Extended-spectrum Beta-lactamase Genes Research Articles

Antibiotypes and genetic characteristics of fluoroquinolone- and beta-lactam-resistant Escherichia coli isolated from food-producing animals

Background and Aim: Farm animals, including cattle, have been implicated as antimicrobial-resistant bacterial pathogen reservoirs. This study aimed to determine the antimicrobial resistance profiles and genetic characteristics of cattle colonized by fluoroquinolone-resistant and extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in Ebonyi state, Nigeria. Materials and Methods: We randomly collected 100 fresh fecal samples from 100 cattle in major abattoirs and analyzed the samples using standard microbiological methods. Isolates were further characterized by polymerase chain reaction (PCR) using 16S rRNA sequence primers. Phenotypic detection of ESBL production was performed using the double disk synergy test. Antimicrobial susceptibility profiles of ESBL-producing Escherichia coli were determined using the disk diffusion method, whereas molecular characterization of ESBL- and fluoroquinolone-resistant genes was performed by PCR using specific primers. Results: A total of 20 (20%) ESBL-producing E. coli were isolated from 100 animal fecal samples. Isolates were generally multidrug-resistant (MDR) with a resistance rate of 100% to 45% to trimethoprim-sulfamethoxazole, tetracycline, amoxicillin, cephalosporins, and ciprofloxacin. The average multiple antibiotic resistance index values of the isolates ranged from 0.5 to 0.8. BlaTEM (75%), followed by blaCTX-M (20%) and blaSHV (5.0%) was the most predominant ESBL gene among the isolates. The Aac-lb-6-cr fluoroquinolone-resistant gene was harbored by 90% of the isolates, whereas Qnr was absent. Conclusion: This study showed a high frequency of MDR ESBL-producing E. coli harboring ESBL and fluoroquinolone-resistant genes in fecal samples of cattle with serious public health consequences if not adequately addressed. Keywords: Escherichia coli, extended-spectrum beta-lactamase genes, fluoroquinolone resistance genes, cattle, multidrug-resistance.

Prevalence and molecular characterization of Multidrug-Resistant ESBL-Producing E. coli in commercial poultry

Background: Rural poultry farmers are gradually changing from the backyard poultry production to intensive production, which made poultry more vulnerable to diseases and lead to indiscriminate use of antimicrobials in poultry production. As per OIE, FAO and WHO, multiple spread antimicrobial resistant pathogenic bacteria are a serious global human and animal health problem. Hence the study has been carried out to know the antimicrobial resistance status of poultry in Tirunelveli, Tamil Nadu, India. Methods: The study was aimed to find the occurrence of extended spectrum beta lactamases (ESBLs) producing E. coli from poultry origin. Caecum samples from the broiler (n=180), desi chicken (n=180) and Japanese quail (n=180) were processed for the isolation and phenotypic identification of ESBL E. coli. The isolates were tested for antimicrobial susceptibility test and characterization of ESBL and virulence genes done by PCR. Result: On phenotypic confirmation of ESBL producing E. coli by combined disk diffusion test (CDDT), a total of 9.63% samples were positive for extended spectrum beta lactamase producer (ESBL). Multi drug resistance (MDR) was observed in 38.23% of the broiler isolates, 27.27% and 57.14% in desi chicken and Japanese quail isolates, respectively. All the phenotypically confirmed ESBL producing E. coli isolates (n=52) were positive for uspA gene by PCR confirming the isolates as E. coli. The overall presence of the blaTEM gene was 57.69%, with broiler isolates having the highest prevalence of 67.47%, followed by desi chicken 36.36% and Japanese quail 42.85%. In contrast, the blaSHV gene was found in only 17.64% of the broiler isolates, but none were positive for the blaCTXM and blaOXA-1 gene.

Emerging Coproduction of AmpC Beta-lactamase in Extended-spectrum Beta-lactamase-producing Escherichia coli Clinical Isolates in Indonesia

