blaVIM Genes Research Articles

Virulence profile of carbapenem-resistant Klebsiella pneumoniae strains by an in vivo model of Galleria mellonella.

Klebsiella pneumoniae is a significant healthcare-associated pathogen, notable for its diverse virulence and antibiotic resistance profiles. This study aimed to characterize the genotypic and phenotypic diversity of K. pneumoniae isolates and evaluate their virulence using the Galleria mellonella model. Biomass production, metabolic activity, capsule formation, and siderophore production were assessed in 27 K. pneumoniae isolates from hospital-associated infections. Lethality curves were generated using the G. mellonella model, with survival monitored hourly from 16 to 48 hours. The most common sequence types (ST) identified were the high-risk clones ST307 (N = 10), ST512 (N = 8), ST101 (N = 7), and ST661 (N = 2). These STs were associated with distinct K-locus, including KL102, KL107, KL17, and KL39. Most isolates belonged to the O2afg locus (N = 18), with the K. pneumoniae carbapenemase genotype detected in 96.3% of strains. None of the isolates were classified as hypervirulent. Phenotypically, ST661 exhibited the highest biomass production despite showing similar metabolic activity to other STs. A positive correlation was observed between biomass and siderophore production, while capsule production was inversely correlated with biomass. In the G. mellonella model, ST661 demonstrated the highest virulence, resulting in 100% mortality by 48 hours, compared to survival rates of 21.4% for ST101, 38.0% for ST307, and 31.2% for ST512. These findings underscore the pathogenic potential of ST661 isolates with enhanced biofilm production. The G. mellonella model may serve as an effective in vivo system for evaluating the virulence of emerging K. pneumoniae lineages.IMPORTANCEWe demonstrate that the Galleria mellonella model is a useful tool to analyze the virulence of carbapenem-resistant Klebsiella pneumoniae strains. Our findings highlight the pathogenicity of carbapenem-resistant K pneumoniae isolates, particularly the role of the ST661 that, despite being a rare lineage, harbors the blaVIM gene and is associated with high biofilm production and the highest mortality rates.

Molecular Investigation of Metallo B-Lactamase genes in Klebsiella Pneumoniae Bacteria from Clinical Isolates

Klebsiella pneumoniae is a significant nosocomial bacteria, that causes a large range of infections e.g. bacteremia, pneumonia, meningitis, and urinary tract infections. The current study intends to detect the existence of specific genes (blaNDM-1, blaNDM-2, blaVIM, blaIMP) associated with metallo-beta-lactamase (MBL) production in multidrug-resistant Klebsiella pneumoniae isolates attained from various clinical isolate. These genes are responsible for conferring resistance to a wide range of antibiotics, making an infection caused by such bacteria difficult to treat. Two hundred and seventy eight samples are collected from patients in Baquba Teaching Hospital during the period between December 2023 and May 2024. The samples included urine, sputums, burn, and wound swabs in addition to blood samples. The isolates are diagnosed microscopically and biochemically and the VITEK-2 compact system was used for diagnosis confirmation. It is found that 39.4%; n=69 is K. pneumoniae with 26%; n=18 of isolates are Multidrug resistant. Also, this study ascertains that multidrug-resistant (MDR) K.pneumoniae has the highest resistance against different types of antibiotics, including: AMP: 100%, AMC73: 36%, PIP: 81.16%, ATM: 72.46%, FEP: 71.01%, CAZ: 62.32%, CRO: 56.5%, IPM: 27.53%, MEM: 26.19%, AK: 47.82%, TOB: 43.47%, GM: 36.23%, LEV: 31.82%, OFX: 28.98%, CIP: 24.63% and SXT: 65.22%. Referring to phenotypic detection of MBL, 11 from 18 of MDR K. pneumoniae (61.11%) indicate a positive result. Also, during the molecular detection of Metallo B-lactamase genes, it was found that (81.81%; n=9) isolates were positive for blaNDM-1, blaNDM-2, and blaVIM gene, while no isolates showed positive results to blaIMP. Objective: Molecular detection of metallo-beta-lactamase genes in the bacteria Klebsiella pneumoniae.

