Extended-spectrum Beta-lactamase Genes Research Articles

In-silico insights of ESBL variants and tracking the probable sources of ESBL-producing Escherichia coli in a small-scale poultry farm

Commercial broiler farms face challenges of extended spectrum beta-lactamase (ESBL)-producing Escherichia coli transmitted from both vertical and horizontal routes. Understanding the dynamics of ESBL-E. coli transmission in compromised biosecurity settings of small-scale rural poultry farms is essential. This study aimed to elucidate the probable transmission pathways of ESBL-E. coli in such settings, employing phylogenetic analysis and molecular docking simulations to explore the catalytic properties of β-lactamase variants. Sampling was conducted on a small-scale poultry farm in West Bengal, India, collecting 120 samples at three intervals during the broiler production cycle. E. coli isolates underwent resistance testing against eight antimicrobials, with confirmation of ESBL production. Genotypic analysis of ESBL genes and sequencing were performed, alongside molecular docking analyses and phylogenetic comparisons with publicly available sequences. Among 173 E. coli isolates, varying resistance profiles were observed, with complete resistance to cefixime and high resistance to amoxicillin and tetracycline. The incidence of ESBL-E. coli fluctuated over the production cycle, with dynamic changes in the prevalence of blaCTX-M-type and blaSHV-type genes. Phylogenetic analysis indicated partial clonal relationships with human clinical strains and poultry strains from the Indian subcontinent. Molecular docking confirmed the catalytic efficiencies of these ESBL variants. The study highlights probable vertical transmission of ESBL-E. coli and emphasizes drinking water as a potential source of horizontal transmission in small-scale poultry farms. Strict biosecurity measures could prevent the spread of antimicrobial-resistant bacteria in birds and their products in a small scale poultry farm.

Coexistence of plasmid-mediated quinolone resistance (PMQR) and extended-spectrum beta-lactamase (ESBL) genes among clinical Pseudomonas aeruginosa isolates in Egypt

BackgroundData about the prevalence of plasmid-mediated quinolone resistance (PMQR) and extended-spectrum beta-lactamase (ESBL) production in P. aeruginosa compared to the Enterobacteriaceae family is limited. The availability of limited therapeutic options raises alarming concerns about the treatment of multidrug-resistant P. aeruginosa. This study aimed to assess the presence of PMQR and ESBL genes among P. aeruginosa strains.MethodsFifty-six P. aeruginosa strains were isolated from 330 patients with different clinical infections. Phenotypically fluoroquinolone-resistant isolates were tested by PCR for the presence of six PMQR genes. Then, blaTEM, blaSHV, and blaCTX-M type ESBL genes were screened to study the co-existence of different resistance determinants.ResultsOverall, 22/56 (39.3%) of the studied P. aeruginosa isolates were phenotypically resistant to fluoroquinolones. PMQR-producing P. aeruginosa isolates were identified in 20 isolates (90.9%). The acc(6')-Ib-cr was the most prevalent PMQR gene (77.3%). The qnr genes occurred in 72.7%, with the predominance of the qnrA gene at 54.5%, followed by the qnrS gene at 27.3%, then qnrB and qnrC at 22.7%. The qepA was not detected in any isolate. The acc(6')-Ib-cr was associated with qnr genes in 65% of positive PMQR isolates. Significant differences between the fluoroquinolone-resistant and fluoroquinolone-susceptible isolates in terms of the antibiotic resistance rates of amikacin, imipenem, and cefepime (P value < 0.0001) were found. The ESBL genes were detected in 52% of cephalosporin-resistant P. aeruginosa isolates. The most frequent ESBL gene was blaCTX-M (76.9%), followed by blaTEM (46.2%). No isolates carried the blaSHV gene. The acc(6')-Ib-cr gene showed the highest association with ESBL genes, followed by the qnrA gene. The correlation matrix of the detected PMQR and ESBL genes indicated overall positive correlations. The strongest and most highly significant correlation was between qnrA and acc(6')-Ib-cr (r = 0.602) and between qnrA and blaCTX-M (r = 0.519).ConclusionA high prevalence of PMQR genes among the phenotypic fluoroquinolone-resistant P. aeruginosa isolates was detected, with the co-carriage of different PMQR genes. The most frequent PMQR was the acc(6')-Ib-cr gene. Co-existence between PMQR and ESBL genes was found, with 75% of PMQR-positive isolates carrying at least one ESBL gene. A high and significant correlation between the ESBL and PMQR genes was detected.

