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147400.9892Simple, rapid, and cost-effective modified Carba NP test for carbapenemase detection among Gram-negative bacteria. PURPOSE: Detection of carbapenemases among Gram-negative bacteria (GNB) is important for both clinicians and infection control practitioners. The Clinical and Laboratory Standards Institute recommends Carba NP (CNP) as confirmatory test for carbapenemase production. The reagents required for CNP test are costly and hence the test cannot be performed on a routine basis. The present study evaluates modifications of CNP test for rapid detection of carbapenemases among GNB. MATERIALS AND METHODS: The GNB were screened for carbapenemase production using CNP, CarbAcineto NP (CANP), and modified CNP (mCNP) test. A multiplex polymerase chain reaction (PCR) was performed on all the carbapenem-resistant bacteria for carbapenemase genes. The results of three phenotypic tests were compared with PCR. RESULTS: A total of 765 gram negative bacteria were screened for carbapenem resistance. Carbapenem resistance was found in 144 GNB. The metallo-β-lactamases were most common carbapenemases followed by OXA-48-like enzymes. The CANP test was most sensitive (80.6%) for carbapenemases detection. The mCNP test was 62.1% sensitive for detection of carbapenemases. The mCNP, CNP, and CANP tests were equally sensitive (95%) for detection of NDM enzymes among Enterobacteriaceae. The mCNP test had poor sensitivity for detection of OXA-48-like enzymes. CONCLUSION: The mCNP test was rapid, cost-effective, and easily adoptable on routine basis. The early detection of carbapenemases using mCNP test will help in preventing the spread of multidrug-resistant organisms in the hospital settings.201728966495
209610.9878Investigation of isepamicin in vitro efficiency in Gram negative bacteria efficacy of isepamicin. CONTEXT: Isepamicin is a new semisynthetic aminoglycoside derived from gentamicin B and it is effective against Gram negative bacteria. Antibiotic resistance is an emerging problem and new options need for the treatment of infections caused by Gram negative bacteria. AIMS: In this study we aimed to investigate the in vitro efficiency in carbapenem susceptible and nonsusceptible Enterobacterales and Pseudomonas aeruginosa. METHODS AND MATERIAL: A total of 214 isolates of Gram-negative bacteria (Enterobacterales n = 129 and P. aeruginosa n = 85). Identification of the bacteria was tested in Vitek MS (Biomeriux, France). Susceptibility of isepamicin, amikacin, gentamicin, tobramycin and netilmicin was determined by Kirby Bauer disc diffusion method. The breakpoints for susceptibility to isepamicin, amikacin, gentamicin, streptomycin, tobramycin and netilmicin were evaluated according to the Comité de l'Antibiogramme dela Société Française de Microbiologie (CA-SFM) and EUCAST, respectively. Aminoglycoside modifying enzyme (AME) genes were investigated by multiplex PCR method. RESULTS: Isepamicin susceptibility was determined as 92.3% for Enterobacterales and 67% for P. aeruginosa and 94.4% for carbapenem resistant Enterobacterales. The most common AME gene was aac (6')-Ib in both Enterobacterales (76%) and P. aeruginosa (14.1%). Seven of the isepamicin intermediate or resistant isolates were positive aac (6')-Ib in Enterobacterales and P. aeruginosa. CONCLUSIONS: In this study, isepamicin showed good efficiency against both susceptible and carbapenem nonsusceptible Enterobacterales. But amikacin was prior to isepamicin P. aeruginosa isolates. Isepamicin could be a therapeutic option for the infections caused by Enterobacterales.202133610258
141420.9878Prevalence and antimicrobial susceptibility of extended-spectrum beta-lactamase-producing bacteria in intensive care units of Sanandaj general hospitals (Kurdistan, Iran). This study focused on analyzing the spread of extended-spectrum beta-lactamase (ESBL) enzymes among Gram-negative bacteria at intensive care units (ICUs). Between January 2007 and January 2008, 301 consecutive clinical isolates of Gram-negative type were isolated. Of these, 66 strains were collected from patients in ICUs in two major hospitals in Sanandaj (Kurdistan, Iran). The isolates were identified, tested for antimicrobial susceptibility, and analyzed for the presence of ESBL using the double-disk synergy test. Isolates with a positive ESBL phenotype were subjected to PCR for SHV, TEM, OXA and CTX-M beta-lactamase gene families. Sixty-six Gram-negative bacteria were isolated from clinical samples of 66 ICU patients. These isolates included 16 Escherichia coli, 28 Enterobacter spp., 5 Pseudomonas spp., 10 Klebsiella pneumoniae, 3 Serratia marcescens and 1 Stenotrophomonas maltophilia. Twenty-three (34.85%) of these isolates were ESBL producing. The ESBL genes detected were SHV, TEM, OXA-1, OXA-2 and CTX-M. The results show the presence of ESBL genes among Gram-negative bacteria in the ICU setting of Sanandaj's hospitals. There is a need to institute a strict hospital infection control policy and regular surveillance of bacterial resistance to antimicrobial agents.200919521074