MOLECULAR CHARACTERIZATION OF MBL IN UROPATHOGENIC E. COLI ISOLATED FROM PATIENTS OF TERTIARY CARE HOSPITAL

Background: Antibiotic resistance is on an increasing trend, particularly in gram-negative bacteria. The production of metallo β-lactamase (MBL) puts the health sector at great risk as it further limits the treatment option for MDR strain. MBL is mostly reported in Enterobacteriaceae, which poses resistance to almost all β-lactam antibiotics except monobactam. The current study aims to determine the prevalence, antibacterial sensitivity pattern, and molecular characterization of MBL in Uropathogenic E. coli from clinical samples of hospitalized patients in Khyber Pakhtunkhwa. Methods: From tertiary care hospitals in Peshawar, 250 Urine samples were collected from indoor hospitalized patients. Gold standard microbiological methods were used to identify UPEC from these clinical samples.For that,urine samples was inoculated onto Cysteine Lactose Electrolyte. Deficient (CLED) agar plate, and MacConkey Agar.Positive growth of E.coli identified through Gram staining, colony morphology, Biochemical Tests and E.coli 16srRNA gene amplification .Antibiotic sensitivity was determined by the disc diffusion method on Muller Hinton agar. For the detection of MBL production double disc synergy, and a combination disc test of the antibiotics were used. Furthermore, multiplex PCR was used for the molecular characterization of the MBL (blaIMP, blaVIM, and blaNDM) genes. Results: Of the 250 samples, only 110 samples were confirmed as Uropathogenic E. coli based on colonial morphology, biochemical testing, and molecular level by targeting the 16SrDNA. Female was found more susceptible to UTI compared to male. High prevalence was found in the age group 45-65 years. UPEC was found highly resistance to Ciprofloxacin (90%), followed by Cefotaxime and Ceftriaxone (86%), Ceftazidime and Augmentin (81%), Tazobactam (61%). while the lowest resistance was reported against Meropenum (20%) Imipenem (18%) and Amikacin (37%). PCR-based confirmed prevalence of MBL encoding genes was blaNDM (42%), blaVIM (32%), and blaIMP (26%). Conclusion: The study proposed a higher prevalence of urinary tract infections (UTIs) in females aged group 54-65 years compared to males. An analysis of antibiotic sensitivity revealed Imipenem and Meropenem to be the most effective antimicrobial agents, while Ciprofloxacin, Cefotaxime and Amoxicillin were found to be the less effective. UPEC were found highly resistance to Ciprofloxacin 91%, and ceftazidime 86%, while comparatively less resistance to meropenem, and imipenem,20% and 18% respectively. Genotype BlaNDM of the MBL is highly prevalent (42%) among UPEC.Furthermore, the presence of MBL genes was detected in over 19% of UPEC, and in different combinations.The upraise of the MBLs resistance in uropathogenic E. coli is an alarming sign for clinicians to decide on treatment options for complicated UTIs.

Prevalence of some metallo β-lactamase enzymes genes in P. aeruginosa isolated from different clinical sample in Baghdad, Iraq

The increasing challenge of carbapenem antibiotics resistance caused by Pseudomonas aeruginosa is one of the global healthcare problems. P. aeruginosa is a significant opportunistic infection. For epidemiological reasons, identifying resistance genes is essential, and it is also desirable to quickly identify the techniques for producing carbapenemase enzymes. So, the study aims to determine the prevalence of beta-lactamase encoding genes (blaIMP, blaVIM, bla GIM, and bla SPM) phenotypically and genotypically in P. aeruginosa isolates to address the epidemiological spread of these genes in Baghdad city. The study identified P. aeruginosa isolates from various clinical sources by chemical characterization and VITEK 2 system, the antibiogram test, phenotypic screening carbapenem resistance by Combined disk synergy test (CDST), and conventional PCR were used to detect presence of VIM, IMP, SPM, and GIM genes. Bacterial susceptibility testing revealed (40 %) of isolates were resistant to Imipenem and (85 %) of them positive to CDST. Genotypic screening on phenotypic Metallo-β-lactamase isolates showed that (100 %) isolates contained blaVIM genes, blaGIM genes (88 %), whereas (65 %) isolates carried blaSPM genes. In this investigation, there was no evidence of blaIMP among carbapenem-resistant isolates. The study appeared high prevalence of multi-drug resistance P. aeruginosa isolates that produce carbapenemase enzymes and having β-lactamase genes in local hospitalized patients compared to global ratio. Expanding the sample size and types of enzymes screening in MDR P. aeruginosa should be the main focus in the future research.

Genomically-supported Redefinition of an Outbreak in a Pediatric Unit Caused by blaVIM -harboring Klebsiella michiganensis.

Klebsiella michiganensis , a member of the Klebsiella oxytoca complex, is an emerging nosocomial pathogen known to frequently carry plasmids with antibiotic-resistance genes, including carbapenemases. Using genomics, this study redefined an outbreak alert of K. michiganensis carrying a blaVIM carbapenemase in a pediatric ward in a Spanish hospital. A total of 31 isolates of Verona integron-encoded metallo-β-lactamase (VIM)-carbapenemase K. oxytoca from suspected outbreak cases and unrelated controls from 2015 to 2022 were analyzed. Whole-genome sequencing (both short and long reads) was applied to determine phylogenetic relationships based on single-nucleotide polymorphisms (SNPs) and identify plasmids and antimicrobial resistance genes. The sequences from 12 isolates identified in 2021 showed pairwise SNP distances ranging from 0 to 16 SNPs, confirming the outbreak. Examination of isolates before and after the study period revealed 7 additional cases, 2 in 2020 and 5 in 2022. The outbreak comprised 18 isolates from 17 patients in 3 different pediatric wards, together with 1 environmental sample. In all outbreak isolates, the blaVIM-1 gene was located within a gene cassette carried by a class 1 integron on an IncFIB(pQil) plasmid. A genomic network based on SNPs revealed 5 unsampled intermediate nodes, suggesting additional subclones that may have involved healthcare staff, patient relatives or environmental reservoirs. Blood and rectal isolates obtained from the same patient were positioned on separate branches of the network, making a direct evolutionary pathway between them unlikely. Our study redefined the full extent of this K. michiganensis -VIM outbreak and highlights the critical importance of genomic analysis in accurately understanding outbreaks in healthcare settings.

