Global diversity and inferred ecophysiology of microorganisms with the potential for dissimilatory sulfate/sulfite reduction

Abstract

Sulfate/sulfite-reducing microorganisms (SRM) are ubiquitous in nature, driving the global sulfur cycle. A hallmark of SRM is the dissimilatory sulfite reductase encoded by the genes dsrAB. Based on analysis of 950 mainly metagenome-derived dsrAB-carrying genomes, we redefine the global diversity of microorganisms with the potential for dissimilatory sulfate/sulfite reduction and uncover genetic repertoires that challenge earlier generalizations regarding their mode of energy metabolism. We show: (i) 19 out of 23 bacterial and 2 out of 4 archaeal phyla harbor uncharacterized SRM, (ii) four phyla including the Desulfobacterota harbor microorganisms with the genetic potential to switch between sulfate/sulfite reduction and sulfur oxidation, and (iii) the combination as well as presence/absence of different dsrAB-types, dsrL-types and dsrD provides guidance on the inferred direction of dissimilatory sulfur metabolism. We further provide an updated dsrAB database including > 60% taxonomically resolved, uncultured family-level lineages and recommendations on existing dsrAB-targeted primers for environmental surveys. Our work summarizes insights into the inferred ecophysiology of newly discovered SRM, puts SRM diversity into context of the major recent changes in bacterial and archaeal taxonomy, and provides an up-to-date framework to study SRM in a global context.

Keywords: dsrAB; dissimilatory sulfite reductase; metagenomics; sulfate reduction; sulfur cycle; sulfur oxidation.

Figure 1.

The pathway of dissimilatory sulfate reduction. The Dsr-pathway includes the enzymes sulfate adenylyltransferase (Sat), adenylyl phosphosulfate reductase (AprAB), dissimilatory sulfite reductase (DsrAB), and the sulfide releasing DsrC protein. The complexes QmoAB(C) and DsrMK(JOP) complement the pathway by transferring reducing equivalents towards AprAB and DsrC, respectively (Pereira et al. , Ramos et al. 2012, Santos et al. 2015). Reducing equivalents required by DsrAB can be delivered by a yet unknown oxidoreductase or DsrL (Löffler et al. 2020). DsrD acts as an allosteric activator of DsrAB in sulfate/sulfite-, thiosulfate-, and organosulfonate reducers as well as sulfur disproportionating microorganisms (Ferreira et al. 2022).

Figure 2.

Phylogeny and Dsr-pathway composition of DsrAB-encoding bacteria. Bacterial genome tree inferred from 120 concatenated proteins as based on the GTDB taxonomy (Parks et al. 2018). The phylogenomic tree was inferred from 902 bacterial (metagenome-assembled) genomes. The scale bar indicates 20% sequence divergence. The maximum likelihood tree was constructed with IQ-TREE (Nguyen et al. 2015) using automatic substitution model selection (LG + F + R10) and ultrafast bootstrap analysis (n = 1000). Bootstrap support is indicated by black dots (≥90%) or black circles (70–90%). Within each lineage, the presence of Dsr-pathway encoding genes was indicated if > 30% of dsrAB-containing genomes carried the respective genes as inferred by an automated hmm search (Zhou et al. , custom-made pHMM for DsrL: https://github.com/AnantharamanLab/Diao_et_al_2023).

Figure 3.

Phylogenetic analysis of bacterial DsrL proteins. The maximum likelihood tree was inferred from 438 DsrL proteins and constructed with IQ-TREE (Nguyen et al. 2015) using automatic substitution model selection (Q.pfam + I + R9) and ultrafast bootstrap analysis (n = 1000). Bootstrap support is shown by black dots (≥90%). DsrL sequences marked with an asterisk were not detected by the custom-made pHMM for DsrL (https://github.com/AnantharamanLab/Diao_et_al_2023), but have been previously described in the literature (Hausmann et al. , Löffler et al. 2020). Additional DsrL sequences (*1) were collected from MAGs with low completeness, which were not included in our MAG analysis.

Figure 4.

