Extended-spectrum Beta-lactamase Genes Research Articles

Epidemiological study of antimicrobial-resistant bacteria in healthy free-ranging bantengs (Bos javanicus) and domestic cattle.

Antimicrobial-resistant microorganisms (ARMs) have been increasing among wild animals. Interactions occurring at the interface between wildlife, humans, and livestock can lead to the transmission of ARMs. Thus, the prevalence of ARMs in wild and domestic animals should be determined to address and prevent this issue. This study aimed to determine the resistance patterns of cefotaxime (CTX)-resistant Escherichia coli and identify the presence of extended-spectrum beta-lactamase (ESBL) genes in ESBL-producing E. coli among a population of wild banteng (Bos javanicus) and domestic cattle kept on farms located close to the Lam Pao non-hunting area, Kalasin province, Thailand. Forty-five fecal samples were taken from wild bantengs inhabiting the Lam Pao non-hunting area in Thailand, alongside 15 samples from domestic cattle. Bacterial culture, triple sugar iron, and motile indole lysine tests were conducted to identify E. coli. A polymerase chain reaction (PCR) was conducted for specific confirmation. MacConkey agar supplemented with 2 μg/mL of CTX was used to identify CTX-resistant E. coli, which would be used to identify ESBL production based on a double-disk synergy test. Extended-spectrum beta-lactamase-producing samples were subjected to disk diffusion tests to determine resistant patterns, and the sizes of PCR bands and DNA sequencing were used to differentiate ESBL gene types. All samples tested positive for E. coli. Forty-five isolates from 15 banteng samples and three isolates from one domestic cattle sample displayed CTX-resistant and ESBL-producing traits. The banteng and domestic cattle populations exhibited nine and three distinct resistant patterns, respectively. The PCR results indicated that the banteng isolates harbored the following genes: Cefotaxime-M1 (n = 38), CTX-M9 (n = 5), and the SHV group (n = 2). All three isolates from the domestic cattle sample contained the CTX-M1 gene. Classification of ESBL genes based on the DNA sequences of the banteng isolates showed the characteristics of CTX-M15 (n = 20), CTX-M55 (n = 6), CTX-M14 (n = 5), and CTX-M79 (n = 1). The three domestic cattle isolates exhibited the characteristics of CTX-M15, CTX-M55, and CTX-M79. Despite no previous antibiotic applications, approximately one-third of the banteng samples displayed CTX resistance, indicating ARM contamination within the ecosystem. The similarity in ESBL genes between the banteng and domestic cattle populations suggests potential gene transmissions between these animal groups. However, the initial source of ARMs remains unclear, as the banteng population exhibited more ESBL genes than the domestic cattle, suggesting the possibility of multiple ARM sources. These findings raise concerns because the banteng population inhabits an area that is an important source of freshwater and nourishes the entire north-east region of Thailand and other South-east Asian countries, including Laos, Cambodia, and Southern Vietnam.

Characterization of Carbapenemase- and ESBL-Producing Gram-Negative Bacilli Isolated from Patients with Urinary Tract and Bloodstream Infections.

A total of 199 Gram-negative bacterial isolates from urinary tract infections and 162 from bloodstream infections were collected from 12 healthcare systems throughout the United States between May 2021 and August 2022. The isolates, phenotypically non-susceptible to 2nd or 3rd generation cephalosporins or carbapenems, were characterized through antimicrobial susceptibility testing and whole genome sequence analysis to obtain a broad snapshot of beta-lactamase-mediated resistance among these two sample types. Overall, 23 different carbapenemase genes were detected among 13 species (20.5% of isolates). The blaKPC-3 and blaKPC-2 subtypes were the most common carbapenemase genes identified, followed by blaNDM and the co-carriage of two different blaOXA carbapenemases by Acinetobacter baumannii isolates. All carbapenemase-producing A. baumannii isolates were mCIM negative. Extended-spectrum beta-lactamase genes were identified in 66.2% of isolates; blaCTX-M-15 was the most common. AmpC genes, both plasmid and chromosomal, were detected in 33.2% of isolates. Importantly, 2.8%, 8.3%, and 22.2% of blaKPC-positive organisms were susceptible to ertapenem, imipenem, and meropenem, respectively. The correlation between broth microdilution and disk diffusion results was high for most drugs except cefepime, where the detection of resistance was statistically lower by disk diffusion. Thus, there were gaps in the accuracy of susceptibility testing for some mechanisms of resistance.

