scholarly journals Molecular diagnosis of E.coliO157:H7 Which Isolated from Children with Diarrhea by using Multiplex PCR

2010 ◽  
Vol 7 (1) ◽  
pp. 207-215
Author(s):  
Baghdad Science Journal
Keyword(s):  
Escherichia Coli ◽  
Diagnostic System ◽  
Latex Agglutination ◽  
Diagnostic Methods ◽  
Bacterial Isolates ◽  
Biochemical Tests ◽  
E Coli ◽  
Stool Samples ◽  
Polymerase Chain ◽  
Positive Results

A total of 96 stool samples were collected from children with bloody diarrhea from two hospitals in Baghdad. All samples were surveyed and examined for the presence of the Escherichia coli O157:H7 and differentiate it from other Non -Sorbitol Fermenting Escherichia coli (NSF E. coli). The Bacterial isolates were identifed by using morphological diagnostic methods, Samples were cultured on liquid enrichment medium, incubated at 37C? for 24 hrs, and then cultured on Cefixime Tellurite -Sorbitol MacConkey Agar (CT- SMAC). 32 non-sorbitol fermenting bacterial isolates were obtained of which 11 were identified as Escherichia coli by using traditional biochemical tests and API20E diagnostic system without differentiation between serotype O157:H7 and other NSF E. coli isolates . Four special biochemical tests were done for serotype O157:H7 differentiation from other NSF bacteria. Only 3 isolates belonging to the serotype O157:H7 were obtained . Latex agglutination test for O157 and H7 showed that the 3 isolates gave positive results with both tests. The Bacterial isolates were identifed by using Multiplex Polymerase Chain Reaction (MPCR) technology for the presence or absence of 4 genes (Stx1, Stx2, hlyA and eaeA) that encode for main virulence factors to diagnose E. coli O157:H7 isolated By using specific primers in MPCR . The result showed that one E. coli O157:H7 isolates contain all 4 genes , other isolates contain 3 genes: Stx2, hlyA & eaeA.

scholarly journals Isolation and characterization E. coli O157:H7 from meat &cheese and detection of Stx1, Stx2, hlyA, eaeA by using multiplex PCR

2011 ◽  
Vol 5 (2) ◽  
pp. 15-24
Author(s):  
Shatha T. Ahmed ◽  
Amina N. Jasim ◽  
Ayad M. Ali
Keyword(s):  
Escherichia Coli ◽  
Diagnostic System ◽  
Latex Agglutination ◽  
Diagnostic Methods ◽  
Bacterial Isolates ◽  
Biochemical Tests ◽  
E Coli ◽  
Isolation And Characterization ◽  
Minced Beef ◽  
Polymerase Chain

total of 189 samples were collected from 74 raw uncooked minced beef meat, 115 local white cheeses from 3 different areas in Baghdad. All samples were surveyed and examined for the presence of the Escherichia coli O157:H7 and differentiate it from other Non -Sorbitol Fermenting Escherichia coli (NSF E. coli). The Bacterial isolates were identified by using morphological diagnostic methods; Samples were cultured on liquid enrichment medium, incubated at 41.5Cº for 6 hrs, and then cultured on Cefixime Tellurite -Sorbitol MacConkey Agar (CT- SMAC). 66 non-sorbitol fermenting bacterial isolates were obtained of which 13 were identified as Escherichia coli from (6 meat and 7 cheese samples). By using traditional biochemical tests and Api20E diagnostic system without differentiation between serotype O157:H7 and other NSF E. coli isolates. Four specific biochemical tests (Cellobiose fermentation, β-Glocuronidase production, KCN and Enterohemolysin production) were done to differentiate serotype O157 differentiation from other NSF bacteria. Only 2 isolates belonging to the serotype O157 were obtained of which one isolate from meat and other isolate from cheese. Latex agglutination test for O157 and H7 showed that the 5 isolates gave positive results with both kits. The Bacterial isolates were identified by using Multiplex Polymerase Chain Reaction (MPCR) technology for the presence or absence of 4 genes (Stx1, Stx2, hlyA and eaeA) that encode for main virulence factors to diagnose E. coli O157:H7 isolated from various sources by using specific primers in mPCR. The result showed that gene content variety in two E. coli O157:H7 isolates, 1 from meat contain all 4 genes and other isolate from cheese contains 2 genes: Stx1 and hlyA .

