Muktiningsih Nurjayadi, Dandy Akbar Juliansyah, Jefferson Lynford Declan, Gladys Indira Putri, Ismaya Krisdawati, Atikah Nur Rahmawati, Maharanianska Azzahra, Irvan Maulana, Gusti Angieta, Fera Kurniadewi, Irma Ratna Kartika, Vira Saamia, I. Made Wiranatha, Bassam Abomoelak, Hesham Ali El-Enshasy
Cronobacter sakazakii has been linked to cause meningitis, necrotizing enterocolitis, and sepsis in infants and newborns, with case fatality rates ranging from 40 to 80%. The most common source of infection has been identified as Cronobacter sakazakii-contaminated infant formula. With a relatively specific target hfq gene, this study aims to develop a real-time PCR method to identify Cronobacter sakazakii in infant formula milk. Real-time PCR is used as a detection method because rt-PCR has higher specificity and sensitivity compared to conventional PCR methods. The real-time PCR method also has a higher level of effectiveness and time efficiency compared to conventional PCR. Cronobacter sakazakii ATCC 29544 genomic DNA was isolated and used in a real-time PCR assay. Cronobacter sakazakii DNA was amplified using a primer targeting the hfq gene, yielding a 145 bp amplicon. The results of the real-time PCR test showed that Cronobacter sakazakii DNA with a concentration of 53 ng/µL could be amplified by the primer pairs of hfq gene with Ct values of 11 respectively then had Tm values of 81.7°C±0.5. The specificity test showed that the hfq primer pairs could differentiate between the target and some non-target bacteria. The sensitivity test showed the ability of the primer to detect the smallest concentration of 3.392 pg/µL with a Ct of 26.16. Based on the results obtained, it can be concluded that the hfq primer has the potential to be used as a fast detection method for Cronobacter sakazakii bacteria in infant formula using real-time PCR. © 2025@ author(s).
Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta, Gedung KH. Hasjim Asj'ari, 6thFloor, Jakarta Timur, 13220, Indonesia; Research Center for Detection of Pathogenic Bacteria, Lembaga Penelitian dan Pengabdian Kepada Masyarakat, Universitas Negeri Jakarta, Jakarta Timur, 13220, Indonesia; Center Forensic Laboratory of the Criminal Investigation, Police of the Republic of Indonesia, Cipambuan Babakan Madang, Bogor, 16869, Indonesia; Arnold Palmer Hospital Pediatric Specialty Diagnostic Laboratory, Orlando, 32806, FL, United States; Innovation Center in Agritechnology for Advanced Bioprocessing (ICA), Universiti Teknologi Malaysia (UTM), Johor, Pagoh, Malaysia; School of Chemical and Energy Engineering, Faculty of Engineering, Universiti Teknologi Malaysia (UTM), Johor Bahru, Skudai, Malaysia; City of Scientific Research and Technology Applications, New Burg Al Arab, Alexandria, Egypt
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