Rapid Detection of Beta-Lactamases Genes among Enterobacterales in Urine Samples by Using Real-Time PCR. - Related Documents




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144501.0000Rapid Detection of Beta-Lactamases Genes among Enterobacterales in Urine Samples by Using Real-Time PCR. The objective of this study was to develop and evaluate newly improved, rapid, and reliable strategies based on real-time PCR to detect the most frequent beta-lactamase genes recorded in clinical Enterobacterales strains, particularly in Tunisia (bla(SHV12) , bla(TEM) , bla(CTX-M-15) , bla(CTX-M-9) , bla(CMY-2) , bla(OXA-48) , bla(NDM-1) , and bla(IMP) ) directly from the urine. Following the design of primers for a specific gene pool and their validation, a series of real-time PCR reactions were performed to detect these genes in 78 urine samples showing high antibiotic resistance after culture and susceptibility testing. Assays were applied to DNA extracted from cultured bacteria and collected urine. qPCR results were compared for phenotypic sensitivity. qPCR results were similar regardless of whether cultures or urine were collected, with 100% sensitivity and specificity. Out of 78 multiresistant uropathogenic, strains of Enterobacterales (44 E. coli and 34 K. pneumoniae strains) show the presence of the genes of the bla group. In all, 44% E. coli and 36 of K. pneumoniae clinical strains harbored the bla group genes with 36.4%, 52.3%, 70.5%, 68.2%, 18.2%, and 4.5% of E. coli having bla(SHV-12) , bla(TEM) , bla(CTX-M 15) , bla(CTX-M-9) , bla(CMY-2) , and bla(OXA-48) group genes, respectively, whereas 52.9%, 67.6%, 76.5%, 35.5%, 61.8, 14.7, and 1.28% of K. pneumoniae had bla(SHV-12) , bla(TEM) , bla(CTX-M 15) , bla(CTX-M-9) , bla(CMY-2) , bla(OXA-48) , and bla(NDM-1) group genes, respectively. The time required to have a result was 3 hours by real-time PCR and 2 to 3 days by the conventional method. Resistance genes of Gram-negative bacteria in urine, as well as cultured bacteria, were rapidly detected using qPCR techniques. These techniques will be used as rapid and cost-effective methods in the laboratory. Therefore, this test could be a good candidate to create real-time PCR kits for the detection of resistance genes directly from urine in clinical or epidemiological settings.202235978630
144610.9999One-Day Prevalence of Extended-Spectrum β-Lactamase (ESBL) and Carbapenemase-Producing Bacteria in Fecal Samples from Surgical Patients: A Concerning Trend of Antibiotic Resistance. PURPOSE: Extended-spectrum β-lactamase (ESBL) and carbapenemase producing bacteria are of increasing concern due to their multidrug resistance and infection potential. This study determines the one-day prevalence of faecal carriage of ESBL and carbapenemase producing Gram-negative bacilli. METHODS: Fecal samples were collected from 30 post-surgery patients (hospitalized for at least 48 hours) in each of the four hospitals involved in the study and were analyzed for antibiotic-resistant bacteria. Identification was done using Maldi Tof mass spectrometry, and antibiotic susceptibility was tested using disk diffusion and specialized tests for ESBL (double disk synergy technique) and carbapenem (NG-TEST CARBA 5) resistance detection. PCR was conducted on isolates to detect betalactam resistance genes, carbapenemase genes and quinolone resistance genes. FINDINGS: Out of the 120 patients enrolled, 38.33% (n = 46) and 49.16.33% (n = 59) were found to carry ESBL- and carbapenemase-producing bacteria, respectively, in their fecal samples. Among the isolates, 51.08% (n = 47) exhibited ESBL production, with Escherichia coli (44.56%) being the most common species. The identification of bacteria with resistance to carbapenems showed a predominance of the species Escherichia coli (44.45%) followed by the species Klebsiella pneumoniae (16.06%) and Acinetobacter baumanii (13.58%). The study of the association of variables shows a high degree of association (p < 0.05) for the factors independent walking and use of a wheelchair with ESBL production. The most frequently detected genes among ESBL producing bacteria were bla(CTXM-1) (91.49%), qnrB (70.21%) and qnrs (63.82%). bla(NDM) (54.68%) was the most detected carbapenemase genes among carbapenemase producing isolates. CONCLUSION: This study demonstrates, for the first time, a significant prevalence of ESBL and carbapenemase producing gram-negative bacteria among surgical patients in Benin, with multiple resistance genes detected. Findings should be interpreted in light of the cross-sectional design and >48-hour hospitalization criterion.202540635768
