Comparison of primer sets for short-read NGS-based characterisation of oomycete communities

Oomycetes have diverse ecological roles spanning threats to food security to vital contributors in nutrient cycling and biological control. Despite their importance, they are often overlooked in environmental microbiome studies. We evaluated several primer sets for their ability to characterize oomycete communities in eDNA samples. The ITS1/3oo primer set performed best among those evaluated in our single-protocol eDNA survey, returning high oomycete richness and broader taxonomic coverage; we note this reflects performance under our experimental conditions rather than an absolute, universally optimal primer. For broader eukarya coverage, the 18S rRNA primer set from the Earth Microbiome Programme outperformed the COI-based primers tested. Although these broad-coverage primer sets detected a wide range of oomycetes, their performance in oomycete-specific coverage was not as strong as oomycete-targeted primer sets. Evaluation of the primer sets were undertaken using eDNA samples collected from soil in which Kunzea robusta (kānuka; Myrtaceae), and Phormium tenax (harakeke; Asphodelaceae) were cultivated. An average of 52 and 54 oomycete ASVs, respectively, were associated with the rhizosphere of these plants, spanning Haptoglossa, Alanopsis, Aphanomyces, Achyla, Pythium, Phytopythium, Globiosporangium, Phytophthora, Myzocytiopsis, Eurychasma and Lagendum lineages. Based on evaluation of a range of oomycete targeting primers on real-world samples, we are able to provide recommendations for primer sets that maximise oomycete detection and provide the best balance of specificity and coverage for eDNA-based community profiling. These primers can be used to capture the diversity and ecological roles of oomycetes in natural and managed ecosystems, enabling more accurate assessments of their impact on plant health, soil and nutrient dynamics, as well as deepening our understanding of the diversity and distribution of this understudied clade of microbial life.