Characteristics and genomic epidemiology of colistin-resistant Enterobacterales from farmers, swine, and hospitalized patients in Thailand, 2014–2017

BackgroundColistin is one of the last resort therapeutic options for treating carbapenemase-producing Enterobacterales, which are resistant to a broad range of beta-lactam antibiotics. However, the increased use of colistin in clinical and livestock farming settings in Thailand and China, has led to the inevitable emergence of colistin resistance. To better understand the rise of colistin-resistant strains in each of these settings, we characterized colistin-resistant Enterobacterales isolated from farmers, swine, and hospitalized patients in Thailand.MethodsEnterobacterales were isolated from 149 stool samples or rectal swabs collected from farmers, pigs, and hospitalized patients in Thailand between November 2014–December 2017. Confirmed colistin-resistant isolates were sequenced. Genomic analyses included species identification, multilocus sequence typing, and detection of antimicrobial resistance determinants and plasmids.ResultsThe overall colistin-resistant Enterobacterales colonization rate was 26.2% (n = 39/149). The plasmid-mediated colistin-resistance gene (mcr) was detected in all 25 Escherichia coli isolates and 9 of 14 (64.3%) Klebsiella spp. isolates. Five novel mcr allelic variants were also identified: mcr-2.3, mcr-3.21, mcr-3.22, mcr-3.23, and mcr-3.24, that were only detected in E. coli and Klebsiella spp. isolates from farmed pigs.ConclusionOur data confirmed the presence of colistin-resistance genes in combination with extended spectrum beta-lactamase genes in bacterial isolates from farmers, swine, and patients in Thailand. Differences between the colistin-resistance mechanisms of Escherichia coli and Klebsiella pneumoniae in hospitalized patients were observed, as expected. Additionally, we identified mobile colistin-resistance mcr-1.1 genes from swine and patient isolates belonging to plasmids of the same incompatibility group. This supported the possibility that horizontal transmission of bacterial strains or plasmid-mediated colistin-resistance genes occurs between humans and swine.

Prevalence, clonal diversity, and antimicrobial resistance of hypervirulent Klebsiella pneumoniae and Klebsiella variicola clinical isolates in northern Japan

Hypervirulent Klebsiella pneumoniae (hvKp) and Klebsiella variicola (hvKv) cause hospital/community-acquired infections, often associated with antimicrobial resistance (AMR). This study aimed to investigate the molecular epidemiology of hvKp and hvKv in northern Japan. A total of 500 K. pneumoniae and 421 K. variicola clinical isolates collected from August to December 2021 were studied. Prevalence of virulence factor-encoding genes, wzi sequence and associated K/KL type, sequence type (ST), and beta-lactamases and their types were characterized. Any virulence gene (rmpA, rmpA2, peg-344, iucA, iutA, and iroB) and/or magA was detected in 25% (n=125) of K. pneumoniae and 1% (n=5) of K. variicola. Among these hvKp/hvKv, 22 wzi types (18 and 4 types, respectively) and 24 STs (20 and 4 STs, respectively) were identified. Sequence types of hvKp were classified into some clonal groups (CGs), among which CG35, including six STs, was the most common (n=59; 47%), followed by CG23, and CG65. ST268 (CG35) associated with wzi95-K20 or wzi720 was the dominant lineage (n=43, 34%), while K1:ST23/ST249 and K2:ST65/ST86 accounted for 26% and 13% of hvKp, respectively. Extended-spectrum beta-lactamase (ESBL) genes (blaCTX-M-2, blaCTX-M-3, blaCTX-M-15, and blaCTX-M-27) were detected in only ST23 and CG35 (ST268 and ST412) hvKp. No isolate was resistant to carbapenems, without detection of the ESBL gene in K. variicola. Phylogenetically, wzi was differentiated into two main clusters of K. pneumoniae and K. variicola. A major clonal group CG347 was identified in K. variicola. Clonal structures were revealed for hvKp and hvKv clinical isolates with their AMR status in northern Japan.

Antimicrobial susceptibility and molecular characteristics of beta-lactam- and fluoroquinolone-resistant E. coli from human clinical samples in Nigeria

This study determined the antimicrobial resistance profiles and molecularly characterized beta-lactam- and fluoroquinolone-resistant E. coli recovered from human clinical samples. Three hundred (300) clinical samples collected from two major tertiary healthcare institutions were analyzed using standard microbiological techniques. Antimicrobial susceptibility testing and 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. In total, 44 (14.7%) ESBL-producing E. coli were recovered from human clinical specimens. The recovered isolates were multidrug-resistant with multiple antibiotics resistance index values ranging from 0.5–0.8. Isolates exhibited resistance to sulfamethoxazole/trimethoprim 44 (100%), cefoxitin 42 (95.5%), tetracycline 42 (95.5 %), cefuroxime 41 (93.2%), ciprofloxacin 40 (90.9%), cefotaxime 37 (84.1%), amoxicillin 37 (84.1%), amoxicillin/clavulanic acid 37 (84.1), and ceftazidime 35 (79.5%). BlaTEM 41 (93.2%) ESBL gene was the most predominant gene among the isolates, followed by blaCTX-M 9 (20.5%), and blaSHV 1 (2.3%) genes. Co-existence of blaTEM +blaSHV, blaTEM + blaCTX-M, and blaTEM +blaSHV + blaCTX-M genes was observed in 1 (2.3%), 7 (15.9%), and 4 (9.1%) isolates, respectively. All the fluoroquinolone-resistant isolates harbored aac-lb-6-cr gene while Qnr gene was absent. Association of fluoroquinolone resistance- and β-lactam resistance genes; aac-lb-6-cr + blaTEM + blaSHV + blaCTM was observed in 5 (11.4%) isolates. Our study showed high frequency of ESBL-producing E. coli harboring ESBL and fluoroquinolone resistance genes from human clinical samples in Ebonyi State, Nigeria. The high frequency of multidrug-resistant E. coli in our study is very worrisome and could have significant public health impact such as treatment failures, and possibly death, if not properly managed. It is therefore imperative to establish strong regulative measures and guidelines that would help in curtailing the increasing dissemination of these superbugs in healthcare institutions in Nigeria.

