Detection of toxigenic bacillus cereus strains in powdered infant formula (PIF) milk by PCR assay

Abstract

Background: In recent years, use of powdered infant formula (PIF) milk for neonates feed is increasing; therefore, the quality control (QC) of PIF products is very important. The aim of present study was detection of toxigenic Bacillus cereus species in PIF milk using PCR assay. Methods: The cross-sectional study was carried out on 125 samples of powdered infant formula milk (PIF) purchased between March 2015 and April 2016 in Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran. Briefly, 0.1 dilutions were prepared and inoculated on Bacillus cereus selective media (MYP) and incubated at 30 °C for 24 hours. The suspicious colonies were verified using biochemical tests based on standard methods. Final confirmation of studied isolates was carried out by ITS gene detection using polymerase chain reaction (PCR) assay. Presence of nonhemolytic enterotoxin (NHE) (linked to diarrhoea syndrome) and emetic toxin (EM) (linked to emetic syndrome) virulence genes were investigated using polymerase chain reaction assay.  Results: In this study, of 125 PIF samples, 84 (67.2%) were contaminated. Of various recovered bacteria from these samples, 110 bacterial isolates were suspected to be Bacillus spp. using phenotypic methods. The ITS PCR results showed that 91.8% of the isolates were B. cereus. Respectively, 53.63 and 79% of B. cereus isolates possessed NHE and EM virulence genes. Conclusion: Our data revealed that near 80% of Bacillus cereus isolates have emetic toxin (EM) gene, as result virulence potency of this isolates is very high. However, the low number of this organisms in foods is very important and food safety protocols for these opportunistic toxigenic bacteria should be revised. Since the pasteurization process is ineffective on B. cereus spores; therefore, spores can remain in PIF milk and the vegetative bacterial cells can cause food poisoning in neonates. Therefore, modification of foods quality control protocols is essential in order to identify virulence genes in this bacterium