Phenotypic Characterization of Extended Spectrum Beta-Lactamase, Class C Cephalosporinase and Carbapenemase-Producing Klebsiella Species Isolated from Patients Consulted at Four Yaounde-Based Hospitals

Background: Klebsiella spp. are bacteria of medical importance for their role in opportunistic infections which are often difficult to treat because of resistance to one or several antimicrobials. The aim of this study was to determine antimicrobial resistance due to Extended Spectrum Beta-lactamase (ESBL), Class C cephalosporinase (AmpC) and carbapenemase enzymes in Klebsiella spp. isolated from patients consulted at four hospitals. Methodology: The study was cross-sectional and descriptive. A total of 4190 non-repetitive patients specimens from 13 types of clinical specimens were analysed from February to November 2020. Two hundred and twenty-five (225) Klebsiella spp isolates were identified using API 20E and antimicrobial susceptibility testing done according to the Kirby Bauer disc diffusion method. ESBL and AmpC phenotypes were determined by the combination disc method and carbapenemases by double disc synergy method, referenced by EUCAST guidelines for the resistance testing. Results: The frequency of the species was Klebsiella pneumoniae (69%, 155/255), K. oxytoca (14%, 31/255), K. ozaenae (12%, 27/225) and K. rhinoscleromatis (5%, 11/225). Isolates were most resistant to sulphomethoxazole trimethoprim (84%, 189/225), cepaholosporins (80%, 180/225), and least resistant to carbapenems (10.7%, 24/225). Two K. oxytoca and one K. pneumoniae were resistant to all antibiotics tested. Klebsiella pneumoniae had the most multidrug resistant isolates (59.4%, 134/225). Most isolates (83.6%, 188/225) expressed at least one enzyme, while 63.6% (143/225) of the isolates expressed at least two enzymes. Some isolates were ESBL (71.6%, 161/225), carbapenemase (10.7%, 24/225) and AmpC (6.6%, 15/225) producers. Three carbapenemases (Klebsiella pneumoniae carbapenemase-KPC, Metallo-Beta Lactamase-MBL and OXA-48) were detected. Conclusion: These results revealed that resistance of Klebsiella spp. to cephalosporins is high and this may be exacerbated by co-expression of AmpC and carbapenemases aggravating associated patient morbidity and mortality. Monitoring of antimicrobial resistance of local strains is necessary for informed decisions on empirical treatment. .

Characterization of Extended Spectrum Beta-lactamase and Carbapenamase Producing Enterobacteriaceae Causing Urinary Tract Infection among Children in Kenya: A Cross-Sectional Study

Introduction: Enterobacteriaceae causing urinary tract infections (UTI) have developed resistance to the commonly used antibiotics due to emergence of Extended Spectrum Beta-Lactamases (ESBLs) and Carbapenamase producing Enterobactericeae which are a public health problem worldwide. This study aims to determine the prevalence and characterize ESBLs and carbapenamase producing Enterobactericeae. Method: A cross-sectional study was carried out in Gertrude’s Children’s Hospital, Nairobi. 238 urine samples were collected from patients with urinary symptoms attending the outpatient department within the period 2020-2021. The urine were examined macroscopically and microscopically. Identification and antimicrobial susceptibility testing were done using VITEK® 2 Compact system (BioMérieux). Double disc synergy test and modified hodge tests were done as confirmatory tests for ESBLs and Carbapenamase phenotypes respectively. Polymerase Chain Reaction was used for the detection of blaCTX-M, blaTEM, blaSHV, blaKPC and blaOXA-48 genes. Results: From the 238 children sampled the prevalence of UTI caused by Enterobactericeae was 22.3%. The Enterobacteriaceae species isolated were Escherichia coli (84.9%), Klebsiella pneumoniae (5.66%), Proteus mirabillis (5.66%), Enterobacter aerogenes (1.89%) and Morganella morganii (1.89%). The isolated species were resistant to ampicillin. Meropenem had the highest susceptibility. Only E. coli species had the ESBLs (26.4%) and carbapenamase (1.9%) phenotypes. 100% had BlaCTX-M while 50% had blaTEM resistant gene. There was a significant association (p Conclusion: Ampicillin resistance resulted to use of alternative drugs and Meropenem was the drug of choice where increased resistance to the recommended drugs was noted. Further research on resistant genes is recommended.

