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
Rapid, specific detection of Enterobacter sakazakii in infant formula using a real-time PCR assay
1Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Plant and Dairy, Foods, and Beverages, College Park, Maryland 20740, USA. kseo@cfsan.fda.gov
Insights
A new real-time PCR assay can rapidly detect Enterobacter sakazakii in infant formula. This method offers a faster and more specific alternative to traditional culture methods, improving food safety for vulnerable infants.
Area of Science:
- Microbiology
- Food Safety
- Molecular Biology
Background:
- Enterobacter sakazakii causes severe neonatal infections, often linked to contaminated infant formula.
- Current detection methods are time-consuming, involving multiple culture and biochemical steps.
Purpose of the Study:
- To develop a rapid and specific assay for detecting Enterobacter sakazakii in infant formula.
- To improve the safety screening of infant formula for regulatory agencies and the food industry.
Main Methods:
- Designed a fluorogenic 5' nuclease (TaqMan) real-time PCR assay targeting the E. sakazakii rpsU and dnaG genes.
- Validated assay specificity against 68 Enterobacter and 55 non-Enterobacter strains.
- Determined assay sensitivity at 100 CFU/ml in pure culture and reconstituted infant formula.
Main Results:
- The real-time PCR assay demonstrated high specificity, accurately distinguishing E. sakazakii from other tested strains.
- Achieved detection of 100 CFU/ml without enrichment in 50 PCR cycles.
- The assay significantly reduces detection time by up to 5 days compared to conventional methods.
Conclusions:
- The developed real-time PCR assay provides a rapid, specific, and efficient method for screening infant formula for Enterobacter sakazakii.
- This assay can enhance food safety protocols, benefiting the food industry and regulatory bodies.
- Accelerated detection minimizes risks associated with E. sakazakii contamination in infant nutrition.
Abstract:
Enterobacter sakazakii is a rare cause of invasive infection with high mortality rates in neonates. Powdered milk-based infant formulas have been associated with the E. sakazakii-related outbreaks in premature or other immunocompromised infants. In this study, an assay was developed for the specific detection of E. sakazakii in infant formula using an application of the fluorogenic 5' nuclease assay (TaqMan). A set of primers and probe was designed using the E. sakazakii partial macromolecular synthesis operon: the rpsU gene 3' end and the primase (dnaG) gene 5' end. The specificity of the assay was evaluated using 68 Enterobacter and 55 non-Enterobacter strains. The newly developed assay enables us to detect 100 CFU/ ml in pure culture and in reconstituted infant formula in 50 cycles of PCR without enrichment. The assay was specific enough to discriminate E. sakazakii from all other Enterobacter and non-Enterobacter strains tested. The developed real-time PCR assay could save up to 5 days and eliminate the need for plating samples on selective or diagnostic agars and for biochemical confirmation steps. The real-time PCR assay could be used to rapidly screen infant formula samples for E. sakazakii and would be a boon to food industries and regulatory agencies.
