Efficacy Test of Prototype Kit for Detection Bacillus cereus and Listeria monocytogenes in Processed Meat using Real-time PCR Method

Authors

  • Muktiningsih Nurjayadi Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Tiara Fahriza Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Adinda Myra Amalia Putri Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Atikah Nur Rahmawati Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Ayu Berkahingrum Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Rosita Gio Anggraeni Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Gladys Indira Putri Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Jefferson Lynford Declan Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Dandy Akbar Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Irvan Maulana Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Maharanianska Azzahra Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Muhammad Arkent Shangkara Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Irma Ratna Kartika Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Fera Kurniadewi Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta
  • Dalia Sukmawati Research Center for Detection of Pathogenic Bacteria, Lembaga Penelitian dan Pengabdian Kepada Masyarakat, Universitas Negeri Jakarta
  • Sri Rahayu Research Center for Detection of Pathogenic Bacteria, Lembaga Penelitian dan Pengabdian Kepada Masyarakat, Universitas Negeri Jakarta
  • Vira Saamia Center Forensic Laboratory of the Criminal Investigation, Police of the Republic of Indonesia
  • I Made Wiranatha Center Forensic Laboratory of the Criminal Investigation, Police of the Republic of Indonesia
  • Bassam Abumoelak Arnold Palmer Hospital Pediatric Specialty Diagnostic Laboratory
  • Hesham El Enshasy Institute of Bioproduct Development, Universiti Teknologi Malaysia (UTM)

DOI:

https://doi.org/10.15408/jkv.v11i1.44280

Keywords:

Bacillus cereus, efficacy test, listeria monocytogenes, processed meat, real-time PCR

Abstract

According to the World Health Organization (WHO), harmful agents such as Bacillus cereus and Listeria monocytogenes are responsible for 600 million cases of disease and 420,000 deaths annually. This research aims to test the effectiveness of the real-time PCR method for developing a prototype kit to detect pathogenic bacteria in processed meat. As a comparison, the and conventional PCR methods were used to obtain the accuracy, specificity, sensitivity, and effectiveness of the real-time PCR method. All the samples were cultured in solid media agar, performed amplification using specific primers cyt-K 2 and hly using PCR and real-time PCR. Meatballs, nuggets, and sausages, five samples each, were found to be positive positively contaminated with all targeted bacteria. However, they did not provide specific results using solid media culture and the PCR method. In addition, the real-time PCR method using prototype kit formulas accomplished that all contaminated samples had a Ct value smaller than the negative control, NTC (No Template Control), and had a similar melting curve to the positive control. This establishes that the real-time PCR method clarifies that all samples were contaminated with target bacteria. A formula was developed in the prototype kit with real-time PCR methods that can be used and applied on a commercial scale efficiently and precisely.

Downloads

Download data is not yet available.

References

1. Grace D. Food safety Food safety is not salient in SDGs but may become more important. J Agric Nutr Heal ILRI. 2017;2015:1-11.

2. Shi C, Kang S. Foodborne Pathogenic Bacteria: Prevalence and Control—Volume I. Foods. 2024;13(10):1531. doi:10.3390/foods13101531

3. Alvarado PE, Barrios RMM, Xóchihua JAM, Hernández JFC. Fast and reliable DNA extraction protocol for products sold on the commercial market. Open Agric. 2017;2:469-472. https://www.degruyter.com/document/doi/10.1515/opag-2017-0051/html

4. Setiani BE, Pramono YB, Purwitasari L. Review on Pathogenic Bacteria Listeria monocytogenes, The Detection and The Sequencing Gene Methods Isolated from Meat Products. J Appl Food Technol. 2019;6(2):44-51. doi:10.17728/jaft.6460

5. Younes O, Ibrahim H, Hassan M, Amin R, Maarouf A. Demonstration of some food borne pathogens in different meat products: a comparison between conventional and innovative methods. Benha Vet Med J. 2019;36(2):219-228. doi:10.21608/bvmj.2019.12557.1008

6. BPOM. Batas Maksimal Cemaran Mikroba Dalam Pangan Olahan Produk. Badan Pengawas Obat dan Makanan. Published online 2019:1-48.