148530.9878Evaluation of Verigene Blood Culture Test Systems for Rapid Identification of Positive Blood Cultures. The performance of molecular tests using the Verigene Gram-Positive and Gram-Negative Blood Culture nucleic acid tests (BC-GP and BC-GN, resp.; Naosphere, Northbrook, IL, USA) was evaluated for the identification of microorganisms detected from blood cultures. Ninety-nine blood cultures containing Gram-positive bacteria and 150 containing Gram-negative bacteria were analyzed using the BC-GP and BC-GN assays, respectively. Blood cultures were performed using the Bactec blood culture system (BD Diagnostic Systems, Franklin Lakes, NJ, USA) and conventional identification and antibiotic-susceptibility tests were performed using a MicroScan system (Siemens, West Sacramento, CA, USA). When a single strain of bacteria was isolated from the blood culture, Verigene assays correctly identified 97.9% (94/96) of Gram-positive bacteria and 93.8% (137/146) of Gram-negative bacteria. Resistance genes mecA and vanA were correctly detected by the BC-GP assay, while the extended-spectrum β-lactamase CTX-M and the carbapenemase OXA resistance gene were detected from 30 cases cultures by the BC-GN assay. The BC-GP and BC-GN assays showed high agreement with conventional identification and susceptibility tests. These tests are useful for rapid identification of microorganisms and the detection of clinically important resistance genes from positive Bactec blood cultures.201626904669
140340.9878Evaluation of the AusDiagnostics MT CRE EU assay for the detection of carbapenemase genes and transferable colistin resistance determinants mcr-1/-2 in MDR Gram-negative bacteria. OBJECTIVES: To evaluate the AusDiagnostics MT CRE EU assay for the detection of carbapenemase and acquired colistin resistance genes in Gram-negative bacteria. METHODS: The assay allows the detection of blaKPC, blaOXA-48-like, blaNDM, blaVIM, blaIMP, blaSIM, blaGIM, blaSPM, blaFRI, blaIMI, blaGES (differentiating ESBL and carbapenemase variants), blaSME and mcr-1/-2. It was evaluated against a panel of isolates including Enterobacteriaceae, Pseudomonas spp. and Acinetobacter spp. retrospectively (n = 210) and prospectively (n = 182). RESULTS: The CRE EU assay was able to detect 268/268 carbapenemase genes, with 239 belonging to the 'big five' families (KPC, OXA-48-like, NDM, VIM and IMP) and 29 carbapenemase genes of the SIM, GIM, SPM, FRI, IMI, SME and GES families. It could distinguish between ESBL and carbapenemase variants of GES. It also allowed detection of mcr-1/-2 colistin resistance genes on their own or in isolates co-producing a carbapenemase. CONCLUSIONS: The AusDiagnostics MT CRE EU assay offered wide coverage for detection of acquired carbapenemase genes. It required minimal hands-on time and delivered results in less than 4 h from bacterial culture.201830189011
146350.9877Identification of colistin resistance and its bactericidal activity against uropathogenic gram negative bacteria from Hayatabad Medical Complex Peshawar. OBJECTIVES: Identification of colistin resistance and its bactericidal activity against gram-negative bacteria isolated from urinary tract infection (UTI) patients. METHODS: This 6-month cross sectional study was conducted in Hayatabad Medical Complex Peshawar from January 2019-June2019.. A total of 2000 urine samples were collected and transported to the Health Research Institute, NIH, Research Centre, Khyber Medical College Peshawar. Samples were streaked on different media and incubated at 37C° for 24hrs. Gram negative bacteria were identified through gram staining and Analytical Profile Index (API) 10s. Gram negative bacteria were subjected under antibiotic sensitivity profile through Kirby-Bauer disc diffusion method. Colistin resistance was found through broth microdilution method. Minimum bactericidal activity was performed to find out the lowest concentration of colistin required to kill gram-negative bacteria. RESULTS: A total of 241(12.05%) uropathogenic gram negative bacteria