Phenotypic and Genotypic Characterization of Extended Spectrum Beta-Lactamases Producing Bacteria Causing Urinary Tract Infections among Expectant Women Attending Antenatal Clinic at Ruiru Sub County Hospital, Kenya

Background: Urinary tract infection (UTI) is a bacterial infection affecting males and females but is more prevalent in expectant women. ESBLs are bacteria with enzymes that make them resistant to many antibiotics, posing a significant health challenge. This study aims to determine the characteristics of ESBL-producing bacteria causing UTIs in expectant women. Methodology: A self-administered survey was carried out;300 expectant women were recruited using a random sampling method. A questionnaire was used to collect socio-demographic information. Urine samples were collected in sterile universal bottles and processed at the JKUAT Zoology laboratory. Urine samples were analyzed using urinalysis, microscopy, culture, and sensitivity testing. ESBL-producing bacteria were identified phenotypically using the double-disc synergy test (DDST) and genotyped for specific resistant genes using PCR. Results: UTI prevalence was 32.7% (98/300). UTI was significantly associated with the history of previous UTI (OR = 0.84, p = 0.02) and multigravida (OR = 0.14 p = 0.01). UTI was common in women aged between 28-37 years in their second trimester. Bacteria isolated were E. coli 57.1% (56/98), S. aureus 21.4% (21/98) K. pneumonia 11.2% (11/98) and Proteus spp 10.4% (10/98). Bacteria antibiotic resistance patterns were E. coli-tetracycline (91.1%), sulfamethoxazole (55.4%), cefotaxime (53.4%) and augmentin (53.4%). S. aureus-sulfamethozaxole (100%) and augmentin (71.4%), K. pneumoniae-sulfame-thoxazole (72.2%) cefotaxime (63.6%), chloramphenicol and tetracycline (54.5%). Proteus spp: tetracycline (100%), nitrofurantoin (90%), cefotaxime and chloramphenicol (50%). The proportion of ESBLs bacterial producers was 37.6% (29/77) and 44.8% (13/29) possessed ESBLs resistant genes;Bla CTX-M 53.8% (7/13), Bla SHV and Bla TEM 23.1% (3/13) each, Bla OXA (0%) was not detected. Conclusion: The study revealed a high proportion of ESBLs producing bacteria responsible for UTI in expectant women. ESBLs screening, routine culture and sensitivity testing will guide on proper management and empirical treatment of UTI patients thus reducing multi-drug resistance.

Appendical Perforation by Infection with Extended-Spectrum Beta-Lactamase (ESBL)-Producing <i>Escherichia coli</i>: Case Report

We report the very rare case of a huge appendical abscess with extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli (E. coli) as the pathogen. There have been several reports of appendical infections such as appendicitis and appendical abscess caused by ESBL-producing bacteria in adults. The treatment of ESBL-producing E. coli infection is specific, and ESBL-producing bacteria have recently been reported as pathogens associated appendicitis in children. To the best of our knowledge, this is the second report of perforated appendicitis with abscess due to ESBL-producing E. coli. We discuss the diagnostic modalities and treatments for appendical abscess with ESBL-producing E. coli. and propose that the patients with perforated appendicitis and abscess formation due to ESBL-producing E. coli should be administered the antibiotic MEPM within 2 weeks to treat the abscess more effec-tively without producing other multidrug-resistant bacteria.

Diagnostic Validity of Cica Beta Test 1 for the Detection of Extended Spectrum Beta-Lactamase (ESBL) Producing Gram Negative Bacteria by Comparing with Phenotypic Method