Quorum Sensing and Genetic Lineages in Carbapenem-Resistant <i>Pseudomonas aeruginosa</i>

Background: Carbapenem-resistant Pseudomonas aeruginosa (CRPA) has become a major global concern. Quorum sensing (QS) regulates the expression of biofilm formation genes and virulence factors. Enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) is widely used in epidemiological molecular studies. Objectives: The purpose of the present study was to determine the QS characteristics and genetic relatedness of CRPA. Methods: A total of 57 non-duplicative CRPA isolates were collected. A microtiter plate assay was used to assess biofilm formation. After DNA extraction, PCR was performed to detect resistance elements and QS-encoded genes. Enterobacterial repetitive intergenic ERIC-PCR was conducted using specific primers. Results: The biofilm formation assay revealed that 10.5%, 19.3%, and 70.2% of isolates formed weak, moderate, and strong biofilms, respectively. Of the isolates, 75.4%, 64.9%, 12.3%, and 8.7% carried the blaIMP, blaVIM, blaNDM, and blaKPC genes, respectively. Additionally, 73.7%, 7.0%, and 1.7% of CRPA isolates carried the blaOXA-48-like, blaOXA-23-like, and blaOXA-20/40-like genes, respectively. The prevalence of the lasR, lasI, rhlI, rhlR, aprR, aprA, and rhlAB genes were 100%, 96.5%, 92.9%, 89.5%, 84.2%, 73.6%, and 63.2%, respectively. Enterobacterial repetitive intergenic ERIC-PCR revealed eight distinct clusters (A, B, C, D, E, F, G, and H) using a similarity cut-off of ≥ 60%. Conclusions: The findings indicate a high prevalence of strong biofilm formation and quorum-sensing genes among CRPA isolates. The study highlights the importance of biofilm production and genetic diversity in CRPA isolates, underscoring the challenges in infection control and treatment strategies.

The Molecular detection of blaVIM in carbapenem-resistant Acinetobacter baumannii-calcoaceticus complex infections in Indonesia

Background: Bacteria of carbapenem-resistant Acinetobacter baumannii-calcoaceticus complex (CRABC) is a significant pathogen causes illnesses acquired via healthcare around the world, however little is known regarding the molecular resistant gene of A. baumannii-calcoaceticus complex bacteria resistant to carbapenem in Indonesian healthcare facilities. This study is aimed to identify the carbapenemase resistant gene of CRABC in two referral care hospitals in the islands of North Sulawesiand East Java, Indonesia. Methods: The Vitek2® system was used to identify the CRABC pathogen that were obtained from standard medical cultures, which included blood, lower respiratory tract, wound and urine samples from patients treated in the medical wards, intensive care unit (ICU), surgical wards, neonatal intensive care unit (NICU), and cardiovascular intensive care unit (CVICU) in two referral care hospitals in Manado, North Sulawesi, and Malang, East Java. Using PCR, we identified the presence of the carbapenemase resistance genes blaGES, blaVIM, blaIMP1, and blaOXA-48 carbapenemase resistance genes. Results: Seventy threeA. baumannii-calcoaceticus complex carbapenem resistance bacteria were collected. The specimens from the lower respiratory tract specimens, the isolates of the carbapenem-resistant A. baumannii-calcoaceticus complex were frequently found.In this study, we detected the blaVIM carbapenemase resistance genes from blood and sputum samples in tertiary care hospital in Malang. Conclusions: We identified the blaVIM carbapenemase resistance genes were found in the referral care hospitals in Malang, East Java, Indonesia. Awareness of the national prevention and surveillance system is isolated by the prevention and reduction of CRABC transmission.