An Improved Rapid Technique for Isolation of Escherichia coli O157:H7 from Foods

1995 ◽  
Vol 58 (1) ◽  
pp. 7-12 ◽  
Author(s):  
STEPHEN D. WEAGANT ◽  
JAMES L. BRYANT ◽  
KAREN G. JINNEMAN
Keyword(s):  
Escherichia Coli ◽  
Comparative Method ◽  
Immunomagnetic Separation ◽  
Latex Agglutination ◽  
Escherichia Coli O157 ◽  
Biochemical Tests ◽  
Isolation Technique ◽  
Selective Enrichment ◽  
E Coli ◽  
Polymerase Chain

A newly revised enrichment and agar-plating system was tested for selectivity and sensitivity in recovery of unstressed and cold-stressed Escherichia coli O157:H7 from foods. Various foods inoculated with known levels of enterohemorrhagic E. coli O157:H7 (EHEC) were tested by enrichment for 6 h at 37°C in modified tryptic soy broth (mTSB) base supplemented with vancomycin, cefsulodin and cefixime, referred to as EHEC enrichment broth (EEB). Subsequently, portions were spread-plated on sorbitol–MacConkey agar supplemented with tellurite and cefixime (TCSMAC). Further selective enrichment was also examined using immunomagnetic separation (IMS) from the EEB prior to spread-plating on TCSMAC agar. These methods were compared to a procedure of enrichment in mTSB (supplemented with novobiocin) at 37°C for 24 h followed by spread-plating of decimal dilutions on hemorrhagic colitis 4–methylumbelliferyl–B–D–glucuronide (HC–MUG) agar. The new enrichment isolation technique was found to be sensitive at a level of one EHEC organism per 10 g of food in four food types. This represents an approximate l00-fold to 1,000-fold enhancement in sensitivity over the comparative method for foods with high levels of competitive microflora. These enrichment-isolation protocols also were compared in analysis of naturally contaminated raw or undercooked ground beef samples implicated in foodborne illness. EEB-TCSMAC with and without IMS were combined with rapid biochemical tests, and with O157 latex agglutination and confirmation of toxin genes by polymerase chain reaction (PCR) to provide a completed test within 30 h of initiating testing. The new system was successful in 15 of 17 samples, where only 6 of 17 were found positive by the comparative technique.

scholarly journals Haemolytic uraemic syndrome and Shiga toxin-producing Escherichia coli infection in children in France

2000 ◽  
Vol 124 (2) ◽  
pp. 215-220 ◽  
Author(s):  
B. DECLUDT ◽  
P. BOUVET ◽  
P. MARIANI-KURKDJIAN ◽  
F. GRIMONT ◽  
P. A. D. GRIMONT ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Prospective Study ◽  
Shiga Toxin ◽  
Haemolytic Uraemic Syndrome ◽  
Serum Samples ◽  
E Coli ◽  
Escherichia Coli Infection ◽  
Stool Samples ◽  
Polymerase Chain

