Specificity of LAMP for genetic diagnostics of Chilean needle grass and serrated tussock

摘要

Chilean needle grass (Nassella neesiana [Trin. & Rupr.] Barkworth) and serrated tussock (Nassella trichotoma [Nees] Hack, ex Arechav.) are related Weeds of National Significance (WoNS) undergoing rapid range expansion in Australia. They are difficultto distinguish from native tussock grasses and emergences of these weeds often remain undetected until after they have established as difficult to control populations. Genetic diagnostics using loop-mediated isothermal amplification (LAMP) may provide asimple cost-efficient means for rapid identification of these weeds, under laboratory and field conditions. Provision of LAMP diagnostics would assist in minimising the time-lag to initial recognition of tussock weed outbreaks in new areas, and subsequently improve the time to response for eradication of the weeds where they are recently emergent.Here we report laboratory development and tests of seven LAMP assays incorporating novel primer suites matched to species-specific DNA barcode sequences of Chilean needle grass and serrated tussock, and three additional suites designed for general LAMP validation. LAMP assays conducted using a real-time PCR platform tested positive for replicates of the target weeds at several of the primer suites and in less than 20 min. Three of the suites are particularly promising (89-100% repeatability and specificity) and following optimisation to further minimise false positive or false negative test outcomes they are likely to be reliable for field use. As promising as these results are, the real challenge will be next-stage development of a simple but effective mobile LAMP platform for use in the field by officers engaged in weed surveillance efforts.