M. Planý, K. Šoltys, J. Budiš, A. Mader, T. Szemes, P. Siekel, T. Kuchta. Potential of high-throughput sequencing for broad-range detection of pathogenic bacteria in spices and herbs. Food Control, 83:118-122. 2018.
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A broad-range culture-independent method was developed and evaluated regarding its sensitivity of detection of pathogenic bacteria in spices and herbs, with focus on paprika powder. The method involved DNA extraction using cetyltrimethylammonium bromide (CTAB), 16S rDNA amplification using universal bacterial polymerase chain reaction, and high-throughput sequencing on Illumina MiSeq platform. The sensitivity of the method was evaluated with series of model samples contaminated at different levels with Salmonella enterica and Escherichia coli (as representatives of Gram- negative bacteria) and Staphylococcus aureus (as a representative of Gram- positive bacteria). For spices (paprika, black pepper), the method had a screening-level sensitivity with limits of detection in the range of 104– 105 CFU/g, and a semi-quantitative response. Low sensitivity (LOD ≥107 CFU/g) was observed with herbs (oregano, parsley). The developed method demonstrated a good potential for microbiological screening of spices, with a prospect of further improvement of sensitivity based on progress in high- throughput sequencing technology.