Rapid Detection of Shigella flexneri on Egg Samples by the Real Time Polymerase Chain Reaction Method
Muktiningsih Nurjayadi1, Noer Azizah2, Ulfi Rahma Efrianti3, Fera Kurniadewi4, Dalia Sukmawati5, Vira Saamia6, I Made Wiranatha7, Lidwina Nastassya8
1Muktiningsih Nurjayadi, Associate Professor, Department of Chemistry, Universitas Negeri Jakarta, Indonesia.
2Noer Azizah, Student, Department of Chemistry, Universitas Negeri Jakarta, Indonesia.
3Ulfi Rahma Efrianti, Student, Department of Chemistry, Universitas Negeri Jakarta, Indonesia.
4Fera Kurniadewi, Senior Lecturer, Department of Chemistry, Universitas Negeri Jakarta, Indonesia.
5Dalia Sukmawati, Senior Lecturer, Department of Biology, Universitas Negeri Jakarta, Indonesia.
6Vira Saamia, Senior Researcher, Pusat Laboratorium Forensik Badan Reserse Kriminal Kepolisian RI, Indonesia.
7I Made Wiranatha, Senior Researcher, Pusat Laboratorium Forensik Badan Reserse Kriminal Kepolisian RI, Indonesia.
8Lidwina Nastassya, Senior Researcher, Balai Besar Uji Standar Karantina Pertanian, Jakarta, Indonesia.
Manuscript received on 10 April 2019 | Revised Manuscript received on 17 April 2019 | Manuscript Published on 02 June 2019 | PP: 166-172 | Volume-8 Issue-6C2 April 2019 | Retrieval Number: F10320486C219/19©BEIESP
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© The Authors. Blue Eyes Intelligence Engineering and Sciences Publication (BEIESP). This is an open-access article under the CC-BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)
Abstract: Shigella flexneri is one of the foodborne pathogen bacteria that caused food poisoning. Detection methods that are sensitive, specific and fast are very necessary in dealing with food poisoning, among these methods is Real Time PCR (RT-PCR). This study aims to apply the RT-PCR method to detect and quantify the Shigella flexneri bacteria in egg samples with the ipaH target gene. The amplicon of the target gene from the amplification process is 188 base pairs. Confirmation test of the ipaH primers pair with DNA template in concentration of Shigella flexneri culture of ±50 ng/-L gave the value of Cycle threshold (Ct) ±12. Primers sensitivity evaluation in detecting target bacteria gives the results that up to the smallest concentration of 8.05 pg/-L with a Ct value of 24.939. Specificity testing shows that the ipaH primers pair can differentiate Shigella flexneri bacteria significantly with some non-target bacteria as negative controls. Quantification of the number of bacteria found in egg samples using the flow of line equations by the RT-PCR method of 15.85 x 10-5 CFU/mL. These results provide more sufficiently information compared to the culture method. Based on the results, it can be concluded that the RT-PCR method was successfully applied in detecting Shigella flexneri bacteria with the target ipaH gene in egg samples quickly, sensitive and specific as well as can determine the number of bacteria accurately.
Keywords: ipaH Primers, Real Time PCR, Shigella Flexneri, Egg Sample.
Scope of the Article: Computational Biology