Evaluation of DNA extraction methods for B acillus anthracis spores isolated from spiked food samples

Aims Nine commercial DNA extraction kits were evaluated for the isolation of DNA from 10‐fold serial dilutions of B acillus anthracis spores using quantitative real‐time PCR ( qPCR ). The three kits determined by q PCR to yield the most sensitive and consistent detection (Epicenter M aster P ure G ram P ositive; M o B io P ower F ood; ABI P rep S eq) were subsequently tested for their ability to isolate DNA from trace amounts of B . anthracis spores (approx. 6·5 × 10 1 and 1·3 × 10 2   CFU in 25 ml or 50 g of food sample) spiked into complex food samples including apple juice, ham, whole milk and bagged salad and recovered with immunomagnetic separation ( IMS ). Methods and Results The M aster P ure kit effectively and consistently isolated DNA from low amounts of B . anthracis spores captured from food samples. Detection was achieved from apple juice, ham, whole milk and bagged salad from as few as 65 ± 14, 68 ± 8, 66 ± 4 and 52 ± 16 CFU , respectively, and IMS samples were demonstrated to be free of PCR inhibitors. Conclusions Detection of B . anthracis spores isolated from food by IMS differs substantially between commercial DNA extraction kits; however, sensitive results can be obtained with the M aster P ure G ram P ositive kit. Significance and Impact of the Study The extraction protocol identified herein combined with IMS is novel for B . anthracis and allows detection of low levels of B . anthracis spores from contaminated food samples.