Molecular characterization of carbapenemase and extended spectrum beta-lactamase producing Acinetobacter baumannii isolates causing surgical site infections in Ethiopia

BackgroundAcinetobacter baumannii is an opportunistic pathogen that can cause a variety of nosocomial infections in humans. This study aimed to molecularly characterize extended-spectrum beta-lactamase (ESBL) producing and carbapenem-resistant Acinetobacter species isolated from surgical site infections (SSI).MethodsA multicentre cross-sectional study was performed among SSI patients at four hospitals located in Northern, Southern, Southwest, and Central parts of Ethiopia. The isolates were identified by microbiological methods and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Antibiotic susceptibility was determined using disk diffusion. The presence of phenotypic ESBL and carbapenemase production was detected by employing standard microbiological tests, including combined disk diffusion (CDT). ESBL and carbapenem resistance determinants genes were studied by polymerase chain reaction (PCR) and sequencing.ResultsA total of 8.7% Acinetobacter species were identified from 493 culture-positive isolates out of 752 SSI wounds. The species identified by MALDI-TOF MS were 88.4% A. baumannii, 4.7% Acinetobacter pittii, 4.7% Acinetobacter soli, and 2.3% Acinetobacter lactucae. Of all isolates 93% were positive for ESBL enzymes according to the CDT. Using whole genome sequencing 62.8% of the A. baumannii harbored one or more beta-lactamase genes, and 46.5% harbored one or more carbapenemase producing genes. The distribution of beta-lactamases among Acinetobacter species by hospitals was 53.8%, 64.3%, 75%, and 75% at JUSH, TASH, DTCSH, and HUCSH respectively. Among ESBL genes, blaCTX−M alleles were detected in 21.4% of isolates; of these 83.3% were blaCTX−M−15. The predominant carbapenemase gene of blaOXA type was detected in 24 carbapenem-resistant A. baumannii followed by blaNDM alleles carried in 12 A. baumannii with blaNDM−1 as the most common.ConclusionsThe frequency of Acinetobacter species that produce metallobetalactamases (MBLs) and ESBLs that were found in this study is extremely scary and calls for strict infection prevention and control procedures in health facilities helps to set effective antibiotics stewardship.

Genetic Analyses of Rare ESBL ST628 Klebsiella pneumoniae Detected during a Protracted Nosocomial Outbreak in the United Kingdom.

Klebsiella pneumoniae (K. pneumoniae) cultures from a hospital-wide outbreak in the UK, which lasted for over 12 months, were sequenced. We sought to sequence and genetically characterise the outbreak strain. Antibiotic Susceptibility Testing (AST) was performed on 65 K. pneumoniae isolates saved from the outbreak. All isolates were sequenced using the Oxford Nanopore Technologies (ONT) MinION flowcell: 10 isolates, including the isolate with the earliest collection date in 2017, were additionally sequenced on the NovaSeq 6000 platform to build high-accuracy nanopore-illumina assemblies. Among the sequenced strains, 60 were typed as ST628. 96.6% (n = 58/60) ST628 strains harboured a large ~247-kb FIB(K) plasmid carrying up to 11 antimicrobial resistance genes, including the extended-spectrum beta-lactamase (ESBL) gene, blaCTX-M-15. Clonality between the outbreak isolates was confirmed using single nucleotide polymorphism (SNP) typing. The outbreak strains were phylogenetically related to clinical ST628 strains identified in 2012, 6 years prior to the outbreak. A rare ESBL K. pneumoniae K2 ST628 strain harbouring a multi-drug resistant (MDR) plasmid encoding the ESBL gene blaCTX-M-15 was detected across multiple independent wards during the protracted nosocomial outbreak. Surveillance of this strain is recommended to prevent future nosocomial outbreaks.

Extended-Spectrum Beta-Lactamase-Producing Escherichia coli and Other Antimicrobial-Resistant Gram-Negative Pathogens Isolated from Bovine Mastitis: A One Health Perspective.

Antimicrobial resistance (AMR) poses an imminent threat to global public health, driven in part by the widespread use of antimicrobials in both humans and animals. Within the dairy cattle industry, Gram-negative coliforms such as Escherichia coli and Klebsiella pneumoniae stand out as major causative agents of clinical mastitis. These same bacterial species are frequently associated with severe infections in humans, including bloodstream and urinary tract infections, and contribute significantly to the alarming surge in antimicrobial-resistant bacterial infections worldwide. Additionally, mastitis-causing coliforms often carry AMR genes akin to those found in hospital-acquired strains, notably the extended-spectrum beta-lactamase genes. This raises concerns regarding the potential transmission of resistant bacteria and AMR from mastitis cases in dairy cattle to humans. In this narrative review, we explore the distinctive characteristics of antimicrobial-resistant E. coli and Klebsiella spp. strains implicated in clinical mastitis and human infections. We focus on the molecular mechanisms underlying AMR in these bacterial populations and critically evaluate the potential for interspecies transmission. Despite some degree of similarity observed in sequence types and mobile genetic elements between strains found in humans and cows, the existing literature does not provide conclusive evidence to assert that coliforms responsible for mastitis in cows pose a direct threat to human health. Finally, we also scrutinize the existing literature, identifying gaps and limitations, and propose avenues for future research to address these pressing challenges comprehensively.

