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Rapid, specific, simple, in‐field detection of Xanthomonas campestris pathovar musacearum by loop‐mediated isothermal amplification

Author:
Hodgetts, J., Hall, J., Karamura, G., Grant, M., Studholme, D.J., Boonham, N., Karamura, E., Smith, J.J.
Source:
Journal of applied microbiology 2015 v.119 no.6 pp. 1651-1658
ISSN:
1364-5072
Subject:
DNA, Musa, Xanthomonas campestris, Xanthomonas vasicola, bacteria, bananas, diagnostic techniques, endophytes, genes, loop-mediated isothermal amplification, microbial detection, pathogens, pathovars, polymerase chain reaction, secretion, Uganda
Abstract:
AIMS: To develop and evaluate a loop‐mediated isothermal amplification (LAMP) assay for Xanthomonas campestris pathovar musacearum (Xcm), the causal agent of banana Xanthomonas wilt, a major disease of banana in Africa. METHODS AND RESULTS: LAMP primers were designed to the general secretion pathway protein D gene and tested against 17 isolates of Xcm encompassing the known genetic and geographic diversity of the bacterium and all isolates were detected. Seventeen other Xanthomonas isolates, including closely related Xanthomonas vasicola, other bacterial pathogens/endophytes of Musa and two healthy Musa varieties gave negative results with the LAMP assay. The assay showed good sensitivity, detecting as little as 51 fg of Xcm DNA, a greater level of sensitivity than that of an Xcm PCR assay. Amplification with the LAMP assay was very rapid, typically within 9 min from bacterial cultures. Symptomatic field samples of Musa from Uganda were tested and all produced amplification in less than 13 min. CONCLUSIONS: The LAMP assay provides rapid, sensitive detection of the pathogen that is ideally suited for deployment in laboratories with basic facilities and in‐field situations. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first LAMP assay for Xcm which provides a significant improvement compared to existing diagnostics.
Agid:
4790834