A recent technique for the detection of Bacillus cereus in milk

Abstract

The occurrence of Bacillus cereus in milk and dairy products is attributed to deficient hygiene protocols, consequently resulting in two gastrointestinal illnesses: rare emetic sickness and diarrheal illness triggered by enterotoxins. The current study aimed to overcome the limitations of PCR in detecting low bacterial concentrations through the incorporation of unmodified gold nanoparticles (GNPs) in polymerase Chain Reaction (PCR). Bacillus cereus in milk was analyzed using the GNPs-assisted PCR method to detect the nheA gene in comparison to conventional PCR and SYBRGreen qPCR. The nheA gene is a key virulence factor that encodes a protein involved in the generation of non-hemolytic enterotoxins. The result showed adding GNPs to PCR reactions boosted DNA yields and enabled detection down to 10 2 copies of Bacillus cereus DNA, versus 10 3 with standard PCR. GNPs-assisted PCR detected Bacillus cereus for spiked milk samples at 10 2 cfu/ml, while conventional PCR required 10 3 cfu/ml. SYBRGreen qPCR also achieved detection at 10 2 DNA copies and 10 2 cfu/ml for spiked milk. GNPs-assisted PCR specifically amplified Bacillus cereus, not other bacteria like Bacillus subtilis, E. coli, Staphylococcus aureus, Listeria monocytogenes, and Salmonella, demonstrating assay specificity. Overall, GNPs improved PCR sensitivity for Bacillus cereus detection.