<p> 1Supplementary Figure legends, Imachi et al.</p><p>2</p><p>3Fig. S1. The procedure for inoculation of the sediment slurry into the sponge carriers. (a) The</p><p>4sponge carriers were submerged into the sediment slurry and then squeezed by hand to fill the</p><p>5sediment slurry. During the filling process, the sediment slurry was flushed by nitrogen gas.</p><p>6(b) The color of sponge materials changed from white to brown. (c) The sponge carriers were</p><p>7inserted into net-like cylindrical plastic covers and placed into the PVC box.</p><p>8</p><p>9Fig. S2. Detailed design of the distributor unit.</p><p>10</p><p>11Fig. S3. Sampling of the sponge carriers from the DHS reactor. (a) During the sampling of the</p><p>12sponge carriers, nitrogen gas was flushed into the sampling port. The sponge carriers were</p><p>13removed from the reactor with tweezers. (b) Sponge carriers after removal. (c) After removing</p><p>14the net-like cylindrical plastic covers, sediments were squeezed from the sponges in anaerobic</p><p>15synthetic seawater. (d) The sediment slurry was used for further analysis and cultivation.</p><p>16</p><p>17Fig. S4. Time course of changes in (a) ORP and (b) pH values of the effluent seawater.</p><p>18</p><p>19Fig. S5. Phylogenetic tree of archaeal phylotypes obtained in this study. (a) A tree for most of</p><p>20methanogenic and euryarchaeotic phylotypes and (b) an expanded bacterial phylogenetic tree</p><p>21for the phylum Crenarchaeota and deeply branching Archaea. Sequences obtained in this</p><p>22study are marked in blue (the inoculum sample) and red (the enrichment samples on the DHS</p><p>23reactor and batch-type enrichment cultures). The number in parentheses after the phylotypes</p><p>24obtained from the inoculum sample indicates the number of identical clones. The number in</p><p>25parentheses after the phylotypes obtained from the enrichment samples on the DHS reactor</p><p>1 1 1indicate the number of identical clones for each phylotype and the frequency of each clone</p><p>2library in the following order; DNA-based clone library from day 357, DNA-based clone</p><p>3library from day 560, RNA-based clone library from day 560, DNA-based clone library from</p><p>4day 761, and RNA-based clone library from day 761. The number in parentheses after the</p><p>5phylotypes obtained from the batch-type enrichment cultures indicates the numbers of</p><p>6identical clones obtained per number of clones analyzed for each phylotype. The accession</p><p>7numbers are also given after each phylotype name. The scale bar represents the estimated</p><p>8number of nucleotide changes per sequence position. The symbols at the nodes show the</p><p>9bootstrap values (>70% indicated only) obtained after 1,000 resampling.</p><p>10</p><p>11Fig. S6. Phylogenetic tree of deduced McrA amino acid sequences showing the phylogenetic</p><p>12positions of clones and isolates obtained in this study. The bar indicates 10% estimated</p><p>13sequence divergence. The meanings of the colored sequences and the numbers in parentheses</p><p>14are the same as in Fig. 3.</p><p>15</p><p>16Fig. S7. Phylogenetic tree of bacterial 16S rRNA gene phylotypes obtained in this study. (a) A</p><p>17large bacterial tree for diverse bacterial group and (b-e) expanded bacterial phylogenetic trees</p><p>18for the phyla (b) Proteobacteria, (c) Planctomycetes, (d) Firmicutes and (e) Chloroflexi. The</p><p>19scale bar represents the estimated number of nucleotide changes per sequence position. The</p><p>20symbols at the nodes show the bootstrap values obtained after 1,000 resampling. The</p><p>21meanings of the colored sequences and the numbers in parentheses are the same as in Fig. 3.</p><p>22</p><p>23Movie S1. The DHS bioreactor system used in this study.</p><p>24</p><p>1 2</p>