EurekaMag
+ Translate
+ Most Popular
Advantages and disadvantages of bordeaux mixture and of lime-sulphur used on apples in the growing season
Observations on the Umaria marine bed
10 years of hearing conservation in the Royal Air Force
Chocolate crumb - dairy ingredient for milk chocolate
Effect of daily gelatin ingestion on human scalp hair
Comparison of rice bran and maize bran as feeds for growing and fattening pigs
The composition of pampas-grass (Cortaderia argentea.)
The Accraian Series:
The mechanism of the Liebermann-Burchard reaction of sterols and triterpenes and their esters
Cerebrovascular Doppler ultrasound studies (cv-Doppler)
Toria: PT-303 - first national variety
Hair growth promoting activity of tridax procumbens
Productivity of Pekin x Khaki Campbell ducks
A stable cytosolic expression of VH antibody fragment directed against PVY NIa protein in transgenic potato plant confers partial protection against the virus
Solar treatment of wheat loose smut
Swimmers itch in the Lake of Garda
Bactofugation and the Bactotherm process
The effects of prefrontal lobotomy on aggressive behavior in dogs
Visual rating scales for screening whorl-stage corn for resistance to fall armyworm
Breakdown of seamounts at the trench axis, viewed from gravity anomaly
Kooken; pennsylvania's toughest cave
Recovery of new dinosaur and other fossils from the Early Cretaceous Arundel Clay facies (Potomac Group) of central Maryland, U.S.A
Zubor horny (Bison bonasus) v prirodnych podmienkach Slovensku
The extended Widal test in the diagnosis of fevers due to Salmonella infection
Hair of the american mastodon indicates an adaptation to a semi aquatic habitat

Detection of enterohemorrhagic Escherichia coli O157:H7 by using a multiplex real-time PCR assay for genes encoding intimin and Shiga toxins


Detection of enterohemorrhagic Escherichia coli O157:H7 by using a multiplex real-time PCR assay for genes encoding intimin and Shiga toxins



Veterinary Microbiology 93(3): 247-260



ISSN/ISBN: 0378-1135

PMID: 12695048

DOI: 10.1016/s0378-1135(03)00039-7

A multiplex real-time PCR (R-PCR) assay was designed and evaluated on the ABI 7700 sequence detection system (TaqMan) to detect enterohemorrhagic Escherichia coli (EHEC) O157:H7 in pure cultures, feces, and tissues. Three sets of primers and fluorogenic probes were used for amplification and real-time detection of a 106-bp region of the eae gene encoding EHEC O157:H7-specific intimin, and 150-bp and 200-bp segments of genes stx1 and stx2 encoding Shiga toxins 1 and 2, respectively. Analysis of 67 bacterial strains demonstrated that the R-PCR assay successfully distinguished EHEC O157:H7 serotype from non-O157 serotypes and provided accurate profiling of genes encoding intimin and Shiga toxins. Bacterial strains lacking these genes were not detected with this assay. The detection range of the R-PCR assay for the three genes was linear over DNA concentrations corresponding from 10(3) to 10(8)CFU/ml of EHEC O157:H7. The R-PCR allowed construction of standard curves that facilitated quantification of EHEC O157:H7 in feces and intestinal tissues. Detection sensitivity of the R-PCR assay ranged from 10(4) to 10(8)CFU/g of feces or tissues without enrichment. Enrichment of feces in a non-selective broth for 4 and 16h resulted in the detection of levels (from 10(0) to 10(3)CFU/g of feces) considered sufficient for infection in humans. The R-PCR assay for eae(O157:H7), stx1, and stx2 proved to be a rapid test for detection of EHEC O157:H7 in complex biological matrices and could also potentially be used for quantification of EHEC O157:H7 in foods or fecal samples.

Please choose payment method:






(PDF emailed within 0-6 h: $19.90)

Accession: 003704872

Download citation: RISBibTeXText

Related references

Detection of shiga-like toxin (stx1 and stx2), intimin (eaeA), and enterohemorrhagic Escherichia coli (EHEC) hemolysin (EHEC hlyA) genes in animal feces by multiplex PCR. Applied and Environmental Microbiology 65(2): 868-872, 1999

Real-time fluorescence PCR assays for detection and characterization of Shiga toxin, intimin, and enterohemolysin genes from Shiga toxin-producing Escherichia coli. Journal of Clinical Microbiology 40(7): 2555-2565, 2002

Development of a multiplex PCR assay for detection of Shiga toxin-producing Escherichia coli, enterohemorrhagic E. coli, and enteropathogenic E. coli strains. Frontiers in Cellular and Infection Microbiology 2: 8, 2012

Multiplex real-time PCR assay for detection of Escherichia coli O157:H7 and screening for non-O157 Shiga toxin-producing E. coli. Bmc Microbiology 17(1): 215, 2017

PCR detection of Shiga toxins, enterohaemolysin and intimin virulence genes of Escherichia coli O157:H7 strains isolated from faeces of Anatolian water buffaloes in Turkey. Zoonoses and Public Health 57(7-8): E33-E37, 2010

Development of a multiplex real-time polymerase chain reaction for simultaneous detection of enterohemorrhagic Escherichia coli and enteropathogenic Escherichia coli strains. Foodborne Pathogens and Disease 7(7): 801-808, 2010

A new pyrosequencing assay for rapid detection and genotyping of Shiga toxin, intimin and O157-specific rfbE genes of Escherichia coli. Journal of Microbiological Methods 109: 167-179, 2015

In vitro transcription inhibition of genes encoding shiga-like toxins and intimin protein in E coli O157H7 strains. Infection Genetics and Evolution 2(4): 295, 2003

Diagnosis of enterohemorrhagic Escherichia coli infection by detection of Shiga toxins. Clinical Microbiology Newsletter 26(7): 49-54, 2004

Detection in Escherichia coli of the genes encoding the major virulence factors, the genes defining the O157:H7 serotype, and components of the type 2 Shiga toxin family by multiplex PCR. Journal of Clinical Microbiology 40(10): 3613-3619, 2002

Detection of Shiga toxins, intimin and enterohemolysin in Escherichia coli strains isolated from children in eastern Slovakia. Folia Microbiologica 47(2): 185-188, 2002

Typing of intimin genes in human and animal enterohemorrhagic and enteropathogenic Escherichia coli: characterization of a new intimin variant. Infection and Immunity 68(1): 64-71, 2000

Rapid and reliable detection of Shiga toxin-producing Escherichia coli by real-time multiplex PCR. Journal of Food Protection 75(4): 643-650, 2012

A Four-Plex Real-Time PCR Assay, Based on rfbE, stx1, stx2, and eae Genes, for the Detection and Quantification of Shiga Toxin-Producing Escherichia coli O157 in Cattle Feces. Foodborne Pathogens and Disease 12(9): 787-794, 2015

Inhibition of the transcription of the Escherichia coli O157:H7 genes coding for shiga-like toxins and intimin, and its potential use in the treatment of human infection with the bacterium. Annals of Tropical Medicine and Parasitology 97(3): 281-287, 2003