ABSTRACT Introduction: Extended-spectrum beta-lactamase (ESBL) and AmpC beta-lactamase (AmpC-BL) in Enterobacteriaceae are a global threat. Coproduction of AmpC-BL and ESBL reduces therapeutic options, with more resistance against cephamycin and beta-lactamase inhibitor combinations. This study determines the proportion, incidence, and distribution of AmpC-BL-resistant genes in clinically isolated ESBL-producing Escherichia coli, completing its scarce data in Indonesia. Methods: The samples in this study were ESBL-producing E. coli from blood and urine specimens, confirmed by BD Phoenix semiautomatic examination combined with cefoxitin disk screening method. Confirmation tests of AmpC-BL used the AmpC-disk method and the polymerase chain reaction method. Results: Thirteen (27.8%) of 108 E. coli isolates were nonsusceptible to cefoxitin, and 3 (11.5%) isolates were confirmed to produce AmpC-BL and contained AmpC-BL CITM gene. Of the 3 AmpC-BL and ESBL coproduction isolates, one isolate was a copresentation of ESBL and AmpC-BL genes, namely blaTEM and CITM. Fisher’s exact test showed that the coproduction of AmpC-BL in ESBL-producing isolates was associated with reduced susceptibility to cefoxitin (P = 0.020) and amoxicillin-clavulanate (P = 0.048) compared to isolates producing ESBL alone. The effect of ESBL and AmpC-BL coproduction on reducing susceptibility to carbapenems needs further investigation. Conclusions: AmpC coproduction was found in 3 (2.8%) of 108 ESBL-producing E. coli isolates, and one isolate copresented AmpC-BL and ESBL coding genes blaTEM and CITM. These three isolates were associated with widened antibiotic resistance to cefoxitin and amoxicillin--clavulanate compared to isolates producing ESBL alone.

Investigation of antibiotic resistance, virulence genes, and biofilm formation of Escherichia coli isolated from sheep feces in Shiraz industrial slaughterhouse, South of Iran.

With the increase in human population, the consumption of livestock products such as sheep meat has also increased. Sheep are the reservoir and shedder of Escherichia coli that can be transmitted to humans. Aims: Characterization of fecal E. coli isolated from sheep in slaughterhouse. Stool specimens were collected from 30 apparently healthy sheep from different flocks in Shiraz industrial slaughterhouse. The resistance of E. coli isolates against 10 antibiotics was determined by disk diffusion method. The presence of three major extended spectrum beta-lactamase (ESBL) genes and five tetracycline resistance genes as well as seven virulence genes were investigated by polymerase chain reaction (PCR) technique. Using the microtiter plate method, the biofilm formation ability of E. coli isolates was investigated. The highest frequency of resistance was to amoxicillin (100%) followed by tetracycline (25%). All E. coli isolates were susceptible to gentamicin and nitrofurantoin, and only one isolate was resistant to the tested third-generation cephalosporins. Multidrug resistance phenotype was observed in 16.7% of the isolates. bla TEM (25%) was the most prevalent ESBL gene and tetA (62.5%) was the most prevalent tetracycline resistance gene in the isolates. crl, csgA, fimH, and bcsA genes were present in all isolates, and the prevalence of papC and afa genes was 95.8% and 83.3%, respectively. In total, 62.5% of the isolates were biofilm producers. According to the concept of One Health, the presence of virulent antibiotic-resistant biofilm producing strains of E. coli in sheep is a risk to public health.

Frequency of Fecal Carriage of ESBL Resistance Genes in Multidrug-Resistant Pseudomonas aeruginosa Isolates from Cancer Patients at Laquintinie Hospital, Douala, Littoral Region, Cameroon.

Background. Opportunistic infections are the second cause of death among cancer patients. This study aimed at determining the antimicrobial profile and the prevalence of extended-spectrum beta-lactamase (ESBL)-gene carriage of Pseudomonas aeruginosa isolates among cancer patients at the Douala Laquintinie Hospital, Littoral Region of Cameroon. Between October 2021 and March 2023, 507 study participants were recruited among whom 307 (60.55%) were cancer patients, compared to 200 (39.45%) noncancer patients. Fifty-eight P. aeruginosa isolates were isolated from fecal samples of forty-five cancer patients and thirteen noncancer patients using Cetrimide agar. The antimicrobial resistance profile of the isolates was determined using the Kirby-Bauer disk diffusion method. The polymerase chain reaction was used to detect the presence of extended-spectrum beta-lactamase genes among P. aeruginosa isolates. P. aeruginosa showed significant resistance rates in cancer patients compared to noncancer patients to imipenem, cefotaxime, and ceftazidime, piperacillin-tazobactam, ticarcillin-clavulanic acid, and ciprofloxacin. The multidrug resistance (MDR) rate was significantly (p < 0.05) higher in cancer patients than in noncancer patients. The frequency of beta-lactamase genes in the 58 ESBL-producing P. aeruginosa isolates was determined as 72.41% for bla TEM, 37.93% for bla OXA, 74.14% for blaCTX-M, and 44.83% for bla SHV genes. The study revealed an alarmingly high prevalence of fecal carriage of ESBL-producing P. aeruginosa with a high rate of MDR among cancer patients. It indicates that regular monitoring and surveillance of ESBL-producing P. aeruginosa among cancer patients are needed to improve the management of patients.

