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Ventilator-associated pneumonia (VAP) is a serious complication in critically ill patients, significantly increasing morbidity and mortality. One concerning organism behind VAP is Acinetobacter baumannii, a multidrug-resistant bacterium has ability to evade treatment, particularly with carbapenems, the last-line antibiotics. This is especially worrisome within the confines of Intensive Care Units (ICUs) of tertiary care hospitals, hubs for high-risk patients and potential reservoirs of antimicrobial resistance. This study focused on identifying carbapenem-resistant Acinetobacter baumannii using both phenotypic and genotypic methods. In 132 isolates of Acinetobacter baumannii, we observed 96% resistance to the cephalosporins while least resistance found to colistin and tigecycline. However, a concerning 51.5% of isolates exhibited carbapenem resistance. Phenotypically confirmation of carbapenem resistance detected in 47% isolates by Combined Disc Test and 51.5% isolates by Modified Hodge Test and E-test. Genotypic analysis with RT-PCR revealed a diverse array of resistance genes: blaIMP (33.82%), blaVIM (25%), blaOXA-Group (20.58%), and blaNDM (8.82%). These findings highlight the alarming prevalence of carbapenem-resistant Acinetobacter baumannii in healthcare settings.

ABSTRACT: Objectives: Pseudomonas aeruginosa (P. aeruginosa) stands out as a key culprit in the colonization of burn wounds, instigating grave infections of heightened severity. In this study, we have performed comparative whole genome analysis of a difficult to treat extensively drug resistant P. aeruginosa isolated from a burn patient in order to elucidate genomic diversity, molecular patterns, mechanisms and genes responsible for conferring antimicrobial resistance and virulence. Method: P. aeruginosa SHNIBPS206 was isolated from an infected burn wound of a critically injured burn patient. Whole genome sequencing was carried out and annotated with Prokka. Sequence type, serotype, antimicrobial resistance genes and mechanisms, virulence genes, metal resistance genes and CRISPR/Cas systems were investigated. Later, pangenome analysis was carried out to find out genomic diversity. Result: P. aeruginosa SHNIBPS206 (MLST 357, Serotype O11) was resistant to 14 antibiotics including carbapenems and harboured all four classes of beta lactamase producing genes: Class A (blaPME-1, blaVEB-9), Class B (blaNDM-1), Class C (blaPDC-11) and Class D (blaOXA-846). Mutational analysis of Porin D gave valuable insights. Several efflux pump, virulence and metal resistance genes were also detected. Pangenome analysis revealed high genomic diversity among different strains of P. aeruginosa. Conclusion: To our knowledge, this is the first report of an extensively drug resistant ST 357 P. aeruginosa from Bangladesh, which is an epidemic high-risk P. aeruginosa clone. Further research and in-depth comprehensive studies are required to investigate the prevalence of such high-risk clone of P. aeruginosa in Bangladesh.

Ventilator-associated pneumonia (VAP) is a serious complication in critically ill patients, significantly increasing morbidity and mortality. One concerning organism behind VAP is Acinetobacter baumannii, a multidrug-resistant bacterium has ability to evade treatment, particularly with carbapenems, the last-line antibiotics. This is especially worrisome within the confines of Intensive Care Units (ICUs) of tertiary care hospitals, hubs for high-risk patients and potential reservoirs of antimicrobial resistance. This study focused on identifying carbapenem-resistant Acinetobacter baumannii using both phenotypic and genotypic methods. In 132 isolates of Acinetobacter baumannii, we observed 96% resistance to the cephalosporins while least resistance found to colistin and tigecycline. However, a concerning 51.5% of isolates exhibited carbapenem resistance. Phenotypically confirmation of carbapenem resistance detected in 47% isolates by Combined Disc Test and 51.5% isolates by Modified Hodge Test and E-test. Genotypic analysis with RT-PCR revealed a diverse array of resistance genes: blaIMP (33.82%), blaVIM (25%), blaOXA-Group (20.58%), and blaNDM (8.82%). These findings highlight the alarming prevalence of carbapenem-resistant Acinetobacter baumannii in healthcare settings. [ABSTRACT FROM AUTHOR]

Multidrug-resistant Enterobacter strains that harbour Klebsiella pneumoniae carbapenemase and Extended Spectrum β-Lactamases genes are a therapeutic challenge in hospitals that have made infection control and antibiotic therapy difficult and use of natural compounds such as thymol is interesting in the treatment of infections. Sixty-seven Enterobacter spp. isolates were obtained from patients admitted to hospitals in Hamadan and Tehran. Antimicrobial susceptibility, ESBLs and KPC genes detection and biofilm production of isolates were measured. The results showed that the highest and lowest antibiotic resistance were related to ceftazidime (82.1%) and tigecycline (3%), respectively. Also, 79.1% of the isolates were MDR. The highest presence of genes was related to TEM (100%), SHV (82%), CTX-M (77%) and KPC (8.96%) genes. The MIC and MBIC of thymol were obtained 8-31 μg/ml and16-62 μg/ml, respectively. Use of thymol, as an aid in antibiotic therapy is recommended. [ABSTRACT FROM AUTHOR]