High prevalence of multidrug-resistant Enterobacterales carrying extended-spectrum beta-lactamase and AmpC genes isolated from neonatal sepsis in Ahvaz, Iran

ObjectivesIn the recent years, multidrug resistant (MDR) neonatal septicemia-causing Enterobacterales has been dramatically increased due to the extended-spectrum beta-lactamases (ESBLs) and AmpC enzymes. This study aimed to assess the antibiotic resistance pattern, prevalence of ESBLs/AmpC beta-lactamase genes, and Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) fingerprints in Enterobacterales isolated from neonatal sepsis.ResultsIn total, 59 Enterobacterales isolates including 41 (69.5%) Enterobacter species, 15 (25.4%) Klebsiella pneumoniae and 3 (5.1%) Escherichia coli were isolated respectively. Resistance to ceftazidime and cefotaxime was seen in all of isolates. Furthermore, all of them were multidrug-resistant (resistant to three different antibiotic categories). The phenotypic tests showed that 100% of isolates were ESBL-positive. Moreover, AmpC production was observed in 84.7% (n = 50/59) of isolates. Among 59 ESBL-positive isolates, the highest percentage belonged to blaCTX−M−15 gene (66.1%) followed by blaCTX−M (45.8%), blaCTX−M−14 (30.5%), blaSHV (28.8%), and blaTEM (13.6%). The frequency of blaDHA, blaEBC, blaMOX and blaCIT genes were 24%, 24%, 4%, and 2% respectively. ERIC-PCR analysis revealed that Enterobacterales isolates were genetically diverse. The remarkable prevalence of MDR Enterobacterales isolates carrying ESBL and AmpC beta-lactamase genes emphasizes that efficient surveillance measures are essential to avoid the more expansion of drug resistance amongst isolates.

Discerning the dissemination mechanisms of antibiotic resistance genes through whole genome sequencing of extended-spectrum beta-lactamase (ESBL)-producing E. coli isolated from veterinary clinics and farms in South Korea

Extended-spectrum beta-lactamase (ESBL)-producing bacteria are resistant to most beta-lactams, including third-generation cephalosporins, limiting the treatment methods against the infections they cause. In this study, we performed whole genome sequencing of ESBL-producing E. coli to determine the mechanisms underlying the dissemination of antibiotic resistance genes. We analyzed 141 ESBL-producing isolates which had been collected from 16 veterinary clinics and 16 farms in South Korea. Long- and short-read sequencing platforms were used to obtain high-quality assemblies. The results showed that blaCTX-M is the dominant ESBL gene type found in South Korea. The spread of blaCTX-M appears to have been facilitated by both clonal spread between different host species and conjugation. Most blaCTX-M genes were found associated with diverse mobile genetic elements that may contribute to the chromosomal integration of the genes. Diverse incompatibility groups of blaCTX-M-harboring plasmids were also observed, which allows their spread among a variety of bacteria. Comprehensive whole genome sequence analysis was useful for the identification of the most prevalent types of ESBL genes and their dissemination mechanisms. The results of this study suggest that the propagation of ESBL genes can occur through clonal spread and plasmid-mediated dissemination, and that suitable action plans should be developed to prevent further propagation of these genes.

Identification of Extended Spectrum Beta Lactamases Producing Pseudomonas aeruginosa from Subclinical Mastitis Milk Samples in an Organized Dairy Farm of SVVU, Andhra Pradesh

