The Potential of grxB Gene for Detection of C. sakazakii in Infant Formula Milk Using Real-Time Polymerase Chain Reaction

¹ Department of Chemistry, Faculty of Mathematics and Natural Science, Universitas Negeri Jakarta, Gedung KH. Hasjim Asj’ari, 6th Floor, Jl. Rawamangun Muka, Jakarta Timur, 13220, Indonesia
² Research Center for Detection of Pathogenic Bacteria, Lembaga Penelitian dan Pengabdian Kepada Masyarakat, Universitas Negeri Jakarta, Jl. Rawamangun Muka, Jakarta Timur 13220, Indonesia
³ Center Forensic Laboratory of the Criminal Investigation, Police of the Republic of Indonesia, Cipambuan Babakan Madang, Bogor, 1681, Indonesia
⁴ Institute of Bioproduct Development, Universiti Teknologi Malaysia (UTM), Skudai, Johor Barhru, Malaysia.
⁵ School of Chemical and Energy Engineering, Faculty of Engineering, Universiti Teknologi Malaysia (UTM), Skudai, Johor Bharu, Malaysia.
⁶ City of Scientific Research and Technology Applications, New Burg Al Arab, Alexandria, Egypt

^* Corresponding author: muktiningsih@unj.ac.id

Abstract

Cronobacter sakazakii is one of the bacteria that causes food poisoning that contaminates infant formula. This pathogen causes necrotizing enterocolitis, sepsis, and meningitis in infants or neonates with reported case fatality rates ranging from 40% to 80%. Therefore, it is necessary to develop fast and accurate detection of C. sakazakii in infant formula milk. This research aims to develop a method for detecting C. sakazakii bacteria using real-time PCR with high sensitivity, specificity, and accuracy. A rapid detection method using real-time PCR with the target gene grxB successfully detects the presence of C. sakazakii DNA in artificially contaminated formula milk. The results of the real-time PCR test showed that C. sakazakii DNA with a concentration of 53 ng/µL could be amplified by the grxB gene primer pair with a Ct value of 12 and a Tm value of 85.8ºC. The specificity test showed that the grxB primer could differentiate between target and some non-target bacteria. The sensitivity test showed the ability of the grxB primer to detect the smallest concentration of 3,392 pg/µL with a Ct of 24,06. Based on the results obtained, it can be concluded that the grxB primer has the potential to be used as rapid detection method for C. sakazakii bacteria in infant formula using real-time PCR.