Copyright of Diyala Journal of Medicine is the property of Republic of Iraq Ministry of Higher Education & Scientific Research (MOHESR) and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Abstract Background Worldwide, come out and dissemination of extended-spectrum beta-lactamases (ESBLs) producing Enterobacteriaceae has been warning the efficacy of antibiotics to treat an infection. Hospital wastewaters were a reservoir of such kind of resistant bacteria. Currently, the predominant antibiotics used for the treatment of hospitalized patients infected by Gram-negative bacteria are the β-lactam antibiotics. Therefore, it is an important source to investigate the magnitude of ESBLs producing bacteria and their antimicrobial susceptibility pattern. This study aimed to determine the occurrence of ESBLs producing Enterobacteriaceae (ESBLs-pE) and their antibiotic susceptibility pattern in wastewater released from five governmental hospitals in Addis Ababa, Ethiopia. Methods A cross-sectional study was conducted from April 1 to May 31, 2020. A total of 100 wastewaters were collected from five governmental hospitals in Addis Ababa using a grap-sampling technique. All Enterobacteriaceae were screened for ESBLs production using cefotaxime and ceftazidime as per 29th CLSI guideline. Each screen positive for ESBLs production was confirmed by the combination disk method (CDT) and their antibiotic susceptibility pattern was done using the Kirby–Bauer disk diffusion method on Muller Hinton agar (MHA). Data were entered and summarized using SPSS version 20 software. Results Of all Enterobacteriaceae, 48.3% were confirmed ESBLs-pE. The highest ratio of ESBLs-PE was observed in the adult ward (66.7%) and laundry unit effluent (58.8%). The highest ESBL producers were E. coli (21.8%) and K. pneumoniae (4.8%). The most elevated resistance level of ESBL producers were observed to cefotaxime (95.8%) and amoxicillin/clavunalate (93%). 64% of tested Enterobacteriaceae isolates were multi drug resistant (MDR). Conclusions Higher magnitude of MDR and ESBLs-pE were present in the hospital wastewater. The majority of them were in the adult ward and laundry unit effluents. The most frequent ESBLs-pE was among E.coli and K. pneumoniae. Hence, Consistent infection prevention and control procedures should be in practice at each ward/unit.

Antibiotic resistance is a global health problem. In particular, chickens are the source of antibiotic resistance and resistance genes. In this study, it was aimed to investigate the Gram-negative bacterial species that cannot ferment lactose from cloacal swap samples and to determine the antibiotic resistance profiles. Identification of the bacteria was performed with MALDI-TOF-MS and multidrug resistance was determined by disk diffusion test. In addition, the presence of ESBL, AmpC and carbapenemase in isolates was investigated by CLSI directions. Totally, 27 isolates were collected and 20 of them were Escherichia coli, 4 of them were E. fergusonii and 1 isolate of Pseudomonas fulva, Aeromonas media, Serratia marcescens. 19 different antibiotic discs from 7 different classes were used in the study and 63% of bacteria had resistance to antibiotics from 3 or more classes. While the highest resistance rate was observed in tetracycline (74.07%), all isolates were found that sensitive to imipenem. While carbapenemase could not be detected in any isolate, it was observed that P. fulva had ESBL and AmpC. Also, PCR was conducted for blaCTX-M, CIT, blaKPC genes in P. fulva. It was found that the bacterium had only blaCTX-M gene. As a result, although it is pleasing to find low presence of beta lactamase, high multidrug resistance has been determined in bacteria. This situation suggests newer therapeutic approaches. In addition, considering the "One Health" concept, antibiotic resistance should be constantly monitored with the interaction of the animal-human-environment. Because the development of resistance is in a constant state of change between bacteria. [ABSTRACT FROM AUTHOR]

The evolution of new biochemical activities frequently involves complex dependencies between mutations and rapid evolutionary radiation. Mutation co-occurrence and covariation have previously been used to identify compensating mutations that are the result of physical contacts and preserve protein function and fold. Here, we model pairwise functional dependencies and higher order interactions that enable evolution of new protein functions. We use a network model to find complex dependencies between mutations resulting from evolutionary trade-offs and pleiotropic effects. We present a method to construct these networks and to identify functionally interacting mutations in both extant and reconstructed ancestral sequences (Network Analysis of Protein Adaptation). The time ordering of mutations can be incorporated into the networks through phylogenetic reconstruction. We apply NAPA to three distantly homologous β-lactamase protein clusters (TEM, CTX-M-3, and OXA-51), each of which has experienced recent evolutionary radiation under substantially different selective pressures. By analyzing the network properties of each protein cluster, we identify key adaptive mutations, positive pairwise interactions, different adaptive solutions to the same selective pressure, and complex evolutionary trajectories likely to increase protein fitness. We also present evidence that incorporating information from phylogenetic reconstruction and ancestral sequence inference can reduce the number of spurious links in the network, whereas preserving overall network community structure. The analysis does not require structural or biochemical data. In contrast to function-preserving mutation dependencies, which are frequently from structural contacts, gain-of-function mutation dependencies are most commonly between residues distal in protein structure.

Tuberculosis is one of the most frequent causes of death in humans worldwide. One of the primary reasons tuberculosis remains a public health threat is that diagnosis can take weeks to months, is often not very sensitive and cannot be accomplished in many remote environments. A rapid, sensitive and inexpensive point-of-care (POC) diagnostic would have a major impact on tuberculosis eradication efforts. The tuberculosis diagnostic system REFtb is based on specific detection of the constitutively expressed β-lactamase (BlaC) in Mycobacterium tuberculosis using a custom fluorogenic substrate designated as CDG-3. REFtb has potential as a diagnostic for tuberculosis that could be very inexpensive (

Genus Pectobacterium bacteria include important agricultural pathogens. Pectobacterium versatile isolates contain a chromosome-borne beta-lactamase, PEC-1. This enzyme is the closest relative of TEM-1, a plasmid-borne beta-lactamase widespread in the Enterobacterales. We performed bioinformatics and phenotypic analyses to investigate the genetic and phenotypic features of PEC-1 and its frequency and ability to spread within genus Pectobacterium. We also compared the characteristics of PEC-1 and TEM-1 and evaluated the likelihood of transfer. We found that blaPEC-1 was present principally in a small number of genetic environments in P. versatile. Identical blaPEC-1 genetic environments were present in closely related species, consistent with the high frequency of genetic exchange within the genus Pectobacterium. Despite the similarities between PEC-1 and TEM-1, their genetic environments displayed no significant identity, suggesting an absence of recent transfer. Phenotypic analyses on clonal constructs revealed similar hydrolysis spectra. Our results suggest that P. versatile is the main reservoir of PEC-1, which seems to transfer to closely related species. The genetic distance between PEC-1 and TEM-1, and the lack of conserved elements in their genetic environments, suggest that any transfer that may have occurred must have taken place well before the antibiotic era. [ABSTRACT FROM AUTHOR]