144720.9999Molecular detection of β-lactamase and integron genes in clinical strains of Klebsiella pneumoniae by multiplex polymerase chain reaction. INTRODUCTION: Infections caused by β-lactamase-producing gram-negative bacteria, such as Klebsiella pneumoniae, are increasing globally with high morbidity and mortality. The aim of the current study was to determine antimicrobial susceptibility patterns and the prevalence of antibiotic resistance genes (β-lactamase and integron genes) using multiplex PCR. METHODS: One-hundred K. pneumoniae isolates were collected from different clinical samples. Antibiotic susceptibility testing was performed with thirteen different antibiotics. Multiplex-PCR was used to detect β-lactamase (bla TEM, bla CTX-M, bla SHV , bla VEB, bla PER, bla GES, bla VIM, bla IMP, bla OXA, and bla KPC) and integron genes (int I, int II, and int III). RESULTS: The highest and lowest rate of resistance was exhibited against amikacin (93%) and imipenem (8%), respectively. The frequency of β-lactamase-positive K. pneumoniae was 37%, and the prevalence of the bla TEM, bla CTX-M, bla SHV , bla VEB, bla PER, bla GES, bla VIM, bla IMP, bla OXA, and bla KPC genes was 38%, 24%, 19%, 12%, 6%, 11%, 33%, 0%, 28%, and 23%, respectively. Of the 100 isolates, eight (8%) were positive for class I integrons; however, class II and III integrons were not detected in any of the strains. CONCLUSIONS: These results indicate co-carriage of a number of β-lactamase genes and antibiotic resistance integrons on the same plasmids harboring multi-drug resistance genes. It seems that these properties help to decrease treatment complications due to resistant bacterial infections by rapid detection, infection-control programs and prevention of transmission of drug resistance.201728700049
144430.9999The Prevalence of bla VIM, bla KPC, bla NDM, bla IMP, bla SHV, bla TEM, bla CTX-M, and class I and II integrons Genes in Aeromonas hydrophila Isolated from Clinical Specimens of Qom, Iran. BACKGROUND: Aeromonas hydrophila is an opportunistic gram-negative bacillus that causes diseases such as gastroenteritis, muscle infections, soft tissue, sepsis, and skin diseases in humans. Today, the prevalence of antibiotic resistance in bacteria has led to treatment failure and prolonged treatment. Therefore, the aim of this study was to evaluate the level of antibiotic resistance in isolates carrying bla VIM, bla KPC, bla NDM, bla IMP, bla SHV, bla TEM, bla CTX-M and class I and II integrons in Aeromonas hydrophila. METHODS: In this cross-sectional study, Aeromonas hydrophila were collected from different clinical specimens in Hazrat Masoumeh Hospital, Qom Province, Iran, from 2018 to 2020. The collected isolates were identified by standard biochemical tests. Then, using specific primers bla VIM, bla KPC, bla NDM, bla IMP, bla SHV, bla TEM, bla CTX-M genes, and class I and II integrons were evaluated by PCR method. Then, data were analyzed using SPSS software and chi-squared tests, and the significance level was determined as p ≤ 0.05. RESULTS: During the sample collection period, 100 Aeromonas hydrophila were collected. Based on the results of the antibiotic resistance pattern, the highest and lowest rate of antibiotic resistance to ampicillin (92%) and azithromycin (4%) were determined for both. Among the 100 isolates, 60 isolates produced broad-spectrum beta-lactamase (ESBL) and 50 isolates produced carbapenemase. Among the studied beta-lactamase genes, the highest and lowest frequencies were related to bla CTX-M (58%) and bla TEM (1%), respectively. The frequency of class I and II integron genes was 27% and 15%, respectively. CONCLUSIONS: The results of the study of antibiotic resistance, beta-lactamase, and carbapenemase genes showed high resistance in Aeromonas hydrophila, which raises concerns with regard to controlling infection in medical centers. Also, the study of antibiotic resistance in the presence of beta-lactamase genes showed that there was only a significant relationship between the presence of bla CTX-M gene and resistance to imipenem (p = 0.037).202336649515
211140.9999Antimicrobial Resistance and Resistance Determinant Insights into Multi-Drug Resistant Gram-Negative Bacteria Isolates from Paediatric Patients in China. INTRODUCTION: The emergence of multi-drug-resistant Gram-negative bacteria (GNB) is a concern in China and globally. This study investigated antimicrobial resistance traits and resistance determinant detection in GNB isolates from paediatric patients in China. METHODS: In the present study, a total of 170 isolates of GNB including the most prevalent Escherichia coli, Klebsiella pneumoniae and Acinetobacter baumannii were collected from Shenzhen Children's Hospital, China. ESBLs production was confirmed by using the combination disc diffusion method, and carbapenemase production was confirmed by using a carbapenem inactivation method followed by antimicrobial susceptibility. In addition, β-lactamase-encoding genes and co-existence of plasmid-borne colistin resistance mcr-1 gene were determined by PCR and sequencing. RESULTS: Overall, 170 etiological agents (GNB) were recovered from 158 paediatric patients. The most prevalent species was E. coli 40% (n=68), followed by K. pneumoniae 17.64% (n=30), and Enterobacter cloacae 14.11% (n=24). Of 170 GNB, 71.76% (n=122) were multi-drug-resistant, 12.35% (n=21) extreme-drug resistant, and 7.64% (n=13) single-drug-resistant, while 8.23% (n=14) were sensitive to all of the studied antibiotics. The prevalence of ESBLs and carbapenemase producers were 60% and 17%, respectively. bla (CTX-M) was the most prevalent resistance gene (59.42%), followed by bla (TEM) (41.17%), bla (SHV) (34.270%), bla (KPC) (34.11%), bla (OXA-48) (18.82%) and bla (NDM-1) (17.64%). CONCLUSION: The present study provides insights into the linkage between the resistance patterns of GNB to commonly used antibiotics and their uses in China. The findings are useful for understanding the genetics of resistance traits and difficulty in tackling of GNB in paediatric patients.201931819545