Extended-spectrum beta-lactamase genes among Gram negative bacilli isolates from Egyptian children with diarrhea

Extended-spectrum beta-lactamase genes among Gram negative bacilli isolates from Egyptian children with diarrhea

Genomic Characterization of Salmonella Isangi: A Global Perspective of a Rare Serovar.

Salmonella Isangi is an infrequent serovar that has recently been reported in several countries due to nosocomial infections. A considerable number of reports indicate Salmonella Isangi multidrug resistance, especially to cephalosporins, which could potentially pose a risk to public health worldwide. Genomic analysis is an excellent tool for monitoring the emergence of microorganisms and related factors. In this context, the aim of this study was to carry out a genomic analysis of Salmonella Isangi isolated from poultry in Brazil, and to compare it with the available genomes from the Pathogen Detection database and Sequence Read Archive. A total of 142 genomes isolated from 11 different countries were investigated. A broad distribution of extended-spectrum beta-lactamase (ESBL) genes was identified in the Salmonella Isangi genomes examined (blaCTX-M-15, blaCTX-M-2, blaDHA-1, blaNDM-1, blaOXA-10, blaOXA-1, blaOXA-48, blaSCO-1, blaSHV-5, blaTEM-131, blaTEM-1B), primarily in South Africa. Resistome analysis revealed predicted resistance to aminoglycoside, sulfonamide, macrolide, tetracycline, trimethoprim, phenicol, chloramphenicol, and quaternary ammonium. Additionally, PMQR (plasmid-mediated quinolone resistance) genes qnr19, qnrB1, and qnrS1 were identified, along with point mutations in the genes gyrAD87N, gyrAS83F, and gyrBS464F, which confer resistance to ciprofloxacin and nalidixic acid. With regard to plasmids, we identified 17 different incompatibility groups, including IncC, Col(pHAD28), IncHI2, IncHI2A, IncM2, ColpVC, Col(Ye4449), Col156, IncR, IncI1(Alpha), IncFIB (pTU3), Col(B5512), IncQ1, IncL, IncN, IncFIB(pHCM2), and IncFIB (pN55391). Phylogenetic analysis revealed five clusters grouped by sequence type and antimicrobial gene distribution. The study highlights the need for monitoring rare serovars that may become emergent due to multidrug resistance.

Abundance and prevalence of ESBL coding genes in patients undergoing first line eradication therapy for Helicobacter pylori.

The spread of extended-spectrum beta-lactamases (ESBLs) in nosocomial and community-acquired enterobacteria is an important challenge for clinicians due to the limited therapeutic options for infections that are caused by these organisms. Here, we developed a panel of ESBL coding genes, evaluated the abundance and prevalence of ESBL encoding genes in patients undergoing H. pylori eradication therapy, and summarized the effects of eradication therapy on functional profiles of the gut microbiome. To assess the repertoire of known beta lactamase (BL) genes, they were divided into clusters according to their evolutionary relation. Primers were designed for amplification of cluster marker regions, and the efficiency of this amplification panel was assessed in 120 fecal samples acquired from 60 patients undergoing H. pylori eradication therapy. In addition, fecal samples from an additional 30 patients were used to validate the detection efficiency of the developed ESBL panel. The presence for majority of targeted clusters was confirmed by NGS of amplification products. Metagenomic sequencing revealed that the abundance of ESBL genes within the pool of microorganisms was very low. The global relative abundances of the ESBL-coding gene clusters did not differ significantly among treatment states. However, at the level of each cluster, classical ESBL producers such as Klebsiella sp. for blaOXY (p = 0.0076), Acinetobacter sp. for blaADC (p = 0.02297) and others, differed significantly with a tendency to decrease compared to the pre- and post-eradication states. Only 13 clusters were common across all three datasets, suggesting a patient-specific distribution profile of ESBL-coding genes. The number of AMR genes detected in the post-eradication state was higher than that in the pre-eradication state, which could be attributed, at least in part, to the therapy. This study demonstrated that the ESBL screening panel was effective in targeting ESBL-coding gene clusters from bacterial DNA and that minor differences exist in the abundance and prevalence of ESBL-coding gene levels before and after eradication therapy.

