Validating metabarcoding-based biodiversity assessments with multi-species occupancy models: A case study using coastal marine eDNA

the minimal level of augmentation required, as described above in the Unknown Species Rich- ness section. Significance of continuous covariates was assessed by determining if the 95% con- fidence intervals of parameter estimates overlapped with zero [31]. To investigate the effects of phylum on the probability of detection of each amplicon, we ran one additional model (Model 9), which included amplicon as a categorical covariate (α2) and group-level effects at the species-level following [31]. In this model, we only included metazoan phyla where at least 2 species were detected. Here, species coefficients arise from community-level parameters that vary by phylum: lpsik � Nðmlpsi½phylum�; slpsi½phylum�Þ lthetak � Nðmltheta½phylum�; sltheta½phylum�Þ lpk � Nðmlp½phylum�; slp½phylum�Þ β1k � Nðmβ1½phylum�; sβ1½phylum�Þ β2k a2 � Nðmβ2½phylum�; sβ2½phylum�Þ β3k � Nðmβ3½phylum�; sβ3½phylum�Þ Results We ran eight multi-species, multi-scale occupancy models with different combinations of covariates (i.e., water depth at the level of occupancy, sequencing depth and amplicon at the Table 2. Model comparison between multi-scale, multi-species occupancy models using four methods (WAIC, CPO, Brier score and Log Score). The covariates (water depth at the sampling site, sequencing depth for each techni- cal replicate, and amplicon sequenced for each technical replicate) included at each level of the model (occupancy: ψ, capture: θ, detection: p) are listed on the left. Bolded values indicate the best model for each method of model comparison. MODELS WAIC CPO Brier Score Log Score

Model 1 ψ(.) θ(.) p(.) 13,340 5,621,109 98 1842 Model 2 ψ(water depth) θ(.) p(sequencing depth, amplicon) 33,626 14,217,497 217 2034 Model 3 ψ(.) θ(.) p(sequencing depth) 12,993 4,061,736 98 1834 Model 4 ψ(.) θ(.) p(amplicon) 35,894 20,793,808 135 2823 Model 5 ψ(water depth) θ(.) p(.) 13,135 2,748,236 143 1759 Model 6 ψ(water depth) θ(.) p(amplicon) 35,649 17,897,824 224 2732 Model 7 ψ(water depth) θ(.) p(sequencing depth) 12,869 1,697,452 142 1753 Model 8 ψ(.) θ(.) p(sequencing depth, amplicon) 33,499 16,555,815 133 2405 https://doi.org/10.1371/journal.pone.0224119.t002 PLOS ONE Multi-species occupancy models for metabarcoding PLOS ONE | https://doi.org/10.1371/journal.pone.0224119 March 19, 2020 7 / 17