Background: Detection of extended spectrum beta lactamase producing bacteria is an important issue in the clinical settings. Objective: The purpose of the present study was to validate the Cica Beta Test 1 for detection of extended spectrum beta-lactamase (ESBL) producing bacteria. Method: This analytical type of cross-sectional study was carried out in the Department of Microbiology and Immunology at Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka from January 2006 to December 2006 for a period of one (01) year. All the patients presented with the clinical features of urinary tract infection and surgical as well as burn wound infection at any age with both sexes were selected as study population. All bacteria were isolated and identified by their colony morphology, staining characters, pigment production, motility and other relevant biochemical tests. Phenotypic confirmation of ESBLs producing isolates were done by inhibitor potentiated disc diffusion test according to CLSI recommendation. The Cica Beta Test 1 was performed according to the manufacturer’s instructions. Result: A total number of 288 Gram negative bacteria were isolated. Among these isolates Cica Beta test 1 was positive in 97 strains and phenotypic confirmatory test was positive in 89 strains. The test sensitivity of Cica Beta Test 1 was 100% (95% CI 95.9% to 100.0%). Specificity of the test was 96.0% (95% CI 92.2% to 98.2%). The positive predictive value (PPV) and negative predictive value (NPV) were 92.7% (95% CI 84.5% to 95.7%) and 100.0% (95% CI 98.0% to 100.0%) respectively. The accuracy of the test was 97.2% (95% CI 95.1% to 99.1%). Area under ROC curve = 0.980 (95% CI 0.964 to 0.996);p value 0.0001. Conclusion: In conclusion, Cica Beta Test 1 is very high sensitivity and specificity for the detection of ESBL from Gram negative bacteria.

Antibiogram of Extended-Spectrum <i>β</i>-Lactamase (ESBL) Producing <i>Escherichia coli</i>

Background: Extended-spectrum β-lactamases (ESBLs) are enzymes capable of hydrolyzing extended-spectrum cephalosporins, penicillins and monobactams but inactive against cephamycins and carbapenems. The ESBL-producing organisms are a breed of multidrug-resistant pathogens. Objectives: This study was aimed to determine the susceptibility pattern of ESBL-producing Escherichia coli to ciprofloxacin, amikacin and imipenem. Methods: A total of 75 ESBL-producing E. coli, were obtained from the tertiary care hospitals of Bangladesh and were studied for susceptibility pattern from October, 2010 to December, 2011. These isolates were identified by double disc synergy test (DDST) and were confirmed phenotypically as ESBL-producer by phenotypic confirmatory disc diffusion test (PCDDT). Minimum inhibitory concentrations (MICs) of ciprofloxacin, amikacin and imipenem among ESBL-producing E. coli were determined using agar dilution method. Results: Out of 75 DDST positive ESBL-producing E. coli, 71 (94.67%) were also positive by PCDDT. All ESBL-producing E. coli, were susceptible to imipenem. About 92.95% ESBL-producing E. coli were susceptible to amikacin but only 14.08% were susceptible to ciprofloxacin. Conclusion: In this study, ESBL-producing E. coli, showed high resistance to ciprofloxacin. Imipenem and amikacin were most effective against ESBL positive strains.

Phenotypic Detection and Susceptibility Pattern for the Detection of Extended Spectrum β-Lactamase-Producing <i>Klebsiella pneumonia </i>Isolates in Nairobi, Kenya

Klebsiella pneumoniae is an opportunistic pathogen that is an important cause of nosocomial infections. Detection of ESBL producers’ poses a special challenge for clinical microbiology laboratories, although ESBL producing pathogens are able to hydrolyze extended-spectrum penicillins, cephalosporins, and aztreonam, the minimum inhibitory concentrations (MIC) of some and perhaps even all of these agents may be within the susceptible range. The third generation cephalosporins have the reputation for being useful against a broad range of bacterial infections. However, resistance to these agents is something that must still be considered and creates obstacles for their clinical use. A total of 80 multi drug resistant clinical isolates of Klebsiella pneumoniae were obtained from a study on anaerobes associated with Pelvic Inflammatory disease (P.I.D), KEMRI S.S.C No.495. The isolates were identified by standard microbiological procedures. Antimicrobial susceptibility testing was carried out by Kirby-Bauer method. Upon identification, the antibiogram profiles of the isolates were determined and those resistant to third-generation cephalosporins were tested for production of ESBL. ESBL production among the multi drug resistant isolates was detected using the phenotypic confirmatory disc diffusion test (PCDDT) and double disk synergy test (DDST). While using standard double disk synergy test (DDST) as screening method for identifying potential ESBL producers, ceftriaxone was the most efficient antimicrobial in screening isolates as potential ESBL producers followed by cefotaxime.