The present study was conducted during August 2021 at Buffalo Research Station, Venkatarammanagudem of West Godavari District, Andhra Pradesh, India and was aimed to detect the presence of extended-spectrum beta-lactamases (ESBL) producing P. aeruginosa from sub-clinical mastitis that are apparently healthy Murrah buffaloes. Inflammation of the udder by microbial infection is one of the leading economic disease in the dairy sector. Most of the pathogens especially Pseudomonas aeruginosa are refractory to the antibiotic theraphy. Nowadays, the antimicrobial resistance owed by the bacteria is at an alarming rate. Pseudomonas aeruginosa, a multi drug resistant pathogen carry the extended spectrum beta lactamase genes in its plasmids and thus become resistant to antibiotic therapy. A total of 276 milk samples were collected from 69 milch animals in the month of August, 2021 comprising from all the four quarters of each buffalo. Nine (n=9) (3.26%) P. aeruginosa isolates were identified from the sub-clinical mastitis milk samples from the total 276 samples collected in the study. The isolates on morphological analysis were typical to P. aeruginosa. Seven isolates were positive for phenotypical beta-lactamases production. A total of 1 and 2 isolates were found reactive of the blaSHV and blaOXA genes, and two isolates harboured both blaSHV and blaOXA genes, respectively on PCR assay. No blaTEM gene was found in the isolates. Antibiogram of the ESBL producing P. aeruginosa isolates (n=7) possessed 100% resistance against most of the commonly used antimicrobials like ampicillin, amoxycillin, clindamycin, co-trimoxazole oxytetracycline, streptomycin and ceftriaxone (85%). The P. aeruginosa isolates were sensitive to amikacin (100%), ciprofloxacin (87.4%), gentamicin (84%), norfloxacin (84.8%) and enrofloxacin (82%).

Occurrence of ESBL‐Producing Klebsiella pneumoniae in Ready‐to‐eat Street Foods in Bouaké, Côte d’Ivoire

Aims: This study aimed to estimate the occurrence of ESBL-producing Klebsiella pneumoniae (K. pneumoniae) in ready-to-eat street foods.&#x0D; Study Design: It's a prospective study&#x0D; Place and Duration of Study: foods samples were collected from February 2016 to June 2017 from street vendors in Bouaké, Côte d’Ivoire&#x0D; Methodology: A total of 300 foods samples were collected from February 2016 to June 2017 from street vendors in Bouaké, Côte d’Ivoire. After a microbiological analysis, bacterial identification was performed according to the conventional microbiological tests. Antibiotic sensitivity of K. pneumoniae isolates to 18 antibiotics was determined by the disk diffusion method and by VITEK 2. Phenotypic and molecular detection of producing extended Spectrum Beta-Lactamase (ESBL) was performed by Chromogenic medium and double disk synergy test and by PCR method.&#x0D; Results: Four percentages (4 %) of foods samples were contaminated by K. pneumoniae. Most of the 12 K. pneumoniae strains isolated expressed resistance to the various classes of antibiotics used. Of the 12 strains, one was ESBL-producing, representing a prevalence of 8 %. This strain was isolated from cooked cow's milk and resistant with all 18 antibiotics tested except gentamicin and sulfamethoxazol-trimepothrim. This strain harbored only blaSHV gene.&#x0D; Conclusion: The results of this study indicate that RTE foods are a reservoir of resistant bacteria. Thus, they may play a role in spreading antimicrobial resistant bacteria and ESBL genes to humans.

ANTIBIOTIC SUSCEPTIBILITY AND DETECTION OF EXTENDED-SPECTRUM B-LACTAMASE GENES IN GRAM-NEGATIVE BACTERIA ISOLATED FROM AUTOMATED TELLER MACHINES

Antibiotic-resistant (AR) bacteria especially from commonly shared surfaces in the environment, pose a serious threat to global public health. The increasing demand for electronic banking and the associated risk of the spread of pathogenic bacteria makes it crucial to assess the antibiotic susceptibility and the presence of extended-spectrum beta-lactamase genes in bacteria isolated from automated teller machines (ATMs). Following standard procedures, a total of 22 samples were collected randomly from different ATMs using sterile cotton swabs soaked in physiologic saline and cultured on selective media. Isolates were characterized biochemically. Antibiotics susceptibility test was carried out on isolates using the Kirby-Bauer disc diffusion method. All Gram-negative isolates were subjected to polymerase chain reaction (PCR) to evaluate the common extended-spectrum beta-lactamase –encoding (ESBL) genes. Bacterial isolates were characterized as Klebsiella pneumoniae (33.33%), Pseudomonas aeruginosa (19.61%), Escherichia coli (15.69%), Staphylococcus aureus (9.80%), Acinetobacter spp. (7.84%), Enterobacter spp. and Bacillus subtilis (5.88%), and Enterococcus faecalis (1.96%). The highest antibiotic resistance pattern was displayed against ceftazidime (45.1%), followed by tetracycline (43.1%), cefixime (41.2%), ciprofloxacin and chloramphenicol (29.4%) and meropenem (9.8%). The TEM gene (54.8%) was recovered most, followed by CTX-M (28.6%) and SHV (19.0%). This study reveals that ATMs could harbour pathogenic bacteria with antibiotic resistance (AR) genes especially ESBL genes (blaTEM) which could be responsible for the widespread resistance to antibiotics. Therefore, adequate personal hygiene by users, proper cleaning regimen to sanitize these facilities regularly and public enlightenment are recommended to reduce the accompanying risks of spreading AR genes within the environment