Detection of Extended Spectrum Beta Lactamase Producing Escherichia Coli and Klebsiella Species Isolated from Urine Samples of UTI Patients by Phenotypic and Genotypic Methods

Background: Escherichia coli and Klebsiella spp. are the most common organisms causing urinary tract infection (UTI) and commonly responsible for extended spectrum beta lactamase (ESBL) production. This study was carried out to detect ESBL producing uropathogenic Escherichia coli and Klebsiella spp. by phenotypic and genotypic method. Rapid and accurate detection of ESBL producing E. coli and Klebsiella spp. has an important role to avoid treatment failure. Materials and Methods: This was a cross sectional observational study and carried out in department of microbiology in Chittagong Medical College, Bangladesh from January to December 2017. Urine was collected from suspected UTI patients and standard microbiological and biochemical tests were carried out. ESBL producing E. coli and Klebsiella spp. were identified by phenotypic confirmatory disc diffusion test (PCDDT). Polymerase chain reaction (PCR) was performed by using standard protocol with specific primers. Results: 448 urine samples were collected. Among them, 140 showed bacterial growth; 72 were E. coli and 35 were Klebsiella spp. Among E. coli 38(52.8%) and in Klebsiella spp. 15(42.9%) were detected as ESBL producers by PCDDT respectively. Among E. coli, 41(56.9%) and in Klebsiella spp. 21(60%) strains produced ESBL genes by PCR. Out of 38 phenotypically positive E. coli, 7 strains do not carry any detectable genes. Similarly, out of 15 phenotypically positive Klebliella spp. 3 isolates did not produce any detectable gene. On the other hand, 10 E. coli isolates and 9 Klebsiella spp. carry detectable genes although these were not phenotypically ESBL producers. Moreover, ESBL producing E. coli and Klebsiella spp. showed more multidrug resistant than Non-ESBL producing E. coli and Klebsiella spp. Conclusion: This study revealed that large portion of E. coli and Klebsiella spp. was ESBL producers. PCR can detect some additional cases of ESBL producing isolates. So, PCR can be used along with phenotypic ......

Resistance profiles and genotyping of extended-spectrum beta-lactamase (ESBL) -producing and non-ESBL-producing E. coli and Klebsiella from retail market fishes.

Studies on antimicrobial resistance (AMR) profiles and epidemiological affirmation for AMR transmission are limited in fisheries and aquaculture settings. Since 2015, based on Global Action Plan on AMR by World Health Organization (WHO) and World Organization for Animal Health (OIE), several initiatives have been under taken to enhance the knowledge, skills and capacity to establish AMR trends through surveillance and strengthening of epidemiological evidence. The focus of this study was to determine the prevalence of antimicrobial resistance (AMR), its resistance profiles and molecular characterization with respect to phylogroups, antimicrobial resistance genes (ARGs), virulence genes (VGs), quaternary ammonium compounds resistance (QAC) genes and plasmid typing in retail market fishes. Pulse field gel electrophoresis (PFGE) to understand the genetic lineage of the two most important Enterobacteriaceae members, E. coli and Klebsiella sp. was performed. 94 fish samples were collected from three different sites viz., Silagrant (S1), Garchuk (S2) and North Guwahati Town Committee (NGTC) Region (S3) in Guwahati, Assam. Out of the 113 microbial isolates from the fish samples, 45 (39.82%) were E. coli; 23 (20.35%) belonged to Klebsiella genus. Among E. coli, 48.88% (n=22) of the isolates were alerted by the BD Phoenix M50 instrument as ESBL, 15.55% (n=7) as PCP and 35.55% (n=16) as non-ESBL. E. coli (39.82%) was the most prevalent pathogen among the Enterobacteriaceae members screened and showed resistance to ampicillin (69%) followed by cefazoline (64%), cefotaxime (49%) and piperacillin (49%). In the present study, 66.66% of E. coli and 30.43% of Klebsiella sp. were categorized as multi drug resistance (MDR) bacteria. CTX-M-gp-1, with CTX-M-15 variant (47%), was the most widely circulating beta-lactamase gene, while other ESBL genes blaTEM (7%), blaSHV (2%) and blaOXA-1-like (2%) were also identified in E. coli. Out of the 23 isolates of Klebsiella, 14(60.86%) were ampicillin (AM)-resistant (11(47.82%) K. oxytoca, 3(13.04%) K. aerogenes), whereas 8(34.78%) isolates of K. oxytoca showed intermediate resistance to AM. All Klebsiella isolates were susceptible to AN, SCP, MEM and TZP, although two K. aerogenes were resistant to imipenem. DHA and LAT genes were detected, respectively, in 7(16%) and 1(2%) of the E. coli strains while a single K. oxytoca (4.34%) isolate carried MOX, DHA and blaCMY-2 genes. The fluoroquinolone resistance genes identified in E. coli included qnrB (71%), qnrS (84%), oqxB (73%) and aac(6)-Ib-cr (27%); however, in Klebsiella, these genes, respectively, had a prevalence of 87%, 26%, 74% and 9%. The E. coli isolates belonged to phylogroup A(47%), B1(33%) and D(14%). All of the 22(100%) ESBL E. coli had chromosome-mediated disinfectant resistance genes viz., ydgE, ydgF, sugE(c), mdfA while 82% of ESBL E. coli had emrE. Among the non-ESBL E. coli isolates, 87% of them showed the presence of ydgE, ydgF and sugE(c) genes, while 78% of the isolates had mdfA and 39% had emrE genes respectively. 59% of the ESBL and 26% of the non-ESBL E. coli had showed the presence of qacEΔ1. The sugE(p) was present in 27% of the ESBL-producing E. coli and in 9% of non-ESBL isolates. Out of the 3 ESBL-producing Klebsiella isolates, 2(66.66%) K. oxytoca isolates were found harboring plasmid-mediated qacEΔ1 gene while one (33.33%) K. oxytoca isolate had sugE(p) gene. IncFI was the most prevalent plasmid type detected in the isolates studied, with A/C (18%), P (14%), X, Y (9% each) and I1-Iγ (14%, 4%). 50% (n=11) of the ESBL and 17% (n=4) of the non-ESBL E. coli isolates harboured IncFIB and 45% (n=10) ESBL and one (4.34%) non-ESBL E. coli isolates harboured IncFIA. Dominance of E. coli over other Enterobacterales and diverse phylogenetic profiles of E. coli and Klebsiella sp. suggests the possibility of contamination and this may be due to compromised hygienic practices along the supply chain and contamination of aquatic ecosystem. Continuous surveillance in domestic markets must be a priority in addressing antimicrobial resistance in fishery settings and to identify any unwarranted epidemic clones of E. coli and Klebsiella that can challenge public health sector.