Prevalence and characteristics of extended spectrum β-lactamase-producing Escherichia coli from bovine mastitis cases in China

The aim of the study was to investigate the prevalence and characterization of extended-spectrum β-lactamase （ESBL）- producing Escherichia coli isolated from bovine mastitis cases in China. ChromID ESBL agar was used to confirm ESBL-producing E. coli. PCR and DNA sequencing were employed to characterize the genotype of ESBL-producers. Antimicrobial susceptibility was measured by disc diffusion. Overall, 73 of 318 E. coli isolates （22.96％） were identified as ESBL-producers. Of these ESBL-producing E. coli, the prevalence of blaCTX-M and blaTEM-1 was 97.26 and 71.23％, respectively. The predominant CTX-M-type ESBL was CTX-M-15 （65.75％）, followed by CTX-M-14 （10.96％）, CTX-M-55 （9.59％）, CTX-M-64 （5.48％）, CTX-M-65 （4.11％） and CTX-M-3 （1.37％）. This study is the first report of CTX-M-64 and CTX-M-65 in E. coli isolated from bovine mastitis. Furthermore, 72 ESBL-producing E. coli isolates （98.63％） were found to be multidrug-resistance. This study noted high prevalence and rates of antimicrobial resistance of ESBL-producing E. coli isolates from bovine mastitis cases in China.

Antimicrobial Resistance and Genotype Analysis of Extended-Spectrum-β-Lactamase-Producing Proteus Mirabilis

To analyse the genotypes of clinical isolates of Extended-Spectrum-β-Lactamase-Producing (ESBL-producing) Proteus mirabilis (P. mirabilis) and the mechanisms of antimicrobial resistance, to guide reasonable use of antibiotics and to avoid nosocomial outbreak infections by ESBL-producing P. mirabilis. 125 clinical isolates of P. mirabilis were collected from the Drug-Resistant Bacteria Surveillance Center of Anhui Province (from Jan 2009 to May 2010). Searching for the genotypes of ESBLs was perfomed by PCR amplification and DNA sequencing, and performed conjugation test simultaneously. Among ESBL-producing strains, CTX-M was the major genotype (3 CTX-M-13 and 1 CTX-M-3). TEM-1b spectrum β-lactamase was also prevalence in P. mirabilis. The diversity of β-lactamases in P. mirabilis and the emergency of multi-drug-resistance clinical strains will present serious threat to clinical therapy and even will lead to outbreak of nosocomial infections. Our study emphasizes the need for enhanced supervision of ESBL-producing P. mirabilis. Timely and reasonable drug-resistance data are indispensable to clinical therapy.

Detection of Beta-Lactamase and Extended-Spectrum Beta-Lactamase of Pathogens Isolated from Pig and Chicken and Their Antibiotic Susceptibility Test

The antibacterial activity of beta-lactam antibiotics or their combinations with inhibitor sulbactum against non-lactamase- producing strains, lactamase-producing and ESBLs-producing isolates was evaluated with twofold dilution method after pathogens isolated from pigs and chickens were detected, respectively, for beta-lactamase and extended-spectrum beta- lactamases （ESBLs）, The results revealed that most of 43 clinically isolated strains could produce beta-lactamase and 3 strains of shigella isolated from chicken samples produced ESBLs. All of 30 lactamase-producing strains isolated and only one of 16 non-lactamase-producing strains were resistant to amoxicillin and ampicillin. MICs of ampicillin against lactamaseproducing isolates decreased 10-40 and 10-20 times respectively, when it was conbined with sulbactam at ration of 1：2 and 1：4. All clinical isolates were susceptible to third-generation cephalosporins. The MICs of third-generation cephalosporins against lactamase-producing isolates did not change when they were conbined with sulbactam. MICs of ceftiofur and ceftriaxone against ESBLs-producing isolates decreased 2-4 times when they were conbined with sulbactam.

