ESBL Detection Research Articles

Enterobacterales Producing ESBLs and AmpC in Fresh Vegetables from Tebessa City, Algeria.

This study aimed to evaluate the contamination levels of fresh products by ESBLs-producing Enterobacterales (ESBLs-E) or AmpC-producing Enterobacterales and characterize ESBLs genes. A total of 132 samples (67 vegetables and 65 fruits) were collected from markets in Tebessa, eastern Algeria. Among the samples, 16 third-generation cephalosporin-resistant Enterobacterales isolates were identified with a prevalence of 19.40% in vegetable samples, while there was no positive finding in fruit samples. Isolates showed resistance to most β-lactams, and all of them displayed multidrug resistance. Phenotypic tests for ESBLs detection, using double-disk synergy test and double-disk test were positive for 14 strains, including Klebsiella pneumoniae (n = 5), Klebsiella oxytoca (n = 4), Klebsiella terrigena (n = 2), Kluyvera spp. (n = 2), and Enterobacter cloacae (n = 1). Two AmpC-producing strains (Citrobacter freundii and E. cloacae) were identified through the AmpC disk test. Contamination rates of vegetables by ESBLs-E and AmpC-producing Enterobacterales were 19.40% and 2.98%, respectively. PCR results showed the presence of at least one ESBL gene in seven selected strains, with the dominance of blaCTX-M gene. Notably, K. pneumoniae strains showed the co-occurrence of two or three genes. Sequencing identified uncommon variants of ESBLs genes for the first time in Algeria, including blaCTX-M-79 (2/7), blaCTX-M-107 (2/7), blaCTX-M-117 (2/7), blaTEM-112 (1/7), blaTEM-125 (2/7), blaTEM-194 (1/7), and blaSHV-176 (3/7).

A worrisome prevalence of extended-spectrum β-lactamase producers in patients with biliary obstruction and cholangitis: Phenotypic and molecular characterization of biliary Escherichia coli and Klebsiella pneumoniae isolates

Background & aimsThe alarming rise of antibiotic resistance presents a substantial and worrisome issue within the context of biliary obstruction, specifically in the treatment of cholangitis. This abovementioned scenario underscores the critical importance of addressing extended spectrum β-lactamase (ESBL) producers in the biliary system to adequately tackle cholangitis using third-generation cephalosporins. Hence, we aimed to determine the frequency of ESBL and carbapenemases among biliary Escherichia coli and Klebsiella pneumoniae isolated from patients with biliary obstruction. MethodsIn this cross-sectional study, bile samples were collected via aspiration from patients diagnosed with biliary obstruction during the endoscopic retrograde cholangiopancreatography (ERCP) procedure. Subsequent culturing of these samples was performed, followed by phenotypic and molecular assessments for the detection of ESBL- and carbapenemase-producing strains of E. coli and K. pneumoniae. ResultsApproximately 23.5 % of patients with biliary obstruction harbored biliary ESBL-producers, with the majority (70.2 %) being diagnosed with cholangitis. Moreover, 2.1 % of patients had biliary carbapenemase-producing K. pneumoniae strains. Molecular analysis confirmed the high prevalence of blaCTX-M and blaTEM in E. coli, and blaTEM and blaSHV in K. pneumoniae. Additionally, the presence of biliary K. pneumoniae harboring blaKPC, blaNDM, and blaIMP was observed. ConclusionOur study reveals a noteworthy observation that over half of patients experiencing biliary obstruction harbor ESBL-producing bacteria in their biliary tract. Notably, we discovered a significant link between ESBL producers and the risk of cholangitis. These findings raise important concerns regarding the suitability of employing third-generation cephalosporins as initial treatment for cholangitis and other similar biliary infections.

