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Detection of Listeria monocytogenes using Dynabeads® anti-Listeria combined with real-time PCR in soybean sprouts

Author:
Wei, Shuai, Park, Byung-Jae, Kim, Se-Hun, Seo, Kun-Ho, Jin, Yong-Guo, Oh, Deog-Hwan
Source:
Lebensmittel-Wissenschaft + [i.e. und] Technologie 2019 v.99 pp. 533-539
ISSN:
0023-6438
Subject:
DNA, Listeria monocytogenes, bacteria, bean sprouts, detection limit, genes, immunomagnetic separation, protocols, quantitative polymerase chain reaction, rapid methods, soybeans
Abstract:
A rapid detection protocol for Listeria monocytogenes in soybean sprout samples was developed and evaluated using immunomagnetic separation (IMS) combined with real-time PCR. Dynabeads® anti-Listeria was mixed with food samples, separated with a particle concentrator, followed by DNA extraction for real-time PCR targeting the hly gene. The amount of Dynabeads® anti-Listeria used to determine conjugation with bacteria ranged from 5 to 160 μL. Immunoreaction times ranged from 10 to 40 min with different concentrations of L. monocytogenes were also evaluated. The real-time PCR standard curve was constructed, and the curve exhibited a R2 value of 0.999. The efficiency of the real-time PCR method was 96.7%. The limit of detection was 4.4 Log CFU/g with 69 evaluated spiked soybean sprout samples. The sensitivity, accuracy, and specificity of the IMS-real-time PCR method were 95.0%, 95.6% and 100%, respectively. Moreover, the IMS-real-time PCR and ISO method using 20 natural soybean sprout samples gave the same results, and no positive sample was detected by both methods. However, the combination of IMS and real-time PCR was superior in terms of the shorter time needed for detection of L. monocytogenes in soybean sprout samples (27 h versus 4–7 days).