Detection of antibiogram pattern and prevalence of blaNDM and blaVIM genes among carbapenem resistant Eschericia coli isolates in a tertiary care hospital

Background: Clinical significance of Eschericia coli is due to its capacity to develop resistance against many antibiotics specially carbapenem by producing carbapenemase which limits treatment options. blaNDM and blaVIM genes are responsible for this multi-drug-resistance. This is an alarming condition for society as well as clinician because they are very difficult to treat due to limited options of antibiotics and its high virulence. Aims and Objectives: The study was conducted to detect antibiogram pattern and prevalence of blaNDM and blaVIM genes among carbapenem resistant E. coli isolates in a tertiary care hospital. Our study will also help to identify molecular basis of Carbapenem resistance including treatment guideline. Materials and Methods: Cross-sectional study was performed during January’2021 to June’2022 time period. Identification of E. coli was followed by standard protocol and antibiotic sensitivity was done by Kirby-Bauer disk-diffusion method following CLSI guideline, using VITEK-2D-SYSTEM. Detection of genes was done by polymerase chain reaction (PCR). Results: Out of 1867 samples, 161 isolates were identified as E. coli, among which 90 were resistant to carbapenem. Among those, 50 (31%) were resistant to all three carbapenems. Among those, 100% were resistant to ampicillin, amoxycillin-clavulanic acid, cefuroxime, cefuroxime-axetil, cefoperazone-sulbactam, ceftriaxone, cefepime, piperacillin-tazobactam, nalidixic acid, ciprofloxacin; 52% were resistant to amikacin; 54% were resistant to gentamicin; 86% were resistant to cotrimoxazole; 22% were resistant to nitrofurantoin and 96% were sensitive to tigecycline; 100% were intermediate sensitive to colistin. blaNDM gene is detected in 17 (34%) isolates and blaVIM gene is detected in 2 (4%) isolates by PCR. Conclusion: E. coli leads to common clinical presentations such as urinary tract infection, blood stream infection, pneumonia, and wound infection. High burden of carbapenem resistant E. coli was associated with urinary indoor samples. Rising incidence of multi-drug-resistance occurs due to lack of infection control measures, irrational use of antibiotics. Continuous surveillance will prevent development of multi-drug-resistance and help to improve treatment guideline.

Detection of antibiogram pattern and prevalence of blaNDM and blaVIM genes among carbapenem resistant Eschericia coli isolates in a tertiary care hospital

Background: Clinical significance of Eschericia coli is due to its capacity to develop resistance against many antibiotics specially carbapenem by producing carbapenemase which limits treatment options. blaNDM and blaVIM genes are responsible for this multi-drug-resistance. This is an alarming condition for society as well as clinician because they are very difficult to treat due to limited options of antibiotics and its high virulence. Aims and Objectives: The study was conducted to detect antibiogram pattern and prevalence of blaNDM and blaVIM genes among carbapenem resistant E. coli isolates in a tertiary care hospital. Our study will also help to identify molecular basis of Carbapenem resistance including treatment guideline. Materials and Methods: Cross-sectional study was performed during January’2021 to June’2022 time period. Identification of E. coli was followed by standard protocol and antibiotic sensitivity was done by Kirby-Bauer disk-diffusion method following CLSI guideline, using VITEK-2D-SYSTEM. Detection of genes was done by polymerase chain reaction (PCR). Results: Out of 1867 samples, 161 isolates were identified as E. coli, among which 90 were resistant to carbapenem. Among those, 50 (31%) were resistant to all three carbapenems. Among those, 100% were resistant to ampicillin, amoxycillin-clavulanic acid, cefuroxime, cefuroxime-axetil, cefoperazone-sulbactam, ceftriaxone, cefepime, piperacillin-tazobactam, nalidixic acid, ciprofloxacin; 52% were resistant to amikacin; 54% were resistant to gentamicin; 86% were resistant to cotrimoxazole; 22% were resistant to nitrofurantoin and 96% were sensitive to tigecycline; 100% were intermediate sensitive to colistin. blaNDM gene is detected in 17 (34%) isolates and blaVIM gene is detected in 2 (4%) isolates by PCR. Conclusion: E. coli leads to common clinical presentations such as urinary tract infection, blood stream infection, pneumonia, and wound infection. High burden of carbapenem resistant E. coli was associated with urinary indoor samples. Rising incidence of multi-drug-resistance occurs due to lack of infection control measures, irrational use of antibiotics. Continuous surveillance will prevent development of multi-drug-resistance and help to improve treatment guideline.