112250.9999Antibiotic resistance profiles of gram-negative bacteria in southern Tunisia: Focus on ESBL, carbapenem and colistin resistance. The main objective of this cross-sectional study was to investigate the prevalence of beta-lactam (cephalosporins or carbapenems) or colistin resistant bacteria. Those were isolated from urine samples in two private polyclinics located in the Sfax region, in southern Tunisia. From September 2021 to August 2022, 116 strains resistant to β-lactams or colistin were isolated, identified by MALDI-TOF, and their antibiotic susceptibility was assessed by disk diffusion method. Resistance genes were detected by real-time PCR, standard PCR, and sequencing. The results revealed that the 116 strains consisted predominantly of Enterobacteriaceae (92.2 %) and non-fermenting bacteria (7.8 %). Among these strains, 21 (18.1 %) were resistant to carbapenems, three (2.7 %) to colistin, including two strains of Klebsiella pneumoniae (1.7 %) exhibiting resistance to both carbapenems and colistin. In Enterobacteriaceae, bla(CTX-A), bla(SHV), and bla(TEM) were found in 79.5 %, 46.7 %, and 40.2 % of strains, respectively. For these strains, the minimum inhibitory concentrations (MICs) of imipenem and ertapenem ranged from >32 to 6 μg/mL and > 32 to 2 μg/mL, respectively, with bla(OXA-48) and bla(NDM) detected in 21.7 % and 19.6 % of isolates, respectively. Seven A. baumannii isolates resistant to imipenem and meropenem (MICs >32 μg/mL and 8 μg/mL, respectively) carried bla(OXA-23) (n = 5) and bla(OXA-24) (n = 2). In addition, mutations in the mgrB gene conferring colistin resistance were identified in two isolates. Two K. pneumoniae were colistin-resistant and carried the bla(OXA-48) gene. These results highlight the urgency of developing new strategies for the identification and surveillance of pathogenic strains in humans to effectively combat this growing public health threat in Tunisia.202540553790
112160.9999Occurrence of the genes encoding carbapenemases, ESBLs and class 1 integron-integrase among fermenting and non-fermenting bacteria from retail goat meat. The present study was planned to detect the genes encoding carbapenemases, ESBLs and class 1 integron-integrase among bacteria obtained from retail goat meat. Fermenting and non-fermenting bacterial isolates (n = 57), recovered from 61 goat meat samples, were identified by 16S rRNA gene sequencing. Antimicrobial susceptibility of isolates was tested by the broth dilution method using ceftazidime, cefotaxime, meropenem and imipenem. Plasmids were isolated and tested for their physical characters. Plasmids were subjected to screening of carbapenemase, ESBL and intI1 gene. Conjugation assay was performed using bla(NDM) -positive isolates as the donor, and Escherichia coli HB101 as the recipient. Isolates showed the high rates of resistance to ceftazidime (77·2%), cefotaxime (70·2%), meropenem (22·8%) and imipenem (17·5%). They showed variability in number and size (~1 to >20 kb) of plasmids. Among all, 1, 4, 13 and 31 isolates showed the bla(KPC) , bla(NDM) , bla(SHV) and bla(TEM) genes, respectively. The bla(KPC-2) gene was observed in one E. coli isolate. The bla(NDM-1) gene was detected in Stenotrophomonas maltophilia (n = 2), Acinetobacter baumannii (n = 1) and Ochrobactrum anthropi (n = 1) isolates. These isolates co-harboured the bla(TEM) and bla(SHV) genes. The intI1 gene was detected in 22 (38·6%) isolates, and 16 of these isolates showed the carbapenemase and/or ESBL genes. The conjugative movement of bla(NDM) gene could not be proved after three repetitive mating experiments. The presence of genes encoding carbapenemases and ESBLs in bacteria from goat meat poses public health risks.202032767781
105170.9999Multi-drug Resistance, β-Lactamases Production, and Coexistence of bla (NDM-1) and mcr-1 in Escherichia coli Clinical Isolates From a Referral Hospital in Kathmandu, Nepal. The ability of pathogenic Escherichia coli to produce carbapenemase enzymes is a characteristic that allows them to resist various antibiotics, including last-resort antibiotics like colistin and carbapenem. Our objectives were to identify rapidly developing antibiotic resistance (AR), assess β-lactamases production, and detect mcr-1 and bla (NDM-1) genes in the isolates. A prospective cross-sectional study was carried out in a referral hospital located in Kathmandu from November 2019 to December 2020 using standard laboratory and molecular protocols. Among 77 total E. coli isolates, 64 (83.1%) of them were categorized as MDR. Phenotypically 13 (20.3%) colistin-resistant, 30 (46.9%) ESBL and 8 (12.5%) AmpC producers, and 5 (7.8%) ESBL/AmpC co-producers were distributed among MDR-E. coli. Minimum inhibitory concentrations (MIC) against the majority of MDR isolates were exhibited at 1 g/L. Of these 77 E. coli isolates, 24 (31.2%) were carbapenem-resistant. Among these carbapenem-resistant bacteria, 11 (45.9%) isolates were reported to be colistin-resistant, while 15 (62.5%) and 2 (8.3%) were MBL and KPC producers, respectively. Out of 15 MBL producers, 6 (40%) harbored bla (NDM-1), and 8 (61.5%) out of 13 colistin-resistant pathogens possessed mcr-1. The resistance by colistin- and carbapenem were statistically associated (P < .001). However, only 2 (18.2%) of the co-resistant bacteria were found to have both genes. Our study revealed the highly prevalent MDR and the carbapenem-resistant E. coli and emphasized that the pathogens possess a wide range of capabilities to synthesize β-lactamases. These findings could assist to expand the understanding of AR in terms of enzyme production.202336741474