were isolated and identified from 2000 urine samples while excluding intrinsically resistant bacteria. After broth microdilution, colistin resistance was found in 48(19.9%) Escherichia coli, 4(1.6%) Klebsiella pneumoniae and 3(1.3%) Pseudomonas aeruginosa respectively. Colistin resistant Escherichia coli were resistant to 77% Cephalosporins, 81% to Fluoroquinolones and 70% to Penicillin combinations. Colistin resistant Klebsiella pneumoniae were 100% resistant to Cephalosporins, Penicillin combinations and Fluoroquinolones while 75% were resistant to Carbapenems and Monobactams. Pseudomonas aeruginosa isolates were sensitive to all used antibiotics. CONCLUSION: E.coli was the mainly responsible uropathogen causing UTIs. Colistin resistance was found in 22.8% gram negative uropathogens. Klebsiella pneumoniae isolates exhibited highest resistance to antibiotics.202235634614
148860.9876Evaluation of an automated rapid diagnostic assay for detection of Gram-negative bacteria and their drug-resistance genes in positive blood cultures. We evaluated the performance of the Verigene Gram-Negative Blood Culture Nucleic Acid Test (BC-GN; Nanosphere, Northbrook, IL, USA), an automated multiplex assay for rapid identification of positive blood cultures caused by 9 Gram-negative bacteria (GNB) and for detection of 9 genes associated with β-lactam resistance. The BC-GN assay can be performed directly from positive blood cultures with 5 minutes of hands-on and 2 hours of run time per sample. A total of 397 GNB positive blood cultures were analyzed using the BC-GN assay. Of the 397 samples, 295 were simulated samples prepared by inoculating GNB into blood culture bottles, and the remaining were clinical samples from 102 patients with positive blood cultures. Aliquots of the positive blood cultures were tested by the BC-GN assay. The results of bacterial identification between the BC-GN assay and standard laboratory methods were as follows: Acinetobacter spp. (39 isolates for the BC-GN assay/39 for the standard methods), Citrobacter spp. (7/7), Escherichia coli (87/87), Klebsiella oxytoca (13/13), and Proteus spp. (11/11); Enterobacter spp. (29/30); Klebsiella pneumoniae (62/72); Pseudomonas aeruginosa (124/125); and Serratia marcescens (18/21); respectively. From the 102 clinical samples, 104 bacterial species were identified with the BC-GN assay, whereas 110 were identified with the standard methods. The BC-GN assay also detected all β-lactam resistance genes tested (233 genes), including 54 bla(CTX-M), 119 bla(IMP), 8 bla(KPC), 16 bla(NDM), 24 bla(OXA-23), 1 bla(OXA-24/40), 1 bla(OXA-48), 4 bla(OXA-58), and 6 blaVIM. The data shows that the BC-GN assay provides rapid detection of GNB and β-lactam resistance genes in positive blood cultures and has the potential to contributing to optimal patient management by earlier detection of major antimicrobial resistance genes.201424705449
140570.9876The threat of carbapenem resistance in Eastern Europe in patients with decompensated cirrhosis admitted to intensive care unit. BACKGROUND: Multidrug-resistant organisms are an increasing concern in patients with decompensated cirrhosis. AIM: We aimed to evaluate the prevalence of infections with carbapenem-resistant Enterobacteriaceae in patients with decompensated cirrhosis. METHODS: Patients with decompensated cirrhosis admitted to ICU were included. The isolated Enterobacteriaceae strains were tested for carbapenemase-producing genes using the Roche LightMix® Modular VIM/IMP/NDM/GES/KPC/OXA48-carbapenemase detection kit. RESULTS: 48 culture-positive infections were registered in 75 patients with acutely decompensated cirrhosis. Thirty patients contracted a second infection. 46% of bacteria isolated at admission and 60% of bacteria responsible for infections identified during ICU-stay were multiresistant. ESBL+ Enterobacteriaceae were predominant at admission, while carbapenem-resistance was dominant in both Enterobacteriaceae and Non-Fermenting-Gram-Negative Bacteria responsible for infections diagnosed during hospitalisation. OXA 48 or KPC type carbapenemases were present in 30% of the analyzed Enterobacteriaceae and in 40% of the phenotypically carbapenem-resistant Klebsiella pneumoniae strains. The length of ICU stay was a risk-factor for a second infection (p=0.04). Previous carbapenem usage was associated with occurence of infections with carbapenem-resistant Gram-negative bacteria during hospitalization (p=0.03). CONCLUSION: The prevalence of infections with carbapenem-resistant Enterobacteriaceae is high in patients with decompensated cirrhosis admitted to ICU. Carbapenemase-producing genes in Enterobacteriaceae in our center are bla(OXA-48) and bla(KPC).202235732546
