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Evaluation of methods for erwinia amylovora detection

Author:
Kałużna, Monika, Puławska, Joanna, Mikiciński, Artur
Source:
Journal of Horticultural Research 2014 v.21 no.2 pp. 65-71
ISSN:
2353-3978
Subject:
Erwinia amylovora, melting, microbial detection, quantitative polymerase chain reaction
Abstract:
Phytosanitary measures to control diseases require sensitive and reliable detection methods. In case of Erwinia amylovora, the causal agent of fire blight, there are several methods developed. We tested six of them: (1) a conventional method; (2) standard PCR; (3) BIO-PCR; (4) Real-time PCR with SYBR Green; (5) Real-time PCR with TaqMan probe; and (6) Loop Mediated Isothermal Amplification (LAMP) in terms of their specificity, sensitivity and repeatability. Of the methods tested, only Real-time PCR with SYBR Green amplified non-specific fragments, which could be however identified by melting curve analysis. Real-time PCRs (both with SYBR Green and TaqMan probe) and LAMP were most sensitive and allowed to detect E. amylovora even in the samples where the concentration of cells was lower than 2 x 10¹ cfu/reaction. The highest repeatability was obtained using conventional method and Real-time PCRs. Of the methods used, only the LAMP technique was very insensitive to plant impurities.
Agid:
5200914