Comparison of Three Chromogenic Media and Evaluation of Two Molecular-Based Identification Systems for the Detection of Enterobacter sakazakii from Environmental Samples from Infant Formulae Factories

Abstract

Enterobacter sakazakii is an occasional contaminant of powdered infant formula that can cause rare but severe foodborne infections in infants. To determine optimal methods for the detection and identification of E. sakazakii, 38 naturally contaminated samples from infant formulae factories were analyzed by two PCR-based methods and by a method (TS 22964/RM 210) developed by the International Organization for Standardization and the International Dairy Federation (ISO-IDF) using three different commercial chromogenic agars. The ISO-IDF method includes two enrichment steps, plating of the second enrichment broth on E. sakazakii isolation agar (a chromogenic selective agar), picking of five typical colonies for transfer onto tryptone soy agar, and subsequent confirmation of yellow-pigmented colonies by biochemical characterization. Twenty-two of the 38 samples were positive by the culture method. E. sakazakii isolation agar (ESIA; AES Laboratoires), COMPASS agar (Biokar Diagnostics), and Druggan-Forsythe-Iversen agar (Oxoid) compared favorably with violet red bile glucose agar (VRBG, a selective medium for Enterobacteriaceae), with positive predictive values of 86.96, 88, and 74.07%, respectively, in contrast to 47.83% for VRBG. One additional positive sample was detected using the nonpatented real-time PCR method evaluated, and those results were in 97.3% concordance with the ISO-IDF results. Some discrepancies between the results of the DuPont Qualicon BAX system and those of the ISO-IDF method could be explained by heterogeneity of contamination and sampling. Thus, both PCR-based systems were suitable for detecting and specifically identifying E. sakazakii within 1 to 2 days, and COMPASS agar and ESIA could be used interchangeably as a first-step medium to isolate presumptive E. sakazakii colonies.

Detection, antibiotic susceptibility and biofilm formation of Cronobacter spp. from various foods in Korea

“…DNA was isolated using DNA tissue kits (Qiagen, Hilden, Germany) according to the manufacturer's instructions. The primers were targeted for the ompA gene and tRNA-23s rRNA intergenic region (Derzelle et al, 2007.;Mohan Nair et al, 2005), and synthesized commercially(-Bioneer, Daejon, Korea). The PCR products were analyzed using 1% Tris-acetate-EDTA buffer with 0.5 mg/ml of ethidium bromide.…”

Mining for sensitive and reliable species-specific primers for PCR for detection of Cronobacter sakazakii by a bioinformatics approach

“…Several targets have been described for PCR detection of Cronobacter genus, including 16S ribosomal (r)DNA (Iversen et al, 2004), 16S-23S rDNA internal transcribed spacer (Liu et al, 2006), 23S rDNA (Derzelle et al, 2007), transfer (t)RNAGlu (Hassan et al, 2007), dnaG (Seo and Brackett, 2005), ompA (Mohan Nair and Venkitanarayanan, 2006), gluA (Iversen et al, 2007), wehC, wehI , wzx (Jarvis et al, 2011), zpx (Jaradat et al, 2009), and others.…”

Cronobacter spp. in Powdered Infant Formula

“…2). Several studies have been conducted to compare the efficiency of various detection methods, and the best method is still a subject of debate (29,53).…”

“…The advantage of using CSB rather than other selective broths used in the enrichment steps of many culture-based detection methods is the fact that all Cronobacter strains can grow in this broth, thus solving the problem associated with some detection media (51). Although, a number of studies have been conducted to compare the effectiveness of various commercially available media, it is difficult to draw conclusions because of differences in the specific medium employed, the study methodology, sample sizes, and the ultimate findings (29,53).…”