We conducted a study to determine the incidence of haemolytic uraemic syndrome (HUS) in children in France and to assess the role of Shiga-toxin-producing Escherichia coli (STEC) infection in the aetiology of HUS. In collaboration with the Société de Néphrologie Pédiatrique we undertook a retrospective review of all cases of HUS hospitalized from January 1993 to March 1995 and a 1-year prospective study (April 1995–March 1996) of epidemiological and microbiological features of cases of HUS. The polymerase chain reaction (PCR) procedure was used to detect stx, eae, e-hlyA genes directly from case stool samples. Serum samples from cases were examined for antibodies to lipopolysaccharide (LPS) of 26 major STEC serogroups. Two hundred and eighty-six cases were reported. The average incidence per year was 0·7/105 children < 15 years and 1·8/105 children < 5 years. During the prospective study, 122/130 cases were examined for evidence of STEC infection using PCR and/or serological assays and 105 (86%) had evidence of STEC infection. Serum antibodies to E. coli O157 LPS were detected in 79 (67%) cases tested. In conclusion, this study showed that STEC infection is an important cause of HUS in children in France, with a high proportion related to the O157 serogroup.

scholarly journals Modified CIM-Test As a Useful Tool to Detect Carbapenemase Activity Among Extensively Drug- Resistant Klebsiella Pneumoniae, Escherichia Coli and Acinetobacter Baumannii

2021 ◽  
Author(s):  
Abed Zahedi bialvaei ◽  
Alireza Dolatyar Dehkharghani ◽  
Farhad Asgari ◽  
Firouzeh Shamloo ◽  
Parisa Eslami ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Acinetobacter Baumannii ◽  
Sensitivity And Specificity ◽  
E Coli ◽  
Carbapenem Resistant ◽  
Polymerase Chain ◽  
Phenotypic Methods ◽  
Encoding Genes ◽  
Positive Results

Abstract Background Timely detection of carbapenemases is essential for developing strategies to control the spread of infections by carbapenem-resistant isolates. The purpose of our study was to determine the epidemiology of carbapenemase genes among carbapenem resistant isolates of Acinetobacter baumannii, Klebsiella pneumoniae and Escherichia coli and to compare efficacy of modified Hodge Test (MHT), Carba NP and modified carbapenem inactivation method (mCIM) tests. Methods A total of 122 carbapenem-resistant clinical isolates including 77 K. pneumoniae, 39 A. baumannii, and six E. coli were collected from hospitalized patients. Three phenotypic methods, including MHT, Carba NP test and mCIM were used for investigation of carbapenemase production. In addition, polymerase chain reaction (PCR) was performed to detect carbapenemase encoding genes. Results The sensitivity and specificity of the MHT were 75.0% and 100% respectively. In addition, CarbaNP displayed 80.8% sensitivity and 100% specificity, whereas the sensitivity and specificity were 90.4% and 100% for the mCIM test, respectively. Among carbapenem-resistant isolates, 70, 84 and 87 isolates exhibited positive results according to MHT, CarbaNP test and mCIM, respectively. PCR indicated the presence of one or more carbapenemase genes in 119 of carbapenem-resistant isolates, with blaKPC and blaVIM being the most commonly encountered. Co-production of ‘KPC and VIM’, ‘KPC and IMP’ and ‘KPC and OXA-48’ was detected in nine, seven and three isolates, respectively. Conclusion Our results confirm that the mCIM test is a useful tool for the reliable detection of carbapenemases-activity in enterobacterial isolates, especially in clinical microbiological laboratories with limited resources.

scholarly journals Prevalence, phylogeny, and antimicrobial resistance of Escherichia coli pathotypes isolated from children less than 5 years old with community acquired- diarrhea in Upper Egypt

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Rasha M. M. Khairy ◽  
Zahra Atef Fathy ◽  
Doaa Mohamed Mahrous ◽  
Ebtisam S. Mohamed ◽  
Soha S. Abdelrahim
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance Genes ◽  
E Coli ◽  
Extended Spectrum ◽  
Causative Agents ◽  
Typical Epec ◽  
Stool Samples ◽  
Polymerase Chain ◽  
Lactamase Gene ◽  
Children Under 5