A Study on Extended Spectrum β-Lactamase Producing (ESBL) and Fluoroquinolone-Resistant Escherichia coli in the Japanese Quails

Antimicrobial resistance (AMR) is a growing public health threat worldwide, and members of the Enterobacteriaceae family are among the clinically important bacteria rapidly developing resistance to available antibacterial agents. The development of extended-spectrum β-lactamase (ESBL) enzymes is the primary cause of resistance to third-generation cephalosporins among Enterobacteriaceae. Quails also represent a source of animal protein deficiency in developing countries including India. This study aimed to detect the presence of extended-spectrum β-lactamases (ESBLs) and fluoroquinolone-resistant Escherichia coli from fecal samples of quails in Puducherry. A total of 36 fecal samples of quails from six different retail outlets were collected. E. coli could be isolated from 27 fecal samples and confirmed as E. coli by polymerase chain reaction by targeting the uspA gene. The isolates were preliminarily screened for ESBL production by indicator antimicrobial disc and phenotypic ESBL production was confirmed by combination disc method. The isolates were subjected to genotypic detection of ESBL (bla CTX-M, bla TEM, and bla SHV) by PCR. Out of 27 isolates, 16 isolates were positive for the presence of ESBL-producing genes, in which, 9 harbored bla CTX-M, 2 harbored bla TEM, and 5 harboured bla SHV genes. Similarly, out of 27 isolates 13 isolates were found positive for fluoroquinolone resistance in which 13 (48.18%) isolates harbored the qnrS gene, and 11 (40.74%) isolates harbored the qnrB gene. Antimicrobial susceptibility test showed that the isolates exhibited a high level of resistance against Cefpodoxime (90%), Amoxycillin/clavulanic acid (70%), Ceftriaxone (60%), Cefotaxime (55%), Ceftazidime (45%), Ciprofloxacin (40%) and Aztreonam (30%).

Can flow cytometry be an alternative method for the detection of antimicrobial resistance?

Background: The inappropriate use of antibiotics may cause antibiotic resistance, cause side effects, and eventually cause an increase in healthcare costs. This study aimed to determine a flow cytometry-based test to detect ESBL E. coli and MDR p. aeruginosa in a short time. Methods: The study included 25 E.coli isolates and 25 P.aeruginosa isolates identified by Phoenix TM 100 automated system [Becton Dickinson, USA]. In the flow cytometric method, the percentages of death cells exposed to cephalosporin including ceftazidime [CAZ] or cefotaxime [CTX] and clavulanic acid [CLA] combination, were compared with death cells exposed only to a cephalosporin [CAZ or CTX]. CLA index values [CAZ-CLA and CTX-CLA indices] were obtained for CTX and CAZ. Index values that were higher than 1.5 just for one cephalosporin were accepted as positive. Results: Cell viability was performed after 1-hour exposure to the drugs. When PI staining was applied to ESBL-treated and MDR-treated bacteria, 65% and 85% had nonviable membranes, indicating the method efficiently identified only cells with damaged membranes. Conclusion: Flow cytometry is a rapid and reliable method for the detection of ESBL and MDR in clinical microbiology laboratories Keywords: Antibiotic resistance; ESBL; flow cytometry; MDR.

A comparative study of a rapid phenotypic antimicrobial susceptibility testing system directly from positive blood cultures to the disk diffusion and VITEK 2 methods

BackgroundSepsis is a life-threatening condition that impacts 49 million people annually and causes 11 million deaths worldwide. Surviving bloodstream infections (BSIs) depends on the rapid administration of effective antimicrobial treatment, underscoring a need for rapid antimicrobial susceptibility testing (AST). AimTo evaluate the performance of Quantamatrix's dRAST v2.5 system (Seoul, South Korea) for AST directly from positive blood cultures as compared to the Disk-Diffusion (DD) and VITEK 2 methods. MethodsThe study included 191 positive blood cultures from clinical samples and spiked blood culture bottles. Following Gram staining and species-level identification, AST was performed by VITEK 2 and standard DD methods using CLSI (2021) interpretation. ResultsdRAST demonstrated very good AST performance for a Gram-negative isolate, and good performance for Gram-positive isolates, meeting CLSI criteria for the acceptance of a new method. Antimicrobials that were not considered verified compared to VITEK 2 and DD were cefazolin, ceftazidime, meropenem, and trimethoprim/sulfamethoxazole for Gram-negatives and clindamycin, erythromycin, penicillin, and oxacillin for Gram-positives. dRAST ESBL detection results were strongly correlated with the ESBL phenotypes obtained with other methods. Additional resistance mechanisms were in concordance with traditional tests. ConclusionsdRAST demonstrated good AST performance, meeting CLSI criteria for most relevant antibiotics. dRAST was associated with a significant reduction in time-to-results, labor, and the subjectivity of result analyses, making it a valuable addition to efforts supporting the treatment of patients with bacteremia.AST (antimicrobial susceptibility test), blood culture, dRAST, rapid methods, sepsis, turnaround time (TAT).