Post-appendectomy pelvic abscess with extended-spectrum beta-lactamase producing Escherichia coli : A case report and review of literature

BACKGROUND Appendicitis, the inflammation of the appendix, is the most common abdominal surgical emergency requiring expedient surgical intervention. Extendedspectrum beta-lactamases(ESBLs) are bacterial enzymes that catalyse the degradation of the betalactam ring of penicillins and cephalosporins(but without carbapenemase activity), leading to resistance of these bacteria to beta-lactam antibiotics. Recent increases in incidence of ESBL-producing bacteria have caused alarm worldwide. Proportion estimates of ESBLEnterobacteriaceae hover around 46% in China, 42% in East Africa, 12% in Germany, and 8% in the United States.CASE SUMMARY The impact of ESBL-producing bacteria on appendiceal abscesses and consequent pelvic abscesses are yet to be examined in depth. A literature review using the search words "appendiceal abscesses" and "ESBL Escherichia coli(E. coli)" revealed very few cases involving ESBL E. coli in any capacity in the context of appendiceal abscesses. This report describes the clinical aspects of a patient with appendicitis whodeveloped a postoperative pelvic abscess infected with ESBL-producing E. coli. In this report, we discuss the risk factors for contracting ESBL E. coli infection in appendicitis and post-appendectomy pelvis abscesses. We also discuss our management approach for postappendectomy ESBL E. coli pelvic abscesses, including drainage, pathogen identification, and pathogen characterisation. When ESBL E. coli is confirmed, carbapenem antibiotics should be promptly administered, as was done efficaciously with this patient. Our report is the first one in a developed country involving ESBL E. coli related surgical complications in association with a routine laparoscopic appendectomy.CONCLUSION Our report is the first involving ESBL E. coli and appendiceal abscesses, and that too consequent to laparoscopic appendectomy.

Microbiologic and Clinical Comparison of Patients Harboring <i>Escherichia coli</i>Blood Isolates with and without Extended-Spectrum <i>β</i>-Lactamases

The clinical and microbiologic characteristics of 34 patients with extended-spectrum β-lactamase (ESBL) positive E. coli isolated from blood were compared to 66 bacteremic patients with ESBL negative E. coli, from January 2007 through December 2009. Of the 21 ESBL positive isolates available for PCR analysis, 13 were positive for CTX-M, 8 for TEM, 4 for SHV β-lactamases, with 6 possessing multiple enzymes. Twenty of 34 (59%) ESBL-positive and 41 of 66 (62%) ESBL-negative blood isolates were considered community-associated. All but one isolate in both groups had MICs of ≤1.0 μg/ml to meropenem. However, when compared to ESBL-negative isolates, ESBL-positive isolates were more frequently resistant to levofloxacin, trimethoprim/sulfamethoxazole and had higher MICs to gentamicin, tobramycin and piperacillin/tazobactam. The use of intravenous and urinary catheters was strongly associated with the isolation of E. coli bloodstream isolates in both groups of patients. Although hospital stay was similar in both groups, appropriate therapy was given in 87% of patients with ESBL positive vs. 98% of patients with ESBL negative isolates and mortality was greater for patients with ESBL positive isolates (26% vs. 17%). Since a large proportion of E. coli blood isolates were ESBL-positive and community-associated, carbapenems should be considered as initial empiric therapy for such infections in our locale.

Prevalence and Potential Risk Factors of Hospital Acquired Extended-Spectrum Beta-Lactamase—Producing <i>Proteus</i>Species

