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A selective differential medium for Enterobacter sakazakii, a preliminary study
Carol Iversen1, Patrick Druggan, Stephen Forsythe
1School of Science, The Nottingham Trent University, Clifton Lane, Nottingham, NG11 8NS, UK.
Abstract:
Enterobacter sakazakii can cause fatal invasive infection of neonates associated with the presence of this organism in powdered infant milk formula. A new chromogenic medium (Druggan-Forsythe-Iversen agar, DFI) is described for the selective detection of this emergent pathogen. The medium is based on the alpha-glucosidase reaction which is detected using 5-bromo-4-chloro-3-indolyl-alpha,D-glucopyranoside (XalphaGlc). Ent. sakazakii hydrolyses this substrate to an indigo pigment, producing blue-green colonies on this medium. DFI was compared with the current method of detection on violet red bile glucose agar (VRBGA) followed by pigment production on tryptone soy agar (TSA) after 48-72 h at 25 degrees C and subsequent biochemical profile determination using Biomerieux API20E. Ninety-five clinical and food strains of Ent. sakazakii were detected on the DFI chromogenic medium 2 days sooner than the alternative method. The characteristics of 148 strains representing 17 genera of non-Ent. sakazakii Enterobacteriaceae were compared using the two methods. Only 16/18 Escherichia vulneris strains, 2/3 strains of Pantoea spp. and 1/8 Citrobacter koseri strains gave false positive results on DFI agar. Eight alpha-glucosidase positive strains were identified as Pantoea using their API20E biochemical profile, but had higher percentage identification as Ent. sakazakii using ID32E. Therefore the DFI medium enables the detection of Ent. sakazakii within mixed cultures of Enterobacteriaceae, whereas the organism could be missed when using VRBGA since the latter is a general Enterobacteriaceae selective medium. In addition, the common use of API20E to check yellow pigmented colonies on TSA may lead to false negative results and consequently the acceptance of a batch of infant formula milk (IFM) that contains Ent. sakazakii.
Insights
A new chromogenic medium, Druggan-Forsythe-Iversen (DFI) agar, selectively detects the pathogen Enterobacter sakazakii in infant formula. This method identifies the bacteria faster and more accurately than current standards, improving infant safety.
Area of Science:
- Microbiology
- Food Safety
- Clinical Diagnostics
Background:
- Enterobacter sakazakii poses a fatal risk to neonates, particularly through contaminated powdered infant milk formula.
- Current detection methods for Enterobacter sakazakii are time-consuming and may lead to false negatives, endangering infant health.
- Selective and rapid detection of this pathogen is crucial for ensuring the safety of infant nutrition products.
Purpose of the Study:
- To introduce and evaluate a novel chromogenic medium, Druggan-Forsythe-Iversen (DFI) agar, for the selective detection of Enterobacter sakazakii.
- To compare the efficacy of DFI agar against the conventional method using violet red bile glucose agar (VRBGA) and biochemical profiling.
- To assess the specificity and sensitivity of DFI agar in identifying Enterobacter sakazakii in clinical and food samples.
Main Methods:
- Development of DFI agar utilizing the alpha-glucosidase reaction with 5-bromo-4-chloro-3-indolyl-alpha,D-glucopyranoside (XalphaGlc) for visual detection.
- Comparison of DFI agar with VRBGA followed by tryptone soy agar (TSA) and Biomerieux API20E for 95 clinical and food strains of Enterobacter sakazakii.
- Evaluation of DFI agar's performance against 148 strains from 17 non-Enterobacter sakazakii Enterobacteriaceae genera.
Main Results:
- DFI agar detected 95 strains of Enterobacter sakazakii two days faster than the conventional VRBGA method.
- DFI agar demonstrated high specificity, with only minor false positive rates among specific non-Enterobacter sakazakii strains (e.g., Escherichia vulneris, Pantoea spp., Citrobacter koseri).
- The DFI medium effectively differentiates Enterobacter sakazakii in mixed cultures, unlike VRBGA, and avoids potential false negatives associated with API20E analysis.
Conclusions:
- The DFI chromogenic medium provides a more rapid and reliable method for the selective detection of Enterobacter sakazakii compared to existing techniques.
- This improved detection capability is vital for preventing outbreaks of neonatal infections linked to contaminated infant formula.
- DFI agar enhances food safety protocols by offering a superior tool for identifying this critical foodborne pathogen.
