Acidithiobacillia class members originating at sites within the Pacific Ring of Fire and other tectonically active locations and description of the novel genus 'Igneacidithiobacillus'.

Recent studies have expanded the genomic contours of the Acidithiobacillia, highlighting important lacunae in our comprehension of the phylogenetic space occupied by certain lineages of the class. One such lineage is 'Igneacidithiobacillus', a novel genus-level taxon, represented by 'Igneacidithiobacillus copahuensis' VAN18-1T as its type species, along with two other uncultivated metagenome-assembled genomes (MAGs) originating from geothermally active sites across the Pacific Ring of Fire. In this study, we investigate the genetic and genomic diversity, and the distribution patterns of several uncharacterized Acidithiobacillia class strains and sequence clones, which are ascribed to the same 16S rRNA gene sequence clade. By digging deeper into this data and contributing to novel MAGs emerging from environmental studies in tectonically active locations, the description of this novel genus has been consolidated. Using state-of-the-art genomic taxonomy methods, we added to already recognized taxa, an additional four novel Candidate (Ca.) species, including 'Ca. Igneacidithiobacillus chanchocoensis' (mCHCt20-1TS), 'Igneacidithiobacillus siniensis' (S30A2T), 'Ca. Igneacidithiobacillus taupoensis' (TVZ-G3 TS), and 'Ca. Igneacidithiobacillus waiarikiensis' (TVZ-G4 TS). Analysis of published data on the isolation, enrichment, cultivation, and preliminary microbiological characterization of several of these unassigned or misassigned strains, along with the type species of the genus, plus the recoverable environmental data from metagenomic studies, allowed us to identify habitat preferences of these taxa. Commonalities and lineage-specific adaptations of the seven species of the genus were derived from pangenome analysis and comparative genomic metabolic reconstruction. The findings emerging from this study lay the groundwork for further research on the ecology, evolution, and biotechnological potential of the novel genus 'Igneacidithiobacillus'.

Genomic evolution of the class Acidithiobacillia: deep-branching Proteobacteria living in extreme acidic conditions.

Members of the genus Acidithiobacillus, now ranked within the class Acidithiobacillia, are model bacteria for the study of chemolithotrophic energy conversion under extreme conditions. Knowledge of the genomic and taxonomic diversity of Acidithiobacillia is still limited. Here, we present a systematic analysis of nearly 100 genomes from the class sampled from a wide range of habitats. Some of these genomes are new and others have been reclassified on the basis of advanced genomic analysis, thus defining 19 Acidithiobacillia lineages ranking at different taxonomic levels. This work provides the most comprehensive classification and pangenomic analysis of this deep-branching class of Proteobacteria to date. The phylogenomic framework obtained illuminates not only the evolutionary past of this lineage, but also the molecular evolution of relevant aerobic respiratory proteins, namely the cytochrome bo3 ubiquinol oxidases.

Defining Culture Conditions for the Hidden Nitrite-Oxidizing Bacterium Nitrolancea.

Nitrification is a key process for N-removal in engineered and natural environments, but recent findings of novel nitrifying microorganisms with surprising features revealed that our knowledge of this functional guild is still incomplete. Especially nitrite oxidation - the second step of nitrification - is catalyzed by a phylogenetically diverse bacterial group, and only recently bacteria of the phylum Chloroflexi have been identified as thermophilic nitrite-oxidizing bacteria (NOB). Among these, Nitrolancea hollandica was isolated from a laboratory-scale nitrifying bioreactor operated at 35°C with a high load of ammonium bicarbonate. However, its distribution remains cryptic as very few closely related environmental 16S rRNA gene sequences have been retrieved so far. In this study, we demonstrate how such thermophilic NOB can be enriched using modified mineral media inoculated with samples from a wastewater side-stream reactor operated at 39.5°C. Distinct cultivation conditions resulted in quick and reproducible high enrichment of two different strains of Nitrolancea, closely related to Nl. hollandica. The same cultivation approach was applied to a complex nitrite-oxidizing pre-enrichment at 42°C inoculated with biomass from a geothermal spring in the Copahue volcano area in Neuquen, Argentina. Here, an additional distinct representative of the genus Nitrolancea was obtained. This novel species had 16S rRNA and nitrite oxidoreductase alpha subunit (nxrA) gene sequence identities to Nl. hollandica of 98.5% and 97.2%, respectively. A genomic average nucleotide identity between the Argentinian strain and Nl. hollandica of 91.9% indicates that it indeed represents a distinct species. All Nitrolancea cultures formed lancet-shaped cells identical to Nl. hollandica and revealed similar physiological features, including the capability to grow at high nitrite concentrations. Growth was optimal at temperatures of 35-37°C and was strongly enhanced by ammonium supplementation. Genomic comparisons revealed that the four Nitrolancea strains share 2399 out of 3387 orthologous gene clusters and encode similar key functions. Our results define general growth conditions that enable the selective enrichment of Nitrolancea from artificial and natural environments. In most natural habitats these NOB apparently are of low abundance and their proliferation depends on the balanced presence of nitrite and ammonium, with an optimal incubation temperature of 37°C.