Objective Antibiotic resistance by beta lactamase expression is a serious and growing threat. We aimed to determine whether beta-lactamase activity is detectable in urine specimens to enable faster identification of resistance. Methods Urine specimens from patients with extended spectrum beta lactamase (ESBL)-expressing urinary infections were incubated with beta lactam antibiotics. Beta lactam hydrolysis was determined by mass spectrometry methods. Results Ceftriaxone hydrolysis was observed in 45 of 45 ESBL-containing specimens from patients not treated with a beta lactamase inhibitor before specimen collection. Ceftriaxone hydrolysis was not observed in 108 of 108 non-ESBL-containing specimens. Spiking studies show that beta lactam hydrolysis can be observed within 30 minutes. Beta lactam hydrolysis is evidenced by mass spectrometry preceded by either liquid chromatography or matrix-assisted laser desorption ionization specimen processing methods. Conclusion Clinically significant beta lactamase activity is detectable directly from urine specimens. The described methods would enable the detection of beta lactam resistance 24 to 48 hours sooner than culture based methods. [ABSTRACT FROM AUTHOR]

Objective: To determine frequency and pattern of extended spectrum beta lactamase in Klebsiella pneumoniae and E. coli in bacterial cultures. Study Design: Cross Sectional study. Setting: Department of Pathology Sahara Medical College Narowal. Period: January to June 2020. Material & Methods: Total 1100 bacterial isolates were studied out of them 655 E.coli and 445 of klebsiella pneumoniae. All samples were subjected to double disc diffusion method using third generation cephalosporin and amoxacillin-clavulanic acid for detection of ESBL. Data was analyzed using SPSS software version 24, and results were calculated in the form of frequency, percentage and standard deviation. P-value =0.05 was taken as statistically non-significant. Results: There were 48.9% male and 51.1% female subjects out of 1100 total cases. E.coli was detected in 59.5% and klebsiella in 40.5% samples. Total 44.2% samples were positive for ESBL enzyme. Of 655 E.coli samples 40.8% and of 445 Klebsiella pneumoniae samples 49.2% were positive for ESBL enzyme. Conclusion: It is necessary to detect ESBL positive Klebsiella pneumoniae and E.coli in laboratory workflow to avoid unnecessary use of antibiotics and development of resistance against them. [ABSTRACT FROM AUTHOR]

Copyright of Al-Mustansiriyah Journal for Pharmaceutical Sciences is the property of Republic of Iraq Ministry of Higher Education & Scientific Research (MOHESR) and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Aim: Carbapenem-resistant (CR-Kp) strains are important nosocomial pathogens worldwide. In this study, we aimed to reveal the antibiotic resistance of clinical CR-Kp strains and determine the presence of KPC, OXA-48, VIM and IMP carbapenemase genes. , TEM-1, SHV-1 extended-spectrum beta-lactamase (ESBL) genes, , , plasmid-mediated quinolone resistance genes and sul1 and sul2 sulfonamide resistance genes provided molecular epidemiological data. Methods: A total of 175 strains were isolated from clinical samples of patients hospitalised in an intensive care unit (ICU) betweent April and October 2017. The strains were identified with conventional methods, with VITEK 2 (BioMerieux, France) and MALDI-TOF MS (Bruker, USA). Antimicrobial susceptibilities were tested using the disc-diffusion method and E-test (BioMerieux, France). Antimicrobial resistance genes were investigated via real-time PCR in strains identified as CR-Kp.Results: High frequencies of (86.36%), (86.36%), and (95.45%) genes were found in CR-Kp strains. Morever, all three ESBL genes coexisted in 77.3% of all strains. was detected in 12 (54.55%) of the strains, and 4 of them which had an MIC> 16 μg/mL to imipenem showed positivity as well. The gene determinant (86.36%) had the highest frequency, and strains carrying showed higher MICs for ciprofloxacin. Conclusion: CR-Kp strains are able to develop different antimicrobial resistance patterns according to regional changes in antimicrobial therapeutic policies. Thus, it is important to monitor the regional molecular epidemiological data for efficient treatment.

A new member of the class metallo-β-lactamase (MBL), New Delhi metallo-beta-lactamase 1 (NDM-1) has emerged recently as a leading threat to the treatment of infections that have spread in all major Gram-negative pathogens. The enzyme inactivates antibiotics of the carbapenem family, which are a mainstay for the treatment of antibiotic-resistant bacterial infections. This review provides information about NDM-1 spatial structure, potential features of the active site, and its mechanism of action. It also enlists the inhibitors/compounds/drugs against NDM-1 in various development phases. Understanding their mode of inhibition and the structure-activity relationship would be beneficial for development, synthesis, and even increasing biological efficacy of inhibitors, making them more promising drug candidates. [ABSTRACT FROM AUTHOR]

In this study, fresh seafood in retail markets was investigated for the antibiotic susceptibility patterns of the faecal indicator Escherichia coli and distribution of important β-lactamase encoding genes. E. coli were isolated from 50 (37 fish and 13 shellfish) fresh seafood samples and studied with respect to the phenotypic and genotypic characters of antibiotic resistance. Of 475 E. coli isolates from fresh seafood, 71.58% exhibited extended-spectrum β-lactamase (ESBL)-positive phenotypes. A high percentage of isolates were resistant to indicator cephalosporins cefotaxime (95%), cefpodoxime (90.88%) and ceftazidime (90.29%). Relatively higher susceptibilities were recorded against imipenem (74.41%), cefoxitin (66.76%) and meropenem (51.18%). The multiple antibiotic resistance (MAR) index of 97.35% of the isolates was above 0.18. The ESBL genes blaCTX-M, blaSHV and blaTEM were detected in 62.37%, 23.35% and 2.6% of E. coli isolates, respectively. The ESBL-producing isolates also harboured the metallo-β-lactamase-encoding genes blaOXA (7.06%), blaNDM (4.42%) and blaVIM (0.88%). This study highlights the risk of dissemination of multidrug resistant E. coli in seafood consumer communities and also the need to improve the hygiene of the coastal waters, landing centres and the retail markets. [ABSTRACT FROM AUTHOR]