142680.9875Phenotypic and genotypic detection of carbapenemase production among gram negative bacteria isolated from hospital acquired infections. OBJECTIVES: To identify the carbapenemase producing Gram-negative bacteria (GNB) by phenotypic methods and to confirm the presence of resistant genes using real-time polymerase chain reaction (PCR). METHODS: This was a prospective study carried out at the Department of Microbiology, Sri Venkata Sai Medical College and Hospital, Mahabubnagar, India, from March 2018-2021. All samples were screened for carbapenem resistance by disc diffusion method and the VITEK(®)2 compact system (bioMérieux, France). Detection of carbapenemase was carried out using RAPIDEC(®)CARBA NP test (Biomeriux Private Limited, South Delhi, India), screening for metallo-β-lactamases (MBL) was carried out by double disk synergy test (DDST), and genotypic characterization by real-time PCR. RESULTS: Among the 1093 Gram-negative bacilli identified, 220 (17.0%) were resistant to carbapenems by both tested methods. Carbapenemase detection using the RAPIDEC(®)CARBA NP test indicated that 207 (94.0%) were carbapenemase producers, of which 189 (91.2%) were MBL producers. The most common carbapenemase genes identified were New Delhi metallo-β-lactamase (NDM; 47.3%), followed by the co-existence of genes in combination of NDM, with Verona integron-mediated metallo-β-lactamase (VIM; 39.6%), VIM and oxacillin hydrolyzing enzymes-48 (OXA-48; 4.3%), and OXA-48 (1.4%).No gene of active on imipenem, Klebsiella pneumonia carbapenemase, VIM, or OXA-48 alone was detected. CONCLUSION: This study suggests routine carbapenem resistance testing among multi-drug resistant-GNBs, as most of these infections occur in hospitals. In addition, there is a possibility that these highly antibiotic-resistant genes could spread to other bacteria resulting in further dissemination.202235256490
143190.9874The using of the polymerase chain reaction for the detection of resistance genes in gram-negative bacteria in routine practice in a pediatric hospital. Objective - assessment of RT-PCR for the detection of carbapenem-resistance genes in gram-negative bacteria. A total, 499 strains of gram-negative microorganisms isolated in two pediatric hospitals in 2019-2020 were studied. Species identification was performed using MALDI-ToF mass-spectrometry (Bruker Daltonics, Germany). Meropenem and imipenem minimal inhibitory concentration (MIC) was determined by E-test method (BioMerieux, France). The presence of acquired carbapenemase genes of IMP, NDM, VIM, KPC, OXA-48, OXA-23, OXA-40, OXA-58-groups was determined by RT-PCR. Klebsiella pneumoniae (34%), Escherichia coli (4%), Serratia marcescens (6%) and other members of Enterobacterales (6%), also gram-negative non-glucose-fermenting bacteria Acinetobacter baumannii (14%), Pseudomonas aeruginosa (36%) were found among selected strains. Carbapenemase production was found in 385 isolates (77%). The main mechanism determining carbapenem resistance in P. aeruginosa was the production of blaVIM (100%). A. baumanii strains harbored OXA-23 (55%) and OXA-40 (45%) carbapenemases. The major determinant of carbapenem resistance in K. pneumoniae isolates was OXA-48 carbapenemase, detected in 63% strains, 13% of the strains possessed blaNDM-group, 16% isolates had a combination of blaNDM-group and blaOXA-48-like. Carbapenemase of KPC-group was found in 8% K. pneumoniae strains. OXA-48 carbapenemase prevailed (95%) among S. marcescens strains. Most of E. coli isolates harbored metallo-beta-lactamase NDM (89%). Other members of Enterobacterales most often had OXA-48 carbapenemase (57%), 39% of the isolates carried blaNDM-group. In one strain, a combination of blaNDM-group and blaOXA-48-like was discovered. RT-PCR is a fast and reliable method for the detection of acquired carbapenemases and can be recommended for routine use in bacteriological laboratories.202235320635