Extended-spectrum beta-lactamase and carbapenemase producing Klebsiella Pneumoniae isolated from patient with suspected urinary tract infection

Urinary tract infection (UTI) is the most common bacterial infections in humans and serious health problem in many parts of the world. Carbapenem resistant Enterobacteriaceae (CRE) are increasingly reported from healthcare facilities in Nepal and around the world. This study is carried out with an objective to assess the Prevalence of ESBL and carbapenemase producing Klebsiella pneumoniae isolate from patient with suspected of UTI. Total 880 urine sample was collected during 6-month period and processed and identified by using conventional microbiological procedure and biochemical test. ESBL screening isolates were done by using Ceftazidime and Ceftazidime with clavulanic acid. Production of ESBL was determine by combination Disc Test. For the confirmation of carbapenemase producing K. pneumoniae Modified Hodge Method, Carbapenem Inactivation Method and EDTA Impregnated Disc test (MBL) was performed. Out of total sample, 9.30% (82) showed significant growth. Prevalence of UTI seen more in male as compare to female patients. Out of positive isolates, 37.8% were K. pneumoniae. K. pneumoniae were resistance with Ceftriaxone (54.8%) followed by 51.61% Ceftazidime, Cotrimoxazole. Multi drug resistance (MDR) was found to be 54.83%. Out of 31 isolates, 10(32%) were confirmed as ESBL positive by Combination Disc Assay. 14 isolates were screened as carbapenemase producers but 2(14.28%) showed positive through Modified Hodge Method, 4(28.57%) Modified Carbapenem Inactivation Method and 7(50%) EDTA impregnated Disc Test (MBL). Analytically, the sensitivity of MHT, CIM, and MBL tests was 42.81%, 58.10%, and 76.96%, respectively, all with 100% specificity. Colistin was the drug of choice, with 93.54% of isolates showing sensitivity to it. Early detection of ESBL and MBL-producing isolates is crucial for reducing mortality rates and preventing the intra-hospital spread of these strains.

Ventilator Associated Pneumonia in Tertiary Care Hospital, Maharajgunj, Kathmandu, Nepal

Introduction: Ventilator Associated Pneumonia (VAP) is the most common nosocomial infection among intensive care unit (ICU) patients and lack of much information in Nepal. So, the aim of this study was to determine prevalence and bacteriological profile of VAP with special reference to multi-drug resistant (MDR), Methicillin-resistant Staphylococcus aureus(MRSA), Metallo-β-Lactamase(MBL), Extended-Spectrum β-Lactamase(ESBL)-producing bacterial strains. Methods: A total 150 tracheal specimens were studied during June 2011 to May 2012 at Department of Microbiology, TUTH as described by American Society for Microbiology (ASM). Combination disk method was done for the detection of ESBL and MBL producing isolates. Results: Prevalence of VAP was found to be 34%. Acinetobactereal coaccticusbaumannii complex (44%) was the commonest isolate, followed by Klebsiellapneumoniae (22%), Pseudomonas aeruginosa (16%) and Staphylococcus aureus (12%). Among MDR Gram negative bacteria (GNB), 39% were MBL and 33% were ESBL-producers. All GNB (61) were sensitive to Polymyxin B and Colistinsulphate, whereas, 48% were found resistant to Carbapenems. Prevalence of MRSA was 75%, which were all sensitive to Vancomycin. Conclusion: High prevalence of VAP, MDR along with MRSA or ESBL or MBL producing strains was found in the study. Thus, suitable control measures must be adopted to cope up this alarming situation with genetic characterization.