SCUBA divers are predisposed to otitis externa caused by Pseudomonas aeruginosa, which is becoming increasingly multi-drug resistant (MDR). The present work assessed the antibiotic resistance profiles of P. aeruginosa obtained from SCUBA divers and their environment in Sodwana Bay, South Africa. Bacterial isolates from a total of 137 random water and ear swab samples were identified using biochemical and molecular methods. P. aeruginosa strains were further evaluated for antibiotic susceptibility using the Kirby–Bauer assay. Double disk synergy test (DDST) to confirm metallo-β-lactamase (MBL) production and PCR amplification of specific antibiotic resistance genes was performed. All (100%) 22 P. aeruginosa isolates recovered were resistant to 6 of the β-lactams tested including imipenem but exhibited susceptibility to trimethoprim–sulfamethoxazole. MBL production was observed in 77% of isolates while the most prevalent extended-spectrum β-lactamase (ESBL) genes present included blaAmpC (86.9%) followed by blaTEM (82.6%). Sulfonamide resistance was largely encoded by sul1 (63.6%) and sul2 (77.3%) genes with a high abundance of class 1 integrons (77.3%) of which 18.2% carried both Intl1 and Intl2. P. aeruginosa found in Sodwana Bay exhibits multi-drug resistance (MDRce) to several pharmaceutically important drugs with the potential to transfer antibiotic resistance to other bacteria if the judicious use of antibiotics for their treatment is not practiced.

Introduction: Resistance to penicillin among Staphylococcus aureus species is very widespread. One of the most important resistance mechanisms toward penicillin in S. aureus is blaZ production. Detection of beta lactamase production is possible using different methods like disk diffusion, iodometric, acidometric and molecular (PCR) methods. The aim of this study was to detection of beta lactamase production among S. aureus isolated from human dental plaque using iodometric and molecular methods. Descriptive statistics were done using ANOVA test and the statistical package, SPSS, Version 15.0. Methods: In this cross sectional study, 40 samples of human dental plaques was collected through forceps. Identification and isolation of S. aureus from samples was carried out using cultural methods, biochemical and molecular tests. Then, antibiotic resistance patterns of studied Staphylococci were determined using Kirby-Bauer method. Also, betalactamase production among the bacteria was evaluated using different iodometric and molecular methods. Results: 30% (12) of studied samples contained Staphylococcus aureus. All Staphylococcus aureus strains had beta lactamase production ability using filter paper iodometric method. Also, all spesies had blaZ gene, which detected by polymerase chain reaction test. The results of Conclusions: The result of this study showed that molecular (PCR) and iodometric (filter paper) methods were more accurate in detection of penicillin resistant S. aureus.

Background: AmpC β-lactamase enzymes cause multi-drug resistance. Aim and Objectives: The purpose of this study is detection of AmpC enzymes in the isolated strains of E. coli and K. pneumonia in three educational hospitals in Hamadan (Iran). Materials and Methods: One hundred and forty strains of E. coli and K. pneumonia were isolated from the hospitals in Hamadan from March to September 2015. To detection of phenotypic AmpC activity, the AmpC detection disks were used. To detect genes encoding for AmpC PCR method was used. Results: Sixty eight isolates (48.5%) were resistant to third generation cephalosporin and of these 61 (43.5%) isolates had MIC ³ 8 μg/mL to cefoxitin. All 68 isolates were analyzed by AmpC detection disks, of which 10 (7.14%) isolates were AmpC- β-lactamase producers. By PCR method, 24 (35.2%) isolates had cit and 48 (70.5%) were positive for fox, 42 (61.7%) for ebc, 46 (67.6%) for mox genes. None of the dha and acc genes were identified. Conclusion: High resistance to cephalosporins has been observed among the clinical isolates. Due to the possibility of plasmid transferring of ampC genes between bacterial, changing consumption patterns of antibiotics and the treatment protocols is necessary.

Extended-spectrum beta-lactamase-producing Escherichia coli isolated from slaughtered broilers in retail market that sell live chickens in Erbil city, Iraq. Forty-one cloacal fecal samples from broiler caecum were investigated from January to April 2016. ESBLs strains were isolated using MacConkey agar supplemented with cefotaxime 1 mg/l and the isolates were identified phynotypically by biochemical tests, TBX agar and VITEK-2 compact system. A total of 34 Escherichia coli and 4 Proteus mirabilis were analysed for determination of ESBL/AmpC by disc diffusion test using antimicrobial 68DC MAST® ESβL discs group including cefpodoxime, cefpodoxime + ESBL inhibitor, cefpodoxime + AmpC inhibitor and cefpodoxime + ESBL inhibitor + AmpC inhibitor and 67DC MAST® ESβL discs group including cefpodoxime, cefpodoxime + clavulanate, ceftazidime, ceftazidime + clavulanate, cefotaxime and cefotaxime + clavulanate. The phenotypic results showed that in group 68DC discs 23.7% E. coli were resistant to cefpodoxime and in group 67DC discs 73.7% of E. coli and 7.9% of P. mirabilis were resistance to one or more of the cefpodoxime, ceftazidime and ceftazidime. Final results revealed that 78.0% of samples were ESBLs/ AmpC positive. This study is the first examination to determine phenorypically E. coli producing ESBLs/AmpC in broiler chickens in Iraq. Conclusion, the healthy broiler can be a major source of ESBLs/AmpC and the possibility that transmitted to humans through the food chain, direct contact and the surrounding environment raises the concerns about public health and safety of poultry meat and the negative consequences of drug therapy that causes the spread of antibiotic resistance.