1428100.9874Carbapenem-resistant Gram-negative bacteria associated with catheter-related bloodstream infections in three intensive care units in Egypt. We aimed to identify the carbapenem-resistant Gram-negative bacteria (GNB) causing catheter-related bloodstream infections (CRBSI) in intensive care units (ICU) in a tertiary care Egyptian hospital, to study their resistance mechanisms by phenotypic and genetic tests, and to use ERIC-PCR for assessing their relatedness. The study was conducted over 2 years in three ICUs in a tertiary care hospital in Egypt during 2015-2016. We identified 194 bloodstream infections (BSIs); 130 (67.01%) were caused by GNB, of which 57 were isolated from CRBSI patients (73.84%). Identification of isolates was performed using conventional methods and MALDI-TOF MS. Antimicrobial susceptibility testing (AST) was done by disc diffusion following CLSI guidelines. Phenotypic detection of carbapenemases enzymes activity was by modified Hodge test and the Carba-NP method. Isolates were investigated for the most common carbapenemases encoding genes bla(KPC), bla(NDM), and bla(OXA-48) using multiplex PCR. Molecular typing of carbapenem-resistant isolates was done by ERIC-PCR followed by sequencing of common resistance genes. The overall rate of CRBSI in our study was 3.6 per 1000 central venous catheter (CVC) days. Among 57 Gram-negative CRBSI isolates, Klebsiella pneumoniae (K. pneumoniae) was the most frequently isolated (27/57; 47.4%), of which more than 70% were resistant to Meropenem. Phenotypic tests for carbapenemases showed that 37.9% of isolates were positive by modified Hodge test and 63.8% by Carba-NP detection. Multiplex PCR assay detected the bla(NDM) in 28.6% of the isolates and bla(KPC) in 26.8%, bla(NDM) and bla(KPC) were detected together in the same isolate in 5.6%, while bla(OXA-48)-like were not detected. ERIC-PCR detected limited genetic relatedness between K. pneumoniae isolates. Elevated resistance rates were observed to all antibiotics including carbapenems among K. pneumoniae isolates causing CRBSI. ERIC-PCR showed that the resistant isolates were mainly polyclonal. Our results call for reinforcement of antimicrobial stewardship and measures to prevent CRBSI.201829936619
1418110.9874Nosocomial infections and antimicrobial susceptibility patterns among patients admitted to intensive care unit of Imam Khomeini hospital in Ilam, Iran. INTRODUCTION: Nosocomial infections (NIs) are a major challenge worldwide. Identification of antibiotic resistance pattern extended spectrum beta-lactamases (ESBLs) and carbapenem-resistant Enterobacteriaceae (CRE) were the objectives of this study. METHODS: In this cross-sectional study, the antimicrobial susceptibility pattern of bacterial isolates collected from patients with NIs in ICU was determined. Overall, 42 Escherichia coli and Klebsiella pneumoniae isolates from different infection sites were used to determine phenotypic tests of ESBLs, Metallo-β-lactamases (MBLs) and CRE. Detection of ESBLs, MBLs and CRE genes were performed by the polymerase chain reaction (PCR) method. RESULTS: From 71 patients with NIs, 103 different bacterial strains were isolated. The most frequently isolated bacteria were E. coli (n = 29; 28.16%), Acinetobacter baumannii (n = 15; 14.56%), and K. pneumoniae (n = 13; 12.26%). Also, the rate of multidrug-resistant (MDR) isolates was 58.25% (60/103). Based on phenotypic confirmation tests, 32 (76.19%) isolates of E. coli and K. pneumoniae produced ESBLs, and 6 (14.28%) isolates were identified as CRE producers. PCR showed the high prevalence of the bla(CTX-M) (n = 29; 90.62%) in ESBL genes. In addition, bla(NDM) was detected in 4 (66.66%), bla(OXA-23) in 3 (50%), and bla(OXA-48) gene in 1 (16.66%) isolates. The bla(VIM), bla(KPC), and bla(IMP) genes were not detected in any of the isolates. CONCLUSION: The Gram-negative bacteria E. coli, A. baumannii, and K. pneumoniae with high resistance levels were the most common bacteria causing NIs in the ICU. This study for the first time identified bla(OXA-11), bla(OXA-23), and bla(NDM-1) genes in E. coli and K. pneumoniae in Ilam city of Iran.202337155016
1430120.9874Prevalence of multidrug-resistant Gram-negative bacteria from blood cultures and rapid detection of beta-lactamase-encoding genes by multiplex PCR assay. INTRODUCTION: This study aimed to determine the prevalence of multidrug-resistant Gram-negative bacteria (GNB) from blood cultures in a tertiary-care hospital and the multiplex PCR assay's ability to detect resistance genes. METHODS: A total of 388 GNB isolates obtained from hospitalized patients between November 2019 and November 2021 were included in the study. Antimicrobial susceptibility testing was done by VITEK 2 system and broth microdilution method. Beta-lactamase-encoding genes were detected by multiplex PCR assays, BioFire-Blood Culture Identification 2 (BCID2) panel (bioMérieux, France). Extended-spectrum beta-lactamases (ESBLs) were detected phenotypically with VITEK AST-GN71 card (bioMérieux, France). The isolates of GNB were