Characterization of extended-spectrum beta-lactamase and carbapenemase genes in bacteria from environment in Burkina Faso.

This study aimed to characterize extended-spectrum beta-lactamase (ESBL) and carbapenemase genes in bacteria from the environment in Bobo-Dioulasso, Burkina Faso. This study was conducted from January 18 to December 31, 2019. Environmental samples were collected from the effluents of Souro Sanou University Hospital Center and the wastewater treatment plant at Bobo-Dioulasso. MacConkey agar media supplemented with 4 µg/mL cefotaxime was used for bacterial growth, and identification of bacteria was performed using API 20E system (BioMerieux SA, Lyon, France). Antibiotic susceptibility testing, synergy test, carbapenem inactivation method and molecular characterization were performed. A total of 180 bacterial isolates were identified from the different sites with a predominance of Klebsiella oxytoca and Klebsiella pneumoniae (27.5%). All 180 bacterial isolates were ESBL producers and 18 (10.0%) of them produced carbapenemases. Out of the 180 bacterial isolates, DNAs of 98.9% (178/180) bacterial isolates were extracted and tested through polymerase chain reaction (PCR) for characterization of resistant genes. The study showed that 89.8% (160/178) carried the bla-CTX-M genes including 54.4 (87/160) from hospital effluents and 45.6 (73/160) from the wastewater treatment plant. Regarding the carriage of carbapenemase genes, 7.9 (14/178) blaNDM-1 was found in all the sites including 71.4% (10/14) from hospital effluents and 28.6 (4/14) from the wastewater treatment plant. blaOXA-48-like was only found in bacteria from hospital effluents and represented 2.2% (4/178). This study highlights the need to build hospital effluent treatment plants to reduce the load of resistant bacteria before discharging the effluents into the urban wastewater system.

Evaluation and clinical practice of pathogens and antimicrobial resistance genes of BioFire FilmArray Pneumonia panel in lower respiratory tract infections

BackgroundExisting panels for lower respiratory tract infections (LRTIs) are slow and lack quantification of important pathogens and antimicrobial resistance, which are not solely responsible for their complex etiology and antibiotic resistance. BioFire FilmArray Pneumonia (PN) panels may provide rapid information on their etiology.MethodsThe bronchoalveolar lavage fluid of 187 patients with LRTIs was simultaneously analyzed using a PN panel and cultivation, and the impact of the PN panel on clinical practice was assessed. The primary endpoint was to compare the consistency between the PN panel and conventional microbiology in terms of etiology and drug resistance, as well as to explore the clinical significance of the PN panel. The secondary endpoint was pathogen detection using the PN panel in patients with community-acquired pneumonia (CAP) or hospital-acquired pneumonia (HAP).ResultsFifty-seven patients with HAP and 130 with CAP were included. The most common pathogens of HAP were Acinetobacter baumannii and Klebsiella pneumoniae, with the most prevalent antimicrobial resistance (AMR) genes being CTX-M and KPC. For CAP, the most common pathogens were Haemophilus influenzae and Staphylococcus aureus, with the most frequent AMR genes being CTX-M and VIM. Compared with routine bacterial culture, the PN panel demonstrated an 85% combined positive percent agreement (PPA) and 92% negative percent agreement (NPA) for the qualitative identification of 13 bacterial targets. PN detection of bacteria with higher levels of semi-quantitative bacteria was associated with more positive bacterial cultures. Positive concordance between phenotypic resistance and the presence of corresponding AMR determinants was 85%, with 90% positive agreement between CTX-M-type extended-spectrum beta-lactamase gene type and phenotype and 100% agreement for mecA/C and MREJ. The clinical benefit of the PN panel increased by 25.97% compared with traditional cultural tests.ConclusionThe bacterial pathogens and AMR identified by the PN panel were in good agreement with conventional cultivation, and the clinical benefit of the PN panel increased by 25.97% compared with traditional detection. Therefore, the PN panel is recommended for patients with CAP or HAP who require prompt pathogen diagnosis and resistance identification.

Genomic analysis of Shigella isolates from Lebanon reveals marked genetic diversity and antimicrobial resistance.