Characteristics of β-Lactamase Synthesis in E. coli and K. pneumanie Strains in Nosocomial Infections

Background: Recently micro-organisms that synthesize extended-spectrum β-lactamase (ESBLs) were increased. The peculiarities of ESBL synthesis of Escherichia coli and Klebsiella pneumoniae strains that cause nosocomial urinary tract infections, surgical site infections and pneumonia in surgical clinic were studied. ESBL synthesis were observed 38.9% of E. coli strains obtained from urine, 92.3% of strains obtained from surgical site infections, and 50% of strains obtained from sputum. ESBL synthesis were observed 37.5% of K. pneumoniae strains obtained from urine, 85.7% of strains obtained from surgical site infections, and 60% of strains obtained from sputum. Different levels of ESBL synthesize of E. coli and K. pneumoniae strains isolated from different pattern is discussed. Conclusion. ESBL synthesis is common in E. coli and K. pneumoniae strains, which cause nosocomial infections. The frequency of occurrence of ESBL s synthesis among of these strains depends on clinical forms of nosocomial infections.

Analysis of AmpC β-lactamase Gene in Pseudomonas aeruginosa

The gene and the amino acid sequence of the structural and regulatory region of the Pseudomonas aeruginosa with different resistance patterns were analyzed. Six strains with different resistance patterns were selected and the AmpC β-lactamase was identified. The objective gene fragment was amplified by colonies PCR. The sequences of the PCR-products were analyzed. The DNA sequence of the structural gene ampC and the regulatory genes ampR, ampD and ampE was detected. The 6 strains and the wild-type Pseudomonas aeruginosa are highly homogeneous in structural and regulatory region. Some new mutant points were found.

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.

Prevalence and Risk Factors of Penicillinase-Type β-Lactamase Producing Neisseria gonorrhoeae Isolated from Patients Attending Health-Facilities in Yaounde, Cameroon

Background and Objectives: Mitigation of antibiotic resistant Neisseria gonorrhoeae has become a priority due to considerable health and economical disabilities it generates. In order to tackle the emergence of resistant Neisseria gonorrhoeae, this study aimed to determine the prevalence and risk factors of penicillinase type β-lactamase-producing Neisseria gonorrheae among patients consulting for genital infectious disorders in two health-facilities in Yaounde, Cameroon. Materials and Method: A cross-sectional descriptive and analytical study was conducted over a 3-month period, from July 2nd to October 2nd, 2022. Vaginal and urethral secretions were collected. Biochemical identification tests were performed on colonies grown on chocolate agar + polyvitex using the Api NH gallery. The detection of penicillinases was equally performed using the API NH gallery and confirmed using the antimicrobial susceptibility testing. The Minimum Inhibitory Concentrations of some antibiotics were determined using the E-Test. Results: The results showed that out of the 198 patients sampled, 16 (8.08%) were positive for Neisseria gonorrhoeae, among which 13/16 (81.25%) were penicillinase-type β-lactamase producers. Antimicrobial susceptibility testing results showed high co-resistances to antibiotics, mainly ciprofloxacin (100%), nalidixic acid (92.31%) and azithromycin (84.62%). Moreover, high Minimum Inhibitory Concentrations of ceftriaxone (ranging from 6 to 24 mg/L) was observed toward Neisseria gonorrhoeae isolates. The risk factors of the carriage of penicillinase-type β-lactamase producing Neisseria gonorrhoeae identified were: a history of Sexually Transmitted infections (p = 0.01) and unprotected sexual intercourse (p = 0.01). Conclusion: The emergence of penicillinase-type β-lactamase producing Neisseria gonorrhoeae is increasing and the situation is becoming worrisome. The identified risk factors can constitute a basic outlook to tackle resistant Neisseria gonorrhoeae, and therefore sustain antibiotic stewardship.

