Integrating 16S rRNA amplicon metagenomics and selective culture for developing thermophilic bacterial inoculants to enhance manure composting

Abstract

Composting is an important method for treating and recycling organic waste, and the use of microbial inoculants can increase the efficiency of composting. Herein, we illustrate an approach that integrate 16S rRNA amplicon metagenomics and selective culture of thermophilic bacteria for the development of inoculants to improve manure composting. The 16S rRNA amplicon sequencing analysis revealed that Firmicutes and Actinobacteria were dominant in the composting mixture, and that different microbial hubs succeeded during the thermophilic stage. All isolated thermophilic bacteria were affiliated with the order Bacillales, such as Geobacillus, Bacillus, and Aeribacillus. These isolated thermophilic bacteria were grouped into 11 phylotypes, which shared ＞99% sequence identity to 0.15% to 5.32% of 16S rRNA reads by the amplicon sequencing. Three of these phylotypes transiently enriched during the thermophilic stage. Six thermophilic bacteria were selected from the three phylotypes to obtain seven microbial inoculants. Five out of seven of the microbial inoculants enhanced the thermophilic stage of composting by 16.9% to 52.2%. Three-dimensional excitation emission matrix analysis further revealed that two inoculants (Thermoactinomyces intermedius and Ureibacillus thermophilus) stimulated humification. Additionally, the 16S rRNA amplicon sequencing analysis revealed that inoculation with thermophilic bacteria enhanced the succession of the microbial community during composting. In conclusion, 16S rRNA amplicon metagenomics is a useful tool for the development of microbial inoculants to enhance manure composting.