Field ready: Development of a rapid LAMP-based colorimetric assay for the causal agent of white pine blister rust, Cronartium ribicola

Abstract

The invasive fungal pathogen Cronartium ribicola causes white pine blister rust which is considered one of the most destructive diseases of five-needle (white) pines in North America. The disease has a life cycle that requires two hosts: white pines and Ribes spp., although other non-Ribes species, including Castilleja and Pedicularis, have been demonstrated as alternate hosts as well. Detection of this disease can be difficult because of the ephemeral nature of sporulation on pine hosts with ambiguity in other symptoms, and the alternate hosts for C. ribicola can also be an alternate host for other pine rust species. We used the previously published C. ribicola genome and species-specific real-time PCR assay to develop a field-ready loop-mediated isothermal amplification (LAMP) specific colorimetric assay for this pathogen. Specificity results across regionally identified pine rust pathogens showed the assay is highly specific to C. ribicola and can detect as little as 40 pg of pathogen DNA. We also developed a simple DNA extraction method that works with several tissue types (bark/phloem, aeciospores, and urediniospores/telia) to prepare the DNA samples for the LAMP assay. The DNA extraction and LAMP assay take ~70 min to complete and require a relatively small investment in equipment. This tool enables quick and efficient detection of white pine blister rust.