Batch-Mode Analysis of Thermophilic Methanogenic Microbial Community Changes in the Overacidification Stage in Beverage Waste Treatment

Biogasification by methane fermentation is an important and effective way to utilize beverage wastes. Beverage wastes are good feedstocks for methane fermentation because of their richness in sugars and proteins, although overacidification and inhibition of methane production caused by high substrate loading often become problematic. This study investigated changes in microbial communities in the overacidification state of the thermophilic methane fermentation process with beverage waste by establishing a simulated batch culture. We assessed 20 mL-scale batch cultures using a simulant beverage waste mixture (SBWM) with different amounts of addition; high cumulative methane production was achieved by adding 5 mL of SBWM (11358 mg-chemical oxygen demand-COD/L of organic loading), and overacidification was observed by adding 10 mL of SBWM (22715 mg-COD/L of organic loading). The results of 16S rRNA amplicon sequence analysis using nanopore sequencer suggested that Coprothermobacter proteolyticus, Defluviitoga tunisiensis, Acetomicrobium mobile, and Thermosediminibacter oceani were predominantly involved in hydrolysis/acidogenesis/acetogenesis processes, whereas Methanothrix soehngenii was the major acetotrophic methane producer. A comparison of microbial population between the methane-producing cultures and overacidification cultures revealed characteristic population changes especially in some minor species under 0.2% of population. We concluded that careful monitoring of population changes of the minor species is a potential indicator for prediction of overacidification.