Coexistence of multidrug resistance and ESBL encoding genes - blaTEM, blaSHV, and blaCTX-M; its amplification and dispersion in the environment via municipal wastewater treatment plant

Municipal wastewater treatment plants (MWWTPs) are a global source of antibiotic resistance genes (ARGs), collecting wastewater from a variety of sources, including hospital wastewater, domestic wastewater, runoff from agricultural and livestock farms, etc. These sources are contaminated with organic and inorganic pollutants, ARGs and antibiotic-resistant bacteria (ARB). Such pollutants aided eutrophication and encouraged bacterial growth. During bacterial growth horizontal gene transfer (HGT) and vertical gene transfer (VGT) of ARGs and extended-spectrum β-lactamase (ESBL) encoding genes may facilitate, resulting in the spread of antibiotic resistance exponentially. The current study investigated the prevalence of multidrug resistance (MDR) and ESBL encoding genes in various treatment units of MWWTP and their spread in the environment. A total of three sampling sites (BUT, BRO, and BFB) were chosen, and 33 morphologically distinct bacterial colonies were isolated. 14 of the 33 isolates tested positive for antibiotic resistance and were further tested for the coexistence of MDR and ESBL production. The selected 14 isolates showed the highest resistance to trimethoprim (85.71%), followed by ciprofloxacin, azithromycin, and ampicillin (71.42%), tetracycline (57.14%), and vancomycin, gentamicin, and colistin sulphate (50%). A total of 9 isolates (64.28%) were phenotypically positive for ESBL production (BUT2, BUT3, BUT5, BRO1, BRO2, BRO3, BRO4, BRO5 and BFB1). The molecular detection of ESBL encoding genes, i.e. blaTEM, blaSHV, and blaCTX-M was carried out. The most prevalent gene was blaTEM (69.23%), followed by blaSHV (46.15%), and blaCTX-M (23.07%). In this study, 9 isolates (64.28%) out of 14 showed the coexistence of MDR and ESBL encoding genes, namely BUT3, BUT4, BUT5, BUT6, BUT7, BRO1, BRO2, BRO4, and BFB1. The coexistence of ESBL encoding genes and resistance to other antibiotic classes exacerbates human health and the environment.

A proof-of-principle study for the point-of-care detection of ESBL (CTX-M) by NG-Test® CTX-M MULTI lateral flow assay in urine samples using a simplified method for use in a resource-limited setting.

The rise of extended-spectrum β-lactamase-producing Enterobacterales (ESBL-E) in low- and middle-income countries limits treatment options, leading to the frequent use of broad-spectrum antibiotics. Reducing time-to-result for a urinary infection can facilitate correct antibiotic treatment and support antimicrobial and diagnostic stewardship measures. This study compared two simplified enrichment methods for detecting CTX-M directly from urine specimens. Two enrichment methods, namely centrifugation of 2 mL urine and filtration of 1 mL urine using the DirecTool adaptor, were compared using 20 culture-positive urine samples (20 suspected ESBL-E and 20 non-ESBL-E). CTX-M production was detected using a lateral flow assay (LFA), NG-Test® CTX-MMULTI. The presence of bla CTX-M genes was confirmed by whole-genome sequencing (WGS). The results of both enrichment methods were identical, with a sensitivity of 87.5% and a specificity of 100%. In 19/20 (95%) of the urine samples, the results of the CTX-M LFA were identical with the phenotypic confirmation and WGS. Both methods could detect ESBL-E bacteriuria with ≥104 cfu/mL. All ESBL-E-negative samples were identified accurately. Both enrichment methods yielded negative results in one ESBL-E-positive (CTX-M-15) sample despite phenotypic and genotypic confirmation of ESBL production. High leukocyte count (>500 cells/µL), the presence of boric acid or polymicrobial samples did not appear to impact the performance of both enrichment methods. Our study underscores the feasibility of directly detecting CTX-M in urine. Simplified enrichment methods, particularly with a filtration kit, enhance the assay's practicality, rendering it suitable for use in primary care, emergency departments or remote laboratories without sophisticated equipment.

Detection of ESBL E.coli That Carried STX1 and STX2 Form Common Carp (Cyprinus carpio) in Salhaldeen Province

Detection of ESBL E.coli That Carried STX1 and STX2 Form Common Carp (Cyprinus carpio) in Salhaldeen Province

Phenotypic detection of ESBL and Amp C beta lactamase among clinical isolates of Enterobacteriaceae

Phenotypic detection of ESBL and Amp C beta lactamase among clinical isolates of Enterobacteriaceae

Phenotypic identification of different β-Lactamases in intrinsic and acquired colistin resistant uropathogenic gram negative bacteria.