ObjectivesTo describe the molecular characteristics of beta-lactamases in bloodstream-infection Escherichia coli isolated from elderly patients, and to determine the genotypic patterns of blaCMY–2 and blaADC–162.MethodsA total of 50 bloodstream-infection E. coli isolates were obtained from patients aged > 50 years at Shanghai Sixth People’s Hospital South Campus during 2015–2018. The isolates were subjected to beta-lactamase detection using phenotypic and molecular methods. Beta-lactamase genes were verified by sequencing and the phylogenetic relationships of the isolates were analyzed by multilocus sequence typing (MLST). The transferability of plasmids carrying blaCMY–2 and blaADC–162 genes was verified by conjugation experiments and plasmid replicon typing.ResultsEight beta-lactamase subtypes were detected in 50 isolates of bloodstream-infection E. coli. blaTEM–1 (21/50) was the most common beta-lactamase gene, followed by blaCTX–M–14 (8/50), blaOXA–27 (5/50), blaCTX–M–27 (3/50), blaCTX–M–65 (1/50), blaADC–162 (1/50), and blaCMY–2 (1/50). Of these, blaADC–162 (ST95-A), and blaCMY–2 (ST95-B2) have not previously been reported in bloodstream-infection E. coli. In 21 isolates, beta-lactamase genes were located on conjugative plasmids belonging to incompatibility groups FrepB (n = 7), FIA (n = 1), FIC (n = 2), K (n = 8), N (n = 1), and I (n = 1), and blaCTX–M was associated with the common elements ISEcp1, IS903, and IS26, but with special sequences (region V, region Y, and region W) for ISEcp1 in 14 isolates.ConclusionTo the best of our knowledge, this study provides the first molecular characterization of beta-lactamase genes in E. coli isolated from the bloodstream in elderly patients. Beta-lactamase genes were detected at a relatively high frequency in elderly patients with bloodstream E. coli infections. Plasmid replicon analysis showed that horizontal dissemination of beta-lactamase genes was mainly mediated by IncK and IncF plasmids, which could encode multidrug resistance genes. The study also provides the first report of ISAba1-blaADC–162-tnpA and ISEcp1-blaCTX–M–14-IS903-blaCMY–2-blc-sugE in E. coli, and demonstrates IncF plasmid-mediated blaADC–162 and blaCMY–2 gene dissemination among bacteria.

Objectives: To describe the molecular characteristics of beta-lactamases in bloodstream-infection Escherichia coli isolated from elderly patients, and to determine the genotypic patterns of bla CMY–2 and bla ADC–162. Methods: A total of 50 bloodstream-infection E. coli isolates were obtained from patients aged > 50 years at Shanghai Sixth People's Hospital South Campus during 2015–2018. The isolates were subjected to beta-lactamase detection using phenotypic and molecular methods. Beta-lactamase genes were verified by sequencing and the phylogenetic relationships of the isolates were analyzed by multilocus sequence typing (MLST). The transferability of plasmids carrying bla CMY–2 and bla ADC–162 genes was verified by conjugation experiments and plasmid replicon typing. Results: Eight beta-lactamase subtypes were detected in 50 isolates of bloodstream-infection E. coli. bla TEM–1 (21/50) was the most common beta-lactamase gene, followed by bla CTX–M–14 (8/50), bla OXA–27 (5/50), bla CTX–M–27 (3/50), bla CTX–M–65 (1/50), bla ADC–162 (1/50), and bla CMY–2 (1/50). Of these, bla ADC–162 (ST95-A), and bla CMY–2 (ST95-B2) have not previously been reported in bloodstream-infection E. coli. In 21 isolates, beta-lactamase genes were located on conjugative plasmids belonging to incompatibility groups FrepB (n = 7), FIA (n = 1), FIC (n = 2), K (n = 8), N (n = 1), and I (n = 1), and bla CTX–M was associated with the common elements IS Ecp 1, IS903, and IS26, but with special sequences (region V, region Y, and region W) for IS Ecp 1 in 14 isolates. Conclusion: To the best of our knowledge, this study provides the first molecular characterization of beta-lactamase genes in E. coli isolated from the bloodstream in elderly patients. Beta-lactamase genes were detected at a relatively high frequency in elderly patients with bloodstream E. coli infections. Plasmid replicon analysis showed that horizontal dissemination of beta-lactamase genes was mainly mediated by IncK and IncF plasmids, which could encode multidrug resistance genes. The study also provides the first report of IS Aba 1- bla ADC–162- tnp A and IS Ecp 1- bla CTX–M–14-IS903- bla CMY–2- blc - sug E in E. coli , and demonstrates IncF plasmid-mediated bla ADC–162 and bla CMY–2 gene dissemination among bacteria. [ABSTRACT FROM AUTHOR]

In this study, fresh seafood in retail markets was investigated for the antibiotic susceptibility patterns of the faecal indicator Escherichia coli and distribution of important β-lactamase encoding genes. E. coli were isolated from 50 (37 fish and 13 shellfish) fresh seafood samples and studied with respect to the phenotypic and genotypic characters of antibiotic resistance. Of 475 E. coli isolates from fresh seafood, 71.58% exhibited extended-spectrum β-lactamase (ESBL)-positive phenotypes. A high percentage of isolates were resistant to indicator cephalosporins cefotaxime (95%), cefpodoxime (90.88%) and ceftazidime (90.29%). Relatively higher susceptibilities were recorded against imipenem (74.41%), cefoxitin (66.76%) and meropenem (51.18%). The multiple antibiotic resistance (MAR) index of 97.35% of the isolates was above 0.18. The ESBL genes blaCTX-M, blaSHV and blaTEM were detected in 62.37%, 23.35% and 2.6% of E. coli isolates, respectively. The ESBL-producing isolates also harboured the metallo-β-lactamase-encoding genes blaOXA (7.06%), blaNDM (4.42%) and blaVIM (0.88%). This study highlights the risk of dissemination of multidrug resistant E. coli in seafood consumer communities and also the need to improve the hygiene of the coastal waters, landing centres and the retail markets.