In this study, we characterized 54 clinical isolates of Shigella collected in North Lebanon between 2009 and 2017 through phenotypic and genomic analyses. The most prevalent serogroup was S. sonnei, accounting for 46.3 % (25/54) of the isolates, followed by S. flexneri (27.8 %, 15/54), S. boydii (18.5 %, 10/54) and S. dysenteriae (7.4 %, 4/54). Only three isolates were pan-susceptible, and 87 % (47/54) of the isolates had multidrug resistance phenotypes. Notably, 27.8 % (15/54) of the isolates were resistant to third-generation cephalosporins (3GCs) and 77.8 % (42/54) were resistant to nalidixic acid. 3GC resistance was mediated by the extended-spectrum beta-lactamase genes bla CTX-M-15 and bla CTX-M-3, which were present on various plasmids. Quinolone resistance was conferred by single point mutations in the gyrA DNA gyrase gene, leading to GyrA S83L, GyrA D87Y or GyrA S83A amino acid substitutions. This is the first study, to our knowledge, to provide genomic insights into the serotypes of Shigella circulating in Lebanon and the various antimicrobial resistance determinants carried by these strains.

Identification of blaSHV and blaTEM extended spectrum beta-lactamase genes in Klebsiella pneumoniae in the dairy wastewater, East Java Province, Indonesia

Abstract. Dameanti FNAEP, Yanestria SM, Effendi MH, Plumeriastuti H, Tyasningsih W, Ugbo EN, Sutrisno R, Safri MAAS. 2023. Identification of blaSHV and blaTEM extended spectrum beta-lactamase genes in Klebsiella pneumoniae in the dairy wastewater, East Java Province, Indonesia. Biodiversitas 24: 6092-6099. Klebsiella pneumoniae infection in dairy cows causes various livestock production hazards and threatens public health because it acts as a Multi-Drug Resistance Bacteria (MDRB), especially Extended-Spectrum Beta-Lactamase (ESBL) strains. Livestock wastewater is a repository for commensal isolates including K. pneumoniae and potentially can be a reservoir for ESBL spreaders for the environment. The aim of this study was to identify the blaSHV and blaTEM ESBL genes in K. pneumoniae from wastewater of dairy cattle farms in East Java Province. This study used a sample of 342 dairy farm wastewater isolated on 2% BPW media from 6 cities/regencies with the highest population of dairy cows in East Java, Indonesia, namely Pasuruan District, Malang District, Tulungagung District, Blitar District, Batu City and Kediri District. Samples were identified on MCA media, Gram staining, EMBA, and biochemical tests. Confirmation of ESBL phenotypically used the Double Disc Synergy Test (DDST) method and the Polymerase Chain Reaction (PCR) test to detect the ESBL gene. The result showed that bacterial isolation and identification were 14.32% (49/342), confirmed K. pneumoniae. DDST confirmation results of ESBL-producing K. pneumoniae isolates from the wastewater of dairy farms in East Java obtained were 5.55% (19/342). The sample was continued with the PCR test showing positive for the blaSHV gene 63.1% (12/19) and blaTEM 31.57% (6/19). In conclusion, the ESBL gene blaSHV and blaTEM identification in K. pneumoniae isolates, indicate that wastewater from dairy farms has the potential to be a reservoir of ESBL carriers in the environment and pose a threat to public health in East Java Province, Indonesia.

Extended spectrum beta-lactamase carriage among elderly residents of a long-term care facility in Beirut

Antimicrobial resistance is an emerging problem worldwide, endangering antimicrobials efficacy and resulting in high rates of morbidity and mortality. It is one of the major concerns that health care facilities are facing nowadays. Mainly, extended-spectrum beta-lactamases (ESBL)-producing Enterobacterales play a role in hydrolyzing β-lactams, specifically the third-generation cephalosporins. This study aimed to investigate the prevalence of fecal carriage and molecular characterization of ESBL-producing Enterobacterales among Lebanese elderly residents in a long-term care facility (Dar Al-Ajaza Al Islamia Hospital). Rectal culture swab specimens were collected from 132 patients at Dar Al Ajaza Al Islamia hospital between January 2019 till June 2020. The phenotype of ESBL producers was confirmed by a modified double disc synergy test and antibiotic susceptibility was determined using the Kirby-Bauer disk diffusion method. Genotypically, multiplex polymerase chain reaction was used to detect the ESBL genes. The main Enterobacterales strain observed was E coli (90.15%) followed by Klebsiella pneumoniae (4.54%) and Klebsiella oxytoca (3.80%). It has been found that the ESBL percentage rate has decreased when compared to a study conducted previously at the same hospital. Moreover, the predominant ESBL gene wasCTX-M (cefotaximase). This study is the first to demonstrate the improved current status of ESBL in one long-term care facility. In addition, the CTX-M is still the major type in ESBL-producing organisms.

