A rapid and specific PCR–ELISA for detecting Salmonella typhi

Seyed Latif Mousavi; Jafar Salimiyan; Ahmad Karimi Rahgerdi; Jafar Amani; Shahram Nazarian; Hassan Ardestani

Archives of Clinical Infectious Diseases

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A rapid and specific PCR–ELISA for detecting Salmonella typhi

Author(s):

Seyed Latif Mousavi^1,*,

Jafar Salimiyan¹,

Ahmad Karimi Rahgerdi¹,

Jafar Amani¹,

Shahram Nazarian¹,

Hassan Ardestani¹

1Department of Biology, Institute of Basic Sciences, Imam-Hossein University, Tehran, Iran

Archives of Clinical Infectious Diseases:Vol. 1, issue 3; e93386

Published online:Jul 30, 2006

Article type:Research Article

Received:May 12, 2019

Accepted:Jul 18, 2006

How to Cite:Seyed Latif MousaviJafar SalimiyanAhmad Karimi RahgerdiJafar AmaniShahram NazarianHassan Ardestaniet al.A rapid and specific PCR–ELISA for detecting Salmonella typhi.Arch Clin Infect Dis.1(3):e93386.

Abstract

Background: Salmonella continues to be a major food borne pathogen for animals and humans. A sensitive and specific PCR–ELISA technique was developed to detect Salmonella typhi.

Materials and methods: The assay was based on the incorporation of digoxigenin-labeled dUTP and a biotin-labeled primers specific for rfbE gene during PCR amplification. The labeled PCR products were bound to streptoavidin-coated wells of a microtiter plate and detected by ELISA. The specificity of the PCR was determined using 4 strains of Entrobacteriaceae family including Escherichia coli, Klebsiella, Proteus, and Enterobacter and 2 strains of salmonella genus including paratyphi A and entritidis.

Results: Among all the strains, only Salmonella typhi was positive. The PCR-ELISA detecting system was able to increase the sensitivity of assay up to 100 fold, compared with a conventional PCR. The detection limit in PCR-ELISA was 2.5ρg in genomic DNA and 20 cells in direct manner per reaction. The entire procedure took about 100 minutes. For further confirmation of the test, internal biotin labeled probe was designed for rfbE gene and detected with streptavidin.

Conclusion: We have developed a rapid and simple PCR-ELISA protocol suitable for routine analysis of viable Salmonella typhi.

Keywords

References

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