Validation of nanopore sequencing on a set of aquaponics biofilter samples that were sequenced using Illumina and nanopore technologies (2 replicates each).

(A) In low abundances, the Expectation-Maximization (EM) algorithm identifies fewer unique references than the “naive” strategy of just selecting the highest scoring read. Each dot denotes the number of unique 16S sequences in the SILVA database that pass the abundance cutoff. (B) Abundances across sequencing protocols. Each dot denotes the abundance of a single taxon at the indicated taxonomic rank. Blue lines denote a linear regression for the taxa found in both sequencing technologies and gray errors denote the 95% confidence interval of the regression. The red boxplot summarizes the distribution of spurious mappings in the nanopore data (absent in the Illumina data). (C) Distribution of false positive mappings (mappings not observed in the Illumina data) in nanopore sequencing. The dashed red line denotes the used abundance cutoff that removed >96% of those spurious mappings. (D) Abundances of nitrifying taxa in both sequencing protocols. Dots denote the sum in the two replicates. Abundances smaller than one denote taxa not detected in Illumina sequencing.

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