Enrichment and isolation of acid-tolerant sulfate-reducing microorganisms in the anoxic, acidic hot spring sediments from Copahue volcano, Argentina.

The geothermal Copahue-Caviahue (GCC) system (Argentina) is an extreme acidic environment, dominated by the activity of Copahue volcano. Environments characterised by low pH values, such as volcanic areas, are of particular interest for the search of acidophilic microorganisms with application in biotechnological processes. In this work, sulfate-reducing microorganisms were investigated in geothermal acidic, anaerobic zones from GCC system. Sediment samples from Agua del Limón (AL1), Las Máquinas (LMa2), Las Maquinitas (LMi) and Baño 9 (B9-2, B9-3) were found to be acidic (pH values 2.1-3.0) to moderate acidic (5.1-5.2), containing small total organic carbon values, and ferric iron precipitates. The organic electron donor added to the enrichment was completely oxidised to CO2. Bacteria related to 'Desulfobacillus acidavidus' strain CL4 were found to be dominant (67-83% of the total number of clones) in the enrichment cultures, and their presence was confirmed by their isolation on overlay plates. Other bacteria were also detected with lower abundance (6-20% of the total number of clones), with representatives of the genera Acidithiobacillus, Sulfobacillus, Alicyclobacillus and Athalassotoga/Mesoaciditoga. These enrichment and isolates found at low pH confirm the presence of anaerobic activities in the acidic sediments from the geothermal Copahue-Caviahue system.

Domestication of Local Microbial Consortia for Efficient Recovery of Gold Through Top-Down Selection in Airlift Bioreactors.

Extreme acidophiles play central roles in the geochemical cycling of diverse elements in low pH environments. This has been harnessed in biotechnologies such as biomining, where microorganisms facilitate the recovery of economically important metals such as gold. By generating both extreme acidity and a chemical oxidant (ferric iron) many species of prokaryotes that thrive in low pH environments not only catalyze mineral dissolution but also trigger both community and individual level adaptive changes. These changes vary in extent and direction depending on the ore mineralogy, water availability and local climate. The use of indigenous versus introduced microbial consortia in biomining practices is still a matter of debate. Yet, indigenous microbial consortia colonizing sulfidic ores that have been domesticated, i.e., selected for their ability to survive under specific polyextreme conditions, are claimed to outperform un-adapted foreign consortia. Despite this, little is known on the domestication of acidic microbial communities and the changes elicited in their members. In this study, high resolution targeted metagenomic techniques were used to analyze the changes occurring in the community structure of local microbial consortia acclimated to growing under extreme acidic conditions and adapted to endure the conditions imposed by the target mineral during biooxidation of a gold concentrate in an airlift reactor over a period of 2 years. The results indicated that operative conditions evolving through biooxidation of the mineral concentrate exerted strong selective pressures that, early on, purge biodiversity in favor of a few Acidithiobacillus spp. over other iron oxidizing acidophiles. Metagenomic analysis of the domesticated consortium present at the end of the adaptation experiment enabled reconstruction of the RVS1-MAG, a novel representative of Acidithiobacillus ferrooxidans from the Andacollo gold mineral district. Comparative genomic analysis performed with this genome draft revealed a net enrichment of gene functions related to heavy metal transport and stress management that are likely to play a significant role in adaptation and survival to adverse conditions experienced by these acidophiles during growth in presence of gold concentrates.

A Deeper Look into the Biodiversity of the Extremely Acidic Copahue volcano-Río Agrio System in Neuquén, Argentina.

The Copahue volcano-Río Agrio system, on Patagonia Argentina, comprises the naturally acidic river Río Agrio, that runs from a few meters down the Copahue volcano crater to more than 40 km maintaining low pH waters, and the acidic lagoon that sporadically forms on the crater of the volcano, which is studied for the first time in this work. We used next-generation sequencing of the 16S rRNA gene of the entire prokaryotic community to study the biodiversity of this poorly explored extreme environment. The correlation of the operational taxonomic units (OTUs)s presence with physicochemical variables showed that the system contains three distinct environments: the crater lagoon, the Upper Río Agrio, and the Salto del Agrio waterfall, a point located approximately 12 km down the origin of the river, after it emerges from the Caviahue lake. The prokaryotic community of the Copahue Volcano-Río Agrio system is mainly formed by acidic bacteria and archaea, such as Acidithiobacillus, Ferroplasma, and Leptospirillum, which have been isolated from similar environments around the world. These results support the idea of a ubiquitous acidic biodiversity; however, this highly interesting extreme environment also has apparently autochthonous species such as Sulfuriferula, Acidianus copahuensis, and strains of Acidibacillus and Alicyclobacillus.