Antimicrobial Susceptibility and Genotyping of Plasmid Mediated- Extended Spectrum β-lactamase, AmpC β-lactamase and Aminoglycoside Modifying Enzymes in Klebsiella pneumoniae

The antimicrobial susceptibility and genotyping of plasmid mediated-extended spectrum β-lactamase (ESBLs), AmpC β-lactamase (pAmpC) and aminoglycoside modifying enzymes (AMES) in Klebsiella pneumoniae isolated in Tianjin, China were investigated in the present study. Sixty strains of Klebsiella pneumoniae isolated from Huanhu Hospital were tested against 21 antimicrobial agents by Microscan MIC method, and the PCR and DNA sequencing were used to determine the genotypes of ESBLs, pAmpC and AMES. The result in antimicrobial susceptibility testing showed that all the 60 strains of bacteria tested were proved to be multi-resistant, and that of genotyping demonstrated different genotypes among these bacterial strains, in which 60 strains belonged to the TEM type; 25 strains to the CTX-M type and 54 strains to the DNA pAmpC type. The AMES genes were found in 59 strains, while the CTX-M type of ESBLs, DHA pAmpC type AMES genes were detected in 22 strains simultaneously. It concludes that the problem of drug-resistance of Klebsiella pneumoniae on hospital in Tianjin is a serious issue, and at least 3 kinds of β-lactamase and 3 AMES genes exist.

Evaluation of Disk Potentiation Test (DPT) and Double Disk Synergy Test (DDST) for The Detection of Metallo-β-Lactamases (MBLs) in Clinical Isolates of Bangladesh

Objective: Increasing the emergence of Metallo-β-lactamase (MBL) producing gram-negative bacteria and their dexterous horizontal transmission demands rapid and accurate detection. This study was conducted to determine a suitable method to promptly detect MBL-producing gram-negative bacteria. Methods: A total of 103 gram-negative bacteria were identified from various clinical samples at a tertiary care hospital in Dhaka city. MBL producers were detected by two phenotypic methods, the Disk Potentiation Test (DPT) and the Double Disk Synergy Test (DDST) based on β-lactam chelator combinations where EDTA/SMA has been used as an inhibitor and Imipenem, Ceftazidime as substrates. Results: 103 isolates which were identified as Escherichia coli spp, Klebsiella spp, Pseudomonas spp, Acinetobacter spp, Proteus spp, Providencia spp were found to be multidrug-resistant in antibiogram test. Isolates showed complete resistance (100%) to Imipenem, Meropenem, and Amoxiclav. The highest carbapenem-resistant etiological agents were Acinetobacter spp 40 (38.8%) followed by Pseudomonas spp 27 (26.2%), Klebsiella spp 26 (25.2%), Escherichia coli 8 (7.8%), Proteus spp 1 (1%) and Providencia spp 1 (1%). DPT method detected significantly (p = 0.000009) a higher number of MBL-producers (Imipenem with 0.5 M EDTA n = 61, 59.2% & Ceftazidime with 0.5 M EDTA n = 56, 54.4%) compared to the DDST method (Imipenem -0.5 M EDTA n = 43, 41.7%, Imipenem – SMA n = 38, 36.9% & Ceftazidime -0.5 M EDTA n = 15, 14.6%). Conclusion: Pieces of evidence suggest that DPT is a more sensitive method than DDST and could be recommended for identifying MBL-producing bacteria in Bangladeshi hospitals for the proper management of patients, to reduce time constraints and treatment costs.