Antimicrobial resistance profiling of bacteria isolated from wastewater and samples of pharmaceutical industries in South India

The study was aimed to determine the phenotypic and genotypic antimicrobial resistance in the isolated bacteria from the influent (25), effluent (15), surface and ground water samples (15) surrounding the pharmaceutical industries located in south India. From 55 samples, 48 isolates of 10 different bacteria were obtained. The identified bacterial isolates were viz. Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterobacter aerogenes, Corynebacterium sp., Acinetobacter sp., Aeromonas punctata, Ralstonia picketti, Staphylococcus aureus, Stenotrophomonas maltophillia, and Citrobacter freundii. The phenotypic profile of resistance through antibiotic susceptibility test was carried out against sixteen different antibiotics. Standard PCR technique was used for the detection of 12 resistance genes encoding carbapenems, quinoline, aminoglycoside, β-lactam belonging blaOXA-58,blaOXA-22,qnrA, qnrB, aac(6)-Ib-cr, aac (3)-XI, mec A, qepA, aadB, blaVIM, blaOXA-48 and blaNDM. Pseudomonas aeruginosa (1: TN/I/2020) showed presence of 3 resistance genes. qnrB (489 bp) gene was present in maximum of 7 isolates while blaVIM (196 bp) gene was present in 6 isolates. The resistance genes blaNDM (621 bp) was present in three different isolates; aac (X):6)-lb-cr (482 bp), qepA (495 bp), aadB (500 bp), blaOXA-58 (843 bp) resistant genes were present in two different isolates each among the bacterial isolates obtained in this study. In phenotypic resistance profiling by AST method, out of 16 antibiotics tested, 14 showed resistance. Similarly, in genotypic resistance profiling, among 12 resistance genes tested, a maximum of three resistance genes were noticed in Pseudomonas aeruginosa. There were positive and negative correlations observed between phenotypic and genotypic resistance among different antibiotics and their resistance genes indicating the variations in the resistance gene expression.

Presence of extended-spectrum beta-lactamase-producing Escherichia coli and carbapenem resistance in ready-to-eat stuffed mussels in Istanbul

AbstractFoodborne pathogens' transmission is essential in the spread of antibiotic resistance, and extended-spectrum beta-lactamase-producing Escherichia coli especially threatens public health. E. coli plays an essential role in the resistance to commonly used beta-lactam group antibiotics. Ready-to-eat (RTE) stuffed mussels are among many restaurants and street vendors, presenting potential health risks of food hygiene origin. 200 RTE stuffed mussels were collected from the Asian and European sides of Istanbul and analysed for the presence of E. coli. As a result of PCR analysis, E. coli was detected in 7 (3.5%) samples. An antibiotic susceptibility test was performed using the disc diffusion method to determine ESBL and carbapenem resistance. All isolates were resistant to ampicillin. The double-disk synergy test was performed as an ESBL phenotypic confirmation test, and no phenotypically ESBL-producing E. coli were detected. The blaTEM gene was detected in one isolate (14.2%) by mPCR, but blaCTX-M, blaSHV, and blaOXA genes were not observed. Meropenem and imipenem were used with the disk diffusion method for carbapenem resistance study, and no resistant isolate was found. Carbapenem resistance genes were investigated by monoplex PCR, and blaNDM-1, blaOXA-48, blaVIM, and blaIMP resistance genes were not detected. This is the first report on ESBL-producing E. coli in RTE stuffed mussels in Türkiye, which draws attention to a public health risk.

Prevalence and Molecular Characterization of Carbapenem and Colistin-Resistance Genes in Clinical Strains of Pseudomonas aeruginosa Isolated from Diabetic Patients

Abstract Introduction Pseudomonas aeruginosa is one of the important opportunistic nosocomial pathogens that are increasingly becoming resistant to many drugs including the drugs of last resort, carbapenems, and colistin. Methods The study involved the examination of clinical isolates of P. aeruginosa recovered from diabetic patients for the presence of acquired genes implied in carbapenem and colistin resistance. Results A total of 100 clinical isolates from diabetic patients' wound and sputum were investigated for their susceptibility to imipenem and colistin followed by phenotypic assay and resistance gene screening. The study revealed 27 sputum isolates being resistant to imipenem further confirmed by a modified Hodge test and 28 wound isolates were resistant to colistin. The resistant isolates of wound and sputum were found to possess blaOXA-48 , blaVIM , mcr-1, and mcr-5 genes. The strains had a coexistence of carbapenem- and colistin-resistant genes. Conclusion Considering the fact that infection by such resistant isolates in diabetics could be frightening, clinical laboratories must employ rapid diagnostic methods to detect resistance which can aid in the treatment decision-making.

Carbapenems and colistin resistance genes isolated in Musca domestica from a garbage dump near a hospital in Lima.

