Additional file 1 of Microbial Tracking-2, a metagenomics analysis of bacteria and fungi onboard the International Space Station

Field: Immunology and Microbiology (Life Sciences)

Additional file 1: Figure S1. Microbial burden assessment using culture. Bacteria were grown on R2A and blood agar plates and the cfu/m2 averaged from each type of plate and then plotted. Fungi were grown on PDA plates. Each dot and corresponding number represents one of the eight surface locations sampled. X-axis labels: “BAC” refers to the bacterial load and “PDA” to the fungal load. Fx refers to one of the four flight sampling events. Figure S2. Alpha diversity metrics for surface samples. The species richness (top row), exponentiated Shannon index (middle row), and inverse Simpson Index (bottom row) are shown for each sample. F4 (green), F5 (yellow), F6 (purple), and F7 (red) samples are grouped by surface location. Figure S3. NMDS plot of MT-1 and MT-2 surface samples. Samples were colored by surface location (A) and flight group (B) to visualize samples clustering. The distance between samples was determined using the Jaccard dissimilarity. Figure S4. Top 12 most abundant bacterial species by flight group. The top 12 most abundance fungal species for F4 (A), F5 (B), F6 (C), and F7 (D) were determined separately. The read counts for each species are shown by surface location. “Other” refers to those bacterial species detected that were not in the top 12. Figure S5. Top 12 most abundant fungal species by flight group. The top 12 most abundance fungal species for F4 (A), F5 (B), F6 (C), and F7 (D) were determined separately. The read counts for each species are shown by surface location. “Other” refers to those fungal species detected that were not in the top 12. Figure S6. Co-occurrence analysis. Co-occurrence analysis was performed using the 50 most abundant genera to determine associations amongst organisms. The x-axis shows the organisms that were included in the analysis and the y-axis shows the percent of either negative, positive or random associations. Gray bars represent random associations, blue bars represent positive associations and yellow bars represent negative associations. The genera are listed in order of least occurrences (left) to most occurrences (right). Figure S7. Heat map of ISS resistome. The metagenomic dataset was analyzed with DeepARG to look for anti-microbial resistance genes. The genes were then grouped into classes of resistance (i.e “beta-lactam”), which is displayed in this heatmap (y-axis). The heat-map shows the relative abundance of each resistance class in each sample collected, with red being the most abundant and green the least. Gray indicates zero counts detected in that sample The samples analyzed (x-axis) are from the non-PMA treated samples. x-axis naming: Fx_xS, with F referring to the flight (i.e., F4) and S referring to the location of the surface sampled (i.e., 1S is location 1, the port panel next to cupola). The AMR counts were normalized based on the 16S rRNA counts. Figure S8. Subsystem category distribution of Pseudomonas fluroescens metagenome assembled genome. The metaWRAP pipeline was used to assemble and annotate draft genomes constructed from the metagenomic data of 32 samples (F4-F7), otherwise known as MAGs- metagenomic assembled genomes. The MAG of Pseudomonas fluorescens was further analyzed with SEED and RAST. Figure S9. Read counts for control samples. The top 12 most abundance species across the control samples are colored. The read counts in the controls were significantly less than those detected in the samples and had different profiles.