Lewin et al., Long-term cellulose enrichment selects for highly cellulolytic consortia and competition for public goods: Supplemental Material
Creators: name: Lewin, Gina
name: Davis, Nicole M.
name: R. McDonald, Bradon
name: J. Book, Adam
name: Chevrette, Marc G.
name: Suh, Steven
name: Boll, Ardina
name: Currie, Cameron R.
Publisher: name: figshare
Year: 2021
Subjects: subject: 60504 Microbial Ecology
subject: FOS: Biological sciences

description: Supplemental methods, Figs S1-S6, Tables S1 and S2, Datasets S1-S3, and supplemental references for the manuscript "Long-term cellulose enrichment selects for highly cellulolytic consortia and competition for public goods" by Gina R. Lewin, Nicole M. Davis, Bradon R. McDonald, Adam J. Book, Marc G. Chevrette, Steven Suh, Ardina Boll, and Cameron R. Currie (2022) <i>mSystems</i>, e01519-21. https://doi.org/10.1128/msystems.01519-21<br><br><b>Fig. S1. Replicates of enrichment lines. </b>At the first transfer, each enrichment line was split into three replicate enrichment lines. Replicate lines were stopped after transfer 20 except for those marked with an asterisk, which were continued for up to 60 transfers (Fig. 1) and used in sequencing analyses. After the first transfer, the filter paper did not break apart in enrichment lines 1AB or 1AC, even after over two months of incubation. <br><br><b>Fig. S2. OTU patterns and correlations among enrichment lines and across transfers. </b>(A) Heatmap showing the percentage of sequences that corresponded to each OTU. OTUs are clustered using Ward’s method, and their assigned taxonomy is indicated. The Spearman correlation of each OTU’s abundance with the NMDS analysis in Fig. 2 is also shown. (B) Network of correlating OTUs. OTUs are indicated by the number in each box. Correlations were determined using Pearson, Spearman, and Kendall correlation metrics, and a correlation (edge) was designed between 2 OTUs (nodes) if significant correlations of their abundances across samples were found using at least two of these methods to be greater than 0.5 (co-occurrence; green solid line) or less than -0.5 (mutually exclusive; red dashed line) after statistical correction. OTU abundance was also correlated with transfer time point and days to degrade the cellulose filter paper. Line thickness indicates average correlation strength. OTUs are colored based on the enrichment lines they were identified in; no OTUs included in this network were found exclusively in enrichment line 1A. OTU size indicates the total read abundance across samples. (C) Frequency of correlation coefficients in network analysis for Pearson, Spearman, and Kendall correlations.<br><br><b>Fig. S3. Taxonomic analysis of <i>Cellulomonas</i>. </b>16S rRNA gene taxonomy of <i>Cellulomonas </i>and <i>Cellulosimicrobium </i>type strains, sequences constructed from the 1A and 2B metagenome data using EMIRGE, and amplicon sequences for OTU2 clustered at 100% identity. The number of reads comprising each amplicon sequence is indicated. Taxonomy was built using RAxML with 100 bootstraps and <i>Streptomyces lividans</i> as an outgroup.<br><br><b>Fig. S4. Taxonomic and functional analysis of <i>Cellvibrio</i>.</b> (A) 16S rRNA gene taxonomy of <i>Cellvibrio </i>type strains, sequences constructed from the 2B and 3A metagenome data using EMIRGE, amplicon sequences for OTU4 clustered at 100% identity, and the Cellvibrio isolate from community 3A at transfer 50 (C. sp. 3A-50a). The abundance of each amplicon sequence in the corresponding sample is indicated in the heat map, and the corresponding transfer time point for the metagenome and isolate samples is indicated with black boxes. Taxonomy was built using RAxML with 100 bootstraps and <i>Rhizobacter </i>and <i>Pseudomonas</i> as outgroups. (B) Representative cellulose filter paper tube of <i>C. </i>sp. 3A-T50a after three days. (C) Quantitative cellulose filter paper degradation of<i> C.</i> sp. 3A-T50a isolate compared with <i>Cellvibrio japonicus</i>, a well-studied cellulolytic strain isolated from soil (18), and enriched community 3A at transfer 50 (Comm. 3A-T50). Error bars indicate standard deviation and N=3.<br><br><b>Fig. S5. Cellulase genes and transcripts annotation and abundance.</b> The metagenome coverage or metatranscriptome read count of endocellulase, exocellulase, and LPMO CAZy genes were summed and normalized to the total coverage of reads in each sample.<br><br><b>Fig. S6. Functional annotation and expression level of the 25 most highly expressed genes in MAGs.</b> (A) Functional annotations were assigned using SEED, TIGRFAM, Pfam, KEGG, and CAZy annotations and similar functional classes were grouped (Database S3). The size of each bubble represents the sum of transcriptomic reads mapping to each functional class for the 25 most highly expressed genes in each MAG, relative to the total number of reads in the metatranscriptome. (B) Hybrid PKS-NRPS biosynthetic gene cluster including two highly expressed PKS genes in the 3A.1 <i>Cellvibrio </i>MAG. (C) NRPS gene cluster in 3A.1 <i>Cellvibrio </i>MAG that includes a highly expressed NRPS gene and is predicted to make a compound similar to the siderophore turnerbactin.<br><br><b>Table S1. </b>(A) Metagenome summary. (B) Metatranscriptome summary.<b><br></b><b>Table S2. </b>(A) MAG quality and metatranscriptome mapping statistics for enrichment line 1A. (B) MAG quality and metatranscriptome mapping statistics for enrichment line 2B. (C) MAG quality and metatranscriptome mapping statistics for enrichment line 3A.<b><br></b><b>Database S1.</b> Gene annotations for all open reading frames in the metagenomes of enrichment lines 1A and 1B at transfer 14 and enrichment lines 2A, 2B, 3A, and 3B at transfer 20. Gene expression information is also included for enrichment lines 1A, 2B, and 3A.<b><br></b><b>Database S2. </b>Gene annotation and expression for MAGs from enrichment lines 1A, 2B, and 3A.<b><br></b><b>Database S3.</b> Additional annotations for the 25 most highly expressed genes from each MAG from enrichment lines 1A, 2B, and 3A.<b></b>