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Background:The emergence and increase in the incidence of Extended-spectrum beta lactamase (ESBL) producing Escherichia coli has become an emerging challenge especially in hospitalized patients with UTI. The aim of the present study was to survey the frequency of bla CTX-M genotype in ESBL producing E. coli isolated from hospitalized patients with UTI and determination of their antibiotic resistance pattern.Material and methodsA total of 135 E. coli isolates were collected from isolated from patients with UTI. The isolates were subjected to confirmatory phenotype tests for the presence of ESBL. 75 E. coli isolates were confirmed as ESBL-positive by means of the Double disc synergy test. In vitro susceptibility of ESBL isolates to 15 antimicrobial agents amoxicillin, penicillin, ceftazidime, cefotaxime, cefoxitin, ceftriaxone, cefixime, cephalexin, co-trimoxazole, gentamicin, nalidixic acid, ciprofloxacin, nitrofourantoin, amikacin and imipenem was performed by Kirby-Bauer’s Disk diffusion method according to Clinical and Laboratory Standards Institute (CLSI, 2012) guideline. PCR method was used to identify bla CTX-M gene in 75 ESBL positive strains.Results:PCR and sequence analysis showed that 75 (55.5%) isolates produced bla CTX-M genes. In vitro susceptibility of ESBL producing E. coli showed that all of them were resistant to amoxicillin and penicillin and The rates of resistance to the majority of tested antibiotics varied between 61% to 100 %, with the exception of amikacin (14.7%) and imipenem (2.7%). Our results showed that the frequency of bla CTX-M was strikingly high (93.3%).Conclusion:These data confirmed that the frequency of bla CTX-M genes were high among E. coli isolated from patients with UTI. The trend of multidrug-resistant profile has been associated with bla CTX-M gene is alarming. Therefore, it is very important to establish a routine screening of ESBL in clinical isolates to prevent dissemination of resistant isolates in health care settings.

The study investigated the prevalence of potentially pathogenic and drug resistant Escherichia coli among drinking water sources in Jordan. A total of 109 confirmed E. coli isolates were analyzed. Antimicrobial susceptibility testing was done using the Kirby Bauer disk diffusion method. Phenotypic identification of extended spectrum beta-lactamase (ESBL) and carbapenemase production was done using the double disk synergy test and the modified Hodge test, respectively. Isolates’ plasmid profiles were determined by gel electrophoresis. PCR was used for detection of virulence and resistance genes. Overall, 22.0% of the isolates were potentially intestinal pathogenic E. coli (IPEC); namely enteroaggregative E. coli (16.5%), enteropathogenic E. coli (2.8%), enteroinvasive E. coli (1.8%), and enterohemorrhagic E. coli (0.9%). A third of the isolates were multi-drug resistant. The highest rates of antimicrobials resistance were observed against ampicillin (93.6%) and sulfamethoxazole/trimethoprim (41.3%). All isolates were susceptible to imipenem, meropenem, doripenem and tigecycline. The prevalence of ESBL and carbapenemase producers was 54.1% and 2.8%, respectively. BlaVIM was the most prevalent resistance gene (68.8%), followed by blaCTX (50.5%), blaTEM (45.9%), blaNDM (11%), blaKPC (4.6%), and blaSHV (0.9%). Fifty-eight (53.2%) isolates contained one or more plasmid ranging from 1.0 to 8.0 kbp. Overall, high prevalence of potentially pathogenic and resistant isolates was observed.

Introduction: Staphylococci release a large number of enzymes. Some of these, such as coagulase, beta lactamase, hemolysins and biofilms are considered indices of pathogenicity. The aim of the current study was based on the isolation and identification of Staphylococcus aureus and coagulase negative Staphylococci (CNS) strains from sheep sub clinical mastitis and examining their biofilm, beta lactamase, hemolysins production and antibiotic resistance pattern. Materials and methods: 55 Staphylococci strains were isolated from seventy cases of sheep subclinical mastitis. Thirty three were determined as Staphylococcus aureus (60%) and 22 (40%) as CNS. The hemolytic activity was evaluated by plating Staphylococci strains on 5% bovine blood agar. The biofilm assay was performed by using micro titer plates. Beta Lactamase production was detected by test tube iodometric technique and susceptibility to antimicrobial agents was determined for isolated strains by the disk diffusion method. Results: Twenty six (78.8%) S. aureus strains were biofilm producers. For CNS (59.9%) strains were positive in biofilm production. Two isolates (6.06%), of S. aureus were α, the same number β and 6 (18.2%) isolates were ∂ hemolysin producers. Six isolates of CNS (27.27%) were α and ten (45.45%) ∂ hemolysin producers. Sixteen S. aureus (48.5%) and five CNS (22.72%) isolates were positive in beta lactamase production. The isolated Staphylococci show a low sensitivity pattern to methicillin and streptomycin. Discussion and conclusion: A high percentage of strains make α toxin that play a role in S. aureus biofilm formation. Twenty one out of 33 (63.63%) isolated Staphylococci were biofilm producers that can have deleterious effects because biofilm formation is thought to play an important role in the survival of virulent strains of Staphylococci. Sixteen out of 33 (48.5%) isolated S. aureus were positive in beta lactamase test, Excluding resistant to methicillin, all of these isolates show a marked sensitivities to other examined beta lactam drugs. High percentage of hemolysins, biofilm and beta lactamase production by isolated Staphylococci, suggest an important role of these virulence factors in the pathogenesis of isolated Staphylococci from mastitis sheep milk samples.

Background: AmpC â-lactamase enzymes cause multi-drug resistance. Aim and Objectives: The purpose of this study is detection of AmpC enzymes in the isolated strains of E. coli and K. pneumoniae in three educational hospitals in Hamadan (Iran). Materials and Methods: One hundred and forty strains of E. coli and K. pneumoniae were isolated from the hospitals in Hamadan from March to September 2015. To detection of phenotypic AmpC activity, the AmpC detection disks were used. To detect genes encoding for AmpC PCR method was used. Results: Sixty eight isolates (48.5%) were resistant to third generation cephalosporin and of these 61 (43.5%) isolates had MIC ³ 8 μg/mL to cefoxitin. All 68 isolates were analyzed by AmpC detection disks, of which 10 (7.14%) isolates were AmpC- â-lactamase producers. By PCR method, 24 (35.2%) isolates had cit and 48 (70.5%) were positive for fox, 42 (61.7%) for ebc, 46 (67.6%) for mox genes. None of the dha and acc genes were identified. Conclusion: High resistance to cephalosporins has been observed among the clinical isolates. Due to the possibility of plasmid transferring of ampC genes between bacterial, changing consumption patterns of antibiotics and the treatment protocols is necessary. [ABSTRACT FROM AUTHOR]