Prevalence of extended-spectrum beta-lactamase and molecular detection of blaTEM, blaSHV, and blaCTX-M genotypes among gram-negative Bacilli isolates from hospital acquired infections in pediatrics, one institutional study

BackgroundGram-negative bacilli represents an important pathogen in hospital-acquired infections (HAIs) worldwide. The emergence of antibiotic resistance in these pathogens warrants attention for the proper management of infections. Extended-spectrum beta-lactamase (ESBL) resistance represents a major therapeutic problem in infections due to Gram-negative bacilli.The present study aimed to study the extended-spectrum beta-lactamase genes blaTEM, blaSHV, and blaCTX-M by multiplex polymerase reaction in isolated Gram-negative bacilli from HAIs in pediatric patients.MethodsThe study included one hundred-five isolates of Gram-negative bacilli from pediatric patients with different types of HAIs. The isolates were subjected to full microbiological identification, antibiotics susceptibility by disc diffusion method, the phenotypic study of ESBL, and the genetic study of ESBL genes by multiplex PCR.ResultsFifty isolates of Gram-Negative bacilli showed ESBL activity by a phenotypic study by double disc diffusion method (50/105). All ESBL producers’ isolates were positive by PCR for ESBL genes. The most frequent gene was blaTEM (64%), followed by blaSHV (30%) and CTX-M (22%). Mixed genes were found in 4 isolates (8%) for blaTEM and blaSHV, blaTEM and CTX-M. There was a significant association between PCR for ESBL genes and phenotypic ESBL detection (P = 0.001). There was significant detection of ESBL genes in E. coli (28%), followed by Enterobacter spp. (26%), Klebsiella spp. (24%), Serratia (14%), Pseudomonas spp. (6%) and Proteus (2%), P = 0.01. There Seventy percent of isolates positive for ESBL production had an insignificant association between MDR and PCR for ESBL genes (P = 0.23).ConclusionThe present study highlights the prevalence of ESBL activity among clinical isolates of Gram-negative bacilli isolated from hospital-acquired infections in pediatric patients. The most common gene responsible for this activity was blaTEM gee followed by blaSHV and blaCTX-M. There was a high prevalence of multiple antibiotic resistance among isolates with ESBL activity. The finding of the present study denotes the importance of screening extended beta-lactamase among Gram-negative bacilli associated with HAIs in pediatric patients.

Transition mutation bias is crucial to adaptive extended spectrum beta lactamase (ESBL) resistance evolution

The importance of slow accumulation of mutation in the nucleic acids of bacteria and the possibility of organismal transcriptional modification to suit their adaptation are key drivers favouring natural selection and subsequently adaptive evolution. This study was aimed at providing insight into factors influencing adaptive extended spectrum beta lactamase (ESBL) resistance evolution. A total of twenty-six (26) extended spectrum beta lactamase-producing Gram-negative bacteria isolated in our previous study were subjected to uniplex and multiplex polymerase chain reaction (PCR) and sequencing prior to mutation discovery analyses, while the maximum likelihood value, transition-transversion ratio, tajimas neutrality test, nucleotide diversity and selection analyses were estimated following standard recommended protocols. Results obtained depict the PCR discriminated ESBL alleles as; TEM (n = 11.5 %), CTX-M (n = 15.4 %), SHV (n = 19.2 %) while the co-occurred ESBL alleles were estimated as CTX-M and TEM (n = 3.85 %), CTX-M and SHV (n = 15.4 %), TEM and SHV (n = 23.1 %), CTX-M, TEM and SHV (n = 11.5 %). The transition, transversion, deletion and insertion distribution pattern were evaluated as; TEM (46, 39,11 and 4 %), SHV (50, 31, 8 and 11 %) and CTX-M (60, 21,12 and 7 %) (p < 0.05) respectively. The maximum likelihood value, transition-transversion ratio, tajimas neutrality test, nucleotide diversity and selection analyses for TEM, SHV and CTX-M were (−8436, 1.18, 5.2, 62 % and 84.9 %,), (−8321, 1.60, 5.4, 68.9 % and 97.8 %) and (−7284, 2.86, 6.0, 55.2 % and 77.2 %) sequentially. Our findings therefore have shown that adaptive ESBL resistance evolution is driven by transition mutation, positive natural selection and ESBL gene flow. It is thus imperative to state that understanding these adaptive resistance driven factors and how they operate is crucial to deciphering the fundamental principles of adaptive evolution that might help circumvent the global challenges of antibiotic resistance.

