Purpose: To compare the performance of four DNA extraction methods in their efficiency to extract Salmonella DNA from pre-enriched broth and selective enriched broths of naturally contaminated chia powder.
Methods: Around 200 g chia powder samples were received from a state lab. Twenty test portions (10 g each) were pre-enriched in 90 ml modified buffer peptone water for 24 h at 35°C. Aliquots of 1.0 and 0.1 ml from the incubated pre-enrichments were subcultured to 10 ml tetrathionate (TT) broth and to 10 ml Rappaport-Vassiliadis (RV) broth, respectively. From pre-enriched and selective enriched broths, DNA was extracted by using Bio-rad InstaGene matrix, Fortius LyseNow DNA extraction kit, Life Technology automated PrepSEQ nucleic acid extraction kit, and automated Qiagen Biosprint 96 one-for-all vet kit. Salmonella qPCR assay developed by FDA was performed using DNA extracted from 24 h pre-enriched broth and 48 h selective enriched broths (RV/TT).
Results: BAM culture result shows all 20 test portions positive. It indicates a high Salmonella contamination level greater than 9.34 MPN/g in this naturally contaminated chia powder sample. qPCR result (1 positive/20 test portions) from PrepSEQ extracted DNA in 24 h pre-enriched broth showed a significantly less detection rate than culture result. The average Ct values from LyseNow method are significantly higher than those from InstaGene and BioSprint one-for-all.
Significance: To compare efficiency of different DNA extraction methods in pre-enriched and selective enriched culture will help improve sensitivity and reliability of the detection of Salmonella by qPCR assay from food products with different Salmonella contamination levels.