MiDAS4, global survey of activated sludge

Biological wastewater treatment and the transition to wastewater resource recovery facilities is fundamental for protection of the environment and human health and to ensure sustainable development. Microorganisms are essential for these processes, but our knowledge of their diversity and function is still poor, partly due to the lack of good reference databases and comprehensive global studies. Here, we first sequenced more than 5 million high-quality full-length 16S rRNA gene sequences from 740 diverse wastewater treatment plants (WWTPs) across the world, and used the sequences to construct MiDAS 4, an amplicon sequence variant resolved full-length 16S rRNA gene reference database with a comprehensive taxonomy from the domain to species-level for all references. MiDAS 4 covers ~32,000 species, which represents the majority of prokaryotes in global WWTPs, and provides a unifying taxonomy for the field, which can act as an anchor point for linking microbial identities with their functions across studies. MiDAS 4 was used for a global-scale microbial community profiling of the activated sludge plants using amplicon sequencing with two sets of primers (V1-V3 and V4). We found that the V1-V3 primers were best suited for microbial community profiling of this ecosystem, and revealed how environmental conditions and biogeography shaped the community composition. In addition, we used the data to identify process-critical taxa (core and conditionally abundant taxa) for different process types across the world, encompassing approximately 900 genera and 1500 species. Most of them are poorly described or novel taxa. Among known functional guilds, such as nitrifiers or polyphosphate accumulating organisms, the same genera were prevalent worldwide, with only a few abundant species in each genus. The abundance of dominant species was primarily shaped by process type, temperature, and continent. Based on MiDAS 4, we have developed an online field guide that presents all data and summarises present knowledge about all taxa in global WWTPs.