Extended-spectrum β-lactamase controversies

G.A.Jacoby 教授毕业于美国哈佛医学院,为国际知名的传染病学、微生物学和免疫学专家,曾长期担任美国麻省总医院传染病科医师,现任麻省 Lahey 医学中心传染病实验室主任,中国感染与化疗杂志特邀编委。长期以来 Jacoby 教授对细菌耐药机制有深入研究,尤其对细菌(肺炎克雷伯菌、大肠埃希菌等)产生质粒介导的超广谱β内酰胺酶(ESBL)以及喹诺酮类的耐药机制研究。曾发表多篇有关上述专题的权威性论著。此次本刊特邀 Jacoby 教授撰写有关 ESBL 的新进展,文中阐述目前 ESBLs 除通常所指质粒介导β内酰胺酶中扩大了酶水解的底物谱导致细菌对头孢噻肟、头孢他啶、氨曲南等抗生素耐药外,还包括许多具有不同特点的β内酰胺酶,其中某些酶产自共生菌。除 ESBL 外,AmpC 酶和各种碳青霉烯β内酰胺酶也可导致细菌对上述抗生素耐药,因此临床微生物实验室准确检出和鉴别各种β内酰胺酶,对于临床选用适宜的抗茵药十分重要。美国 CLSI(过去称为 NCCLS)推荐采用两步法(筛选及确证)检测细菌产 ESBL;但有的学者认为测定抗茵药物对细菌的 MIC,如用药后 T>MIC 在50%以上即可达到满意疗效,无需检测细菌是否产 ESBL。目前认为碳青霉烯类抗生素治疗产 ESBL 菌感染的疗效最为满意,但由此可能引起细菌对该类药物耐药,值得关注。采用大剂量头孢吡肟或哌拉西林-三唑巴坦治疗产ESBL 菌感染是否有效尚有争论,临床上对β内酰胺类最为耐药的菌株往往对几乎所有现用抗菌药耐药。黏菌素(或多黏菌素 B)曾成功的用于治疗此种多重耐药菌感染。此外替莫西林(temocillin,一种对β内酰胺酶稳定的青霉素类)与替加环素(tigecvcline,为米诺环素衍生物)体外对产 ESBL 菌有抗菌作用,但尚无临床研究资料。

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.

L1 β-Lactamase催化反应机理研究

用混合量子力学和分子力学(QM/MM)方法和密度泛函理论讨论了L1β-Lactamase催化Nitrocefin水解的过程,研究结果表明,反应为多步反应:第一步亲核进攻反应为反应的决速步骤,并且伴随着酰胺键的断裂,第二步反应为质子迁移反应.同时讨论了金属锌在反应中的作用.

T4-like coliphage ΦKAZ14 virulent to pathogenic and extended spectrum β-lactamase-producing Escherichia coli of poultry origin

Dear Editor,Bacteriophages(otherwise called phages)are a type of virus that infect bacteria.This viral type has found useful applications in the control of bacterial pathogens in foods and food processing environments.In addition,phages may be useful to prevent colonization and shedding of bacteria into the surrounding environment.

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.

产AmpC酶的猪源奇异变形杆菌分离鉴定及生物学特性分析

奇异变形杆菌(Proteus mirabilis)是一种可导致多种动物感染的条件致病菌,目前有关产AmpC酶的动物源奇异变形杆菌研究较少。为探究广西某猪场母猪流产、死胎的原因,从流产猪胎肝脏中分离到病原菌,对分离株进行纯化培养、染色镜检、生化试验和16S rRNA测序确定为奇异变形杆菌,命名为GX-Y9251株。对GX-Y9251株进行药敏试验、耐药基因检测、AmpC酶基因检测、毒力基因检测、小鼠致病性试验和生物膜半定量检测。药敏试验结果显示,该菌株对8种药物敏感,对17种抗生素耐药。GX-Y9251株存在多种耐药基因,且产AmpC酶,为blaDHA基因。GX-Y9251株存在6个毒力基因,其对小鼠的半数致死量(LD50)为5.4×107CFU;具有生物被膜形成能力,为中等黏附型(++)。研究结果为奇异变形杆菌感染的防治提供了理论依据。

Comparative study on occurrence of class A and class Cβ-lactamase genes and their co-occurrence in Indian Enterobacteriaceae during years 2009 and 2010

Objective:To determine the occurrence of class A and class Cβ-Iactamase genes and their cooccurrence in Indian Enterobacteriaceae.Methods:52 third generation cephalosporin resistant isolates were phenotypically detected by combination disk method and screened by PCR to identify class A and class C typeβ-lactamase genes.Results:Of the 52 isolates,94.2%（49） were found harboring any of the bla?,blaCTX-M,blaSHV and blaTEM were present in 82.6%（43/52）, 59.6%（31/52）,and 42.3%（22/52） isolates,respectively.Of the 49 ESBL positive isolates 57.1% （28/49） showed co-occurrence of blaampCwith bla?.On the contrary,the collection from 2009 showed their co-occurrence in 81.4%isolates.Conclusions:The comparative study shows a downward trend for co-existence of bla? with blaampC from 2009 to 2010.Further large scale studies are needed to address the co-occurrence of class A and class Cβ-lactamases in India and the resistance trend occurring over a period of time.