Abstract Background Diarrhoea, affecting children in developing countries, is mainly caused by diarrheagenic Escherichia coli (DEC). This study principally aimed to determine the prevalence of DEC pathotypes and Extended-spectrum β-lactamase (ESBL) genes isolated from children under 5 years old with diarrhea. Methods A total of 320 diarrhoea stool samples were investigated. E. coli isolates were investigated for genes specific for enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC), enteroaggregative E. coli (EAEC), enteroinvasive E. coli (EIEC) and enterohemorrhagic E. coli (EHEC) using polymerase chain reaction (PCR). Furthermore, antimicrobial susceptibility testing, detection of antibiotic resistance-genes and phylogenetic typing were performed. Results Over all, DEC were isolated from 66/320 (20.6%) of the children with diarrhoea. EAEC was the predominant (47%), followed by typical EPEC (28.8%) and atypical EPEC (16.6%). Co-infection by EPEC and EAEC was detected in (7.6%) of isolates. However, ETEC, EIEC and EHEC were not detected. Phylogroup A (47%) and B2 (43.9%) were the predominant types. Multidrug-resistance (MDR) was found in 55% of DEC isolates. Extended-spectrum β-lactamase (ESBL) genes were detected in 24 isolates (24 blaTEM and 15 blaCTX-M-15). Only one isolate harbored AmpC β-lactamase gene (DHA gene). Conclusion The study concluded that, EAEC and EPEC are important causative agents of diarrhoea in children under 5 years. MDR among DEC has the potential to be a big concern.

Molecular detection of Escherichia coli (E. coli) from Diarrheal stool samples from children in Quetta, Balochistan, Pakistan.

2019 ◽  
Vol 2 (2) ◽  
pp. 27-31
Author(s):  
Tauseef M Asmat
Keyword(s):  
Escherichia Coli ◽  
Developing Countries ◽  
Disease Transmission ◽  
Causes Of Death ◽  
Growth Media ◽  
Huge Number ◽  
E Coli ◽  
Stool Samples ◽  
Polymerase Chain ◽  
Control Disease

Diarrhea is one of the major causes of death in children, particularly in developing countries. Rapid detection and treatment is necessary to control disease transmission in the community and thus limiting the huge number of death toll. The major cause of diarrhea in developing countries is Escherichia coli (E. coli).This study was aimed to isolate E. coli from diarrheal stool samples from children aged 05 months to 05 years visited/ hospitalized in Quetta due to acute/persistent diarrhea. Diarrheal stool samples from 200 children were collected from Lady Sandeman Hospital Quetta and cultured on nutrient agar and later transferred to E. coli specific growth media for initial detection. For further confirmation the colonies were subjected to polymerase chain reaction (PCR). The PCR results revealed that 44(22%) samples out of 200 samples were positive for E. coli. These results indicate a high proportion of E. coli infection among children suffering with diarrhea.

scholarly journals Modified CIM test as a useful tool to detect carbapenemase activity among extensively drug-resistant Klebsiella pneumoniae, Escherichia coli and Acinetobacter baumannii

2021 ◽  
Vol 71 (1) ◽  
Author(s):  
Abed Zahedi Bialvaei ◽  
Alireza Dolatyar Dehkharghani ◽  
Farhad Asgari ◽  
Firouzeh Shamloo ◽  
Parisa Eslami ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Klebsiella Pneumoniae ◽  
Acinetobacter Baumannii ◽  
Sensitivity And Specificity ◽  
E Coli ◽  
Carbapenem Resistant ◽  
Polymerase Chain ◽  
Phenotypic Methods ◽  
Encoding Genes ◽  
Positive Results