Identification of MBL, AmpC and ESBLs in colistin intrinsic and acquired resistant uropathogenic gram negative bacteria. Urine samples were collected from Hayatabad Medical Complex, Peshawar during 17 January to 30 June 2019. Collected urine samples were aseptically transported microbiology lab of Health Research Institution (HRI), National Institute of Health (NIH), Khyber Medical College, Peshawar and streaked on different media. Positive growth was identified by API-10s. Antibiotic sensitivity profile was done by Modified Kirby Bauer disc diffusion method. Detection of metallo βlactamases (MBL) production by Imipenem EDTA synergy test, Double Disc Synergy Test (DDST) for detection of ESBLs and D-test for the detection of inducible AmpC beta lactamases test was used. Colistin resistance was identified via broth micro dilution according to CLSI manual. Colistin resistant bacteria was divided in two categories; acquired and intrinsic resistant bacteria according to CLSI manual. Out of 2000 urine samples, 281(14%) gram-negative bacteria were isolated. Among positive samples, acquired colistin resistant bacteria were 241 and intrinsic resistant bacteria were 40 isolates. MBL was produce by twenty one (11.7%) E.coli and seventeen (40.5%) Pseudomonas aeruginosa. E. coli, Pseudomonas aeruginosa, Klebsiella Pneumoniae, Serratia Oderifora and Proteus Marblis were ESBLs producing bacteria. AmpC production was prevalent in fourteen (7.8%) E. coli and twelve (28.6%) Pseudomonas aeruginosa. Fifty-five samples showed resistance to colistin out of 241 samples. In colistin resistant bacteria, two E.coli were MBL, ESBLs, while one E.coli was ESBLs, AmpC co-producing bacteria. The most prevalent extended drug resistant bacteria were Pseudomonas aeruginosa (28.6%) and Escherichia coli (6.1%), While 155(86.6%) Escherichia coli, 25 (59.5%) Pseudomonas aeruginosa and 22 (95.7%) Serratia Oderifora was multi drug resistant bacteria. Current study concluded that ESBL, MBL AmpC enzymes and their co-expression was observed with colistin resistance in E.coli and Pseudomonas aeruginosa.

Comparative Study of Biofilm and Non-Biofilm Producing Klebsiella pneumoniae with Special Reference to Metallo-Beta-Lactamase Production

Klebsiella pneumoniae is one of the most common bacteria among all biofilm-producing as well as the beta-lactamase producing strains, which is responsible for multi-drug resistance. For better therapeutic applications, it is important to detect the biofilm production by Klebsiella pneumoniae and their antibiogram along with the ability to produce ESBL, MBL, and AmpC β-lactamases. The aim of the study was to determine the prevalence of biofilm formation and ESBL, MBL, AmpC β-lactamase phenotypes in K. pneumoniae as well as the antibiogram of all (biofilm and MBL-producing and non-producing) isolates of K. pneumoniae. Isolates of K. pneumoniae were tested for biofilm formation by the Congo-red agar method. ESBL, MBL, and AmpC β-lactamase detection were done by both screening and confirmatory tests as per CLSI guidelines. The antibiogram was obtained by the Kirby-Bauer disc diffusion method. Among the total 100 isolates of K. pneumoniae, 40% were biofilm-producing. Most of them were from urine specimens. Out of biofilm-producing isolates, ESBL – 28%, MBL- 47% and AmpC β-lactamase- 25.8% producers were observed. K. pneumoniae isolates were seen to have maximum resistance to ceftazidime and maximum sensitivity to nitrofurantoin. Study findings suggest the importance of assessment of biofilm formation for better treatment. The scenario further worsens if such biofilm-producing isolates are also MBL-positive leading to limited therapeutic options.