Role of CTX-M-15 gene in spread of extended-spectrum beta-lactamases among immunocompetent patients in Ghana.

Patients with faecal carriage of extended-spectrum beta-lactamases (ESBL)-producing Enterobacterales serve as reservoirs and sources of dissemination and infection. This report examined immunocompetent patients for faecal carriage of ESBL-producing Enterobacterales in a district care hospital setting in Ghana. Between March 2019 and May 2020, cross-sectional sampling was performed to enrol patients and conduct questionnaire-structured interviews for factors that predispose patients to ESBL faecal carriage. Faecal samples from study patients were quantified for ESBL-producing Enterobacterales. The ESBL genes were characterised by polymerase chain reaction and sequencing. The overall proportion of ESBL faecal carriage was 35.5% (n = 38/107). The blaCTX-M gene, mostly CTX-M-15, was detected in 89.5% (n = 34/38) of the ESBL-producing isolates. The other ESBL types included blaSHV (n = 3) and blaOXA (n = 1). The CTX-M-15-positive isolates, when present in a faecal sample compared to the non-ESBL-CTX-M-15 isolates, constituted the predominant faecal Enterobacterales, with significantly higher colony counts than all other enterobacteria in that sample. In multivariate regression, independent risk factors for faecal carriage of ESBL-producing Enterobacterales were hospitalisation in the past year, infections since admission, use of antibiotics in the past 6 weeks, and admission from another hospital. The study found that CTX-M-15-producing isolates were the predominant faecal Enterobacterales, and that further investigations are needed to determine the reasons behind this dominance. The CTX-M-15-producing isolates dominance in this study shows the misuse and abuse of antibiotics in an African medical facility and indicates the potential role of immunity in controlling ESBL spread, which is to be investigated further.

The diagnostic accuracy of the GeneXpert ESBL-ampC prototype assay for rapid PCR-based detection of extended-spectrum beta-lactamase genes directly from urine.

Early identification of complicated urinary tract infections caused by ESBL-producing Enterobacterales has the potential to limit the use of carbapenems to those patients without alternative antibiotic options and avoid the empirical use of carbapenems in patients without ESBL-producing bacteria. The purpose for such a test will differ by setting and ESBL prevalence rates. Countries with low ESBL rates and cephalosporins as empiric treatment (e.g., The Netherlands) will need a rule-in test to decide to use carbapenems, while countries with high ESBL rates and empiric carbapenem treatment will need a rule-out test for ESBLs to de-escalate therapy early. Anyway, such as a test would-at least theoretically-improve patient care and reduce selective pressure for the emergence of carbapenem resistance.

Detection of Antimicrobial Susceptibility Pattern and Molecular Detection of Resistance Genes in Klebsiella pneumoniae Isolated from Urine Samples of Suspected UTI Patients Attending Public Hospitals in Nasarawa South Senatorial District, Nasarawa State, Nigeria

Aims: The aim of the current study was to determine antimicrobial susceptibility patterns and the incidence of resistance genes (β-lactamase-encoded genes) among clinical isolates of K. pneumoniae.&#x0D; Study Design: Investigative.&#x0D; Place and Duration of Study: Department of Microbiology, Nasarawa State University, Keffi, between November 2022 and July 2023.&#x0D; Methodology: A total of 375 clinical samples were randomly collected from patients and isolation and identification was done using standard microbiological methods. The antibiotic susceptibility testing for the isolates was carried out in accordance with the Clinical and Laboratory Standards Institute protocol. The detection of extended-spectrum beta-lactamase (ESBL) production in K. pneumoniae isolates was carried out using double disc synergy test. In addition, molecular detection of ESBL genes was carried out using Polymerase Chain Reaction (PCR) method.&#x0D; Results: The overall incidence rate of K. pneumoniae in the 375 clinical samples from Nasarawa South Senatorial District was 38 (10.13%). Resistance was high to ampicillin (94.74%) and amoxicillin-clavulanic acid (73.6%) but low to cefotaxime (13.16%), ceftriaxone (18.42%), ceftazidime and cefoxitin (23.60%) and gentamicin (28.94%). Most of the isolates were multi-drug resistant (MDR; 81.5%) and none was Pan Drug resistant. All 15 isolates jointly resistant to cefotaxime and/or ceftazidime and ciprofloxacin 100.0% were ESBL producers, distributed as follows: blaCTX-M-1 7(46.6%); blaCTX-M-34(26.6%); blaSHV 6(40.0%); blaTEM6 (40.0%); and blaOXA 5(33.3%).&#x0D; Conclusion: K. pneumoniae isolated from the urine of patients attending public hospitals in Nasarawa South were less resistant to cefotaxime, ceftazidime, gentamicin, ceftriaxone, and cefoxitin in the study location. This implies that the antibiotics are useful in the treatment of infection caused by K. pneumoniae. Also, ESBL-positive K. pneumoniae isolates harbored ESBL genes, with blaCTX-M as the most common.