Vibriosis in South Asia: A systematic review and meta-analysis

ObjectivesSouth Asia remains home to foodborne diseases caused by the Vibrio species. We aimed to compile and update information on the epidemiology of vibriosis in South Asia. MethodsFor this systematic review and meta-analysis, we searched PubMed, Web of Science, EMBASE, and Google Scholar for studies related to vibriosis in South Asia published up to May 2023. A random-effects meta-analysis was used to estimate the pooled isolation rate of non–cholera-causing Vibrio species. ResultsIn total, 38 studies were included. Seven of these were case reports and 22 were included in the meta-analysis. The reported vibriosis cases were caused by non-O1/non-O139 V. cholerae, V. parahaemolyticus, V. fluvialis, and V. vulnificus. The overall pooled isolation rate was 4.0% (95% confidence interval [CI] 3.0-5.0%) in patients with diarrhea. Heterogeneity was high (I2 = 98.0%). The isolation rate of non-O1/non-O139 V. cholerae, V. parahaemolyticus, and V. fluvialis were 9.0 (95% CI 7.0-10.0%), 1.0 (95% CI 1.0-2.0%), and 2.0 (95% CI: 1.0-3.0%), respectively. Regarding V. parahaemolyticus, O3:K6 was the most frequently isolated serotype. Cases peaked during summer. Several studies reported antibiotic-resistant strains and those harboring extended-spectrum beta-lactamases genes. ConclusionsThis study demonstrates a high burden of infections caused by non–cholera-causing Vibrio species in South Asia.

Occurrence of Extended-Spectrum Beta-Lactamase and Quinolone Resistance Genes among Escherichia coli and Klebsiella pneumoniae Isolated from Poultry, Domestic Pigs and Environ in Msimbazi River Basin in Tanzania

Occurrence of Extended-Spectrum Beta-Lactamase and Quinolone Resistance Genes among Escherichia coli and Klebsiella pneumoniae Isolated from Poultry, Domestic Pigs and Environ in Msimbazi River Basin in Tanzania

Antimicrobial Susceptibility and Characterization of Extended-Spectrum β-Lactamase-Producing Escherichia coli Isolated from Stools of Primary Healthcare Patients in Ethiopia.

Antimicrobial resistance of Escherichia coli is a growing problem in both developed and developing countries. This study aimed to investigate the phenotypic antimicrobial resistance of E. coli isolates (n = 260) isolated from the stool specimen of patients attending public health facilities in Addis Ababa and Hossana. This study also aimed to characterize phenotypically confirmed extended-spectrum beta-lactamase (ESBL)-producing E. coli isolates (n = 22) using whole-genome sequencing. Resistance to 18 different antimicrobials was assessed using the disc diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The highest resistance rate among the E. coli isolates was found for ampicillin (52.7%), followed by trimethoprim-sulfamethoxazole (29.6%). Of all isolates, 50 (19.2%) were multidrug-resistant and 22 (8.5%) were ESBL producers. ESBL genes were detected in 94.7% of the sequenced E. coli isolates, and multiple β-lactamase genes were detected in 57.9% of the isolates. The predominant ESBL gene identified was blaCTX-M-15 (78.9%). The blaTEM-1B gene was detected in combination with other ESBL genes in 57.9% of the isolates, while only one of the sequenced isolates contained the blaTEM-1B gene alone. The blaCTX-M-3 gene was detected in three isolates. The genes blaCTX-M-15 and blaTEM-1B as well as blaCTX-M-15 and blaTEM-169 were confirmed to coexist in 52.6% and 10.5% of the sequenced E. coli isolates, respectively. In addition, blaOXA-1 was identified together with blaCTX-M-15 and blaTEM-1B in one isolate, and in one isolate, blaTEM-169 together with blaCTX-M-15 and blaTEM-1B was found. The results obtained show that measures need to be taken to reduce the spread of drug resistance and ensure the long-term use of available antimicrobials.