classified into multidrug-resistant, extensively-drug-resistant, and pandrug-resistant categories, and their prevalence and distribution in different wards, including coronavirus diseases 2019 (COVID-19) intensive care units (ICU), were calculated. RESULTS: Results revealed that all isolates of Acinetobacter baumannii and Pseudomonas aeruginosa were multidrug-resistant as well as 91.6% of Enterobacter cloacae, 80.6% of Proteus mirabilis, and 76.1% of Klebsiella pneumoniae, respectively. In fermentative bacteria, bla(OXA-48-like) (58.1%), bla(NDM) (16.1%), bla(KPC) (9.7%) and bla(VIM) (6.5%) genes were detected. More than half of Enterobacter cloacae (58.3%) and Klebsiella pneumoniae (53.7%) produced ESBLs. Among non-fermenters, the bla(NDM) gene was carried by 55% of Pseudomonas aeruginosa and 19.5% of Acinetobacter baumannii. In the COVID-19 ICU, Acinetobacter baumannii was the most common isolate (86.1%). CONCLUSIONS: This study revealed high proportions of multidrug-resistant blood isolates and various underlying resistance genes in Gram-negative strains. The BCID2 panel seems to be helpful for the detection of the most prevalent resistance genes of fermentative bacteria.202238021186
1483130.9873Clinical Evaluation of the iCubate iC-GPC Assay for Detection of Gram-Positive Bacteria and Resistance Markers from Positive Blood Cultures. The iC-GPC Assay (iCubate, Huntsville, AL) is a qualitative multiplex test for the detection of five of the most common Gram-positive bacteria (Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Enterococcus faecalis, and Enterococcus faecium) responsible for bacterial bloodstream infections, performed directly from positive blood cultures. The assay also detects the presence of the mecA, vanA, and vanB resistance determinants. This study comparatively evaluated the performance of the iC-GPC Assay against the Verigene Gram-positive blood culture (BC-GP) assay (Luminex Corp., Austin, TX) for 1,134 patient blood culture specimens positive for Gram-positive cocci. The iC-GPC Assay had an overall percent agreement with the BC-GP assay of 95.5%. Discordant specimens were further analyzed by PCR and a bidirectional sequencing method. The results indicate that the iC-GPC Assay together with the iCubate system is an accurate and reliable tool for the detection of the five most common Gram-positive bacteria and their resistance markers responsible for bloodstream infections.201829899000
1436140.9873Characterisation of carbapenem-resistant Gram-negative organisms from clinical specimens in Yola, Nigeria. OBJECTIVES: This study aimed to identify carbapenem-resistant Gram-negative bacteria from clinical specimens of patients in Yola, Nigeria. METHODS: Routine clinical specimens were screened for the presence of carbapenem-resistant Gram-negative bacteria using chromogenic agar plates. Susceptibility of all presumptive isolates to carbapenems was tested by MIC and disk diffusion methods. Real-time PCR was used to test for the presence of carbapenemase genes. RESULTS: Screening of 1741 clinical specimens yielded 119 (6.8%) presumptive carbapenem-resistant Gram-negative bacteria. Antimicrobial susceptibility testing confirmed carbapenem resistance in 105 of these isolates. New Delhi metallo-β-lactamase (bla(NDM)) gene was detected in 26 isolates and Verona integron-encoded metallo-β-lactamase (bla(VIM)) gene was detected in four. The mechanism of resistance could not be identified in approximately two thirds of the carbapenem-resistant isolates. CONCLUSION: While bla(NDM) and bla(VIM) accounted for 28.6% of the resistance seen, further molecular-based studies are needed to characterise the other mechanisms of carbapenem resistance in these isolates.202031472281
1464150.9873Detection of TEM and CTX-M genes from ciprofloxacin resistant Proteus mirabilis and Escherichia coli isolated on urinary tract infections (UTIs). The multidrug resistant Gram negative bacteria (MDRGNB) is an emerging burden and now represents a daily challenge for the management of antimicrobial therapy in healthcare settings. The present study was aimed to detect the prevalence of TEM and CTX-M type genes from GNB on urinary tract infection (UTIs). The ciprofloxacin resistant uropathogens were detected by HEXA UTI 5 disc diffusion method. The phenotypic detection of uropathogens producing extended spectrum beta lactamases (ESBLs) was confirmed by double disc combination test (DDCT) and phenotype confirmation test (PCT). The prevalence of TEM and CTX-M genes of uropathogens was identified by multiplex PCR analysis. The