Genome Analysis of Klebsiella pneumoniae Reveals International High-Risk Pandemic MDR Clones Emerging in Tertiary Healthcare Settings in Uganda.

Klebsiella pneumoniae is a threat to public health due to its continued evolution. In this study, we investigated the evolution, convergence, and transmission of hypervirulent and multi-drug resistant (MDR) clones of K. pneumoniae within healthcare facilities in Uganda. There was high resistance to piperacillin (90.91%), cefuroxime (86.96%), ceftazidime (84.62%), cefotaxime (84.00%), amoxicillin/clavulanate (75%), nalidixic acid (73.68%), and nitrofurantoin (71.43%) antibiotics among K. pneumoniae isolates. The isolates were genetically diverse, consisting of 20 different sequence types (STs) and 34 K-serotype groups. Chromosomal fosA (for fosfomycin) and oqxAB efflux pump genes were detected in all isolates. Two carbapenem resistance genes, blaNDM-5 and blaOXA-181 plus extended-spectrum beta-lactamase (blaCTX-M-15) gene (68.12%), quinolone-resistant genes qnrS1 (28.99%), qnrB1 (13.04%), and qnrB6 (13.04%) and others were found. All, except three of the isolates, harbored plasmids. While the isolates carried a repertoire of virulence genes, only two isolates carried hypervirulent genes demonstrating a low prevalence (2.90%) of hypervirulent strains. Our study demonstrated genetically diverse populations of K. pneumoniae, low levels of carbapenem resistance among the isolates, and no convergence of MDR and hypervirulence. Emerging high-risk international pandemic clones (ST11, ST14, ST147, ST 86 and ST307) were detected in these healthcare settings which are difficult to treat.

Extended-Spectrum Beta-Lactamases Genes in Klebsiella pneumoniae Isolates Obtained from Patients in Intensive Care Units

Background: Drug-resistant hospital-acquired infections (HAIs) are a growing concern in modern medicine throughout the world. Klebsiella pneumoniae is one of the most prominent causative agents of multidrug-resistant nosocomial infections. It is also widely recognized for having a high resistance level to many antibiotic classes, particularly beta-lactams. Carbapenemase-producing K. pneumoniae has been identified as a major global cause of HAIs with adverse clinical outcomes. Therefore, it is of the utmost importance to have an in-depth understanding of the antimicrobial resistance (AMR) genetic determinants of this bacterium to stop the spread of highly resistant K. pneumoniae in healthcare facilities and the resulting patient morbidity and mortality. Objectives: This study aimed to investigate the AMR pattern of K. pneumoniae isolates obtained from intensive care units (ICUs), with a focus on extended-spectrum beta-lactamases (ESBLs) genes blaCTX-M, blaGES, and blaIMP. Methods: A total of 105 K. pneumoniae isolates obtained from the sputum samples of ICU patients were identified and confirmed using standard microbiological tests and 16S rRNA polymerase chain reaction (PCR). The antibiotic susceptibility test was performed for all the isolates. The presence of ESBL genes was determined phenotypically and by PCR. Results: The highest level of resistance was observed against ceftazidime (100%), cefotaxime (99%), and imipenem (93.3%). Approximately 87.6% and 39% of the isolates were sensitive to colistin and gentamicin, respectively. Phenotypic ESBL production was observed in 16 isolates, and the prevalence of blaCTX-M was 86.7%. No blaGES and blaIMP genes were detected. Conclusions: Periodic investigation of AMR-mediating genes is essential due to the high prevalence of ESBL genes in HAIs. The presence of other ESBL genes needs to be investigated for a more accurate understanding of the AMR status of K. pneumoniae in healthcare settings.