Nicotinamide phosphoribosyl-transferases (NAMPT) are enzymes that play a role in targeting cancer metabolism, while beta lactamases are involved in bacterial resistance to beta-lactam antibiotics. Many protein inhibitors exhibit such property which is often correlated with their cellular potency. In order to understand such a phenomenon, the present article conducts an analysis of the dynamic behavior of complexes formed by the inhibitors, that is indolizine derivatives, with the studied enzymes. Both docking and molecular dynamics led to identification of their interactions and showed the mechanism of inhibition of the two studied enzymes. The differences in the behavior of ligand at the active sites of beta lactamases and nicotinamide phosphoribosyl-transferases are indicated by structural and enthalpy values. [ABSTRACT FROM AUTHOR]

Recently, the quick spreads of broad-spectrum beta-lactams antibiotic resistance in pathogenic strains of bacteria have become a major global health problem. These new emerging resistances cause ineffectiveness of antibiotics and increasing the severity of diseases and treatment costs. Among different and diverse resistance targets, we chose a class A beta lactamase, CTX-M-9, with the aim of identifying new chemical entities to be used for further rational drug design. Based on this purpose, a set of 5000 molecules from the Zinc database have been screened by docking experiments using AutoDock Vina software. The best ranked compound, with respect of the previously proved inhibitor compound 19 , was further tested by molecular dynamics (MD) simulation. Our molecular modeling analysis demonstrates that ZINC33264777 has ideal characteristics a potent beta lactamase CTX-M-9 inhibitor. In the free form of beta lactamase, NMR relaxation studies showed the extensive motions near the active site and in the Ω-loop. However, our molecular dynamics studies revealed that in the compound 1: beta lactamase complex, the flexibility of Ω-loop was restricted. [ABSTRACT FROM AUTHOR]

Introduction: Pseudomonas aeruginosa is one of the most important causes of opportunistic infections. As Pseudomonas aeruginosa is prevalent, its resistance to antibiotics is especially significant. This study aimed to determine the prevalence of antibiotic resistance in extended spectrum of beta lactamase (ESBL) strains. Material and Method: From a total of 499 samples of urine cultures, blood cultures, wounds, pulmonary secretions and other samples, 88 isolates of P. aeruginosa were collected in hospitals of Zahedan in 2014. Initially, ESBLproducing strains were identified and their antibiotic resistance was determined using disk diffusion method according to CLSI standards. Results: Of the 88 isolates of Pseudomonas aeruginosa, 51 isolates (96.62%) were beta-lactamase producing, with the highest resistance to Cefoxitin (30μg) and the least resistance to Aztreoname (30μg). Most ESBL strains were separated from women, that is, a significant relationship was observed between the presence of resistant strains and gender of patients admitted to hospital. Furthermore, a significant relationship was observed between resistance to some antibiotics and strains of Pseudomonas aeruginosa. However, no relationship was found between isolates taken from patients in various wards, types of sample and the isolated bacterium (P≤0.05). Conclusion: Because the prevalence of resistant strains of Pseudomonas aeruginosa is high, it is necessary to prescribe medication for and treat infections with greater care. [ABSTRACT FROM AUTHOR]

The aim of the study was to investigate vancomycin and high-level aminoglycoside resistance (HLAR) in Enterococcus species by phenotypic and genotypic methods. A hundred Enterococcus strains were included in the study. Antimicrobial susceptibilities of strains were investigated by automated system, betalactamase production was investigated by nitrocefin disks, vancomycin resistance and HLAR were investigated by gradient diffusion method (GDM) and disk diffusion method, respectively. For detection of vancomycin and high-level gentamicin resistance (HLGR) genes, polymerase chain reaction was used. Teicoplanin linezolid, vancomycin, ampicillin, penicillin are the most susceptible antibiotics and strains were detected not to produce beta lactamase. Vancomycin resistance was detected in ten isolates by automated system and in only five isolates by GDM. Five isolates carrying VanA gene were determined. The ratio of HLGR and high-level streptomycin resistance was found 40 and 63% respectively. aac (6’)-1eaph (2’’)-1a gene was detected in 58% of strains. E. faecium strains were found more resistant to the antibiotics than the other species. Beta lactamase was detected in none of strains. The automated system detected vancomycin resistance in more strains than GDM. Therefore it is concluded that strains, which were detected to be resistant to vancomycin, should be confirmed by GDM. The ratio of VanA gene in strains is consistent with other studies. The HLAR ratio was found in about half of strains. The ratio of aac(6’)-1e-aph(2’’)-1a gene, which is the most reported gene in our country and other countries and one of the HLGR genes investigated in our study, was detected 58%.

Over the past two decades Branhamella catarrhalis (now known as Moraxella catarrhalis) has come out as a major pathogenic bacteria in humans. Different researches have uncovered its association in respiratory tract infections (e.g., otitis media, sinusitis, pneumoniaandbronchitis). In children eye infections while in adultlaryngitis, bronchitis, and pneumonia commonly occurred. There is rising number of beta lactamase positive strains in this genus, we designed a study project in which we evaluated the sensitivity pattern of Moraxella catarrhalis isolated from sputum. Total 124 sputum samples were analysed. Confirmation was done by using chocolate agar a typical golden yellow which can be distinguished by hockey puck like colony. Further identifications were done by gram staining, oxidase test &other biochemical tests. Antibiogram was done by CLSI method. Sensitivity pattern showed Amoxicillinclavunate v 99%, Teicoplanin 49%, Ceftrioxone 01%, Erythromycin 30%, Trimethoprim -sulphamethoxazole 04%, Ciproflaxacin 50%,Cefexatin 84%. To control the resistivity of this organism some preventive measurements should be taken.

A 5-year old Holstein Friesian cross breed cow was presented to Madras Veterinary College Teaching Hospital with the history of reduced milk yield. Clinical examination of udder revealed normal milk color and soft udder. The milk pH was 7.0, with California Mastitis Test score 3+, Electrical Conductivity 270U, and Somatic Cell Count as 328,000. Isolation and identification of causative agent revealed Methicillin Resistant Staphylococcus aureus (MRSA) from the sub-acute mastitis sample. Agar disc diffusion method for antimicrobial susceptibility revealed that the MRSA was sensitive to Enrofloxacin, Gentamicin, Oxytetracycline and Amoxicillin+Sulbactam. On the other hand, the isolate was resistance to Amoxicillin, Penicillin G, Ceftriaxone and Methicillin. The isolate was positive for β-lactamase resistance by Nitrocefin test. The MRSA was confirmed for the presence of mecA and blaZ target genes by polymerase chain reaction (PCR). The cow was treated with Enrofloxacin, Vitamin E and inorganic Selenium, and was recovered after 5 days of post-treatment.