Genomic and functional portrait of multidrug-resistant, hydrogen sulfide (H2S)-producing variants of Escherichia coli.

Atypical Escherichia coli forms exhibit unusual characteristics compared to typical strains. The H2S-producing variants of some atypical E. coli strains cause a wide range of illnesses in humans and animals. However, there are sparse reports on such strains worldwide. We performed whole-genome sequencing (WGS) and detailed characterization of four H2S-producing E. coli variants from poultry and human clinical sources in Dhaka, Bangladesh. All four isolates were confirmed as E. coli using biochemical tests and genomic analysis, and were multidrug-resistant (MDR). WGS analysis including an additional Chinese strain, revealed diverse STs among the five H2S-producing E. coli genomes, with clonal complex ST10 being detected in 2 out of 5 genomes. The predominant phylogroup detected was group A (n = 4/5). The blaTEM1B (n = 5/5) was the most predominant extended-spectrum beta-lactamase (ESBL) gene, followed by different alleles of blaCTX-M (blaCTX-M -55,-65,-123; n = 3/5). Multiple plasmid replicons were detected, with IncX being the most common. One E. coli strain was classified as enteropathogenic E. coli. The genomes of all five isolates harbored five primary and four secondary function genes related to H2S production. These findings suggest the potential of these isolates to cause disease and spread antibiotic resistance. Therefore, such atypical E. coli forms should be included in differential diagnosis to understand the pathogenicity, antimicrobial resistance and evolution of H2S-producing E. coli.

Genotypic characterization of extended spectrum beta-lactamase in gram negative bacterial contaminants of some door handles in Olabisi Onabanjo University Teaching Hospital, Sagamu, Ogun state.

Nosocomial infections due to cross-transmission by microorganisms from inanimate surfaces have become a recognized public health problem. This research was carried out to detect bacterial contaminants of door handles and their production of extended spectrum beta-lactamase (ESBL) in Olabisi Onabanjo University Teaching Hospital. Seventy door handles from twenty-seven sections of Olabisi Onabanjo University Teaching Hospital were swabbed and inoculated on different culture media. The isolates were identified using standard microbiological procedures. Susceptibility of the isolates to eleven antibiotics was determined by Kirby- Bauer disc diffusion test. Phenotypic detection of ESBL was determined by modified double disc synergy test (MDDST) followed by PCR amplification of ESBL- encoding genes. Forty-four (62.9%) out of 70 door swabs were positive for bacterial growth. S . aureus was the most common isolate (22.9%). Multidrug resistance was observed in 15(79.0%) and 20(69%) of Gram positive and Gram-negative isolates, respectively. Coagulase negative staphylococci, E. coli, Acinetobacter and Shigella species were totally resistant to ceftazidime (100%). PCR amplification of ESBL-genes revealed the presence of blaTEM blaCTX -M and blaSHV in 4(14.3%), 2(7.2%) and 1(3.6%) isolate, respectively. This study revealed that door handles of different sections in hospital environment were contaminated by multidrug resistant Gram-negative bacteria encoding ESBL genes.

