Page 234 - Microbial methane cycling in a warming world From biosphere to atmosphere Michiel H in t Zandt

Chapter 10. Co-culturing methanogens and methanotrophs in an MBR
the KEGG Automatic Annotation Server (KAAS – last update April 3, 2015) (Moriya et al. 2007) for pathway analyses. Genome annotations were examined using the Artemis genome browser release 16.0.0 (Rutherford et al. 2000). For 16S rRNA gene analysis, raw Illumina HiSeq reads were mapped against the SILVA SSU non-redundant database version 128 and de novo assembled as described in in ‘t Zandt et al. (2018).
Nucleotide sequence accession numbers
All sequencing data were submitted to the GenBank databases under BioProject PRJNA434352. The genome bins were submitted as genome data under BioSample accession number SAMN08554708 and SAMN08554709.
Results
Start-up of the oxygen limited bioreactor
A membrane bioreactor was inoculated with aerobic methanotrophs and fed with a gas mixture containing methane and oxygen. A stable co-culture of anaerobic methanogens and aerobic methanotrophs could be obtained for over two months. Optical density (OD) was followed over time (Fig. 2). The initial phase with only aerobic methanotrophs indicated a rapid growth based on an OD600 increase from 0.3 to 0.85 within 55 days. The methanotrophs were put under oxygen limitation, and after the dissolved oxygen levels dropped below the detection limit (≤ 3 μM), the OD600 was diluted to 0.5 and the reactor was inoculated on t = 92 days with the methanogenic archaeon M. barkeri. Oxygen levels remained below the detection limit for the entire experiment. At t = 105 days, methane inflow was reduced to 1.25 mL min-1 to restrict methanotrophic growth. Co-culture OD values increased to almost 1 within 19 days. At t = 113 days, additional biomass was removed and a bleed was installed to remove 1/30th of the reactor volume per day to target an OD600 of around 0.5. From t = 113 days to the end of reactor operation at t = 337 days OD600 values ranged from 0.4 to 0.7.
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