PREVALENCE OF BLATEM, BLASHV, AND BLACTX-M GENES AMONG ESBL-PRODUCING ESCHERICHIA COLI ISOLATED FROM THE BLOOD SAMPLES OF ICUS PATIENTS OF UNIVERSITY HOSPITALS IN SANA'A CITY, YEMEN

Aim and Objective: With the emergence of organisms such as Enterobacteriaceae that produce extended-spectrum β-lactamase (ESBL), which are resistant to multiple medications (multidrug-resistance), concerns about how best to treat infections have significantly increased. The current study examined the molecular features of ESBL in clinical isolates of Escherichia coli that resulted in bloodstream infections as well as the pattern of antibiotic resistance to gather useful data on the infection's epidemiology among Yemeni ICU patients. Subjects and methods: A cross-sectional study was conducted on sepsis patients admitted in intensive care units at four hospitals in Sana'a, Yemen, between January, 2021 and April, 2022. Blood cultures were used on patients suspected of having sepsis. Standard laboratory procedures were then used to isolate and identify possible bacterial infections, and the disk diffusion method was used to test for microbial susceptibility. All strains were tested for Extended Spectrum Beta-Lactamase (ESBL) production using the Modified Double Disc Synergy Test (MDDST). Following analysis, polymerase chain reaction, β-lactamase genes (blaTEM, blaSHV, and blaCTX-M) were identified. Results: The results of the conventional PCR experiment revealed that 33.3% of the blaCTX-M genes, 0.0% of blaSHV, and 100% of blaTEM were present in the strains of ESBL-producing E. coli that were collected. It was discovered that the E. coli isolates' patterns of antibiotic resistance to 23 different antibiotics differed greatly. The bulk of the E. coli isolates were found to be multi-drug resistant (MDR). Furthermore, MDR characteristics were observed in 85% of E. coli isolates. Conclusion: Control and surveillance of antibiotic resistance depend on an understanding of the resistance genes and patterns of antimicrobial resistance of bacterial pathogens within a given geographic area. The current study's findings showed that MDR was very common. According to the current study's findings, TEM was significantly more common than other ESBL gene types. Peer Review History: Received: 7 November 2023; Revised: 12 December; Accepted: 6 January 2024, Available online: 15 January 2024 Academic Editor: Dr. A.A. Mgbahurike, University of Port Harcourt, Nigeria, amaka_mgbahurike@yahoo.com Received file: Reviewer's Comments: Average Peer review marks at initial stage: 6.5/10 Average Peer review marks at publication stage: 8.0/10 Reviewers: Dr. Rola Jadallah, Arab American University, Palestine, rola@aauj.edu Dr. Wadhah Hassan Ali Edrees, Hajja University, Yemen, edress2020@gmail.com

Carbapenemase and extended-spectrum beta-lactamase-producing bacteria in waters originating from a single landfill in Slovenia.

Groundwater, rainwater, and leachate associated with a single landfill were analysed to detect extended-spectrum beta-lactamase (ESBL)-producing and carbapenemase (CP)-producing bacteria. After cultivation on three commercial selective-differential media, 240 bacterial isolates were obtained and identified by matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS). Isolates from clinically relevant species were further genotyped by enterobacterial repetitive intergenic consensus polymerase chain reaction, and tested for antibiotic susceptibility and presence of CPs and ESBL enzymes. Two ESBL-producing isolates and two isolates producing CPs were detected in rainwater, groundwater, and leachate: Klebsiella oxytoca complex with the gene for the ESBL enzyme CTX-M-1 and the gene for the CP OXA-48, Serratia fonticola with the gene for the ESBL enzyme FONA-2, and Pseudomonas aeruginosa with the gene coding Verona integron-encoded Metallo-beta-lactamases (VIM) metallo-beta-lactamase. Our study indicates that bacteria with ESBL and CP genes can be present in landfill-associated waters.

Prevalence of blaCTX-M and blaTEM Genes in Cefotaxime-Resistant Escherichia coli Recovered from Tertiary Care at Central Nepal: A Descriptive Cross-Sectional Study.

Urinary tract infections (UTIs) are highly prevalent globally, and various antibiotics are employed for their treatment. However, the emergence of drug-resistant uropathogens towards these antibiotics causes a high rate of morbidity and mortality. This study was conducted at the Microbiology Laboratory of Grande International Hospital from November 2021 to May 2022 and aimed to assess the prevalence of UTI caused by Escherichia coli and their antibiotic susceptibility pattern with a focus on extended-spectrum beta-lactamases (ESBLs) and the prevalence of two genes (blaCTX-M and blaTEM) in cephalosporin-resistant E. coli. Altogether, 1050 urine samples were processed to obtain 165 isolates of E. coli. The isolates were identified by colony morphology and biochemical characteristics. Antimicrobial susceptibility tests (ASTs) were determined by the Kirby-Bauer disk diffusion method, and their ESBL enzymes were estimated by the combined disk method (CDM). Two ESBL genes (blaCTX-M and blaTEM) were investigated by polymerase chain reaction (PCR) in cefotaxime-resistant E. coli. Among the 1050 urine samples that were processed, 335 (31.9%) were culture-positive with 165 (49.2%) identified as E. coli. The age group ≥60 years (30.3%) had greater susceptibility to bacterial infections. AST revealed that meropenem was highly effective (95.7% susceptibility), while ampicillin showed the least sensitivity (42.4%). Among the E. coli isolates, 86 were multidrug resistant (MDR) and 10 were extensively drug resistant (XDR). Of these, 46 MDR (96%) and 2 XDR (4%) were ESBL producers. The prevalence of ESBL genes (blaCTX-M and blaTEM) was 49.3% and 54.8%, respectively. The overall accuracy of CDM as compared to PCR for the detection of the blaCTX-M gene was 55.26%. The prevalence of MDR E. coli harboring the blaCTX-M and blaTEM genes underscores the imperative role of ESBL testing in accurately identifying both beta-lactamase producers and nonproducers.

