Page 80 - Microbial methane cycling in a warming world From biosphere to atmosphere Michiel H in t Zandt

Chapter 4. Robustness of coal microbial community after nutrient amendment
several studies focused on characterizing and enriching microorganisms from coal (Faiz and Hendry 2006; Midgley et al. 2010; Papendick et al. 2011; Robbins et al. 2016a). However, detailed data on the effects of nutrient amendments on microbial communities in these systems are scarce but highly needed to successfully optimize coalbed methane production.
In this study, we analyzed methane production, cell counts, physicochemical properties of the well water, and acetate degradation in a sub-bituminous coal well over the course of 25 months. We compared this data to the microbial community composition at 17 months when acetate consumption and methane production were observed and at 25 months, the final time point, using 16S rRNA gene amplicon sequencing. For the 25-month sample, the functional potential of the microorganisms was investigated using a metagenome data set. This data set was also interrogated to study the fungal, archaeal and bacterial communities using ribosomal rRNA gene-based phylogeny. By combining this set of complementary experiments, we could reconstruct the complex microbial food web and assess the suitability of nutrient and acetate amendment to transform non-producing coal seams into gas production sites.
Materials and methods
Sample site
The two compared coal wells are located at the Lithgow State Coal Mine in the Western Coal Fields of New South Wales, Australia (Fig. S1). 30.6% of the raw sub-bituminous coal in this study consisted of volatile hydrocarbons; over 50% consisted of fixed carbon (Mark Wainwright Analytical Centre, UNSW, Sydney, Australia). NH4Cl and Na2PO4 for the amendments were purchased from Ajax Finechem, Scoresby VIC, Australia and Sigma- Aldrich, Castle Hill NSW, Australia, respectively. For the nutrient and acetate additions, a 100 m x 0.5 mm (length x diameter) silicon tube was lowered to the base of the well (80 m below ground level) and concentrated nutrient solutions were gravity fed (siphoned) into the subsurface in syringe-fed portions of 100 ml. During the feed, the tube was repeatedly manually raised and lowered from the bottom to the top of the water column (approximately 20 m) ensuring even distribution over the height of the water column. Lateral mixing was achieved by passive diffusion and disturbances imposed in sampling operations. By gravity flow the feeding rate was about 0.5 L min-1. Water sampling was performed using a well bladder pump
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