Background: Multidrug resistance and production of extended spectrum β-lactamases (ESBLs) by a large group of bacterial agents in hospitals are to be a matter of scientific concern. Objective: This cross-sectional study was aimed to investigate the prevalence of ESBL producing Proteus species and risk factors associated with hospital acquired infection in addition to study the antibiotics susceptibility patterns of all bacterial isolates from inpatients of four Yemeni general hospitals. Methods: A total of 740 consecutive non-repeat culture isolates were obtained from admitted patients of Al-Kuwait University Hospital, Al-Thowra General Hospital, Al-Jumhori Teaching Hospital, and Military General Hospitals Sana’a city. We used Kirby-Bauer disk diffusion method to detect antimicrobial susceptibility and establish the presence of ESBLs-producing bacteria according to the Clinical and Laboratory Standards Institute guidelines. Results: Out of 740 isolate, 233 (31.5%) were Escherichia coli followed by Staphylococcus aureus 188 (25.4%), Pseudomonas aeruginosa 149 (20.1%), Klebsiella sp. 107 (14.5%), Enterococcus faecalis 25 (3.4%) and Proteus spp. 38 (5.1%). The highest frequencies of ESBLs producing among Proteus sp. were Proteus mirabilis 26 out 38 (68.4%) and Proteus vulgaris 12 out 38 (31.6%). The most effective of antimicrobial susceptibility pattern among Proteus spp. were Imipenem (100%) followed by Pipracillin-Tazobactam (92.3%) for P. mirabilis and (83.3%) for P. vulgaris, while the Amikacin (80.8%) for P. mirabilis and P. vulgaris with (91.7%). Amoxicillin and Cefotaxime were the highest for both species (100%). Conclusion: The prevalence of ESBL-producing Proteus spp. detected in this study is of great concern for public health authorities and a strict adherence of infection control policies and procedures with continuous antibiotics resistance surveillance including antimicrobial management and routine detection of ESBL-producing isolates are very important to prevent nosocomial infections.

Detection of Extended Spectrum β-Lactamase Producing Klebsiella pneumoniae and Escherichia coli in Two Hospitals in the Federal Capital Territory, Abuja, Nigeria

In this study, the prevalence of Extended Spectrum Beta-lactamase (ESBL) producing Klebsiella pneumoniae and Escherichia coli isolates from the University of Abuja Teaching Hospital and the National Hospital was determined. A total of two hundred and fifteen (215) clinical isolates were examined, of which 60% were E. coli and 40% K. pneumoniae respectively. The isolates were collected from various samples namely: Stool, Urine, Pus, High Vagina Swab, Sputum and Wound swab. Out of these isolates, 54 of K. pneumoniae were screened to be ESBL negative and 32 as ESBL positive isolates, while 88 and 40 E. coli were also screened as ESBL negative and ESBL positive isolates respectively. These represent 37.9% of all K. pneumoniae isolates and 31.25% of E. coli isolates respectively. The prevalence of ESBL among the species was not however statistically different (p > 0.05). Multiple resistance in these isolates was common and there is the need for routine screening of ESBL in our hospitals to guide rational and effective use of antibiotics.

羊腹泻大肠杆菌整合子及ESBLs携带情况检测

为了确定内蒙古自治区羊腹泻大肠杆菌的耐药机制,本试验采用PCR方法检测呼和浩特市周边牧场分离菌株中整合酶基因(Ⅰ、Ⅱ、Ⅲ型)、耐药基因盒和编码超广谱β-内酰胺酶(ESBLs)基因的携带情况。结果显示,108株羊腹泻大肠杆菌中,Ⅰ型整合酶基因和耐药基因盒插入区扩增呈阳性的菌株为42株,检出率为38.9%,未检出Ⅱ、Ⅲ型整合酶基因;菌株中有5类基因盒流行,主要携带甲氧苄氨嘧啶(dfr)和氨基糖苷类(aadA)耐药基因;分离菌株的bla_(TEM)基因检出数为83株,占比76.9%,bla SHV和bla_(CTX)基因PCR扩增均为阴性;质粒DNA上成功扩增出携带bla TEM型ESBLs基因的Ⅰ型整合子阳性菌株40株。结果表明,内蒙古自治区羊腹泻大肠杆菌Ⅰ型整合子的携带率较高,菌株对磺胺类、甲氧苄氨嘧啶和氨基糖苷类药物的耐受性与其携带的耐药基因盒密切相关,分离菌株中ESBLs的产生和传播由菌株自身质粒所介导,这可能是其对β-内酰胺类抗菌药物耐药的主要原因,且Ⅰ型整合子在ESBLs阳性菌株中的广泛分布,也使得菌株携带的耐药基因更易散播。本试验通过对羊腹泻大肠杆菌耐药性的产生和传播进行分析,以指导临床合理用药。

Prevalence of Extended Spectrum Beta-Lactamase-Producing Escherichia coli in Broiler Chickens in YaoundéCapital City of Cameroon

