Browsing by Author "Yinur, Degisew"

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    Application and Evaluation of LAMP Assay for Detection of Enterohemorrhagic Escherichia Coli (O157:H7)
    (Addis Ababa University, 2019-02-05) Yinur, Degisew; Sisay, Tesfaye (PhD)
    O157:H7 E. coli are one of the enterohemorrhagic E. coli strains which cause bloody diarrhea and hemorrhagic colitis as well as hemolytic uremic syndrome in humans. This pathogen has been implicated in the worldwide outbreak of food and water borne diseases. Cost effective, sensitive rapid time detection of this pathogen is beneficial. The objective of the current study was to design and evaluate LAMP diagnostic assay for detection of EHEC O157:H7. In this study, putative fimbria protein coding gene (Z3276) was used to design a loop mediated isothermal amplification (LAMP) assay for the rapid and specific detection of EHEC O157:H7. A total number of 40 pathogenic and non-diarrheic E. coli as well as non E. coli bacteria were used to evaluate the assay. The LAMP amplified DNA samples were visualized as turbid DNA with naked eye as well as using gel electrophoresis and staining. The assay performed with 100% sensitivity, 97.05 % specificity, as well as 97.5% efficiency. The assay was also 10 times more sensitive than the conventional PCR assay; sensitivity evaluation was done through serial dilution. Additionally, the two assay results showed very high agreement (k = 0.97) on detection of the studied isolates. However, faecal samples spiked with the bacteria (DNA extracted by crude lysate) were not successfully detected by both LAMP assay and PCR method.
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    Application and Evaluation of LAMP Assay for Detection of Enterohemorrhagic Escherichia Coli (O157:H7)
    (Addis Ababa University, 2019-02-03) Yinur, Degisew; Sisay, Tesfaye (PhD)
    O157:H7 E. coli are one of the enterohemorrhagic E. coli strains which cause bloody diarrhea and hemorrhagic colitis as well as hemolytic uremic syndrome in humans. This pathogen has been implicated in the worldwide outbreak of food and water borne diseases. Cost effective, sensitive rapid time detection of this pathogen is beneficial. The objective of the current study was to design and evaluate LAMP diagnostic assay for detection of EHEC O157:H7. In this study, putative fimbria protein coding gene (Z3276) was used to design a loop mediated isothermal amplification (LAMP) assay for the rapid and specific detection of EHEC O157:H7. A total number of 40 pathogenic and non-diarrheic E. coli as well as non E. coli bacteria were used to evaluate the assay. The LAMP amplified DNA samples were visualized as turbid DNA with naked eye as well as using gel electrophoresis and staining. The assay performed with 100% sensitivity, 97.05 % specificity, as well as 97.5% efficiency. The assay was also 10 times more sensitive than the conventional PCR assay; sensitivity evaluation was done through serial dilution. Additionally, the two assay results showed very high agreement (k = 0.97) on detection of the studied isolates. However, faecal samples spiked with the bacteria (DNA extracted by crude lysate) were not successfully detected by both LAMP assay and PCR method.