Background: The resistance of Pseudomonas aeruginosa strains to broad spectrum cephalosporins may be mediated by extended spectrum b-lactamases (ESBLs). These enzymes are encoded by different genes located either on chromosome or plasmids. In this study, we determined the antimicrobial resistance patterns of P. aeruginosa isolates and screened for ESBL production. Methods: After isolation from burn patients in Tehran Hospital, identification of P. aeruginosa isolates were assessed using biochemical tests. We then performed disk agar diffusion (DAD) according to CLSI guidelines to determine the pattern of antimicrobial resistance. The frequency of ESBLs and prevalence of the OXA-10 and PER-1 genes were determined with combined disk and polymerase chain reaction (PCR) methods, respectively. Results: One hundred strains of P. aeruginosa were isolated. The resistance of these strains to cephpodoxime, aztreonam, ciprofloxacin, ofloxacin, ceftazidime, cefepime, imipenem, meropenem, cefotaxime, levofloxacin, piperacilin- tazobactam and ceftriaxon was 100%, 90%, 83%, 92%, 85%, 88%, 63%, 66%, 98%, 89%, 70% and 91%, respectively. Of these, 40 strains (40%) were ESBL positive, 29 strains (29%) were OXA-10 positive and 18 strains (18%) were PER-1 positive. Conclusion: Our results confirm the need for proper antimicrobial therapy in burn hospitals, considering the resistance pattern and frequency of strains producing ESBLs and the presence of the OXA-10 and PER-1 genes. Since an increase in the prevalence of ESBL in P. aeruginosa strains might lead to the transfer of these ESBL genes to other gram-negative bacteria, we recommend the use of appropriate drugs, especially cephalosporins, in burn hospitals.

Some Acinetobacter species have emerged as very important opportunistic pathogens in humans. We investigated Acinetobacter spp. from the polluted urban riverine environment in Croatia in regard to species affiliation, antibiotic resistance pattern, and resistance mechanisms. Considerable number of isolates produced acquired extended-spectrum β-lactamase(s) (ESBLs), CTX-M-15 solely or with TEM-116. By Southern blot hybridization, bla was identified on plasmids ca. 10, 3, and 1.2 kb in Acinetobacter junii, A. gandensis, and A. johnsonii. The bla-carrying plasmid in A. gandensis was successfully transferred by conjugation to azide-resistant Escherichia coli J53. A. radioresistens isolate also carried an intrinsic carbapenemase gene bla with IS Aba1 insertion sequence present upstream to promote its expression. Majority of ESBL-producing isolates harbored integrases intI1 and/or intI2 and the sulfamethoxazole resistance gene sul1. Almost all isolates had overexpressed resistance-nodulation-cell division (RND) efflux system, indicating that this mechanism may have contributed to multidrug resistance phenotypes. This is the first report of environmental CTX-M-15-producing Acinetobacter spp. and the first identification of CTX-M-15 in A. johnsonii, A. junii, A. calcoaceticus, A. gandensis, A. haemolyticus, and A. radioresistens worldwide. We identified, also for the first time, the environmental Acinetobacter-producing TEM ESBLs, highlighting the potential risk for human health, and the role of these bacteria in maintenance and dissemination of clinically important antibiotic resistance genes in community through riverine environments. [ABSTRACT FROM AUTHOR]

The aim of the study was to investigate vancomycin and high-level aminoglycoside resistance (HLAR) in Enterococcus species by phenotypic and genotypic methods. A hundred Enterococcus strains were included in the study. Antimicrobial susceptibilities of strains were investigated by automated system, betalactamase production was investigated by nitrocefin disks, vancomycin resistance and HLAR were investigated by gradient diffusion method (GDM) and disk diffusion method, respectively. For detection of vancomycin and high-level gentamicin resistance (HLGR) genes, polymerase chain reaction was used. Teicoplanin linezolid, vancomycin, ampicillin, penicillin are the most susceptible antibiotics and strains were detected not to produce beta lactamase. Vancomycin resistance was detected in ten isolates by automated system and in only five isolates by GDM. Five isolates carrying VanA gene were determined. The ratio of HLGR and high-level streptomycin resistance was found 40 and 63% respectively. aac (6')-1eaph (2")-1a gene was detected in 58% of strains. E. faecium strains were found more resistant to the antibiotics than the other species. Beta lactamase was detected in none of strains. The automated system detected vancomycin resistance in more strains than GDM. Therefore it is concluded that strains, which were detected to be resistant to vancomycin, should be confirmed by GDM. The ratio of VanA gene in strains is consistent with other studies. The HLAR ratio was found in about half of strains. The ratio of aac(6')-1e-aph(2")-1a gene, which is the most reported gene in our country and other countries and one of the HLGR genes investigated in our study, was detected 58%. [ABSTRACT FROM AUTHOR]

Gene duplication is important in evolution, because it provides new raw material for evolutionary adaptations. Several existing hypotheses about the causes of duplicate retention and diversification differ in their emphasis on gene dosage, subfunctionalization, and neofunctionalization. Little experimental data exist on the relative importance of gene expression changes and changes in coding regions for the evolution of duplicate genes. Furthermore, we do not know how strongly the environment could affect this importance. To address these questions, we performed evolution experiments with the TEM-1 beta lactamase gene in Escherichia coli to study the initial stages of duplicate gene evolution in the laboratory. We mimicked tandem duplication by inserting two copies of the TEM-1 gene on the same plasmid. We then subjected these copies to repeated cycles of mutagenesis and selection in various environments that contained antibiotics in different combinations and concentrations. Our experiments showed that gene dosage is the most important factor in the initial stages of duplicate gene evolution, and overshadows the importance of point mutations in the coding region. [ABSTRACT FROM AUTHOR]