104780.9999Biofilm formation and antibiotic resistance profiles of water-borne pathogens. Water sources (surface water, drinking water, rivers, and ponds) are significant reservoirs for transmitting antibiotic-resistant bacteria. In addition, these waters are an important public health problem because they are suitable environments for transferring antibiotic resistance genes between bacterial species. Our study aimed to assess the prevalence of Extended-spectrum beta-lactamase (ESBL) producing isolates in water samples, the susceptibility of the isolates to the specified antibiotics, the determination of biofilm ability, antibiotic resistance genes, and the molecular typing of the isolates. For this purpose, Polymerase chain reaction (PCR) and Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) analyses were used. Out of 70 isolates, 15 (21%) were ESBL producing, and sent for the MALDI-TOF analysis, where Escherichia coli, Acinetobacter calcoaceticus, Enterobacter bugandensis, Acinetobacter pittii, Pseudomonas aeruginosa, Acinetobacter junii, Pseudomonas oleovorans, and Enterobacter ludwigigii were identified. Moreover, colistin resistance genes (mcr 1/2/6, mcr 4, mcr 5, mcr 3/7, and mcr 8), ESBL-encoding genes (bla(SHV), bla(TEM), and bla(CTX-M)) and carbapenemase genes (bla(NDM), bla(OXA-48), and bla(KPC)) using molecular analysis (PCR) were confirmed. The colistin resistance gene was detected at 80% (12/15) in the isolates obtained. The distribution of these isolates according to resistance genes was found as mcr 1/2/6 4 (20%), mcr 3/7 3 (13%), and mcr 5 (40%). Additionally, the isolates harbored bla(SHV)(6.6%) and bla(TEM) (6.6%) genes. However, bla(NDM), bla(OXA-48), bla(KPC), and bla(CTX-M) genes were not detected in any isolates. According to the Congo red agar method, seven (46.6%) isolates showed negative biofilm ability, and eight (53.3%) showed moderate biofilm ability. However, the microplate method detected weak biofilm in 53.3% of the isolates. In conclusion, this study provides evidence for the existence of multidrug-resistant bacteria that co-exist with mcr and ESBL genes in water sources. These bacteria can migrate to other environments and pose increasing threats to public health.202337004897
112490.9999Molecular Identification of Extended-Spectrum β-lactamase and Integron Genes in Klebsiella Pneumonia. INTRODUCTION: Infections caused by Gram negative bacteria, producing extended-spectrum β-lactamase, including Klebsiella pneumoniae are increasing all over the world with high morbidity and mortality. The aim of the present study was determined antimicrobial profile susceptibility and the prevalence of antibiotic resistance genes by multiplex PCR. METHODS: In the present study, we obtained one-hundred isolates of K. pneumoniae from different clinical samples. The antibiotic susceptibility testing was done in thirteen antibiotic and, therefore, M-PCRs were conducted using the DNA amplification for detection of ESBLs (blaTEM, blaCTX-M, blaSHV) and int (I, II, III) genes. RESULTS: The results of resistance to amoxicillin/clavulanate, ciprofloxacin, amikacin, trimethoprim-sulfamethoxazole, cefotaxime, ampicillin, aztreonam, imipenem, gentamicin, ceftazidime, Cefepime, ceftriaxone and levofloxacin were obtained 37%, 37%, 93%, 84%, 52%, 87%, 59%, 8%, 24%, 67%, 52%, 43% and 26%, respectively. The frequency of the extended-spectrum β-lactamase K. pneumoniae was obtained 37%. The prevalence of resistance genes of ESBLs in the M-PCR method showed that the blaTEM, blaCTX and blaSHV were 38%, 24% and 19%, respectively, however, only 8 (8%) out of 100 isolates were found to have positive outcomes for the existence of class 1 integrons and there were no detected class 2 or class 3 integrons. CONCLUSIONS: Our results recommend the likely co-carriage of some ESBLs genes and antibiotic resistance integrons on the same plasmids harboring the MDR genes.201627935927