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.

Comparative Study of Three β Lactamase Test Methods in <i>Staphylococcus</i>aureus Isolated from Two Nepalese Hospitals

Background: β lactamase is a plasmid-encoded enzyme that hydrolyzes β lactam ring of β lactam antibiotics rendering them ineffective. These enzymes, produced by Staphylococcus aureus along with many other organisms, have hindered the use of many useful and once life-saving β lactam antibiotics from clinical practice. Methods: This study was aimed to compare three test methods-chromogenic, acidimetric and iodometric-for the detection of β lactamase enzyme produced by 404 nosocomial induced S. aureus isolated from two Nepali hospitals, Kathmandu based hospital (KBH) and Lalitpur based Hospital (LBH). The study was carried out following standard methodology during November 2007 to June 2009 in the Department of Microbiology, Institute of Medicine, Kathmandu, Nepal. Sensitivity, specificity, efficiency, positive predictive value, and negative predictive values of the tests were calculated taking penicillin resistance and sensitivity as the standard. Results: Chromogenic method was found to be the most sensitive (98.93%) and efficient (98.51%) test and had a high positive predictive value (99.46%). Sensitivity (98.4%) and efficiency (98.27%) of iodometric method was found to be comparable to chromogenic test;its specificity (96.55 %) and positive predictive value (99.73%) were the highest among the 3 tests. Acidimetric test was the least sensitive (97.33%) and efficient (96.78%). Of note, the sensitivity and specificity of these test methods have been compromised due to the negativity of few penicillin resistant isolates and positivity of some penicillin sensitive isolates, respectively. Conclusion: Chromogenic method was found comparatively to be the best test method for the detection of β lactamase production. However, in contrast to the other two test methods whose reagents can be locally and economically prepared, chromogenic test’s use has been impeded by its cost and unavailability in the local Nepali market.

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.

Antimicrobial Resistance and β-Lactamase Production among Hospital Dumpsite Isolates

Metallo-β-Lactamases (MBLs) and Extended Spectrum β-Lactamses (ESBLs) have emerged world-wide as a significant source of β-lactam resistance. The emergence of MBLs and ESBLs encoded on plasmids among Gram-negative pathogens in hospital dumpsites was investigated. Soils of different government and private hospitals were collected and processed following standard bacteriological techniques. Antimicrobial susceptibility testing was carried out by the disk-diffusion technique using Ceftazidime (30 μg), Cefuroxime (30 μg), Cefotaxime (30 μg), Cefixime (5 μg), Trimethprim-sulfamethoxazole (25 μg), Gentamycin (100 μg) Amoxicillin-Clavunalate (30 μg), Ciprofloxacin (5 μg), Ofloxacin (5 μg), Nitrofurantoin (300 μg) and Imipenem (10 μg). The role of plasmids in resistance was evaluated by subjecting isolates to curing using Sodium Dodecyl Sulfate (SDS). ESBLs production by Double-Disk Synergy Test (DDST) was carried out. Isolates resistant to Imipenem were subjected to a confirmatory test using Modified Hodge’s test and to MBLs production by DDST. Eighty-two Gram-negative isolates comprising of 32 (39.02%) Escherichia coli, 20 (24.39%) Serratia marcescens, 14 (17.07%) Klebsiella pneumonia, 10 (12.28%) Proteus mirabilis and 6 (7.32%) Enterobacter aerogenes were obtained. Susceptibility results revealed a 100% resistance of all isolates to Ceftazidime, Cefuroxime, Cefixime, Amoxycillin-clavulanate and Cefotaxime. A total of 66 (80.48%) isolates harboured plasmids out of which 26 (31.71%) isolates were ESBL producers. MBLs production was observed in 8 (25.00%) E. coli, 2 (2.41%) Klebsiella pneumonia and 2 (2.41%) Proteus mirabilis isolates. All MBLs producing isolates were ESBLs producers. The finding of highly resistant isolates producing ESBLs and MBLs in a hospital environment is quite disturbing and should be addressed urgently.