Background: Escherichia coli are ubiquitous bacteria colonising both humans and animals. Extended spectrum β-lactamase-producing E. coli has been selected as a suitable indicator for the monitoring and surveillance of antimicrobial resistance. Death due to resistant bacteria is continuously rising in Cameroon, but the contribution of the aviary sector is not well studied. Therefore, this study aimed to investigate the resistance profile of extended spectrum beta-lactamases-producing Escherichia coli strains, isolated from faeces of broiler chickens in Yaoundé, capital city of Cameroon. Methods: A cross-sectional descriptive study was carried out from February to June 2020. Escherichia coli were isolated from samples of broilers in poultry farms in Yaoundé and submitted to the extended spectrum β-lactamase screening. The logistic regression was used to assess the statistical association of a significance threshold p-value of 0.05. Results: Out of 385 faecal samples collected in broiler farms, 114 Escherichia coli isolates were obtained out of which 30 (26.32%) were Extended Spectrum Beta-Lactamases-producing Escherichia coli. These isolates revealed high resistance to all antibiotic families. Poor storage conditions for feeds and the proximity to latrines, the troughs on the ground, the lack of foot bath and uniforms, the inadequate treatment of faeces, the poor usage of preventive antibiotics and the lack of water treatment have been identified as risk factors to faecal carriage of ESBL-producing Escherichia coli. Conclusion: This work reveals the emergence of Extended Spectrum Beta-Lactamases-producing Escherichia coli in poultry farms in Yaoundé and the failure in the biosecurity system. As such, the awareness of poultry breeders on the respect of biosecurity measures may be an effective tool to tackle antimicrobial resistance, specifically in livestock industries using a One Health approach.

Unravelling Antimicrobial Resistance Phenotypes and Carriage of Extended-Spectrum β-Lactamase Genes in Escherichia coli Isolated from Dairy Farms in Kiambu County, Kenya

The use of antibiotics for prophylaxis and growth enhancement in livestock farming is on the increase globally. This practice has led to the emergence and spread of antimicrobial-resistant bacteria in livestock. Only limited research has been done to establish the role of cattle farming in antimicrobial resistance. The current study sought to establish the carriage of multi-drug resistance and extended-spectrum beta-lactamase genes in Escherichia coli from farmers, their cattle, and cattle slurry within Kiambu County. A total of 286 (81%) E. coli isolates were recovered from 352 samples analysed. Antibiotic resistance profiles showed 114 (40%) isolates were resistant to ≥3 antimicrobial classes and were considered multidrug-resistant. Among multidrug-resistant (MDR) E. coli strains, 40 (14%) were resistant to 3 different antimicrobial classes, while 71 (25%) were resistant to between 4 and 7 antibiotic classes. Extended-spectrum β-lactamase resistance was found in 18 isolates: human (n = 14), cattle (n = 2), and environmental (n = 2). Both the blaCTX-M and blaTEM genes were detected in 10 and 15 strains, respectively. Sequence analysis showed that the isolates carried the blaTEM-116 (n = 7), blaTEM-1 (n = 5), and blaCTX-M-15 (n = 8) genes. Genotyping MDR isolates using (GTG) 5 PCR demonstrated that the isolates were not clonal. This data shows antimicrobial resistance profiles and different types of resistance genes in the E. coli population on dairy farms. As a result, more effective, targeted public health policies and measures need to be put in place to control and prevent the emergence and spread of resistant bacteria.

Phenotypic and Genotypic Characterization of Metallo-Beta-Lactamase and Extended-Spectrum Beta-Lactamase among Enterobacteria Isolated at National Public Health Laboratory of Brazzaville

