Cueva de la Mora is a permanently stratified acidic pit lake and a model system for extreme acid mine drainage (AMD) studies. Using a combination of amplicon sequencing, metagenomics and metatranscriptomics we performed a taxonomically resolved analysis of microbial contributions to carbon, sulfur, iron, and nitrogen cycling. We found that active green alga Coccomyxa onubensis dominated the upper layer and chemocline. The chemocline had activity for iron(II) oxidation carried out by populations of Ca. Acidulodesulfobacterium, Ferrovum, Leptospirillium, and Armatimonadetes. Predicted activity for iron(III) reduction was only detected in the deep layer affiliated with Proteobacteria. Activity for dissimilatory nitrogen cycling including nitrogen fixation and nitrate reduction was primarily predicted in the chemocline. Heterotrophic archaeal populations with predicted activity for sulfide oxidation related to uncultured Thermoplasmatales dominated in the deep layer. Abundant sulfate-reducing Desulfomonile and Ca. Acidulodesulfobacterium populations were active in the chemocline. In the deep layer, uncultured populations from the bacterial phyla Actinobacteria, Chloroflexi, and Nitrospirae contributed to both sulfate reduction and sulfide oxidation. Based on this information we evaluated the potential for sulfide mineral precipitation in the deep layer as a tool for remediation. We argue that sulfide precipitation is not limited by microbial genetic potential but rather by the quantity and quality of organic carbon reaching the deep layer as well as by oxygen additions to the groundwater enabling sulfur oxidation. Addition of organic carbon and elemental sulfur should stimulate sulfate reduction and limit reoxidation of sulfide minerals.

Cueva de la Mora (CM) is an acidic, meromictic pit lake in the Iberian Pyrite Belt characterized by extremely high metal(loid) concentrations and strong gradients in oxygen, metal, and nutrient concentrations. We hypothesized that geochemical variations with depth would result in differences in community composition and in metal resistance strategies among active microbial populations. We also hypothesized that metal resistance gene (MRG) expression would correlate with toxicity levels for dissolved metal species in the lake. Water samples were collected in the upper oxic layer, chemocline, and deep anoxic layer of the lake for shotgun metagenomic and metatranscriptomic sequencing. Metagenomic analyses revealed dramatic differences in the composition of the microbial communities with depth, consistent with changing geochemistry. Based on relative abundance of taxa identified in each metagenome, Eukaryotes (predominantly Coccomyxa) dominated the upper layer, while Archaea (predominantly Thermoplasmatales) dominated the deep layer, and a combination of Bacteria and Eukaryotes were abundant at the chemocline. We compared metal resistance across communities using a curated list of protein-coding MRGs with KEGG Orthology identifiers (KOs) and found that there were broad differences in the metal resistance strategies (e.g., intracellular metal accumulation) expressed by Eukaryotes, Bacteria, and Archaea. Although normalized abundances of MRG and MRG expression were generally higher in the deep layer, expression of metal-specific genes was not strongly related to variations in specific metal concentrations, especially for Cu and As. We also compared MRG potential and expression in metagenome assembled genomes (MAGs) from the deep layer, where metal concentrations are highest. Consistent with previous work showing differences in metal resistance mechanisms even at the strain level, MRG expression patterns varied strongly among MAG populations from the same depth. Some MAG populations expressed very few MRG known to date, suggesting that novel metal resistance strategies remain to be discovered in uncultivated acidophiles.

Deep chlorophyll maxima (DCM) and metalimnetic oxygen maxima (MOM) are outstanding biogeochemical features of acidic pit lakes (APL). However, knowledge of the eukaryotic phototrophs responsible for their formation is limited. We aimed at linking the dynamics of phototrophic communities inhabiting meromictic APL in Spain with the formation of these characteristic layers. Firstly, the dynamics of DCM and MOM and their relation to physico-chemical parameters (photosynthetically active radiation (PAR), pH, dissolved ferric iron concentration, temperature), pigments and nutrient distribution is described; secondly, the phototrophic community composition is studied through a combination of microscopy, biomolecular and “omics” tools. Phototrophic communities of the studied APL show a low diversity dominated by green microalgae, specifically Coccomyxa sp., which have been successfully adapted to the chemically harsh conditions. DCM and MOM are usually non-coincident. DCM correspond to layers where phototrophs have higher chlorophyll content per cell to cope with extremely low PAR (<1 µmol m⁻² s⁻¹), but where photosynthetic oxygen production is limited. MOM correspond to shallower waters with more light, higher phytoplankton biomass and intense photosynthetic activity, which affects both oxygen concentration and water temperature. The main drivers of DCM formation in these APL are likely the need for nutrient uptake and photo-acclimation.

Acidophilic sulphate‐reducing bacteria (aSRB) are widespread anaerobic microorganisms that perform dissimilatory sulphate reduction and have key adaptations to tolerate acidic environments (pH <5.0), such as proton impermeability and Donnan potential. This diverse prokaryotic group is of interest from physiological, ecological, and applicational viewpoints. In this review, we summarize the interactions between aSRB and other microbial guilds, such as syntrophy, and their roles in the biogeochemical cycling of sulphur, iron, carbon, and other elements. We discuss the biotechnological applications of aSRB in treating acid mine drainage (AMD, pH <3), focusing on their ability to produce biogenic sulphide and precipitate metals, particularly in the context of utilizing microbial consortia instead of pure isolates. Metal sulphide nanoparticles recovered after AMD treatment have multiple potential technological uses, including in electronics and biomedicine, contributing to a cost‐effective circular economy. The products of aSRB metabolisms, such as biominerals and isotopes, could also serve as biosignatures to understand ancient and extant microbial life in the universe. Overall, aSRB are active components of the sulphur and carbon cycles under acidic conditions, with potential natural and technological implications for the world around us.