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Evaluation of DNA extraction methods for B acillus anthracis spores isolated from spiked food samples
Author(s) -
Thomas M.C.,
Shields M.J.,
Hahn K.R.,
Janzen T.W.,
Goji N.,
Amoako K.K.
Publication year - 2013
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1111/jam.12206
Subject(s) - bacillus anthracis , spore , dna extraction , immunomagnetic separation , biology , microbiology and biotechnology , food science , extraction (chemistry) , serial dilution , chromatography , polymerase chain reaction , bacteria , chemistry , biochemistry , medicine , genetics , gene , alternative medicine , pathology
Abstract 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.