Pisciculture and Fish Parts as Non-clinical Source of CTX-M and TEM Extended Spectrum Beta-lactamases Producing Escherichia coli in Southeastern Nigeria

Background and Objectives: Antimicrobial agent use in pisciculture exerts a selective pressure, resulting in a reservoirs of extended spectrum beta-lactamase producing Escherichia coli in the aquatic environment. Here we screened pisciculture and fish parts as non-clinical source of CTX-M and TEM Extended Spectrum Beta-lactamases (ESBL) producing Escherichia coli in Southeastern Nigeria Methodology: A total of sixty fish parts [fish gills, body part and intestinal swab samples] were collected from pisciculture farm A, C, F and G within Abakaliki metropolis. The samples were bacteriologically analyzed using standard microbiological techniques for isolation and identification. ESBL enzymes were phenotypically detected in E. coli, isolates using the double disk diffusion technique. The presence of TEM and CTX-M genes was determined by polymerase chain reaction using specific primers. Results: Escherichia coli and Extended spectrum beta-lactamase producing Escherichia coli accounted for overall occurrence rate of 27(45.0 %) and 17(28.3 %) respectively. There was no statistically significant difference in the occurrence of Escherichia coli and ESBL-E. coli among the different samples P>0.05. The presence of TEM and CTX-M gene 80 % and 100 % respectively were confirmed among the isolates. Conclusion: Our findings reports the presence of ESBL gene in pisciculture and fish parts and it is important in understanding the mechanism of resistance operating in these common pathogens, which are also endemic in most pisciculture area. Strict rules and monitoring/surveillance of antimicrobial agent used in pisciculture activities combined with food safety training of farmer owners/breeders on various aspects of good hygiene practices are strongly recommended.

Characterization of blaTEM and blaCTX-M beta-lactam resistance genes in chronic rhinosinusitis

BackgroundChronic sinusitis is one of the most challenging health problems of contemporary society. Although several treatment methods have been defined, a comprehensive understanding of the underlying causes (e.g., antibiotic resistance) is still elusive. The aim of this study was to characterize two of the main extended-spectrum beta-lactamase genes—i.e., blaTEM and blaCTX-M genes—and investigate antimicrobial resistance in bacteria isolated from chronic sinusitis. Samples from 70 chronic sinusitis patients and 20 healthy individuals (controls) were analyzed for the presence of blaTEM and blaCTX-M resistance genes using the polymerase chain reaction (PCR) test, followed by gene sequence analysis.ResultsPhenotypic and genotypic beta-lactam resistance was observed in 58.7% and 61.54% of the gram-negative isolates, respectively, with 38.46% carrying the blaTEM gene and 34.62% harboring the blaCTX-M gene. Sequencing data indicated high heterogeneity in blaCTX-M genes (69–100% similarity to reported sequences) and lower heterogeneity in blaTEM genes (93–99%).ConclusionBroad-spectrum beta-lactam resistance is a major pathogenesis factor in chronic rhinosinusitis, and careful consideration is required for antimicrobial therapy. High blaCTX-M heterogeneity could mean high horizontal transfer rate of this gene and warrant a surveillance program.

Investigation of Delafloxacin Resistance in Multidrug-Resistant Escherichia coli Strains and the Detection of E. coli ST43 International High-Risk Clone.

Delafloxacin is a novel fluoroquinolone agent that is approved for clinical application. In this study, we analyzed the antibacterial efficacy of delafloxacin in a collection of 47 Escherichia coli strains. Antimicrobial susceptibility testing was performed by the broth microdilution method and minimum inhibitory concentration (MIC) values were determined for delafloxacin, ciprofloxacin, levofloxacin, moxifloxacin, ceftazidime, cefotaxime, and imipenem. Two multidrug-resistant E. coli strains, which exhibited delafloxacin and ciprofloxacin resistance as well as extended-spectrum beta-lactamase (ESBL) phenotype, were selected for whole-genome sequencing (WGS). In our study, delafloxacin and ciprofloxacin resistance rates were 47% (22/47) and 51% (24/47), respectively. In the strain collection, 46 E. coli were associated with ESBL production. The MIC50 value for delafloxacin was 0.125 mg/L, while all other fluoroquinolones had an MIC50 value of 0.25 mg/L in our collection. Delafloxacin susceptibility was detected in 20 ESBL positive and ciprofloxacin resistant E. coli strains; by contrast, E. coli strains that exhibited a ciprofloxacin MIC value above 1 mg/L were delafloxacin-resistant. WGS analysis on the two selected E. coli strains (920/1 and 951/2) demonstrated that delafloxacin resistance is mediated by multiple chromosomal mutations, namely, five mutations in E. coli 920/1 (gyrA S83L, D87N, parC S80I, E84V, and parE I529L) and four mutations in E. coli 951/2 (gyrA S83L, D87N, parC S80I, and E84V). Both strains carried an ESBL gene, blaCTX-M-1 in E. coli 920/1 and blaCTX-M-15 in E. coli 951/2. Based on multilocus sequence typing, both strains belong to the E. coli sequence type 43 (ST43). In this paper, we report a remarkable high rate (47%) of delafloxacin resistance among multidrug-resistant E. coli as well as the E. coli ST43 international high-risk clone in Hungary.