1050100.9999Antibiotic resistance and β-lactam resistant genes among bacterial isolates from clinical, river water and poultry samples from Kathmandu, Nepal. OBJECTIVE: To assess the antibiotic resistance and beta-lactam resistance genes among bacterial isolates from clinical, river water and poultry samples. METHODS: Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa were isolated from clinical, poultry and river water samples collected during 2020-22. They were subjected to antimicrobial susceptibility tests following the CLSI guidelines. The bacteria were screened for β-lactam resistance genes bla (TEM), mcr-1, mecA and bla(NDM-1) . RESULTS: Among 2835 clinical samples, E. coli was the most frequently isolated bacterium (10.3%, 292), followed by S. aureus (6.0%, 169) and P. aeruginosa (4.0%, 143). Of the E. coli isolates, 64.4% exhibited multidrug resistance (MDR) and 43.8% were extended-spectrum β-lactamase (ESBL) producers, with 44.5% and 16.4% harbouring the blaTEM and mcr-1 genes, respectively. Among S. aureus isolates, 80.9% of methicillin-resistant strains (MRSA) carried the mecA gene, while 30.1% of metallo-β-lactamase (MBL)-producing P. aeruginosa were positive for the blaNDM-1 gene. In poultry samples, 30.4% of E. coli isolates harboured the blaTEM gene among 128 ESBL producers, and the prevalence of colistin-resistant isolates carrying mcr-1 was higher than in clinical samples. In contrast, the occurrence of ESBL-producing E. coli and MRSA, along with their associated resistance genes, was lower in water samples. CONCLUSIONS: This study demonstrated widespread multidrug resistance (MDR) and ESBL production among clinical, poultry and river water bacterial isolates in the Kathmandu valley. Colistin-resistant E. coli carrying the mcr-1 gene, methicillin-resistant S. aureus (MRSA) with mecA and metallo-β-lactamase (MBL)-producing P. aeruginosa harboring blaNDM-1 were detected across sources. These findings emphasize an urgent One Health approach to curb the growing threat of antimicrobial resistance in the region.202541113068
945110.9998Extended Spectrum Beta Lactamase (ESBL), bla(TEM),bla(SHV) and bla(CTX-M), Resistance Genes in Community and Healthcare Associated Gram Negative Bacteria from Osun State, Nigeria. BACKGROUND: Extended Spectrum Beta Lactamase (ESBL) production in gram negative bacteria confers multiple antibiotic resistance, adversely affecting antimicrobial therapy in infected individuals. ESBLs result from mutations in β-lactamases encoded mainly by the bla(TEM),bla(SHV) and bla(CTX-M) genes. The prevalence of ESBL producing bacteria has been on the increase globally, especially its upsurge among isolates from community-acquired infections has been observed. AIM: To determine ESBL prevalence and identify ESBL genes among clinical isolates in Osun State, Nigeria. MATERIAL AND METHODS: A cross-sectional study was carried out from August 2016 - July 2017 in Osun State, Nigeria. Three hundred and sixty Gram-negative bacteria recovered from clinical samples obtained from both community and healthcare-associated infections were tested. They included 147 Escherichia coli (40.8%), 116 Klebsiella spp (32.2%), 44 Pseudomonas aeruginosa (12.2%) and 23 Proteus vulgaris (6.4%) isolates. Others were Acinetobacter baumannii, Serratia rubidae, Citrobacter spp, Enterobacter spp and Salmonella typhi. Disk diffusion antibiotic susceptibility testing was carried out, isolates were screened for ESBL production and confirmed using standard laboratory procedures. ESBLs resistance genes were identified by Polymerase Chain Reaction (PCR). RESULTS: All isolates demonstrated multiple antibiotic resistance. Resistance to ampicillin, amoxicillin with clavulanate and erythromycin was 100%, whereas resistance to Imipenem was very low (5.0%). The overall prevalence of ESBL producers was 41.4% with Klebsiella spp as the highest ESBL producing Enterobacteriacaea. ESBL producers were more prevalent among the hospital pathogens than community pathogens, 58% vs. 29.5% (p=0.003). ESBL genes were detected in all ESBL producers with the bla(CTX-M) gene predominating (47.0%) followed by bla(TEM) (30.9%) and bla(SHV) gene was the least, 22.1%. The bla(CTX-M) gene was also the most prevalent in the healthcare pathogens (62%) but it accounted for only 25% in those of community origin. CONCLUSION: A high prevalence of ESBL producing gram-negative organisms occurs both in healthcare and in the community in our environment with the CTX-M variant predominating. Efforts to control the spread of these pathogens should be addressed.202132729432
924120.9998Screening of Antimicrobial Resistance Genes and Epidemiological Features in Hospital and Community-Associated Carbapenem-Resistant Pseudomonas aeruginosa Infections. INTRODUCTION: Researching carbapenem-resistant isolates enables the identification of carbapenemase-producing bacteria and prevents their spread. METHODS: P. aeruginosa isolates were recovered from Medicine Faculty of Recep Tayyip Erdoğan University and identified by conventional methods and the automated Vitek 2 Compact system. Antimicrobial susceptibility experiments were performed in accordance with CLSI criteria and the automated Vitek 2 Compact system. The PCR method was investigated for the presence of β-lactamase resistance genes. PFGE typing was performed to show clonal relation among samples. RESULTS: Seventy P. aeruginosa isolates were isolated from seventy patients. Of the patients, 67.1% had contact with the health service in the last 90 days and 75.7% of the patients had received antimicrobial therapy in the previous 90 days. Twenty-four isolates were carbapenem resistant, 2 isolates were multidrug-resistant