in vitro antimicrobial susceptibility of E. coli producing ESBL (26), 21 isolates of P. mirabilis, 17 P. aeruginosa, 14 K. pneumoniae and 6 Enterobacter sp. were detected. Based on the extension of the cephalosporin zone edge towards augmentin disc in the DDST method proved 84% of the isolates were ESBL positive. Similar results were obtained in phenotypic confirmatory test (PCT) by the increases of ≥5 mm zone of inhibition in the combination disc when compared with ceftazidime disc alone. The prevalence of TEM and CTX-M genes were determined from multidrug resistance uropathogens (MDU) respectively as 83%, 75%, 71%, 63%, 60%, 55%, 54%, 50%. The most prevalent (TEM + CTX-M) genes were also detected in ciprofloxacin resistant strains P. mirabilis BDUMS1 (KY617768) and E. coli BDUMS3 (KY617770). Due to the increase of ESBL genes in uropathogens, sustained supervision for using favorable antibiotics and decreasing the infection is essential.201829778819
1461160.9873Phenotypic and Genetic Characterization of Carbapenemase and ESBLs Producing Gram-negative Bacteria (GNB) Isolated from Patients with Cystic Fibrosis (CF) in Tehran Hospitals. BACKGROUND: Cystic Fibrosis (CF) is an autosomal recessive genetic disorder in white populations caused by mutation in a gene that encodes Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. Since frequent respiratory tract infections are the major problem in patients with CF, obligation to identify the causative bacteria and determining their antibiotic resistance pattern is crucial. The purpose of this project was to detect Gram-negative bacteria (GNB) isolated from sputa of CF patients and to determine their antibiotic resistance pattern. MATERIALS AND METHODS: The sputum of 52 CF patients, treated as inpatients at hospitals in Tehran, was obtained between November 2011 and June 2012. Samples cultured in selective and non-selective media and GNB recognized by biochemical tests. Antimicrobial susceptibility testing to cephalosporins, aminoglycosides and carbapenems was performed by disk diffusion method and MICs of them were measured. For phenotypic detection of carbapenemase and ESBLs production, the Modified Hodge test, double disk synergy test and the combined disk methods were performed. Subsequently, the genes encoding the extended spectrum beta-lactamases (blaPER, blaCTX-M) and carbapenemases (blaIMP-1, blaGES, blaKPC, blaNDM, blaVIM-1, blaVIM-2, blaSPM, blaSIM) in Gram negative bacteria were targeted among the resistant isolates by using PCR. PFGE was used to determine any genetic relationship among the Pseudomonas aeruginosa isolated from these patients. RESULTS: Fifty five GNB were isolated from 52 sputum samples including Pseudomonas aeruginosa, Klebsiella ozaenae, Alcaligenes xylosoxidans, Achromobacter denitrificans, Klebsiella pneumonia and Stenotrophomonas maltophilia. The rates of resistance to different antibiotic were as follows: cefixime (%80), ceftriaxone (%43), ceftazidime (%45) and meropenem (%7). The prevalence of genes encoding the ESBLs and Carbapenemases among the the phenotypically positive strains were as follows: blaCTX-M (19), blaIMP-1 (2), blaVIM-1 (2) and blaVIM-2 (3) genes respectively. No other genes were detected. PFGE analysis revealed 8 genotypes. Six isolates had mutually 3 similar patterns. CONCLUSION: This study showed the existence of important ESBLs and carbapenemases genes among the GNB isolated from patients with CF. Continuous surveillance of ESBLs and Carbapenemases, also identification of their types, in bacteria isolated from these patients have an important clinical impact, since, it can often provide valuable information for effective infection control measures and for the choice of appropriate antimicrobial therapy.201424596716
2118170.9873Gram-negative bacteria as causative agents of ventilator-associated pneumonia and their respective resistance mechanisms. Ventilator-associated pneumonia (VAP) is a serious and common complication in patients admitted to intensive care unit (ICU) and contributes to mortality. Multidrug Gram-negative bacteria such as Acinetobacter baumannii, Pseudomonas aeruginosa, and Klebsiella pneumoniae are frequently associated with VAP in ICU. A prospective study was set up in three ICUs of the University Hospital Center Zagreb and one ICU in General Hospital Pula from September 2017 to March 2018. Antibiotic susceptibility was determined by broth microdilution method. Production of extended-spectrum β-lactamases (ESBLs) was determined by double-disk synergy test and carbapenemases by Hodge and