The improper use of antimicrobials against infectious diseases has allowed microorganisms to develop defense mechanisms that give them insensitivity to these agents. All bacteria are concerned by this phenomenon. This work aimed to assess prevalence of beta-lactamase produced by enterobacterial isolates. Then, disc diffusion, double disc synergy test (DDST) and combined disc test (CDT) were respectively used for antimicrobial resistance, detection of Extended-Spectrum Beta-Lactamases (ESBL) and Metallo-Beta-Lactamases (MBL). bla genes were detected by PCR. A total of 132 enterobacterial strains were studied. Resistance to antibiotic families was observed with a greater frequency than 50%. Gentamicin was the least active beta-lactam antibiotic, with a resistance rate of 88%. 40.9% of strains show an ESBL phenotype and 16.6% were MBL. An overall prevalence of 74% (40/54) and respectively rates of 29.6%, 27.7% and 16.7% for blaSHV, blaCTX and blaTEM genes were observed. SHV, CTX, CTX/SHV/TEM, CTX/TEM, SHV/TEM and CTX/SHV were different ESBL genotypes observed. ESBL-producing enterobacteria isolation worried about the future of antimicrobial therapy in the Republic of Congo. This is a public health problem that requires careful monitoring and implementation of a policy of rational antibiotics use.

Activity of Fosfomycin in Extended-Spectrum Beta-Lactamases Producing Klebsiella pneumonae from Hospital Acquired Urinary Tract Infections

Treatment of hospital acquired urinary tract infections (UTIs) caused by extended-spectrum beta-Lactamases producing Klebsiella pneumonae is a major problem. This organism expresses a high level of resistance to many groups of antibiotics. Fosfomycin is an agent which is recommended for treatment of UTIs caused by ESBLs producers. The aim of this study is to determine the sensitivity pattern of ESBLs producing urinary K. pneumonae to antimicrobial agents including fosfomycin in patients of MUHs and determine the prevalence of fosfomycin resistance mediated by plasmid mediated fosfomycin modifying enzymes fosA, fosB and fosA3. Methods: Klebsiella pneumonae urinary isolates were collected from patients with hospital acquired UTIs in Mansoura University Hospitals (MUHs). The susceptibility pattern was determined by Kirby Baur method. Isolates resistant to extended spectrum cephalosporins were tested for ESBLs production by double disc diffusion method. Fosfomycin resistance was determined by broth dilution method. Isolates resistant to fosfomycin were tested for fosA, fosB and fosA3 by PCR. Results: A total of 128 ESBLs producing K. pneumonae isolates were collected. The highest sensitivity was to imipenem (94.5%). The lowest was to trimethoprime-sulphamethoxazole (21.8%). Co-resistance of ESBLs isolates with fosfomycin was 23.2%. Eighteen fosfomycin resistant isolates (18/30) were positive to fosA. Conclusion: ESBLs producing urinary Klebsiella pneumonae express moderate sensitivity to fosfomycin. Resistance is mainly mediated by plasmid mediated fosfomycin modifying enzymes fosA.

Molecular Identification of Co-Existence of Carbapenemase and Extended-Spectrum <i>β</i>-Lactamase Genes in <i>Klebsiella pneumoniae</i>Clinical Isolates, and Their Phylogenetic Patterns in Kenya

The increasing incidence of multidrug-resistant Klebsiella pneumoniae strains has become a serious global healthcare problem. Additionally, the carriage of both extended-spectrum ß-lactamase and carbapenemase genes on plasmid and genomic DNA in K. pneumoniae clinical isolates has not been documented in Kenya. This study aimed to assess the presence of extended spectrum β-lactamase (ESBL) and carbapenemase genes on genomic and plasmid DNA in K. pneumoniae, and classify these super-bug clinical isolates based on their phylogenetic patterns. The identification of Klebsiella-like clinical isolates (n = 20) collected from Kenyatta National Hospital in Nairobi was performed using API 20E Kit. Screening and confirmation for ESBL and carbapenemase phenotypes were conducted using Kirby-Bauer disk diffusion susceptibility test protocol. Conventional PCR technique was used to characterize ESBL and carbapenemase resistant genes on both genomic and plasmid DNA. Subsequently, 16S rRNA gene amplification and sequencing were performed. The 16S rRNA gene contiguous sequences of the bacterial isolates were analyzed using the ChromasPro. The gene sequence was compared with the sequences in GenBank database, using the BLAST program of NCBI to obtain the nearest phylogenetic neighbours from the databases. Then, the sequences of MDR K. pneumoniae and its relatives were aligned using ClustalW. The evolutionary history was inferred by using the maximum likelihood algorithm in MEGA MX. The phenotypic data of antibiotic susceptibility testing revealed that 2/20 (10%) clinical isolates were resistant both to imipenem and meropenem and producers of carbapenemase. These isolates were carbapenemase producers but not extended β-lactamases. However, 3/20 (15%) isolates that co-harboured blaNDM-1, blaIMP, blaTEM, and bla-OXA were identified during genotypic analysis. The positive control used separately yielded the expected band sizes for blaIMP (275 bp), blaOXA-48 (438 bp), and BlaKPC (798). The phylogenetic analysis showed the dual ESBL and carbapenemase producing Klebsiella pneumoniae could be classified as K. pneumoniae strain DSM 30104 and K. pneumonia subsp. pneumoniae strain GMH1080. This study confirmed the co-existence of ESBL and carbapenemase genes in Klebsiella pneumoniae on both bacterial genomic and Plasmid DNA, and demonstrated that the isolates are evolutionarily distinct. These findings raise a concern about the genotypic diversity of antibiotic resistance genes in bacterial isolates and their location. We, therefore, recommend an alternative management approach to combat these MDR bacterial isolates as well as frequent molecular surveillance programs to support antimicrobial stewardship.