except colistin, and none of the samples had colistin resistance. The gene encoding β-lactamase or metallo-β-lactamase was found in a total of 36 isolates. The bla (VEB) and bla (PER) genes were identified in 1 and 5 isolates alone or 17 and 13 isolates in combination with other resistance genes, respectively. The bla (NDM) was the most detected metallo-β-lactamase encoding gene (n=18), followed by bla (KPC) (n=12). bla (IMP) and bla (VIM) were detected in 5 and 1 isolates, respectively. Also, the association of bla (VEB)-bla (PER) and bla (VEB)-bla (KPC)-bla (NDM) was found to be very high. Much more resistance genes and co-occurrence were detected in hospital-acquired samples than community-acquired samples. No difference was found between the community and hospital-associated isolates according to PFGE results. Simultaneously from 6 patients, other microorganisms were also isolated and 5 of them died. CONCLUSION: The average length of stay (days) was found to be significantly higher in HAI group than CAI group. The death of 5 patients with fewer or no resistance genes showed that the co-existence of other microorganisms in addition to resistance genes was important on death.202133907430
1127130.9998Extended spectrum beta-lactamase and aminoglycoside modifying enzyme genes in multi drug resistant Gram-negative bacteria: A snapshot from a tertiary care centre. BACKGROUND: This study aims to enhance the existing knowledge of the prevalence of genes responsible for beta-lactam resistance and aminoglycoside resistance in gram negative organisms by molecular detection of extended spectrum beta-lactamase and aminoglycoside modifying enzymes in multidrug-resistant gram-negative bacteria. METHODS: Out of 864 gram-negative isolates, 710 were phenotypically identified as multidrug-resistant by antibiotic susceptibility testing. From the above isolates, 102 representative isolates as per sample size calculated were selected for further molecular studies. The presence of blaTEM, blaCTX-M blaSHV, and five AmpC genes was detected by real-time polymerase chain reaction (PCR). Conventional PCR was performed to detect seven aminoglycoside modifying enzyme genes namely aac(6')-Ib, aac(6')-Ic, aac(3)-Ia, aac(3)-Ib, aac(3)-IIa, ant(2'')-Ia, and ant(4'')-IIa. RESULTS: Most common multidrug-resistant isolate was Klebsiella pneumoniae (35%) followed by Escherichia coli (30%). Among the 102 selected isolates all harboured blaTEM gene, 71 (69.6%) harboured blaCTX-M gene and 48 (47%) blaSHV gene. Among the selected isolates 60% showed the presence of AmpC genes. Most common aminoglycosie modifying enzyme gene was AAC 6' Ib (51%) followed by ANT 2" Ia (36%). CONCLUSION: This study suggests a wider use of molecular methods using specific PCR amplification of resistance genes. It would be beneficial to perform the molecular identification of antimicrobial resistance genes to effectively monitor and manage antibiotic resistance, administer appropriate antimicrobial medication, practice antimicrobial stewardship and improve hospital infection control procedures.202439734850
1443140.9998Wastewater Surveillance Detected Carbapenemase Enzymes in Clinically Relevant Gram-Negative Bacteria in Helsinki, Finland; 2011-2012. Antimicrobial resistance profiling of pathogens helps to identify the emergence of rare or new resistance threats and prioritize possible actions to be taken against them. The analysis of wastewater (WW) can reveal the circulation of antimicrobial-resistant bacteria (ARB) and antimicrobial resistance genes (ARG) among the catchment communities. Here, we analyzed WW influent samples to determine the prevalence of carbapenemase genes-carrying Gram-negative bacteria (Carba-GNB) in Helsinki, Finland. This study set important historical reference points from the very early stage of the carbapenemase era, during the period 2011-2012. A total of 405 bacterial isolates grown on CHROMagarKPC (n = 195) and CHROMagarESBL (n = 210) from WW influent samples were collected between October 2011 and August 2012 and were analyzed. The bacterial DNA from the isolates was extracted, and the prevalence of carbapenemases genes bla (KPC), bla (NDM), bla (GES), bla (OXA-48), bla (IMP), bla (IMI), and bla (VIM) were screened with multiplexed PCR. All carbapenemase-positive isolates were identified taxonomically to species or genus level with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). The nucleic acid extraction was successful for 399 isolates, of which 59 (14.8%) were found to carry carbapenemase genes. A total of 89.8% of the carbapenemase positive isolates (53 out of 59) were obtained from CHROMagarKPC plates and only 10.2% (six out of 59) were obtained from CHROMagar ESBL plates. Among the Carba-GNB isolates, 86.4% were bla (GES) (51 out of 59), 10.2% were bla (KPC) (six out of 59), and 3.4% were bla (VIM) (two out of 59). The most common carba-gene, bla (GES), was carried by 10 different bacterial species, including Aeromonas spp., Enterobacter spp., and Kluyvera spp.; the bla (KPC) gene was carried by Escherichia coli, Klebsiella pneumoniae, and Kluyvera cryocescens; and the bla (VIM) gene was carried by Aeromonas hydrophila/caviae and Citrobacter amalonaticus. This study emphasizes that wastewater surveillance (WWS) can be an additional tool for monitoring antimicrobial resistance (AMR) at the population level.202235722284
