Long-term preserved and rapidly revived methanogenic cultures: Microbial dynamics and preservation mechanisms

Bioaugmentation with specialized inocula has been proven a feasible way to remediate under-performing anaerobic digestion (AD) processes. However, a major bottleneck for successful and cost-effective bioaugmentation is the lack of ready-to-use, specialized methanogenic cultures when required. The reason is the slow growth of the anaerobic consortia and the high cost of maintaining them active in the necessary amounts for successful bioaugmentation applications. This study offers an effective procedure where customized AD inocula could be preserved and used on-demand for remediation of ammonia inhibited AD reactors. Additionally, it introduces the biological and physicochemical mechanisms that render the long-term preservation of the AD inocula possible. Specifically, two different preservation carriers (i.e. agar gel and liquid basic anaerobic medium) were assessed at two different temperatures (i.e. 4 °C and 24 °C) using an ammonia tolerant methanogenic consortium. The results from methane production, lag-phase, maximum methane production rate and cell viability indicate that the consortium preserved for 168 days in agar gel at 24 °C performed best compared to the other tested preservation conditions. Meanwhile, 16S rRNA sequencing analysis indicated that Methanosarcina soligelidi and Methanoculleus palmolei shown a high revival rate and metabolic activity after long-term preservation. Thus, this successful long-term preservation method of ready-to-use AD consortia could render the successful bioaugmentation in full-scale biogas reactors economically possible in the near future.