The notable relatedness between ESBL producing Enterobacteriaceae isolated from clinical samples and asymptomatic fecal carriers

BackgroundThe investigation of the presence of extended-spectrum beta-lactamase (ESBL) within Enterobacteriaceae in both fecal carriers and patients is an essential matter. Furthermore, the assessment of distinct characteristics exhibited by resistant bacteria obtained from fecal carriers and patients, as well as the comparison of these characteristics between the two groups, could provide a deeper understanding of how the resistant isolates can remain concealed within a dormant reservoir and intensify antimicrobial resistance. The aim of the present study was to concentrate on the comparison of the antimicrobial resistance pattern and molecular features between strains obtained from clinical and carrier sources.Material and methodsA total of 142 clinical samples and 120 rectal swabs were collected from June to October 2016. ESBL screening was performed using the double-disk synergy test. PCR was done for the detection of ESBL genes. Assessment of biofilm formation, virulence factor genes, and MLVA was performed for K. pneumonae isolates. Phylogroup typing was performed for E. coli isolates.ResultsOf 146 samples, 67.6% were E. coli, and 32.4% were K. pneumoniae. The rate of blaCTXM-15 was 89.4%. In K. pneumoniae type D, ompk35 and fimH were the highest. All the K. pneumoniae isolates were classified into 12 mini clusters and the clinical isolates were characterized into 7 mini clusters. The phylogroup B2 in ESBL-EC was the highest (56.2%).DiscussionComparison of molecular characteristics and clonal relatedness between fecal carriers and patients showed noticeable relatedness and similarity which may indicate that ESBL-KP can be colonized with the same profiles in different settings and, therefore, may be widely distributed in both community and hospital settings. Therefore, implementation of control protocols, including surveillance of the fecal carriers, could impressively reduce silent reservoirs without clinical symptoms as well as patient rates.

Characterization of integrons, extended spectrum beta lactamases and genetic diversity among uropathogenic Escherichia coli isolates from Kerman, south east of Iran.

The study aimed to investigate the distribution of genes encoding integrons, extended spectrum beta-lactamase (ESBL) in E. coli isolated from UTIs, as well as the genetic diversity among the isolates. E. coli isolates were recovered from the patients with UTI in Kerman Iran. Antibiotic susceptibility was done according to CLSI guidelines. The presence of ESBL genes and integrons was evaluated using PCR. PCR and sequencing were applied for the evaluation of cassette content of integrons. Genotyping of the isolates was performed by multiple-locus variable-number tandem repeat analysis (MLVA). Imipenem was the most effective antibiotic, while the highest resistance was observed to streptomycin. In total 40.2% of isolates were ESBL producers. Of 69 integron-positive isolates, 59 only had class I integrons, 4 only had class II integrons and 6 had both types. The most common gene cassette found within class I integrons was dfrA17-aadA5 (n=27). The E. coli isolates were divided into 16 MLVA clusters. The current study demonstrated the simultaneous presence of class I integrons and ESBLs involved in the resistance of UPEC isolates to antibacterial agents. Our finding also revealed that the E. coli isolates belonged to diverse clones.

The Prevalence of Genetic Determinants of Antibiotic Resistance, which are of Particular Epidemiological Consequences, in the Microbiota of the Oropharyngeal Swabs in Patients with Cystic Fibrosis

Relevance. Antibiotic resistance of microorganisms can contribute to the chronicity of the inflammatory process, lead to an increase in the cost of treating patients and make it difficult to eradicate the pathogen. Patients with cystic fibrosis constantly require medical supervision, regular visits to medical institutions, and therefore there is a high risk of infection with nosocomial antibiotic- resistant strains. In addition, the necessary intake of antibacterial drugs provides an advantage for the reproduction of resistant microorganisms.Aim. Comparison of the frequency of detection of antibiotic resistance determinants in oropharyngeal swabs in children with cystic fibrosis and conditionally healthy children using molecular biological methods.Materials and methods. A PCR study of oropharyngeal discharge from 100 children with cystic fibrosis and 100 children from the control (healthy comparison subject) group was performed. Genetic antibiotic resistance locus: metallo-b-lactamases of the VIM, IMP and NDM groups; carbapenemase genes of the KPC and OXA-48 groups; extended-spectrum beta-lactamase genes of the CTX-M group and the mecA gene were detected by polymerase chain reaction (PCR) with hybridization-fluorescence detection.Results and discussion. As a result of the analysis, a statistically significant increase in the frequency of detection of genetic determinants of antibiotic resistance in the microbiota of the oropharyngeal discharge in children with cystic fibrosis was found compared with healthy children (p&lt;0.001). The chances of detecting antibiotic resistance loci in the discharge of the oropharynx among children with cystic fibrosis are 38.5 times higher than among healthy children (95% CI: 5.1-289.5). In 28% of children with cystic fibrosis, DNA of the genetic determinants of antibiotic resistance was detected in the microbiome of the discharge of the oropharynx. A high percentage of the presence of genetic determinants of antibiotic resistance may be the reason for the ineffectiveness of antibiotic therapy.Conclusion. Due to the high occurrence in the microbiome of the oropharyngeal discharge of patients with cystic fibrosis of genetic antibiotic resistance locus that are of particular clinical and/or epidemiological significance, and the high risk of the spread of antibiotic-resistant strains outside medical institutions, it is necessary to include this group of patients in regular epidemiological monitoring.

