MiDAS 4: A global catalogue of full-length 16S rRNA gene sequences and taxonomy for studies of bacterial communities in wastewater treatment plants

Abstract

Microbial communities are responsible for biological wastewater treatment, but our knowledge of their diversity and function is still poor. Here, we sequence more than 5 million high-quality, full-length 16S rRNA gene sequences from 740 wastewater treatment plants (WWTPs) across the world and use the sequences to construct the ‘MiDAS 4’ database. MiDAS 4 is an amplicon sequence variant resolved, full-length 16S rRNA gene reference database with a comprehensive taxonomy from domain to species level for all sequences. We use an independent dataset (269 WWTPs) to show that MiDAS 4, compared to commonly used universal reference databases, provides a better coverage for WWTP bacteria and an improved rate of genus and species level classification. Taking advantage of MiDAS 4, we carry out an amplicon-based, global-scale microbial community profiling of activated sludge plants using two common sets of primers targeting regions of the 16S rRNA gene, revealing how environmental conditions and biogeography shape the activated sludge microbiota. We also identify core and conditionally rare or abundant taxa, encompassing 966 genera and 1530 species that represent approximately 80% and 50% of the accumulated read abundance, respectively. Finally, we show that for well-studied functional guilds, such as nitrifiers or polyphosphate-accumulating organisms, the same genera are prevalent worldwide, with only a few abundant species in each genus.

Reevaluation of the Phylogenetic Diversity and Global Distribution of the Genus “CandidatusAccumulibacter”

“…The alignment used for the tree applied a 20% conservational filter to remove hypervariable positions, giving 1,250 aligned positions. Coverage of the fluorescence in situ hybridization (FISH) probes designed in this study is indicated with black brackets and is based on the MiDAS4 database ( 29 ). Probe coverage of widely applied probes for the “ Ca.…”

“…Despite the lack of resolution of 16S rRNA gene amplicon studies for these lineages, the new MiDAS global reference database of full-length 16S rRNA gene sequences ( 29 ) ensures that amplicon analysis is still a powerful tool to analyze their geographical distribution. On a global scale, MiDAS-defined “ Ca.…”

“…Accumulibacter” is needed for confident application in future studies. This could benefit from the use of comprehensive and ecosystem-specific full-length 16S rRNA gene reference databases, such as MiDAS3 ( 27 , 28 ) and MiDAS4 ( 29 ), which facilitate the analysis of the microbial diversity in WWTPs with species-level resolution.…”

Disentangle genus microdiversity within a complex microbial community by using a multi‐distance long‐read binning method: example of Candidatus Accumulibacter

“…1.3 (McIlroy et al, 2017). This affiliation was confirmed with online 16S rRNA databases MiDAS v4.8 (Dueholm et al, 2021) and EzBioCloud (Yoon et al, 2017), but GTDB-tk v.1.4.1 by (Chaumeil et al, 2020) places ACC004 within the genus Propionivibrio, with an ANI value of 78.94%, an AF value of 0.39 and a RED value of 0.94. Only three different rRNA gene sequences were extracted from the hybrid co-assemblies, ACC002, ACC006 and ACC008.…”

“…The 16S rRNA sequence of ACC004 was affiliated with a Ca. Accumulibacter with 99.0% of identity in MiDAS and EzBiocloud (Yoon et al, 2017;Dueholm et al, 2021). The identity threshold for 16S rRNA gene identity between the members of a genus proposed by Yarza et al (2014) is here fully respected.…”