Organization of dsr gene clusters in MAGs encoding both reductive and oxidative bacterial-type DsrAB.

Figure 5.

Phylogeny and Dsr-pathway composition of DsrAB-encoding archaea. Archaeal genome tree inferred from 122 concatenated proteins as based on the GTDB taxonomy (Rinke et al. 2021). The phylogenetic tree was inferred from 48 archaeal (metagenome assembled) genomes. The scale bar indicates 20% sequence divergence. The maximum likelihood tree was constructed with IQ-TREE (Nguyen et al. 2015) using automatic substitution model selection (LG + F + R6) and ultrafast bootstrap analysis (n = 1000). Bootstrap support is indiacted by black dots (≥90%) or black circles (70%–90%). Within each lineage, the presence of Dsr-pathway encoding genes was indicated if > 30% of dsrAB-containing genomes carried the respective genes as inferred by an automated pHMM search (Zhou et al. 2022).

Figure 6.

Maximum likelihood phylogeny of DsrAB sequences derived from (metagenome assembled) genomes and environmental surveys. Clades represented by a majority of DsrAB-encoding (metagenome assembled) genomes not affiliated to canonical SRM or SOM are shown in magenta. The coverage of inferred phylogenetic clades by published broad-range PCR primers (≥75% of sequences in a clade; 1 mismatch allowed) is indicated by colored dots. The binding positions of the evaluated primers is indicated at the bottom using dsrAB of Desulfovibrio vulgaris or Allochromatiom vinosum as model organism of dsrAB primers designed to target bacterial-type dsrAB encoding the reductive or oxidative enzyme version, respectively. The maximum likelihood tree was constructed using deduced DsrAB amino acid sequences with IQ-TREE (Nguyen et al. 2015) using automatic substitution model selection (LG + R10) and ultrafast bootstrap analysis (n = 1000). Bootstrap support is indicated by black dots (≥90%) or black circles (70–90%). The tree was inferred from 613 representative DsrAB sequences with an indel filter covering 571 amino-acid positions: 346 representative DsrAB sequences were taken from a curated DsrAB database including 7921 pure culture and environmental sequences (as based on Müller et al. 2015) and amended with 267 DsrAB sequences derived from (metagenome assembled) genomes representing novel phylogenetic clades. Scale bar indicates 50% sequence divergence. Clades containing taxonomically resolved uncultured family-level DsrAB lineages are indicated by a superscript number based on the following denomination: 1, uncultured family-lineage 1; 3, uncultured family-lineage 3; 5, uncultured family-lineage 5; 6, uncultured family-lineage 6; 8, uncultured family-lineage 8; 9, uncultured family-lineage 9; 10, uncultured family-lineage 10; 11, uncultured family-lineage 11; 13 uncultured family-lineage 13. Please note that the numbers in brackets behind candidate phylum CG2-30-53–67 represent the two diverging dsrAB copies carried by the single MAG representing this phylum. LA-dsrAB, laterally acquired dsrAB.

Figure 7.

Comparison of phylogenomic and DsrAB trees for microorganisms representing all inferred DsrAB-encoding archaeal and bacterial lineages. The phylogenomic tree was inferred from a set of 43 conserved single-copy marker genes obtained with CheckM (Parks et al. 2015) using 38 representative archaeal and 207 representative bacterial (metagenome-assembled) genomes. The phylogenomic maximum likelihood tree was constructed with IQ-TREE (Nguyen et al. 2015) using automatic substitution model selection (LG + R10) and ultrafast bootstrap analysis (n = 1000). The DsrAB tree was constructed using 269 deduced DsrAB amino acid sequences, which were extracted from 245 representative (metagenome-assembled) genomes. The DsrAB maximum likelihood tree was constructed with IQ-TREE (Nguyen et al. 2015) using automatic substitution model selection (LG + R8) and ultrafast bootstrap analysis (n = 1000). Bootstrap support is indicated by black dots (≥90%) or black circles (70%–90%). For both trees, the scale bars indicates 20% sequence divergence.