A medium for the presumptive detection of Enterobacter sakazakii in infant formula: interlaboratory study.
A standard method for the detection of Enterobacteriaceae was modified for the presumptive detection of Enterobacter sakazakii, and the modified method was validated in an interlaboratory trial with 16 laboratories from 8 European countries. The modification included a differential-elective medium for the isolation of E. sakazakii, consisting of nutrient agar (NA) supplemented with 4-methyl-umbelliferyl alpha-D-glucoside (alpha-MUG). A 25 g sample was added to 225 mL buffered peptone water. After incubation at 35 degrees or 37 degrees C for 16 or 20 h, 10 mL nonselective enrichment was transferred into 90 mL selective enrichment. The selective enrichment was streaked on violet-red bile glucose agar (VRBGA) and incubated at 37 degrees C for 24 h. It was streaked in parallel on NA plates supplemented with alpha-MUG at 50 mg/L and incubated at 25 degrees C for 16 h, and afterwards for an additional 24 h at room temperature in the dark. E. sakazakii appeared as vivid yellow colonies under normal light and showed blue/violet fluorescence under UV light on NA + alpha-MUG plates. Validation samples represented powdered infant formula without E. sakazakii (blanks) and with low (1-10 colony-forming units [CFU]/25 g) and medium (1-10 CFU/g) contamination levels. All samples contained Pseudomonas aeruginosa and Lactobacillus spp. as background flora. The specificity for blank samples was 100%. The sensitivity of the low contamination level was similar for VRBGA and NA + alpha-MUG, i.e., 66.7% (66.7% accordance, 53.9% concordance). For the medium level the sensitivities were 96.7% (93.3% accordance, 93.5% concordance) for VRBGA and 98.3% (96.9% accordance, 96.9% concordance) for NA + alpha-MUG.
Central Science Laboratory, Department for Environment, Food and Rural Affairs, Sand Hutton, York YO41 1LZ, United Kingdom. renata.leuschner@cec.eu.int
Renata G K
RG
Leuschner
Jan
J
Bew
D002648:Child; D003470:Culture Media; D004754:Enterobacter / Q000737:chemistry*; D004755:Enterobacteriaceae / Q000737:chemistry; D005060:Europe; D005612:Freeze Drying; D005960:Glucosides / Q000737:chemistry; D006801:Humans; D006923:Hymecromone / Q000031:analogs & derivatives* / Q000737:chemistry; D007202:Indicators and Reagents; D007223:Infant; D007225:Infant Food / Q000382:microbiology*; D007778:Lactobacillus / Q000737:chemistry; D012015:Reference Standards; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence
D003470:Culture Media; D005960:Glucosides; D007202:Indicators and Reagents; C014110:4-methylumbelliferyl glucoside; D006923:Hymecromone