to the Clinical and Laboratory Standards Institute by disk diffusion method, and microdilution method, respectively. For isolates with colistin MIC ≥4 μg mL-1, PCR was performed for the detection of mcr-1 to mcr-4 genes. Moreover, nucleotide sequences of mgrB, phoP, phoQ, pmrA, and pmrB genes were determined by sequencing. Finally, the transcriptional level of pmrK and pmrC genes was evaluated by quantitative reverse transcription PCR (RT-qPCR). None of the E. coli isolates were resistant to colistin while 21 out 195 K. pneumoniae isolates were identified as resistant, 19 of which carried mutation in the mgrB gene. Three different mutations were observed in the pmrB gene in 3 K. pneumoniae isolates. None of the ColR isolates showed alternations in pmrA, phoP, and phoQ genes. Furthermore, none of the plasmid-encoding genes were detected. Transcriptional level of the pmrK gene increased in all ColR isolates meanwhile, pmrC overexpression was detected in 16 out 21 (76.19%) isolates. Eventually, all ColR isolates were susceptible to tigecycline. Our results demonstrated that the alternation of mgrB gene is the main mechanism related to colistin resistance among ColR K. pneumoniae isolates in this study.202235113039
933180.9998Molecular characterization and diversity of carbapenemases in Gram-negative bacteria in Libyan hospitals. INTRODUCTION: Antimicrobial resistance has become a major threat to public health, especially in developing countries, due to the uncontrolled consumption of antibiotics. This study aims to characterize antibiotic resistance genes in different bacteria recovered in different healthcare facilities in Libya. METHODOLOGY: 379 samples were recovered from various sources from different sites. 210 samples were able to grow on culture media. 133 Gram-negative carbapenem-resistant strains were recovered from clinical specimens (n = 64), and hospital environments (n = 69). Antibiotic susceptibility tests were performed to select carbapenem-resistant strains. Colistin resistance was tested by the UMIC method to determine the minimum inhibitory concentration. RT-PCR was conducted to detect the incidence of carbapenemases-encoding genes. RESULTS: Gram-negative bacteria showed a low susceptibility to carbapenems. Molecular investigations indicated that NDM-1 was the most prevalent in Enterobacteriaceae isolated from patients and hospital environment (n = 26, n = 41), followed by blaOXA-48 (n = 16, n = 15) and blaVIM (n = 3) from patients and blaKPC (n = 1) from hospital environment. Concerning A. baumannii, blaOXA-23 was detected in strains isolated from patients (n = 8) and hospital environment (n = 6), followed by blaNDM (n = 9) from patients and one from hospital environment. Carbapenem resistance in P. aeruginosa was encoded by modification in OprD encoding gene, such as IS (ISpa26), polymorphism, and a premature stop codon. CONCLUSIONS: Several carbapenem resistant Gram-negative bacteria were identified by the expression of different carbapenemases and the alteration of OprD.202540720466
1435190.9998Epidemiology, Phenotypic and Genotypic Characterization of Carbapenem-Resistant Gram-Negative Bacteria from a Libyan Hospital. Antimicrobial resistance, particularly resistance to carbapenems, has become one of the major threats to public health. Seventy-two isolates were collected from patients and hospital environment of Ibn Sina Hospital, Sirte, Libya. Antibiotic susceptibility tests, using the disc diffusion method and E-Test strips, were performed to select carbapenem-resistant strains. The colistin (CT) resistance was also tested by determining the minimum inhibitory concentration (MIC). RT-PCR was conducted to identify the presence of carbapenemase encoding genes and plasmid-mediated mcr CT resistance genes. Standard PCR was performed for positive RT-PCR and the chromosome-mediated CT resistance genes (mgrB, pmrA, pmrB, phoP, phoQ). Gram-negative bacteria showed a low susceptibility to carbapenems. Molecular investigations indicated that the metallo-β-lactamase New Delhi metallo-beta-lactamases-1 was the most prevalent (n = 13), followed by Verona integron-encoded metallo-beta-lactamase (VIM) enzyme (VIM-2 [n = 6], VIM-1 [n = 1], and VIM-4 [n = 1]) that mainly detected among Pseudomonas spp. The oxacillinase enzyme OXA-23 was detected among six Acinetobacter baumannii, and OXA-48 was detected among one Citrobacter freundii and three Klebsiella pneumoniae, in which one coharbored the Klebsiella pneumoniae carbapenemase enzyme and showed resistance to CT (MIC = 64 μg/mL) by modification in pmrB genes. In this study, we report for the first time the emergence of Pseudomonas aeruginosa carrying the bla(NDM-1) gene and belonging to sequence type773 in Libya. Our study reported also for the first time CT resistance by mutation in the pmrB gene among Enterobacteriaceae isolates in Libya.202337145891