Motivation for the study. The presence of antibiotic resistance genes in bacteria isolated from common flies is a potential public health hazard because it facilitates the presence and spread of antibiotic resistance genes in the environment. Main findings. Thirty-eight bacterial strains identified in 14 species were isolated from within the fly bodies, of which 31 strains showed resistance to carbapenems and 26 strains showed resistance to colistin. Seven bacterial strains showed carbapenem resistance genes and one Escherichia coli strain had resistance to KPC, OXA-48 and mcr-1. Implications. This is the first report of antibiotic resistance genes in bacteria carried by common flies in Peru. The objective was to determine the presence of carbapenem resistance genes and plasmid resistance to colistin (mcr-1) in bacteria isolated from Musca domestica in a garbage dump near a hospital in Lima, Peru. Bacteria with phenotypic resistance to carbapenemics were isolated on CHROMagar mSuperCARBATM medium and colistin resistance profiling was performed using the colistin disk elution method. Detection of blaKPC, blaNDM, blaIMP, blaOXA-48, blaVIM and mcr-1 genes was performed by conventional PCR. The antimicrobial susceptibility profile was determined using the automated MicroScan system. We found that 31/38 strains had phenotypic resistance to carbapenemics and 26/38 strains had phenotypic resistance to colistin with a minimum inhibitory concentration ≥ 4 µg/ml. Finally, we identified seven bacterial strains with carbapenem resistance genes (OXA-48 and KPC) and one bacterial strain with plasmid resistance to colistin (mcr-1). One Escherichia coli strain had three resistance genes: KPC, OXA-48 and mcr-1.

Characterisation of carbapenemase-producing Gram-negative bacilli among clinical isolates in a tertiary care centre in Kerala, South India

Background: Infections caused by carbapenem-resistant Gram-negative bacteria are a cause for concern due to the limited choice of antibiotics available for their treatment. Aims, Settings and Design: This study screened multidrug-resistant (MDR) Gram-negative bacilli isolated from clinical samples over a period of one year, for carbapenem resistance and characterised them using phenotypic methods such as combined disc diffusion test (CDDT), modified Hodge test (MHT), E-test for metallo-beta-lactamase (MBL) and molecular method, PCR. Materials and Methods: Two hundred and ten MDR Gram-negative bacilli were screened for carbapenem resistance using Imipenem and Meropenem disc diffusion. These were further checked for carbapenemase production by CDDT, MHT and E-test for MBL. Those positive by E-test were subjected to PCR. Uniplex PCR for New Delhi metallo-beta-lactamase-1 was used for Escherichia coli and Klebsiella pneumoniae , and multiplex PCR for Imipenemase and Verona imipenemase was used for Pseudomonas aeruginosa and Acinetobacter baumannii isolates. Results: Twenty-three (11%) isolates were found to be carbapenem-resistant and included E. coli (six) K. pneumoniae (three), P. aeruginosa (five) and A. baumannii (nine). Seventeen (74%) isolates were positive by phenotypic methods and were subjected to PCR. Out of eight Enterobacteriaceae isolates subjected to PCR, all were positive for bla NDM gene. All were negative for bla KPC gene. All five A. baumannii isolates subjected to PCR were found to contain bla VIM gene. Two out of four P. aeruginosa isolates were positive for bla IMP , one was positive for bla VIM gene. One P. aeruginosa isolate was positive for both bla IMP and bla VIM gene. Conclusions: In view of the increasing resistance of Gram-negative bacilli to carbapenems, rational use of antibiotics needs to be emphasised.

Resistance and Co-Resistance of Metallo-Beta-Lactamase Genes in Diarrheal and Urinary-Tract Pathogens in Bangladesh.

Carbapenems are the antibiotics of choice for treating multidrug-resistant bacterial infections. Metallo-β-lactamases (MBLs) are carbapenemases capable of hydrolyzing nearly all therapeutically available beta-lactam antibiotics. Consequently, this research assessed the distribution of two MBL genes and three β-lactamases and their associated phenotypic resistance in diarrheal and urinary-tract infections (UTIs) to guide future policies. Samples were collected through a cross-sectional study, and β-lactamase genes were detected via PCR. A total of 228 diarrheal bacteria were isolated from 240 samples. The most predominant pathogens were Escherichia coli (32%) and Klebsiella spp. (7%). Phenotypic resistance to amoxicillin-clavulanic acid, aztreonam, cefuroxime, cefixime, cefepime, imipenem, meropenem, gentamicin, netilmicin, and amikacin was 50.4%, 65.6%, 66.8%, 80.5%, 54.4%, 41.6%, 25.7%, 41.2%, 37.2%, and 42.9%, respectively. A total of 142 UTI pathogens were identified from 150 urine samples. Klebsiella spp. (39%) and Escherichia coli (24%) were the major pathogens isolated. Phenotypic resistance to amoxicillin-clavulanic acid, aztreonam, cefuroxime, cefixime, cefepime, imipenem, meropenem, gentamicin, netilmicin, and amikacin was 93.7%, 75.0%, 91.5%, 93.7%, 88.0%, 72.5%, 13.6%, 44.4%, 71.1%, and 43%, respectively. Twenty-four diarrheal isolates carried blaNDM-1 or blaVIM genes. The overall MBL gene prevalence was 10.5%. Thirty-six UTI pathogens carried either blaNDM-1 or blaVIM genes (25.4%). Seven isolates carried both blaNDM-1 and blaVIM genes. MBL genes were strongly associated with phenotypic carbapenem and other β-lactam antibiotic resistance. blaOXA imparted significantly higher phenotypic resistance to β-lactam antibiotics. Active surveillance and stewardship programs are urgently needed to reduce carbapenem resistance in Bangladesh.