Abstract Purpose Timely detection of carbapenemases is essential for developing strategies to control the spread of infections by carbapenem-resistant isolates. The purpose of this study was to determine the epidemiology of carbapenemase genes among carbapenem-resistant isolates of Acinetobacter baumannii, Klebsiella pneumoniae, and Escherichia coli. In addition, the efficacy of the modified Hodge test (MHT), Carba NP test, and modified carbapenem inactivation method (mCIM) were compared. Methods A total of 122 carbapenem-resistant clinical isolates including 77 K. pneumoniae, 39 A. baumannii, and six E. coli were collected from hospitalized patients. Three phenotypic methods, including the MHT, Carba NP test, and mCIM were used for investigation of carbapenemase production. In addition, polymerase chain reaction (PCR) was performed to detect carbapenemase-encoding genes. Result The sensitivity and specificity of the MHT were 75.0% and 100%, respectively. In addition, Carba NP displayed 80.8% sensitivity and 100% specificity, whereas the sensitivity and specificity were 90.4% and 100% for the mCIM test, respectively. Among carbapenem-resistant isolates, 70, 84, and 87 isolates exhibited positive results according to the MHT, Carba NP test, and mCIM, respectively. PCR indicated the presence of one or more carbapenemase genes in 119 of carbapenem-resistant isolates, with blaKPC and blaVIM being the most commonly encountered. Co-production of ‘KPC and OXA-48’, ‘KPC and VIM’, and ‘KPC and IMP’ was detected in three, nine, and seven isolates, respectively. Conclusion Our results confirm that the mCIM test is a useful tool for the reliable detection of carbapenemase activity in enterobacterial isolates, especially in clinical microbiological laboratories with limited resources.

scholarly journals PHYLOGENETIC GROUP OF ESCHERICHIA COLI IN URINE OF CHILDREN ATTENDING A MATERNAL AND CHILD CENTRE, LAGOS

2020 ◽  
Vol 7 ◽  
pp. 76
Author(s):  
Anotu Mopelola Deji-Agboola ◽  
Esther Abieyuwa Utomwen ◽  
Olubunmi Adetokunbo Osinupebi
Keyword(s):  
Escherichia Coli ◽  
Bacterial Infections ◽  
Phylogenetic Group ◽  
Bacterial Isolates ◽  
Chain Reaction ◽  
Phylogenetic Groups ◽  
E Coli ◽  
Antibiotic Susceptibility Test ◽  
Polymerase Chain ◽  
Very High

Urinary tract infection (UTI) is one of the most common bacterial infections causing significant morbidity in children. This study determines the phylogenetic group of Escherichia coli  isolated from the urine of children attending Maternal and Child Centre,  Amuwo-Odofin, Lagos State. Clean voided midstream urine sample collected  from 215 children aged five years and below . The urine samples were culture on Mac Conkey and blood agar, bacterial 5counts greater than or equal to 1 x 10CFU/ml were regarded as positive for UTI. Identification of isolates and antibiotic susceptibility test was performed using standard methods. Polymerase Chain Reaction was used to classify the isolated E. coli into A, B1, B2 and D phylogenetic groups using presence or absence of ChuA, Yja A and TspE4C2. The prevalence of UTI was 51.2% with preponderance in male 62.7% aged 24 – 35months 75.0%. Staphylococcus aureus 19.1% followed by Pseudomonas aeruginosa 12.7% and Escherichia coli10.0% were common bacterial isolates. The isolates were highly resistant to Augmentin (71.8%) and Ampicillin 91.8%; the Escherichia coli were resistant to Augmentin 54.5% and Ampicillin 100%. The E. coli were classed into B1,A, and D phylogenetic groups with percentages of 54.5%27.3% and 18.2% respectively. , The prevalence of bacteria UTI among children in this study was very high, the isolates were highly resistant to the antibiotics tested, the E.coli belong to phylogenetic groups A, B1 and D and all were resistant to Ampicillin.

scholarly journals Isolation, Molecular Detection and Antimicrobial Susceptibility Profile of ‎Escherichia coli O157:H7 in Household - reared Small Ruminants in Zaria ‎Metropolis, Kaduna State, Nigeria

2020 ◽  
Vol 17 (4) ◽  
pp. 16-23
Author(s):  
R. O. Yakubu ◽  
M. K. Lawan ◽  
J. K. P. Kwaga ◽  
J. Kabir
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Susceptibility ◽  
Small Ruminants ◽  
Resistance Index ◽  
Latex Agglutination ◽  
Escherichia Coli O157 ◽  
Biochemical Tests ◽  
Cross Sectional ◽  
E Coli ◽  
Test Kit