carbapenem inactivation method (CIM). The genes encoding ESBLs, carbapenemases of class A, B and D and qnr genes were determined by PCR. In total 97 Gram-negative bacteria isolates were analyzed. P. aeruginosa demonstrated high resistance rates for imipenem and meropenem with 74% and 68% of resistant strains, respectively. Moderate resistance rates were observed for ceftazidime andpiperacillin/tazobactam, ciprofloxacin and gentamicin (44%). All except three A. baumannii isolates, were resistant to carbapenems and to all other antibiotics apart from colistin and amikacin. Eight A. baumannii isolates were positive for bla(OXA-23) and 12 for bla(OXA-24) genes. Four K. pneumoniae and two E. cloacae strains were ESBL positive and harboured group 1 of CTX-M β-lactamases. Three P. mirabilis strains were positive for plasmid-mediated ampC β-lactamase of CMY family. Two carbapenem-resistant K. pneumoniae harboured OXA-48 and one carbapenem-resistant E. cloacae VIM-1. A high proportion of multidrug-resistant P. aeruginosa, K. pneumoniae and extensively resistant A. baumannii was reported. Acquired resistance mechanisms, mainly production of carbapenemases and ESBLs were dominant in A. baumannii and K. pneumoniae, respectively. Resistance of P. aeruginosa isolates was more likely due to upregulation of efflux pumps or porin loss. A marked diversity of β-lactamases was identified in Enterobacteriaceae.202032729399
1404180.9872Evaluation of a DNA microarray for rapid detection of the most prevalent extended-spectrum β-lactamases, plasmid-mediated cephalosporinases and carbapenemases in Enterobacteriaceae, Pseudomonas and Acinetobacter. The dissemination of Gram-negative bacteria (GNB) producing extended-spectrum β-lactamases (ESBLs), plasmid-encoded cephalosporinases (pAmpCs) and carbapenemases is a matter of great clinical concern. In this study, we evaluated a new low-density DNA array 'Check-MDR CT103 XL' (Check-Points, Wageningen, The Netherlands) that identifies the most clinically relevant β-lactamase genes of ESBLs (blaTEM, blaSHV, blaCTX-M, blaBEL, blaPER, blaGES and blaVEB), pAmpCs (blaCMY-2-like, blaDHA, blaFOX, blaACC-1, blaACT/MIR and blaCMY-1-like/MOX) and carbapenemases (blaKPC, blaOXA-48, blaVIM, blaIMP, blaNDM, blaGIM, blaSPM and blaOXA-23, -24 and -58) in cultured bacteria. In total, 223 GNB isolates with well-characterised resistance mechanisms to β-lactams were analysed. A specificity and sensitivity of 100% were recorded for most bla genes, with a slightly lower signal observed for blaIMP. The Check-MDR CT103 XL array proved highly accurate for the identification of epidemiologically relevant ESBL, pAmpC and carbapenemase genes harboured in Enterobacteriaceae, Pseudomonas and Acinetobacter spp. The Check-MDR CT103 XL assay is a significant improvement compared with Check-MDR CT103 and it highlights the ability of this array to evolve rapidly to adjust to the current needs for the detection of resistance mechanisms to β-lactam agents.201627374747
1429190.9872Detection of blaKPC and blaGES Carbapenemase Genes in Klebsiella pneumoniae Isolated from Hospitalized Patients in Kashan, Iran. INTRODUCTION: Klebsiella pneumoniae carbapenemase (KPC)-producing bacteria are among the highly antimicrobial resistant gram negative bacteria and infections due to them are an increasingly major health problem worldwide. METHODS: In this study we have detected the blaKPC and blaGES carbapenemase genes in Klebsiella pneumoniae isolated from hospitalized patients in Kashan, Iran. In a cross-sectional study, a total of 181 K. pneumoniae isolates were recovered from clinical specimens during November 2013 to October 2014. RESULT: Antimicrobial susceptibility profiles were determined using disk diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and CLSI guidelines. Carbapenem-resistant K. pneumoniae isolates were identified. PCR method and sequencing were used for detection of blaKPC and blaGES carbapenemase genes. Of the 181 K. pneumoniae isolates, 35 (19.3%) were found to be resistant to imipenem and 150 (82.9%) were identified as MDR strains. Among carbapenems, the most resistant rate 39 (21.5%) was seen against ertapenem using disk diffusion method. Of K. pneumoniae isolates 21 (11.6%) and 42 (23.2%) carried blaKPC and blaGES genes, respectively and 19(10.5%) carried both genes simultaneously. CONCLUSION: The data of current study revealed that the frequency of resistance to carbapenems and production of carbapenemase enzymes especially GES type was high among clinical isolates of K pneumoniae in Kashan, Iran.201627527726