Enrichment of Methanothrix species via riboflavin-loaded granular activated carbon in anaerobic digestion of high-concentration brewery wastewater amidst continuous inoculation of Methanosarcina barkeri.

Effective treatment of high-concentration brewery wastewater through anaerobic digestion (AD) has always been a challenging issue. Enhancing direct interspecies electron transfer (DIET) was demonstrated to increase methane production during AD under high organic loading rate (OLR). Herein, the feasibility of enhancing DIET with the addition of riboflavin-loaded granular activated carbon (RF-GAC) as well as co-addition with Methanosarcina barkeri (Rf-GAC+M.barkeri) was investigated (M.barkeri is well-known to be capable of DIET with electroactive bacteria). During the whole process, the Rf-GAC and the Rf-GAC+M.barkeri group both achieved average COD removal rates above 97 %, which was 14 % higher than that of the control. The average methane production in the Rf-GAC group and the Rf-GAC+M.barkeri group respectively reached 0.334 ± 0.02 L(stp)/g COD and 0.345 ± 0.02 L(stp)/g COD, 1.35 and 1.39 times higher than the 0.247 ± 0.03 L(stp)/g COD reached by the control. The control reactor deteriorated at an OLR of 12 kg COD/(m·d), whereas the Rf-GAC and the Rf-GAC+M.barkeri group maintained stable as the OLR reached as high as 17.5 kg COD/(m·d) and the volatile fatty acids concentration was consistently below 10 mM. The RF-GAC performed better than Rf-GAC+M.barkeri in enriching Methanothrix, whose relative abundance was 60.6 % in the former group. Metabolic pathway analysis revealed the addition of RF-GAC upregulated genes related to DIET in Methanothrix species, including hdrA and fpoD. Furthermore, Methanothrix remained the dominant archaea even continuously inoculating pure strains of M.barkeri during the entire operational period. Pure culture experiments proved that GAC inhibited M.barkeri growth. The results of this study can be optimized for practical application of AD treating high-concentration brewery wastewater.