Genome-centric metagenomic analysis revealed the microbial superiority of the developed anaerobic digestion startup strategy based on combination of raw and concentrated digestate.

Concentrating the digestate as inoculum to initiate new anaerobic digestion (AD) reactors has been proven to be feasible. However, functional microorganisms after centrifugal concentration have poor activity in the initial stage, resulting in a long startup period. In this study, AD was initiated through addition of centrifugal concentrated digestate combined with different proportions of raw digestate (0%, 9%, 18%, and 27%, in terms of total working volume), aiming to restore the metabolic activity rapidly with a minimum amount of raw digestate to reduce the startup cost. The result showed that Cen-18% (bioaugmentation of 18% raw digestate on the top of concentrated digestate) was the most suitable startup strategy in terms of methane production rate and methane yield. Metagenomic absolute quantitative analysis revealed that methane metabolism (ko00680) was enhanced in Cen-18% (2.9 × 1010 copies/g V.S. (1.5-1.8) × 1010 copies/g in other groups) along with the AD progress by upregulating the abundance of key functional genes in carbohydrate metabolism (gapA, fbaA and eno) and amino acid metabolism (carB, argG and argH). Moreover, genome-centric metagenomic analysis implied that the Cen-18% accelerated carbon conversion by enriching higher abundance of the identified key carbohydrate and protein degraders (i.e., Bacteroidales_MAG2 and Bacteroidales_MAG8) in Cen-18% than other experimental groups. Hydrogenotrophic methanogenesis performed by Methanomicrobiales_MAG1 and Methanomicrobiales_MAG2 was the dominant methanogenic pathway in Cen-18%, which was highly symbiotic with the acetate oxidation bacteria DUMP01_MAG3, DUMP01_MAG1, and Thermosediminibacterales_MAG2. This study provides practical support for the startup of newly built biogas digesters.