Escherichia coli O157:H7 is a zoonotic enteric pathogen of public health significance worldwide. A cross-sectional study was carried out during which 384 faecal samples of household-reared small ruminants and water used in the various houses where the animals are reared were collected. The samples were enriched on tryptone soya broth and cultured on EMB and CT-SMAC to isolate E. coli and E. coli O157:H7 respectively; subjected to conventional biochemical tests and E. coliO157:H7 was confirmed using Wellcolex latex agglutination test kit. E. coli O157:H7 isolates were subjected to antimicrobial susceptibility test and multiplex PCR was carried out to detect the presence of virulence genes stx1, stx2, eaeA and hlyA. The results of the isolation showed isolation rate of E. coli O157:H7 of 4.69% (9/192), 0.52% (1/192) which were obtained from faeces and water samples respectively. The results of the characterisation showed that one of the E. coli O157:H7 isolated harboured the eaeA and hlyA genes but was negative for stx1 and stx2 genes. The highest number of isolates showed resistance to erythromycin (90.9%) while the least was to gentamicin (6.3%). About 97.7% (43/44) of the isolates had multiple antibiotic resistance index greater than 0.2. In conclusion, household-reared small ruminants in the study area were found to be reservoirs of E. coli O157:H7 and humans living within these households are at risk of infection. The multiple antibioticresistance recorded in this study suggests widespread use of antimicrobial drugs in the study area.

scholarly journals Molecular detection of a new pathotype enteroaggregative haemorrhagic Escherichia coli (EAHEC) in Indonesia, 2015

2020 ◽  
Author(s):  
Wahyu Setyarini ◽  
Dadik Raharjo ◽  
Radita Yuniar Arizandy ◽  
Zakaria Pamoengkas ◽  
Subijanto Marto Sudarmo ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Polymerase Chain Reaction Method ◽  
Unique Combination ◽  
Haemolytic Uremic Syndrome ◽  
E Coli ◽  
Virulence Potential ◽  
Uremic Syndrome ◽  
Stool Samples ◽  
Polymerase Chain ◽  
Enterohemorrhagic E.Coli

Enteroaggregative haemorrhagic Escherichia coli (E. Coli, EAHEC) has been identified as the agent responsible for one of the largest outbreaks of gastroenteritis and Haemolytic-uremic syndrome (HUS) that is transmitted through food in Germany in 2011. The hypervirulent pathotype has a unique combination of two pathogens namely enterohemorrhagic E.coli strain (EHEC) which produces shiga/verotoxin and enteroaggregative E.coli toxins (EAEC) which produces toxins similar to ST and hemolysin. The toxin produced by the EAHEC strain is a hybrid pathotype that combines the virulence potential of the EAEC and EHEC strains that will damage the microcirculation, cause vasculitis and other toxic effects. The purpose of this study was to determine the percentage of samples infected with enteroaggregative hemorrhagic E. coli bacteria (EAHEC) in pediatric diarrhea patients at DR. Soetomo Hospital, Surabaya, Indonesia, 2015. This study used PCR (Polymerase Chain Reaction) method to detect enteroaggregative E. coli strains (CVD432 and aaic genes) and enterohemorrhagic E.coli (eae gene).The results showed that 33 out of 40 (82,5%) stool samples examined were detected enteroaggregative E. coli (EAEC), 4 out of 40 (10%) enterohemorrhagic E. coli (EHEC) and 3 out of 40 (7,5%) enteroaggregative haemorrhagic E. coli bacteria (EAHEC) , which caused diarrhea in pediatric diarrhea patients at Dr. Soetomo General Hospital. The unique combination of genomic features of the Surabaya outbreak strain, containing characteristics from pathotypes EAEC and EHEC, suggested that it represents a new pathotype enteroaggregative haemorrhagic E. coli (EAHEC). It is expected that development of specific primer design and sequencing are needed to continue in this research.

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