(A) Tiled view of an ESOM map constructed using all 51 metagenome bins assembled from the samples collected in this study, with the white square encompassing.

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(A) Tiled view of an ESOM map constructed using all 51 metagenome bins assembled from the samples collected in this study, with the white square encompassing the Nitrospira-like metagenome bin. (A) Tiled view of an ESOM map constructed using all 51 metagenome bins assembled from the samples collected in this study, with the white square encompassing the Nitrospira-like metagenome bin. Some metagenome bins expand over the edge of a single ESOM grid. Hence, a tiled view consisting of four copies of the ESOM grid is shown to allow for visualization of metagenome bins at the edge as contiguous clusters. This results in all metagenome bins included in the ESOM analyses appearing four times in the tiled view. (B) Enlarged view of panel A indicating three scaffold fragments that were outliers based on ESOM analyses. (C) Enlarged view of panel A showing fragments of scaffold 158 containing ammonia oxidation genes that were binned with the Nitrospira metagenome. The ESOM binning procedure and contents of the three outlier scaffolds/scaffold fragments are presented in Text S1 in the supplemental material. (D) RAxML-based maximum-likelihood tree constructed using amino acid sequences of the amoA gene in the Nitrospira metagenome bin and pmoA/amoA sequences from a range of ammonia-oxidizing bacteria/archaea and methane-oxidizing bacteria, including the Nitrospira comammox. The tree was built from a trimmed muscle alignment using the Dayhoff model for protein evolution, gamma distribution model, and 500 bootstraps using the archaeal amoA gene of Nitrosopumilus maritimus as the outgroup. Branches are colored according to phylogenetic affiliation, and node support of >70% is indicated. This placement of the amoA gene from the Nitrospira-like genome and overall tree topology were also confirmed by neighbor-joining analysis (500 bootstraps) and the unweighted pair group method with arithmetic mean (UPGMA) (500 bootstraps) in Geneious and Bayesian phylogeny inference (20,000 generations) (Fig. S3). Ameet J. Pinto et al. mSphere 2016; doi:10.1128/mSphere.00054-15