7. Atmanto Y, Asri L, Kadir N. Media Pertumbuhan Kuman. J Med Hutama. 2022;4(1):3072-3073. http://jurnalmedikahutama.com

8. Stingl K, Heise J, Thieck M, Wulsten IF, Pacholewicz E, Iwobi AN, Govindaswamy J, Zeller-Peronnet V, Scheuring S, Luu HQ, Fridriksdottir V, Golz G, Priller F, Gruntar I, Jorgensen F, Koene M, Kovac J, Lick S, Reperant E, Rohlfing A, Huber I. Challenging the “gold standard” of colony-forming units - Validation of a multiplex real-time PCR for quantification of viable Campylobacter spp. in meat rinses. Int J Food Microbiol. 2021;359:109417. doi:10.1016/j.ijfoodmicro.2021.109417

9. Kim DH, Chon JW, Kim H, Kim HS, Choi D, Kim YJ, Yim, JH, Moon JS, Seo KH. Comparison of Culture, Conventional and Real-time PCR Methods for Listeria monocytogenes in Foods. Korean J Food Sci Anim Resour. 2014;34(5):665-673. doi:10.5851/kosfa.2014.34.5.665

10. Widyastuti DA. Deteksi Molekuler Mikroorganisme Patogen pada Bahan Pangan dengan Metode RT-PCR. J Ilmu Pangan dan Has Pertan. 2017;1(1):54-63. doi:10.26877/jiphp.v1i1.1356

11. Nurjayadi M, Maulana I, Pramudiyasih NA, Kusumawati RN, Azzahra M, Liman NK, Sangkara MA, Wibisono E, Kurniadewi F, Saamia V, Saputro DAO, Wiranatha M, El-Enshasy HA. Isolation and PCR Validation of Bacillus cereus and Listeria monocytogenes Bacteria for the Development of Prototype Kit Detection of Foodborne Pathogens Diseases. AIP Conf. Proc. 2023: 2614, 030015. https://doi.org/10.1063/5.0126315

12. Charis M. Galanakis, ed. Sustainable Food Systems from Agriculture to Industry. Elsevier; 2018. doi:10.1016/C2016-0-02157-5

13. Raplong HH, Odeleye PO, Hammuel C, Idoko MO, Asanato JI, Odeke EH. Production of Alpha Amylase by Bacillus cereus in Submerged Fermentation. Aceh Int J Sci Technol. 2014;3(3):124-130. doi:10.13170/aijst.3.3.1592

14. FDA. BAM Chapter 14: Bacillus cereus.

15. Law JWF, Ab Mutalib NS, Chan KG, Lee LH. An insight into the isolation, enumeration, and molecular detection of Listeria monocytogenes in food. Front Microbiol. 2015;6. doi:10.3389/fmicb.2015.01227

16. Mira P, Yeh P, Hall BG. Estimating microbial population data from optical density. Omri A, ed. PLoS One. 2022;17(10):e0276040. doi:10.1371/journal.pone.0276040

17. Brian C M. Differences in bacterial optical density measurements between UV-Visible spectrophotometers. Thermo Sci. 2019;(Mic).

18. Azzahra M, Nurjayadi M, Pramudiyasih NA, Kusumawati RN, Maulana I, Declan JL, Putri GI, Juliansyah DA, Krisdawati I, Kurniadewi F, Kartika IR, Sukmawati D, Nastassya L, Saamia V, Oktaviani DA, Wiranatha SM, El-Enshasy HA, Abomoelak B. Primer concentration and Pre-denaturation Time Effect on cyt-K Bacillus cereus Detection using Real-Time PCR Method. E3S Web Conf. 2023;400:0-4. doi:10.1051/e3sconf/202340004011

19. Utaminingsih S, Sophian A. Analysis of Purity and Concentration of DNA Isolation Results on Chondroitin Samples. BiosciED J Biol Sci Educ. 2022;3(2):56-61. doi:10.37304/bed.v3i2.5425

20. Azzahra M. Validasi Formula Kit Deteksi Bakteri Patogen Bacillus Cereus & Listeria Monocytogenes Penyebab Keracunan Pangan dengan Real-Time Polymerase Chain Reaction. Universitas Negeri Jakarta; 2022.

Downloads

Published

31-05-2025

Issue

Section

Jurnal Kimia VALENSI, Volume 11, No. 1, May 2025

How to Cite

Efficacy Test of Prototype Kit for Detection Bacillus cereus and Listeria monocytogenes in Processed Meat using Real-time PCR Method. (2025). Jurnal Kimia Valensi, 11(1), 50-59. https://doi.org/10.15408/jkv.v11i1.44280