Genomic analysis of extended-spectrum beta-lactamase (ESBL) producing Escherichia coli colonising adults in Blantyre, Malawi reveals previously undescribed diversity.

Escherichia coli is one of the most prevalent Gram-negative species associated with drug resistant infections. Strains that produce extended-spectrum beta-lactamases (ESBLs) or carbapenemases are both particularly problematic and disproportionately impact resource limited healthcare settings where last-line antimicrobials may not be available. A large number of E. coli genomes are now available and have allowed insights into pathogenesis and epidemiology of ESBL E. coli but genomes from sub-Saharan Africa (sSA) are significantly underrepresented. To reduce this gap, we investigated ESBL-producing E. coli colonising adults in Blantyre, Malawi to assess bacterial diversity and AMR determinants and to place these isolates in the context of the wider population structure. We performed short-read whole-genome sequencing of 473 colonising ESBL E. coli isolated from human stool and contextualised the genomes with a previously curated multi-country collection of 10 146 E. coli genomes and sequence type (ST)-specific collections for our three most commonly identified STs. These were the globally successful ST131, ST410 and ST167, and the dominant ESBL genes were bla CTX-M, mirroring global trends. However, 37 % of Malawian isolates did not cluster with any isolates in the curated multicountry collection and phylogenies were consistent with locally spreading monophyletic clades, including within the globally distributed, carbapenemase-associated B4/H24RxC ST410 lineage. A single ST2083 isolate in this collection harboured a carbapenemase gene. Long read sequencing demonstrated the presence of a globally distributed ST410-associated carbapenemase carrying plasmid in this isolate, which was absent from the ST410 strains in our collection. We conclude there is a risk that carbapenem resistance in E. coli could proliferate rapidly in Malawi under increasing selection pressure, and that both ongoing antimicrobial stewardship and genomic surveillance are critical as local carbapenem use increases.

Frequency of Extended-Spectrum Beta-Lactamases-Producing Escherichia coli Among Out- and In-patients in Rafsanjan City, Iran

Background: The frequency of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli strains among clinical isolates has been steadily increasing, leading to limited treatment options. Objectives: This study aimed to assess the antibiotic sensitivity of E. coli and the frequency of ESBL isolates among both out-patients and in-patients. Methods: A total of 390 E. coli isolates were received at the Ali-Ebn-e-Abitaleb Hospital laboratory in Rafsanjan. The antibiogram, as well as the phenotypic and genotypic detection of ESBL isolates, were conducted using Kirby-Bauer, combination disk confirmatory, and polymerase chain reaction tests, respectively. Results: Of all the E. coli isolates, 45.6% exhibited ESBL production. Among these isolates, 46.1% were obtained from hospital wards, while 42.5% were from outpatients. Meropenem and imipenem displayed sensitivities of 97.2% and 93.3%, respectively, whereas amikacin and nitrofurantoin showed sensitivities of 89.7% and 85.6%, respectively, for all isolates. Notably, ceftriaxone, cefotaxime, cefixime, ampicillin, co-trimoxazole, and nalidixic acid demonstrated high resistance rates, surpassing 50%. ESBL-producing isolates were more frequently observed in blood samples (65%) and wounds (60%) compared to other tested isolates. Approximately 8.6% of isolates carried a single type of ESBL gene, while 38.5% carried multiple ESBL genes. Conclusions: The data indicate a prevalence of ESBL-producing E. coli isolates among both out-patients and in-patients, with some of them acquiring two or three types of ESBL enzymes. As a result, their ability to hydrolyze antibiotics has increased, leading to their higher occurrence in clinical samples.