Fecal carriage of extended spectrum beta-lactamase and fluoroquinolone resistant gene in non-typhoidal Salmonella enterica isolates from food-producing animals and humans

This study seeks to determine the feacal carriage of extended spectrum beta-lactamase and fluoroquinolone resistant non-typhoidal Salmonella enterica isolates from food-producing animals and humans. A total of three hundred (300) fecal samples were collected using sterile universal containers from food-producing animals namely (Chicken [100], Pig [100] and humans (100) from Onicha Local Government Area of Ebonyi State and analyzed for the presence of non-typhoidal Salmonella enterica using standard microbiological techniques. Phenotypic detection of extended-spectrum beta-lactamase (ESBL) were done by disc diffusion and Double Disk Synergy Test. Molecular characterization for ESBL and fluoroquinolone-resistant genes were done by PCR with specific primers. The result shows that non-typhoidal Salmonella species (NTS) accounted for 25 % and 17 % in poultry and pig fecal sample respectively while 60 % and 40% were phenotypic ESBL producers respectively. When compared statistically there is significant difference among isolates confirmed ESBL-positive (P˂ 0.05). Also, none of the 16 (58 %) NTS isolated from humans harbored ESBL phenotype. PCR analysis with β-lactam specific primer detected the presence of blaOXA 50 % and 50 %, blaSHV 36 %, and 64 %, blaTEM 43 % and 57 %, blaCTX-M 36 % and 64 % in poultry and pig respectively. Fluoroquinolone resistant gene QnrA was present in 0 and 100 % of poultry and pig respectively. QnrB was 40 % and 60 % in poultry and pig isolates respectively. QnrS was present in 64 % isolates of poultry and 13 % isolates in pig. The high prevalence of genes encoding beta-lactamases and fluoroquinolone resistance (TEM, SHV, CTX-M and OXA, (qnrA, qnrB and qnrS) were present more in poultry and pig than in humans and demonstrate a significant public health threat from consumption of food-producing animal harboring such pathogenic resistant genotype if not properly controlled.&#x0D; Keywords: Extended spectrum beta-lactamase, Fluoroquinolone, non-typhoidal Salmonella enterica, feacal carriage

Antimicrobial Resistance and Extended-Spectrum Beta-Lactamase Genes in Enterobacterales, Pseudomonas and Acinetobacter Isolates from the Uterus of Healthy Mares.

Antibiotic-resistant bacteria are a growing concern for human and animal health. The objective of this study was to determine the antimicrobial resistance and extended-spectrum beta-lactamase genes in Enterobacterales, Pseudomonas spp. and Acinetobacter spp. isolates from the uterus of healthy mares. For this purpose, 21 mares were swabbed for samples, which were later seeded on blood agar and MacConkey agar. The isolates were identified using MALDI-TOF and the antimicrobial susceptibility test was performed using the Kirby-Bauer technique. To characterize the resistance genes, a polymerase chain reaction (PCR) scheme was performed. Of the isolates identified as Gram-negative, 68.8% were Enterobacterales, represented by E. coli, Enterobacter cloacae, Citrobacter spp., and Klebsiella pneumoniae; 28.1% belonged to the genus Acinetobacter spp.; and 3.1% to Pseudomonas aeruginosa. A 9.3% of the isolates were multidrug-resistant (MDR), presenting resistance to antibiotics from three different classes, while 18.8% presented resistance to two or more classes of different antibiotics. The diversity of three genes that code for ESBL (blaTEM, blaCTX-M and blaSHV) was detected in 12.5% of the strains. The most frequent was blaSHV, while blaTEM and blaCTX-M were present in Citrobacter spp. and Klebsiella pneumoniae. These results are an alarm call for veterinarians and their environment and suggest taking measures to prevent the spread of these microorganisms.