Prevalence and characterization of quinolone resistance and integrons in clinical Gram-negative isolates from Gaza strip, Palestine.

Gram-negative bacteria with quinolone resistance and extended-spectrum beta-lactamases (ESBLs) present significant treatment challenges. This study evaluated the prevalence and characteristics of quinolone resistance in Gram-negative strains, investigating the relationship between plasmid-mediated quinolone resistance (PMQR), ESBLs, and integrons. We collected 146 Gram-negative isolates from patients in three Palestinian hospitals. For quinolone resistance isolates, the presence and characterization of PMQR, β-lactamase genes and integrons were studied by PCR and sequencing. Out of 146 clinical isolates, 64 (43.8%) were resistant to quinolones, with 62 (97%) being multidrug-resistant (MDR) and 33 (51.5%) ESBL-producers. PMQR-encoding genes were present in 45 (70.3%) isolates, including aac(6')-Ib-cr (26.6%), qnrA (18.8%), qnrS1 (20.8%), and qnrB (6.4%). BlaCTX-M genes were detected in 50% (32/64) of isolates, with blaCTX-M-15 being the most common. BlaTEM-1, blaSHV-1 and blaVIM genes were found in 13, 6, and 4 isolates, respectively. Class I integrons were found in 31/64 (48%) of isolates, with 14 containing gene cassettes conferring resistance to trimethoprim (dhfr17, dfrA12, dfrA1) and aminoglycosides resistance genes (aadA1, aadA2, aadA5, and aadA6). This study found a high rate of quinolone resistance, ESBL and integrons in clinical Gram-negative isolates from our hospitals. Urgent measures are crucial, including implementing an antimicrobial resistance surveillance system, to control and continuously monitor the development of antimicrobial resistance.

Occurrence of carbapenem-resistant hypervirulent Klebsiella pneumoniae in oysters in Egypt: a significant public health issue

BackgroundThe global dissemination of critical-priority carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp) via food sources represents a significant public health concern. Epidemiological data on CR-hvKp in oysters in Egypt is limited. This study aimed to investigate the potential role of oysters sold in Egypt as a source for carbapenem-resistant K. pneumoniae (CRKP), hypervirulent K. pneumoniae (hvKp), and CR-hvKp and assess associated zoonotic risks.MethodsA sample of 330 fresh oysters was randomly purchased from various retail fish markets in Egypt and divided into 33 pools. Bacteriological examination and the identification of Klebsiella pneumoniae were performed. Carbapenem resistance in K. pneumoniae isolates was determined by phenotypic and molecular methods. Additionally, the presence of hypervirulent K. pneumoniae was identified based on virulence gene markers (peg-344, rmpA, rmpA2, iucA, and iroB), followed by a string test. The clustering of CR-hvKp strains was carried out using R with the pheatmap package.ResultsThe overall prevalence of K. pneumoniae was 48.5% (16 out of 33), with 13 isolates displaying carbapenem resistance, one intermediate resistance, and two sensitive. Both carbapenem-resistant K. pneumoniae and carbapenem-intermediate-resistant K. pneumoniae strains exhibited carbapenemase production, predominantly linked to the blaVIM gene (68.8%). HvKp strains were identified at a rate of 62.5% (10/16); notably, peg-344 was the most prevalent gene. Significantly, 10 of the 13 CRKP isolates possessed hypervirulence genes, contributing to the emergence of CR-hvKp. Moreover, cluster analysis revealed the clustering of two CR-hvKp isolates from the same retail fish market.ConclusionThis study provides the first insight into the emergence of CR-hvKp among oysters in Egypt. It underscores the potential role of oysters as a source for disseminating CR-hvKp within aquatic ecosystems, presenting a possible threat to public health.

Phenotypic and molecular characterization of extended spectrum- and metallo- beta lactamase producing Pseudomonas aeruginosa clinical isolates from Egypt