923150.9998Prevalence of Oxacillinase Genes in Clinical Multidrug-Resistant Gram-Negative Bacteria. BACKGROUND: The emergence of OXA-type beta-lactamases has become a significant threat to public healthcare systems and may lead to prolonged hospital stays and increased mortality rates among affected patients. This study aimed to determine the prevalence of oxacillinase resistance (OXA) genes in multidrug-resistant (MDR) Gram-negative bacteria. METHODS: One hundred and six clinical isolates were collected from a stock of Gram-negative isolates and were identified and tested for antibiotic susceptibility and presence of OXA genes using polymerase chain reaction (PCR). RESULTS: The most common detected isolate was Klebsiella pneumoniae (36.8%), followed by Escherichia coli (33%), Pseudomonas aeruginosa (16%), and Acinetobacter baumannii (14.2%). Out of these isolates, 97.4%, 87.2%, 84.6%, and 79.5% were resistant to ampicillin/sulbactam, cefotaxime, ceftazidime, and aztreonam, respectively. PCR results confirmed the presence of one or more OXA genes in 34% of the samples studied. The blaOXA-1 and blaOXA-10 genes were the most highly detected genes, followed by blaOXA-4 and blaOXA-51. The total number of Pseudomonas aeruginosa isolates was confirmed to carry at least one OXA gene (70.6%), whereas Acinetobacter baumannii, Klebsiella pneumoniae, and Escherichia coli were confirmed to carry at least one OXA gene (53.3, 28.2, and 22.9%, respectively). There was a significant association (p < 0.05) between the resistance genes and the type of isolate. CONCLUSIONS: Pseudomonas aeruginosa and Acinetobacter baumannii are the most common MDR Gram-negative strains carrying OXA-type beta-lactamase genes. Monitoring of MDR pathogens in Gram-negative bacteria must be continuously undertaken to implement effective measures for infection control and prevention.202540066541
1071160.9998Characterization of Beta-Lactamase and Fluoroquinolone Resistance Determinants in Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa Isolates from a Tertiary Hospital in Yola, Nigeria. Infections due to antimicrobial resistant gram-negative bacteria cause significant morbidity and mortality in sub-Saharan Africa. To elucidate the molecular epidemiology of antimicrobial resistance in gram-negative bacteria, we characterized beta-lactam and fluoroquinolone resistance determinants in Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa isolates collected from November 2017 to February 2018 (Period 1) and October 2021 to January 2022 (Period 2) in a tertiary medical center in north-eastern Nigeria. Whole genome sequencing (WGS) was used to identify sequence types and resistance determinants in 52 non-duplicate, phenotypically resistant isolates. Antimicrobial susceptibility was determined using broth microdilution and modified Kirby-Bauer disk diffusion methods. Twenty sequence types (STs) were identified among isolates from both periods using WGS, with increased strain diversity observed in Period 2. Common ESBL genes identified included bla(CTX-M), bla(SHV,) and bla(TEM) in both E. coli and K. pneumoniae. Notably, 50% of the E. coli in Period 2 harbored either bla(CTX-M-15) or bla(CTX-M-1 4) and phenotypically produced ESBLs. The bla(NDM-7) and bla(VIM-5) metallo-beta-lactamase genes were dominant in E. coli and P. aeruginosa in Period 1, but in Period 2, only K. pneumoniae contained bla(NDM-7), while bla(NDM-1) was predominant in P. aeruginosa. The overall rate of fluoroquinolone resistance was 77% in Period 1 but decreased to 47.8% in Period 2. Various plasmid-mediated quinolone resistance (PMQR) genes were identified in both periods, including aac(6')-Ib-cr, oqxA/oqxB, qnrA1, qnrB1, qnrB6, qnrB18, qnrVC1, as well as mutations in the chromosomal gyrA, parC and parE genes. One E. coli isolate in Period 2, which was phenotypically multidrug resistant, had ESBL bla(CTX-M-15,) the serine carbapenemase, bla(OXA-181) and mutations in the gyrA gene. The co-existence of beta-lactam and fluoroquinolone resistance markers observed in this study is consistent with widespread use of these antimicrobial agents in Nigeria. The presence of multidrug resistant isolates is concerning and highlights the importance of continued surveillance to support antimicrobial stewardship programs and curb the spread of antimicrobial resistance.202337999619
