Linking Community Structure and Gene Expression in Methanogen Populations to Methane Emission from Stored Liquid Dairy Manure with Different Inoculant Levels.

Microbial communities in dairy manure inoculants and their effects on methane (CH₄) emissions have seldom been studied. We examined the effects of different levels of inoculant on the structure and function of methanogenic and bacterial communities in stored dairy slurries. Six pilot-scale (3.8×1.73×1.8 m³) manure storage tanks filled (in batch or gradually) with fresh dairy slurry containing 0%, 10%, or 20% residual slurry level were studied over 120 days of storage (June-2_September-24, 2016). While CH₄ flux was monitored continuously at the site, slurry samples were collected (day 0, 65 and 120), nucleic acids extracted, and analyzed for methanogenic and bacterial communities using quantitative real-time PCR and Illumina MiSeq sequencing of DNA and RNA transcripts. Significantly higher (P<0.05) abundance and activity of methanogens and bacteria were observed in inoculant compared to fresh slurry. With batch or gradual filling, higher levels of inoculum resulted in increased abundance and activity (up to 8.5-fold) of methanogenic and bacterial communities during the initial 65 days of storage. In the presence of inoculum, batch filled storage tank showed lower CH₄ flux but higher activity of methanogens (up to 4.7-fold). The absence of inoculant however, lowered both methanogenic activity and CH₄ flux from gradually filled tank. No significant variation in the diversity of methanogenic or bacterial communities were observed between fresh and inoculant slurries, and hence among treatments. Methanogens of the order Methanomicrobiales were predominant in all samples. During peak CH₄ flux, however, RNA data revealed the metabolically diverse methanogens of the order Methanosarcinales as the predominant players in CH₄ production. Results suggest that higher levels of inoculum might not provide new storages with unique phylotypes but large numbers of active methanogens that could result in increased CH₄ flux. With higher methanogenic activity in slurries, studies that involve batch filling of tanks might underestimate CH₄ flux estimates.