BackgroundAntimicrobial resistance among Pseudomonas aeruginosa (P. aeruginosa), a leading cause of nosocomial infections worldwide, is escalating. This study investigated the prevalence of extended-spectrum β-lactamases (ESBLs) and metallo-β-lactamases (MBLs) among 104 P. aeruginosa clinical isolates from Alexandria Main University Hospital, Alexandria, Egypt.MethodsAntimicrobial susceptibility testing was performed using agar dilution technique, or broth microdilution method in case of colistin. ESBL and MBL prevalence was assessed phenotypically and genotypically using polymerase chain reaction (PCR). The role of plasmids in mediating resistance to extended-spectrum β-lactams was studied via transformation technique using plasmids isolated from ceftazidime-resistant isolates.ResultsAntimicrobial susceptibility testing revealed alarming resistance rates to carbapenems, cephalosporins, and fluoroquinolones. Using PCR as the gold standard, phenotypic methods underestimated ESBL production while overestimating MBL production. Eighty-five isolates (81.7%) possessed only ESBL encoding genes, among which 69 isolates harbored a single ESBL gene [blaOXA-10 (n = 67) and blaPER (n = 2)]. Four ESBL-genotype combinations were detected: blaPER + blaOXA-10 (n = 8), blaVEB-1 + blaOXA-10 (n = 6), blaPSE + blaOXA-10 (n = 1), and blaPER + blaVEB-1 + blaOXA-10 (n = 1). Three isolates (2.9%) possessed only the MBL encoding gene blaVIM. Three ESBL + MBL- genotype combinations: blaOXA-10 + blaAIM, blaOXA-10 + blaVIM, and blaPER + blaOXA-10 + blaAIM were detected in 2, 1 and 1 isolate(s), respectively. Five plasmid preparations harboring blaVEB-1 and blaOXA-10 were successfully transformed into chemically competent Escherichia coli DH5α with transformation efficiencies ranging between 6.8 × 10 3 and 3.7 × 10 4 CFU/μg DNA plasmid. Selected tested transformants were ceftazidime-resistant and harbored plasmids carrying blaOXA-10.ConclusionsThe study highlights the importance of the expeditious characterization of ESBLs and MBLs using genotypic methods among P. aeruginosa clinical isolates to hinder the development and dissemination of multidrug resistant strains.

Carbapenem-resistant Enterobacterales in wastewater resources and healthy carriers: A survey in Iran

ABSTRACT The carbapenem-resistant Enterobacterales (CRE) pose a pressing public health concern. Here, we investigated the frequency of CRE bacteria, carbapenemase-encoding genes, and the molecular epidemiology of carbapenemase-resistant Escherichia coli in wastewater resources and healthy carriers in Iran. Out of 617 Enterobacterales bacteria, 24% were carbapenem-resistant. The prevalence of CRE bacteria in livestock and poultry wastewater at 34% and hospital wastewater at 33% was significantly higher (P ≤ 0.05) than those in healthy carriers and municipal wastewater at 22 and 17%, respectively. The overall colonization rate of CRE in healthy individuals was 22%. Regarding individual Enterobacterales species, the following percentages of isolates were found to be CRE: E. coli (18%), Citrobacter spp. (24%), Klebsiella pneumoniae (28%), Proteus spp. (40%), Enterobacter spp. (25%), Yersinia spp. (17%), Hafnia spp. (31%), Providencia spp. (21%), and Serratia spp. (36%). The blaOXA-48 gene was detected in 97% of CRE isolates, while the blaNDM and blaVIM genes were detected in 24 and 3% of isolates, respectively. The B2 phylogroup was the most prominent group identified in carbapenem-resistant E. coli isolates, accounting for 80% of isolates. High prevalence of CRE with transmissible carbapenemase genes among healthy people and wastewater in Iran underscores the need for assertive measures to prevent further dissemination.

Neonatal sepsis due to NDM-1 and VIM-2 co-producing Pseudomonas aeruginosa in Morocco.

Carbapenem-resistant Pseudomonas aeruginosa are being increasingly described worldwide. Here, we investigated the molecular mechanisms underlying carbapenem resistance in an extremely drug-resistant P. aeruginosa isolate from a neonatal intensive care unit in Morocco. P. aeruginosa strain O82J1 was identified using MALDI-TOF-MS. Carba NP, immunochromatographic assay NG Carba5 and antimicrobial susceptibility testing using disc diffusion and microbroth were performed. Whole-genome sequencing using the Illumina and MinION technologies and different software packages available at the Center of Genomic Epidemiology were used to predict the resistome, sequence type and plasmid types. P. aeruginosa O82J1 co-expressed two metallo-β-lactamases, blaNDM-1 and blaVIM-2, and was susceptible to colistin and apramycin only. It belonged to ST773 that is frequently reported worldwide as a high-risk P. aeruginosa clone. The blaVIM-2 gene was integron-borne on a IncP-2 465-kb plasmid, whereas the blaNDM-1 gene was chromosomally encoded and embedded in an integrative conjugative element, probably at the origin of its acquisition. A total of 23 antimicrobial resistance genes were detected including a blaPER-1 ESBL gene, and an 16S-rRNA methyltransferase gene rmtB. The isolation of XDR P. aeruginosa isolates expressing several carbapenemases in a neonatal intensive care unit is of great concern due to the reduced treatment options, relying only on colistin, but not recommended in neonates, and apramycin, not yet approved for human therapy. Concerns were further elevated due to the resistance to cefiderocol and ATM/AVI, two novel and last-resort antibiotics recommended to treat infections caused by Gram-negative bacteria, particularly XDR P. aeruginosa in adults.