A Study of Genotypic Characterization of ESBL and MBL Genes of β-Lactamase Producing Pseudomonas aeruginosa in Various Clinical Samples

Pseudomonas aeruginosa, a Gram-negative bacterium, presents a substantial challenge in healthcare due to its adaptability and resistance. This study delves into its genotypic characteristics, focusing on ESBL and MBL genes. The prevalence of P. aeruginosa in nosocomial infections, the research aims to decipher resistance mechanisms crucial for tailored interventions. The study includes 170 non-repetitive clinical samples with protocols. Antibiotic susceptibility testing reveals diverse resistance patterns. Molecular detection of ESBL and MBL genes involves DNA isolation, PCR amplification, and gel electrophoresis. The study examined 170 P. aeruginosa samples, revealing gender-specific variations with 65.91% male and 34.09% female isolates. Antimicrobial testing displayed resistance in Ceftazidime (59%) and Ciprofloxacin (48%), while Ticarcillin-clavulanic acid showed promising sensitivity (58%). Molecular identification unveiled diverse resistance genes across sample types, emphasizing genetic complexity. The study underscores the urgency for targeted therapeutic interventions and novel antimicrobial strategies against P. aeruginosa infections. As antimicrobial resistance complexities persist, this research guides efforts toward a profound understanding of clinical interventions and strategic antimicrobial management.

MAST® D72C test: a novel option for ESBL, AmpC and carbapenemase detection.

The MAST® D72C test is a phenotypical test which can detect ESBL and AmpC production in Enterobacterales. It can also identify the suspected presence of carbapenemase. The aim of the present study was to assess the sensitivity and specificity of this test and to discuss its usefulness in laboratories, especially those that use only an automated AST system. The performance of the MAST® D72C test was assessed against a collection of 119 non-redundant Enterobacterales isolates characterized for their content in β-lactamases, and compared with that of the reference double disk synergy test. β-lactamase content was established from phenotypic and genotypic analyses to collect a broad diversity of resistance mechanisms and bacterial strains, including 30 ESBL-producing strains, 32 strains overproducing chromosomal AmpC, 10 strains producing plasmid-encoded AmpC, 12 carbapenemase-producing strains, 13 strains combining the production of several β-lactamases, and 22 strains that produced other β-lactamases. The sensitivity and specificity for ESBL-detection were comparable with those of the synergy test, 75 versus 72.5%, and 94.9 versus 93.7%, respectively. The sensitivity and specificity for AmpC-detection were 71.7% and 100%, respectively, and sensitivity reached 78.7% if we excluded carbapenem-resistant isolates. Carbapenemase-detection sensitivity was 90%. These results show that the MAST® D72C test can be a useful tool for the detection of ESBL- and AmpC-production in clinical laboratories.

Survival of highly related ESBL- and pAmpC- producing Escherichia coli in broiler farms identified before and after cleaning and disinfection using cgMLST

BackgroundBroiler chickens are frequently colonized with Extended-Spectrum Beta-Lactamase- (ESBL-) and plasmid mediated AmpC Beta-Lactamase- (pAmpC-) producing Enterobacterales, and we are confronted with the potential spread of these resistant bacteria in the food chain, in the environment, and to humans. Research focused on identifying of transmission routes and investigating potential intervention measures against ESBL- and pAmpC- producing bacteria in the broiler production chain. However, few data are available on the effects of cleaning and disinfection (C&D) procedures in broiler stables on ESBL- and pAmpC- producing bacteria.ResultsWe systematically investigated five broiler stables before and after C&D and identified potential ESBL- and pAmpC- colonization sites after C&D in the broiler stables, including the anteroom and the nearby surrounding environment of the broiler stables. Phenotypically resistant E. coli isolates grown on MacConkey agar with cefotaxime were further analyzed for their beta-lactam resistance genes and phylogenetic groups, as well as the relation of isolates from the investigated stables before and after C&D by whole genome sequencing. Survival of ESBL- and pAmpC- producing E. coli is highly likely at sites where C&D was not performed or where insufficient cleaning was performed prior to disinfection. For the first time, we showed highly related ESBL-/pAmpC- producing E. coli isolates detected before and after C&D in four of five broiler stables examined with cgMLST. Survival of resistant isolates in investigated broiler stables as well as transmission of resistant isolates from broiler stables to the anteroom and surrounding environment and between broiler farms was shown. In addition, enterococci (frequently utilized to detect fecal contamination and for C&D control) can be used as an indicator bacterium for the detection of ESBL-/pAmpC- E. coli after C&D.ConclusionWe conclude that C&D can reduce ESBL-/pAmpC- producing E. coli in conventional broiler stables, but complete ESBL- and pAmpC- elimination does not seem to be possible in practice as several factors influence the C&D outcome (e.g. broiler stable condition, ESBL-/pAmpC- status prior to C&D, C&D procedures used, and biosecurity measures on the farm). A multifactorial approach, combining various hygiene- and management measures, is needed to reduce ESBL-/pAmpC- E. coli in broiler farms.