1069170.9998High Prevalence of Antimicrobial Resistance in Gram-Negative Bacteria Isolated from Clinical Settings in Egypt: Recalling for Judicious Use of Conventional Antimicrobials in Developing Nations. This study was designed to investigate, at the molecular level, the antimicrobial resistance mechanisms of different antimicrobial resistance genes, including, extended-spectrum β-lactamases, AmpC β-lactamases, class 1 and 2 integrons, and plasmid-mediated quinolone resistance genes of Gram-negative bacteria isolated from clinical settings in Egypt. A total of 126 nonduplicate Gram-negative isolates were recovered from different clinical samples taken from hospitalized patients in Egypt in 2014. Antimicrobial susceptibility testing showed that, 93.6% (118/126) of the isolates had a multidrug-resistant phenotype. Interestingly, we reported a high level of antimicrobial resistance nearly for all tested antibiotics; to our knowledge, this is the first report from Egypt indicating very high level of antibiotic resistance in Egypt. Polymerase chain reaction screening and DNA sequencing revealed that, 75.4% (95/126) of the isolates harbored at least one extended-spectrum β-lactamase-encoding gene, with bla(CTX-M) being the most prevalent (65.9%), followed by bla(SHV) (46.8%). The AmpC β-lactamase, bla(CMY), was detected in 7.1% (9/126) of bacterial isolates, with bla(CMY-42) being the most prevalent. Class 1 integrons were detected in 50.8% (64/126) of the isolates, and class 2 integrons were detected in 2.4% (3/126) of the isolates. The plasmid-mediated quinolone resistance gene, qnr, was detected in 58.7% (74/126) of the tested isolates, with qnrS being the most prevalent. Several antimicrobial resistance determinants were identified in Egypt for the first time, such as SHV-27, SHV-28, SHV-33, SHV-63, SHV-71, SHV-82, SHV-142, CMY-42, CMY-6, and the new CMY-72 like. This study highlights the importance of the conscious use of conventional antimicrobials to overcome the multidrug resistance problem.201930681401
2174180.9998Frequency of Beta-Lactamase Antibiotic Resistance Genes in Escherichia Coli and Klebsiella pneumoniae. BACKGROUND: This cross-sectional study was performed on isolates of Klebsiella pneumoniae, and E.coli from clinical specimens of patients admitted to Sayyad Shirazi Hospital by census sampling method in 2019. Antibiogram testing was performed using the disk diffusion method as defined by the Clinical and Laboratory Standards Organization for performing this test. Finally, the abundance of genes was evaluated by PCR using specific primers. Frequency, percentage, mean±SD were used to describe the data. Chi-square and Fisher's exact tests were used to compare the presence and absence of the studied genes alone and in the presence of each other. RESULT: This study was performed on 130 positive samples, isolated from 32 (24.6%) males and 98 (65.4%) females with a mean age of 43.78 ± 21.72. From the total number of 130 isolates, 84 (64.6%) consisted of E.coli, and 46 (35.4%) were Klebsiella. Most of the cultures were urine and vaginal (61.5%). The highest antibiotic resistance in isolates was cephalexin and cefazolin (67.9% in E.coli & 63% in Klebsiella). Colistin was identified as the most effective antibiotic (100%) in both. AMPC extendedspectrum β-lactamase genes were present in 40 (30.8%) isolates. The highest frequency about the gene pattern of AMPC positive β-lactamase bacteria was correlated to DHA, FOX, and CIT genes, while none of the samples contained the MOX β-lactamase gene. E.coli and Klebsiella beta-lactamase-producing AMPC isolates were also significantly correlated with antibiotic resistance to the cephalosporin class (P <0.05). CONCLUSION: This study indicated a high percentage of resistance to third and fourth generation cephalosporins. Hence, careful antibiogram tests and prevention of antibiotic overuse in infections caused by AMPC-producing organisms and screening of clinical samples for the resistance mentioned above genes and providing effective strategies to help diagnose and apply appropriate treatments and change antibiotic usage strategies can partially prevent the transmission of this resistance.202134483624
1048190.9998Characterizing the co-existence of metallo-β-lactamase-producing and extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella pneumoniae isolates in community wastewater samples of Dhaka, Bangladesh. Escherichia coli and Klebsiella pneumoniae isolates with multiple antibiotic-resistance genes in wastewater pose serious public health risks, as they can potentially contaminate the food and water supply. The main aim of this study was to isolate and identify E. coli and K. pneumoniae from community wastewater samples, and determine their antibiotic-resistance profiles and their antibiotic-resistant genes. From the northern part of Dhaka, Bangladesh, 36 wastewater samples were collected across 11 different areas, which were then serially diluted, and cultured using selective media. Isolates were identified via polymerase chain reaction. Out of the 197 isolates identified, E. coli and K. pneumoniae accounted for 55.8% (n = 110) and 44.2% (n = 87), respectively. Antibiotic susceptibility tests revealed multidrug resistance (MDR) in 30% of E. coli and 35.56% of K. pneumoniae isolates. Among E. coli, the prevalence of antibiotic-resistance genes included bla(NDM-1) (8.9%), bla(SHV) (13.9%), and bla(CTX-M) (7.6%). In K. pneumoniae, the percentages were bla(NDM-1) (12.8%), bla(SHV) (4.3%), and bla(CTX-M) (5.0%). Co-existence of multiple antibiotic-resistance genes was observed in 4.54% of E. coli isolates (n = 5) and 5.74% of K. pneumoniae isolates (n = 5). This suggests the escalating issue of infectious species becoming increasingly resistant to antibiotics in wastewater systems.202540298266