DETECTION OF VIRULENCE GENES phoP AND phoQ IN Salmonella spp. USING IN SILICO POLYMERASE CHAIN REACTION

Authors

  • Stalis Norma Ethica Magister Study Program, Medical Laboratory Science, Universitas Muhammadiyah Semarang, Indonesia
  • Hayatun Fuad Magister Study Program, Medical Laboratory Science, Universitas Muhammadiyah Semarang, Indonesia
  • Nur Hidayah Magister Study Program, Medical Laboratory Science, Universitas Muhammadiyah Semarang, Indonesia
  • Sri Sinto Dewi Diploma 4 Study Program, Medical Laboratory Technology, Universitas Muhammadiyah Semarang, Indonesia
  • Aditya Rahman Ernanto Diploma 4 Study Program, Medical Laboratory Technology, Universitas Muhammadiyah Semarang, Indonesia
  • Ayu Rahmawati Sulistyaningtyas Diploma 3 Study Program, Medical Laboratory Technology, Universitas Muhammadiyah Semarang, Indonesia
  • Richard David Silvestre Medical Technology Study Program, St. Dominic College of Asia, Phillipines
  • Sri Darmawati Magister Study Program, Medical Laboratory Science, Universitas Muhammadiyah Semarang, Indonesia

DOI:

https://doi.org/10.33061/rsfu.v4i1.3414

Abstract

Detection of Salmonella bacteria based on their virulence genes is among essential steps in the eradication of clinical infection by bacteria. In this study, two pair of primers, PhoPF-PhoPR: 5’- CCGCGCAGGAAAAACTCAAA-3’ and 5’-ATCTGTTCCAGCATCACCGG -3’ as well as PhoQF-PhoQR: 5’-AGAGATGATGCGCGTACTGG-3’ and 5’- CAGACGCCCCATGAGAACAT-3’, had been successfully designed using Primer3Plus to detect the presence of phoP and phoQ genes in Salmonella spp. Using genomic DNA of 44 genomic data of Salmonella spp. as templates, PhoPF-PhoPR could produce 520-bp amplicon, while PhoQF-PhoQR could result in 598-bp amplicon. Results of in silico PCR showed that both pairs of primers PhoPF-PhoPR and PhoQF-PhoQR could detect only Salmonella enterica species, and no Salmonella bongori species could be detected based on phoP and phoQ sequences. Both pairs of PhoPF-PhoPR and PhoQF-PhoQR primers were also able to detect the virulence genes in most of the studied subspecies of Salmonella enterica available in silico database unless Arizona subspecies. As conclusion, based on this in silico study, phoP and phoQ genes appeared to be biomarkers for Salmonella enterica species. Both pairs of primers designed in this study has potential to be used as detection tool to differentiate species Salmonella enterica from Salmonella bongori, and also to distinguish S.enterica subsp. enterica from subsp. Arizonae.
Keywords: Gene detection, bacterial virulence, phoP, phoQ, Salmonella spp.

References

Andino A, Hanning I. Salmonella enterica: survival, colonization, and virulence differences among serovars. The Scientific World Journal. 2015;2015.

Bellemain, E., Carlsen, T., Brochmann, C., Coissac, E., Taberlet, P., & Kauserud, H. (2010). ITS as an environmental DNA barcode for fungi: an in silico approach reveals potential PCR biases. BMC microbiology, 10(1), 189.

Bikandi, J., San Millán, R., Rementeria, A., and Garaizar, J. 2004. In silico analysis of complete bacterial genomes: PCR, AFLP-PCR, and endonuclease restriction. Bioinformatics 20:798-9. DOI: 10.1093/bioinformatics/btg491

Chen Y, Ye W, Zhang Y, Xu Y. High speed BLASTN: an accelerated MegaBLAST search tool. Nucleic acids research. 2015 Aug 6;43(16):7762-8.

Choi E, Groisman EA, Shin D. Activated by different signals, the PhoP/PhoQ two-component system differentially regulates metal uptake. Journal of bacteriology. 2009 Dec 1;191(23):7174-81.

Ethica SN, Nataningtyas DR, Lestari P, Istini I, Semiarti E, Widada J, Raharjo TJ. Comparative evaluation of conventional versus rapid methods for amplifiable genomic DNA isolation of cultured Azospirillum sp. JG3. Indonesian Journal of Chemistry. 2013 Dec 18;13(3):248-53.

Ethica SN, Semiarti E, Widada J, Oedjijono O, Joko Raharjo T. Characterization of moaC and a nontarget gene fragments of food‐borne pathogen Alcaligenes sp. JG3 using degenerate colony and arbitrary PCRs. Journal of food safety. 2017 Nov;37(4):e12345.

Ethica SN, Sulistyaningtyas AR, Darmawati S. In-silico Specificity Comparison between GMF-GMR and JMF-JMR Primers for Detecting moaC Genes of Food Spoilage Bacteria Pseudomonas spp. In IOP Conference Series: Earth and Environmental Science 2019 Jun (Vol. 292, No. 1, p. 012033). IOP Publishing.

Ethica SN. Detection of genes involved in glycerol metabolism of Alcaligenes sp. JG3 (Doctoral dissertation, Universitas Gadjah Mada).

Gupta, S., Dongre, A., Saxena, J., & Jyoti, A. (2017). Computation and in silico validation of a real-time PCR array for quantitative detection of pathogens isolated from blood sample in sepsis patients.

Priyadharsini, J. V., Girija, A. S., & Paramasivam, A. (2018). In silico analysis of virulence genes in an emerging dental pathogen A. baumannii and related species. Archives of oral biology, 94, 93-98.

Raharjo TJ, Rizki RA, Ethica SN, Rustanti E, Nugroho LH. Characterization of 0.58 kb DNA Stilbene Synthase Encoding Gene Fragment from Melinjo Plant (Gnetum gnemon). Indonesian Journal of Chemistry. 2011 Dec 20;11(3):246-52.

Relman DA. Detection and identification of previously unrecognized microbial pathogens. Emerging infectious diseases. 1998 Jul;4(3):382.

Downloads

PlumX Metrics

Published

2020-01-11

Issue

Vol. 4 No. 1 (2020): RESEARCH FAIR UNISRI

Section

Artikel

License

  1. Hak publikasi atas semua materi informasi yang tercantum dalam situs jurnal ini dipegang oleh dewan redaksi/editor dengan sepengetahuan penulis. Pengelola Jurnal akan menjunjung tinggi hak moral penulis.
  2. Aspek legal formal terhadap akses setiap informasi dan artikel yang tercantum dalam situs jurnal ini mengacu pada ketentuan lisensi Creative Commons Atribusi-NonCommercial-No Derivative (CC BY-NC-ND), yang berarti bahwa hanya dengan izin penulis, informasi dan artikel Jurnal PKM dapat didistribusikan ke pihak lain dengan tanpa merubah bentuk aslinya untuk tujuan non-komersial.
  3. Setiap terbitan Jurnal PKM, baik cetak maupun elektronik, bersifat open access untuk tujuan pendidikan, penelitian, dan perpustakaan. Di luar tujuan tersebut, penerbit atau pengelola jurnal tidak bertanggung jawab atas terjadinya pelanggaran hak cipta yang dilakukan oleh pembaca atau pengakses.