Abstract
Phytophthora is one of the most important and aggressive plant pathogenic genera in agriculture and forestry. Early detection and identification of its pathways of infection and spread are of high importance to minimize the threat they pose to natural ecosystems. eDNA was extracted from soil and water from forests and plantations in the north of Spain. Phytophthora-specific primers were adapted for use in high-throughput Sequencing (HTS). Primers were tested in a control reaction containing eight Phytophthora species and applied to water and soil eDNA samples from northern Spain. Different score coverage threshold values were tested for optimal Phytophthora species separation in a custom-curated database and in the control reaction. Clustering at 99% was the optimal criteria to separate most of the Phytophthora species. Multiple Molecular Operational Taxonomic Units (MOTUs) corresponding to 36 distinct Phytophthora species were amplified in the environmental samples. Pyrosequencing of amplicons from soil samples revealed low Phytophthora diversity (13 species) in comparison with the 35 species detected in water samples. Thirteen of the MOTUs detected in rivers and streams showed no close match to sequences in international sequence databases, revealing that eDNA pyrosequencing is a useful strategy to assess Phytophthora species diversity in natural ecosystems.
Highlights
In recent years the increase of global plant trade and human movement have promoted the risk of introduction of invasive plants and exotic pathogens [1,2,3]
For the control reaction data, clustering at 99% of similarity was the minimum value able to separate all of the species with the minimum number of reads lost
Values below 99% were insufficient to separate P. gonapodyides and P. taxon PgChlamydo in separate Molecular Operational Taxonomic Units (MOTUs)
Summary
In recent years the increase of global plant trade and human movement have promoted the risk of introduction of invasive plants and exotic pathogens [1,2,3]. Biological invasions operate globally and are considered to be the second cause of biodiversity loss after direct habitat alteration and destruction. In this context, Phytophthora species are of particular importance worldwide as they are major pathogens in agriculture, horticulture and forestry causing important economic and ecological losses. Amplicon Pyrosequencing for Phytophthora Discovery Using eDNA funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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