A Powerful LAMP Weapon against the Threat of the Quarantine Plant Pathogen Curtobacterium flaccumfaciens pv. flaccumfaciens

Abstract

<i>Curtobacterium flaccumfaciens</i> pv. <i>flaccumfaciens</i> (<i>Cff</i>) is a Gram-positive phytopathogenic bacterium attacking leguminous crops and causing systemic diseases such as the bacterial wilt of beans and bacterial spot of soybeans. Since the early 20th century, <i>Cff</i> is reported to be present in North America, where it still causes high economic losses. Currently, <i>Cff</i> is an emerging plant pathogen, rapidly spreading worldwide and occurring in many bean-producing countries. Infected seeds are the main dissemination pathway for <i>Cff</i>, both over short and long distances. <i>Cff</i> remains viable in the seeds for long times, even in field conditions. According to the most recent EU legislation, <i>Cff</i> is included among the quarantine pests not known to occur in the Union territory, and for which the phytosanitary inspection consists mainly of the visual examination of imported bean seeds. The seedborne nature of <i>Cff</i> combined with the globalization of trades urgently call for the implementation of a highly specific diagnostic test for <i>Cff</i>, to be routinely and easily used at the official ports of entry and into the fields. This paper reports the development of a LAMP (Loop-Mediated Isothermal Amplification) specific for <i>Cff</i>, that allows the detection of <i>Cff</i> in infected seeds, both by fluorescence and visual monitoring, after 30 min of reaction and with a detection limit at around 4 fg/μL of pure <i>Cff</i> genomic DNA.