Municipal wastewater treatment plant showing a potential reservoir for clinically relevant MDR bacterial strains co-occurrence of ESBL genes and integron-integrase genes

Municipal wastewater treatment plants (MWWTPs) are a milieu for co-occurrence of multiple antibiotic resistance genes (ARGs). This facilitates mixing and genetic exchange; and promotes dissemination of multidrug resistance (MDR) to wastewater bacterial communities which is hazardous for the effluent receiving environment. This study investigated the co-occurrence of extended-spectrum beta-lactamase (ESBL) genes (blaTEM, blaCTX-M, blaSHV, blaOXA), and integron-integrase genes (intI1, intI2, intI3) in MDR bacteria isolated from the Bharwara MWWTP in Lucknow, India. Thirty-one MDR bacterial colonies resistant to three or more antibiotics were isolated from three treatment stages of this MWWTP. Six of these: Staphylococcus aureus, Serratia marcescens, Salmonella enterica, Shigella sonnei, Escherichia coli, and Bacillus sp. Had co-occurrence of ESBL and integron-integrase genes. These six isolates were examined for the occurrence of MDR efflux genes (qacA, acrB) and ARGs (aac(3)-1, qnrA1, tetA, vanA) and tested for resistance against 12 different antibiotics. The highest resistance was against penicillin-G (100%) and lowest for chloramphenicol (16.66%). Bacillus sp. Isolate BWKRC6 had the highest co-occurrence of antibiotic resistance-determining genes and was resistant to all the 12 antibiotics tested. The co-occurrence of ESBL, integron-integrase, antibiotic resistance-determining and MDR efflux genes in bacteria isolated from the Bharwara MWWTP indicates that the wastewaters of this treatment plant may have become a hotspot for MDR bacteria and may present human and environmental health hazards. Therefore, there is need for a rapid action to limit the spread of this threat. Public regulatory authorities must urgently implement measures to prevent MWWTPs becoming reservoirs for evolution of antibiotic resistance genes and development of antibiotic resistance.

Different OXA-Carbapenemases in Genetically Unrelated Klebsiella pneumoniae Strains Isolated in a North Italian Hospital During Multidrug Resistance Screening.

Klebsiella pneumoniae is one of the main opportunistic pathogens that cause a broad spectrum of diseases with increasingly frequent acquisition of resistance to antibiotics, namely carbapenems. This study focused on the characterization of 23 OXA-48-like carbapenemase-producing K. pneumoniae isolates using phenotypic and molecular tests. Phenotypic determination of the presence of β-lactamases was performed using the extended-spectrum beta-lactamase (ESBL) NP test, and phenotypic determination of the presence of carbapenemase was based on the Carba NP test. Antimicrobial susceptibility tests were performed to assess the resistance against carbapenems. Molecular characterization of ESBL genes and carbapenemase genes (blaOXA-48, blaKPC, blaVIM, and blaNDM) was performed using polymerase chain reaction (PCR) techniques. In addition, K. pneumoniae strains were analyzed for their relatedness using multilocus sequence typing PCR analysis based on the Institut Pasteur protocol, which produces allelic profiles that contain their evolutionary and geographic pattern. Following further Sanger sequencing of the blaOXA-48 genes, no genetic mutations were found. Some OXA-48-producing K. pneumoniae isolates coharbored blaKPC, blaNDM, and blaVIM genes, which encode other carbapenemases that can hydrolyze carbapenem antibiotics. The final part of the study focused on the characterization of the plasmid profiles of all isolates to better understand the spreading of the IncL/M blaOXA-48 plasmid gene. The plasmid profile also revealed other incompatibility groups, suggesting that other plasmid genes are spreading in K. pneumoniae isolates, which can coharbor and spread different carbapenemases simultaneously.