Using feeding regime as a microbial selective pressure to optimise biogas production and digestate sanitisation from slurry-based anaerobic digestion

Abstract Background The urgent need to adopt sustainable agricultural practices has positioned anaerobic digestion (AD) as a pivotal technology. Indeed, slurry-based AD can mitigate agricultural pollution by capturing greenhouse gas from stored slurry and converting it into biomethane, a valuable source of renewable energy, while generating digestate that can be used as fertiliser. For such a strategy to be effectively and widely deployed however, AD must be optimised. To this end, efforts have typically focused solely on biogas yields, yet improvements in pathogen load reduction may potentially negate the need for a costly pasteurisation step. Hence, optimisation of AD for sanitisation as well as improved biogas output is desirable. To address this, we set up triplicate 10-L CSTR bioreactors, which were fed with a combination of slurry and fats, oils and grease for 216 days. An organic loading rate (OLR) of 2 g VS L −1 d −1 was used throughout the trial, with a retention time of 21 days. For the first 98 days, bioreactors were fed each weekday (Monday to Friday), with 3 × feedstock on Fridays to maintain the OLR over the weekend. On Day 99 and for the remainder of the trial, the feeding regime was changed to every three days, still maintaining the 2 g VS L −1 d −1 OLR. The change in feeding regime was prompted by a noticeable increase in E. coli removal on Mondays, indicating that feeding regime could potentially function as a controllable ecological selection pressure. Results After an initial period of adaptation to the new operating conditions (from day 99–150), the change in feeding regime resulted in improved E. coli removal, achieving consistently the required reduction in numbers to satisfy EU sanitisation standards (< 1000 CFU g −1 ). Additionally, methane production increased significantly in all bioreactors with an average of 58% higher methane yield per gram VS fed when compared to the previous 5-day feeding regime. Interestingly, process optimisation led to a more tailored microbial community as revealed by metagenomics. Specifically, we observed selection for improved carbon oxidation, syntrophic acetate oxidation and methanogenesis, as well as overall reduced microbial richness and decreased functional diversity. This could potentially lead to a reduced ecosystem stability however the emergence of Methanosarcina prevalence, known for its robustness, together with the detection of the two main methanogenic pathways—acetoclastic and hydrogenotrophic—after process optimisation might confer some resistance against future perturbations. The impact of microbial shifts on ecosystem stability needs to be further assessed experimentally. Conclusions Taken together, we demonstrate that feeding regime can function as a microbial selection pressure in anaerobic digestion. The switch from a 5-day to a 3-day feeding regime led to shifts in microbial pathways, underpinning the simultaneous improvement in methane production and E. coli removal. While further research is required to assess the impact of the observed microbial community dynamics on system stability, our findings suggest that full scale on-farm AD operators could explore the effects of feeding intervals on their process performance.