Related Experiment Video
Updated: Aug 16, 2026
Detection and Genogrouping of Noroviruses from Children's Stools By Taqman One-step RT-PCR
Published on: July 22, 2012
Real time PCR using TaqMan and SYBR Green for detection of Enterobacter sakazakii in infant formula
Yin Liu1, Xiaoning Cai, Xia Zhang
1Department of Biochemistry and Molecular Biology, College of Life Science, Nankai University, Tianjin 300071, China.
Insights
Rapidly detect Enterobacter sakazakii in infant formula using new real-time PCR assays. These methods are highly sensitive, specific, and faster than traditional tests, ensuring infant safety.
Area of Science:
- Microbiology
- Food Safety
- Molecular Biology
Background:
- Enterobacter sakazakii is an emerging pathogen causing severe infections in neonates.
- Infant formula contamination with E. sakazakii has led to outbreaks in vulnerable infants.
Purpose of the Study:
- To develop and validate rapid, sensitive real-time PCR assays for E. sakazakii detection.
- To improve upon traditional microbiological methods for pathogen identification in infant formula.
Main Methods:
- Development of two real-time PCR assays (TaqMan and SYBR Green) following selective enrichment.
- Validation using 35 E. sakazakii strains and 88 non-E. sakazakii strains.
- Novel primer design for SYBR Green assay supporting multiplexing.
Main Results:
- Both assays accurately identified all E. sakazakii strains and yielded negative results for non-E. sakazakii strains.
- Detection limit of 1.1 CFU/100 g infant formula achieved.
- Assays provide results within 2 business days, offering a significant time advantage over traditional methods.
Conclusions:
- The developed real-time PCR assays are sensitive, specific, and rapid for detecting E. sakazakii.
- These methods enhance food safety by enabling quicker identification of this critical pathogen in infant formula.
- The novel primer design offers flexibility for multiplex real-time PCR applications.
Abstract:
Enterobacter sakazakii is an emerging pathogen that causes meningitis, bacteremia, sepsis, and necrotizing enterocolitis in neonates and children. Powdered milk-based infant formulas have been associated with the E. sakazakii-related outbreaks in premature or other immunocompromised infants. In this study, we developed two real time PCR assays using TaqMan and SYBR Green to identify the pathogen after selective enrichment in mLST and BHI. The accuracy of two detections was tested by 35 strains of E. sakazakii and 88 non-E. sakazakii bacterial strains. The results showed that all of these E. sakazakii strains were positive reaction to the detections and all of the non-E. sakazakii strains were negative. The newly developed assays enable us to detect 1.1 CFU/100 g infant formula. And both of the assays can be accomplished within 2 business days. Compared to the traditional detection, the real time PCR procedures are quicker and simpler. In this study, we also developed a new method to design the primers, which can support multiple real time PCR with one pair of primers in SYBR Green detection. The detection methods are more sensitive and effective based on Two-Tm-Value of PCR.