Microbiology of Klebsiella with extended-spectrum β-lactamases production

Extended-spectrum β-lactamases (ESBL) producing Klebsiella represents one of the greatest challenges in therapeutic management of the patients. Hence, the objective was to determine ESBL producing Klebsiella species and its molecular characterization in clinical samples of tertiary-care hospital. Klebsiella isolated from various samples were included and by using disc diffusion method ESBL detection amongst the Klebsiella isolates along with molecular detection of blaTEM, blaSHV, blaCTX-M genes which are associated with ESBL production was done. Results revealed that out of 3194 samples, 200 Klebsiella species were isolated. The ratio of Klebsiella species isolated from inpatient and outpatient departments was 75.5% and 24.5% respectively. The maximum isolates were obtained from men consisting of 57.50% and aged between 31-40 years with 26.50%. Majorly Klebsiella species were isolated from sputum including 30.5%, followed by urine consisting of 28% and pus samples with 19.5%. From the 200 isolates, 197 were Klebsiella pneumonia and 32.5% were ESBL producers with majorly having the blaTEM gene incorporated. Being Multidrug and Pan-drug resistant, Klebsiella with ESBL strains are a threat to mankind. This can be overcome by adhering to strict hospital policies to avoid irrational use of antibiotics, strictly following institutional and National antibiotic policies and by prescribing antibiotics according to the culture and sensitivity reports.

Investigation of Fosfomycin Resistance and Plasmid-mediated Fosfomycin Resistance Genes in Enterobacterales Isolates

The study aimed to investigate the fosfomycin resistance in Enterobacterales isolates and the presence of plasmid-mediated fosfomycin resistance genes fosA3 and fosC2 in fosfomycin-resistant Enterobacterales isolates. A total of 2095 Enterobacterales isolates obtained from urine samples were included in the study. Identification of isolates was performed in the Vitek MS. The antimicrobial susceptibility of the isolates and detection of ESBL was determined using the Vitek 2 Compact. In 185 fosfomycin-resistant isolates, fosA3 and fosC2 genes were investigated by PCR. In addition, carbapenemase genes were investigated in carbapenem-resistant isolates. In 2095 Enterobacterales, 65.8% of the isolates were E. coli, and fosfomycin resistance was determined as 22%. ESBL production was defined as 36.5%, 58.1%, and 32.5% in all isolates, K. pneumoniae, and E. coli isolates, respectively. K. pneumoniae (56.2%) were the most frequently isolated bacteria among 185 fosfomycin-resistant Enterobacterales isolates. In these isolates, 66.95% were ESBL-producing Enterobacterales, and 35.7% were carbapenem-resistant isolates. According to the PCR results, two of the 185 Enterobacterales isolates were positive for the fosA3 gene, fosC2 gene was not detected. According to PCR for carbapenemase genes results with these isolates, blaOXA–48 positivity was 66.7%, and blaNDM positivity was 3%. blaKPC and blaVIM positivity could not be detected. In our study, the prevalence of fosA3 was 1.1%. The occurrence of fosA3 gene positivity in Enterobacterales isolates is important for the future treatment of UTIs.