临床产ESBLs细菌的耐药特征和基因分型的研究

目的了解产超广谱β-内酰胺酶(ESBLs)细菌的流行和耐药特点,初步对产ESBLs细菌进行基因分型,确立耐药基因的位点和种类.方法收集分离鉴定菌株,通过K-B法药敏实验初筛以确认产ESBLs菌株,并对ESBLs基因进行初步分型.结果分离到产ESBLs细菌88株,检出率为38.9%,其中肺炎克雷伯菌阳性率为43.1%、大肠埃希菌阳性率为33.3%;产ESBLs细菌对青霉素类、氨曲南及头孢菌素类耐药率约为90%～100%,加用酶抑制剂克拉维酸或他唑巴坦后耐药率降为18.7%～46.8%;PCR初步分型结果表明:大多数产ESBLs细菌携带TEM型和SHV型β-内酰胺酶基因,其中单独携带TEM型基因的占42%,单独携带SHV型基因的占12.5%,携带两种基因的占31.8%.结论产ESBLs细菌具有多重耐药的特点;产ESBLs的大肠埃希菌和肺炎克雷伯菌大都携带TEM和(或)SHV型β-内酰胺酶基因,其中绝大部分产TEM型β-内酰胺酶.

产ESBLs鲍曼不动杆菌的耐药特性、质粒谱及耐药基因型

目的了解并研究产超广谱β-内酰胺酶(ESBLs)鲍曼不动杆菌的流行、耐药特点及质粒图谱,对产ESBLs菌株进行基因分型。方法先后用nitrocefin纸棒、最低抑菌浓度(M IC)初筛法和纸片扩散确认法从临床分离的鲍曼不动杆菌中检测产ESBLs菌株,用凝胶电泳分析其质粒图谱,并用聚合酶链反应(PCR)对产ESBLs基因进行分型。结果93株鲍曼不动杆菌中,对β-内酰胺酶类抗菌药物耐药47株(79.66%),中度敏感12株(20.34%);产ESBLs菌株有16株,占17.20%。产ESBLs鲍曼不动杆菌对头孢曲松、头孢他啶、头孢唑肟、头孢吡肟、氨曲南、庆大霉素、妥布霉素、磺胺甲唑、环丙沙星和阿米卡星的耐药率均显著高于非产ESBLs菌株(P<0.05)。保留的15株产ESBLs菌具有4种质粒谱,主要为Ⅰ型和Ⅲ型;它们均携带1～2种TEM型或SH V型或PER型β-内酰胺酶基因,且80%携带OXA-23型碳青霉烯酶基因。结论产ESBLs鲍曼不动杆菌常为多重耐药菌,耐药率与产ESBLs密切相关;同一克隆株在同一病房有流行趋势;本院流行株均携带2～3种耐药基因型,主要为TEM型、PER型β-内酰胺酶基因和OXA-23型碳青霉烯酶基因。