In-vitro activity of oral third-generation cephalosporins plus clavulanate against ESBL-producing Enterobacterales isolates from the MERINO trial

Extended-spectrum-beta-lactamase (ESBL)-producing Enterobacterales as a cause of community-acquired uncomplicated urinary tract infection (UTI) is on the rise. Currently, there are minimal oral treatment options. New combinations of existing oral third-generation cephalosporins paired with clavulanate may overcome resistance mechanisms seen in these emerging uropathogens. Ceftriaxone-resistant Escherichia coli and Klebsiella pneumoniae containing CTX-M-type ESBLs or AmpC, in addition to narrow-spectrum OXA and SHV enzymes, were selected from blood culture isolates obtained from the MERINO trial. Minimum inhibitory concentration (MIC) values of third-generation cephalosporins (cefpodoxime, ceftibuten, cefixime, cefdinir), both with and without clavulanate, were determined. One hundred and one isolates were used with ESBL, AmpC and narrow-spectrum OXA genes (e.g. OXA-1, OXA-10) present in 84, 15 and 35 isolates, respectively. Susceptibility to oral third-generation cephalosporins alone was very poor. Addition of 2 mg/L clavulanate reduced the MIC50 values (cefpodoxime MIC50 2 mg/L, ceftibuten MIC50 2 mg/L, cefixime MIC50 2 mg/L, cefdinir MIC50 4 mg/L) and restored susceptibility (33%, 49%, 40% and 21% susceptible, respectively) in a substantial number of isolates. This finding was less pronounced in isolates co-harbouring AmpC. In-vitro activity of these new combinations may be limited in real-world Enterobacterales isolates co-harbouring multiple antimicrobial resistance genes. Pharmacokinetic/pharmacodynamic data would be useful for further evaluation of their activity.

Characteristic of extended substrate-spectrum beta-lactamases producing Enterobacteriales isolated from pork meat

Antibiotic resistance among bacteria is an ever-growing problem in both human and veterinary medicine. Livestock and meat from their slaughter are often sources of drug-resistant bacteria. The ability to produce extended-spectrum beta-lactamases (ESBLs) is common among Enterobacteriales, including commensal bacteria. These bacteria represent a largely ignored reservoir of ESBL genes. The aim of this study was to assess the drug susceptibility of non-pathogenic Enterobacteriales isolates obtained from meat used in food production. Among the strains tested, ESBL-type resistance was demonstrated - 13 of 18 (72%) isolates tested were found to be ESBL-producing strains. Of these, eight possessed the CTX-M gene, three the TEM gene and one the SHV gene.

Molecular Detection of Extended Spectrum Beta-Lactamase Gene in Clinical Isolates of Klebsiella Species in Jos North Central of Nigeria

Aims: The aim of the study was to determine the antimicrobial susceptibility of Klebsiella species, prevalence of Extended Spectrum Beta-Lactamases (ESBLs) and the gene coding ESBL production in Klebsiella Species obtained from clinical specimens of patients attending Plateau State Specialist and Jos University Teaching Hospitals both in Jos North Central of Nigeria.
 Study Design: This was hospital based study conducted in the Department of Medical Microbiology unit of Plateau State Specialist and Jos University Teaching Hospital (JUTH) during the periods of January to April 2020.
 Methodology: Out of 156 non-motile, oxidase negative gram negative organisms collected, 137 isolates were identified to be Klebsiella species using oxoid Microbact 24E kit for Enterobacteriaceae identification and Microsoft for bacteria identification. Antimicrobial Susceptibility Testing (AST) and ESBL screening was done by Kirby-Bauer disk diffusion method using oxoid Single disk antibiotics which include Ceftazidime (30µg), Cefotaxime (30µg), Imipenem (10µg), Ciprofloxacin (5µg), Gentamicin (10µg), Ampicillin (10 µg), Aztreonam (30 µg), and Azithromycin (15 µg) on Mueller Hinton Agar (oxoid).Break point was determined by CLSI AST guidelines. The isolates that showed resistance or reduced susceptibility to Ceftazidime and Cefotaxime are suspected to be ESBL positive Klebsiella. Phenotypic confirmation of ESBL positive isolates was carried out using Double Disk Synergy Test (DDST) with Ceftazidime 30µg (oxoid), Cefotaxime 30µg (oxoid) and Amoxicillin Clavulanic acid 20/10ug (Oxoid).Genotypic detection of ESBL gene was done using Polymerase Chain Reaction (PCR).
 Results: Antimicrobial Susceptibility Test showed Imipenem to be highly sensitive to ESBL positive Klebsiella pneumonia with 100% sensitivity. Gentamicin, Ampicillin, and Azithromycin showed reduced susceptibility. Genotypic results showed presence of bla TEM, bla SHV genes at 868bp and 972bp band size with no occurrence of bla CTX-M gene. There was multiple occurrence of gene in some of the isolates tested while some recorded no ESBL gene. The ESBL prevalence of 15.3% was recorded in this study.
 Conclusions: The prevalence of 15.3% recorded in this study is low compared to other studies from some other states within the country. Efforts should be put in place to ensure non-proliferation of this ESBL Klebsiella strain as it creates not only a therapeutic challenge but health problem.