Dissemination of mcr-1 and β-lactamase genes among Pseudomonas aeruginosa: molecular characterization of MDR strains in broiler chicks and dead-in-shell chicks infections

ObjectivesPseudomonas aeruginosa (P. aeruginosa) is one of the most serious pathogens implicated in antimicrobial resistance, and it has been identified as an ESKAPE along with other extremely significant multidrug resistance pathogens. The present study was carried out to explore prevalence, antibiotic susceptibility phenotypes, virulence-associated genes, integron (int1), colistin (mcr-1), and β-lactamase resistance' genes (ESBls), as well as biofilm profiling of P. aeruginosa isolated from broiler chicks and dead in-shell chicks.DesignA total of 300 samples from broiler chicks (n = 200) and dead in-shell chicks (n = 100) collected from different farms and hatcheries located at Mansoura, Dakahlia Governorate, Egypt were included in this study. Bacteriological examination was performed by cultivation of the samples on the surface of both Cetrimide and MacConkey’s agar. Presumptive colonies were then subjected to biochemical tests and Polymerase Chain Reaction (PCR) targeting 16S rRNA. The recovered isolates were tested for the presence of three selected virulence-associated genes (lasB, toxA, and exoS). Furthermore, the retrieved isolates were subjected to phenotypic antimicrobial susceptibility testing by Kirby–Bauer disc diffusion method as well as phenotypic detection of ESBLs by both Double Disc Synergy Test (DDST) and the Phenotypic Confirmatory Disc Diffusion Test (PCDDT). P. aeruginosa isolates were then tested for the presence of antibiotic resistance genes (ARGs): int1, mcr-1, and ESBL genes (OXA-10, OXA-2, VEB-1, SHV, TEM, and CTX-M). Additionally, biofilm production was examined by the Tube Adherent method (TA) and Microtiter Plate assay (MTP).ResultsFifty –five isolates were confirmed to be P. aeruginosa, including 35 isolates from broiler chicks and 20 isolates from dead in-shell chicks. The three tested virulence genes (lasB, toxA, and exoS) were detected in all isolates. Antibiogram results showed complete resistance against penicillin, amoxicillin, ceftriaxone, ceftazidime, streptomycin, erythromycin, spectinomycin, and doxycycline, while a higher sensitivity was observed against meropenem, imipenem, colistin sulfate, ciprofloxacin, and gentamicin. ESBL production was confirmed in 12 (21.8%) and 15 (27.3%) isolates by DDST and PCDDT, respectively. Antibiotic resistance genes (ARGs): int1, mcr-1, and ESBL genes (OXA-10, SHV, TEM, and CTX-M), were detected in 87.3%, 18.2%, 16.4%, 69.1%, 72.7%, and 54.5% of the examined isolates respectively, whereas no isolate harbored the OXA-2 or VEB-1 genes. Based on the results of both methods used for detection of biofilm formation, Kappa statistics [kappa 0.324] revealed a poor agreement between both methods.Conclusionsthe emergence of mcr-1 and its coexistence with other resistance genes such as β-lactamase genes, particularly blaOXA-10, for the first time in P. aeruginosa from young broiler chicks and dead in-shell chicks in Egypt pose a risk not only to the poultry industry but also to public health.Graphical

A Comparative Evaluation of ESBL Production and Carbapenem Resistance in UTIs Before and During COVID-19

Background: The COVID-19 pandemic has heightened the urgency to address antibiotic resistance, particularly ESBL and carbapenem-resistant UTIs. Our study seeks to assess the pandemic's effect on the prevalence of these resistances, comparing data from before and during COVID-19. By identifying trends in resistance patterns, we aim to enhance antibiotic stewardship and inform healthcare policies. Understanding the pandemic's impact on ESBL and carbapenem resistance is vital for guiding clinical practices and public health initiatives in tackling antibiotic resistance. Methods: This retrospective study, conducted at King Fahad Medical City in Riyadh, Saudi Arabia, analyzed positive urine cultures from January 2018 to December 2022 to assess the prevalence of antibiotic resistance among UTIs. Using WHO-recommended semiquantitative culturing methods on CLED and blood agar, bacterial isolates were identified via the BD Phoenix system and antimicrobial susceptibility was assessed following CLSI guidelines. ESBL detection employed the Double-Disc Synergy Test. Data analysis involved SPSS for statistical evaluation, utilizing chi-square and t-tests for categorical and continuous variables, respectively, and logistic regression to explore associations between variables and antibiotic resistance, with significance set at p