Dryland microbiomes reveal community adaptations to desertification and climate change.

Drylands account for 45% of the Earth's land area, supporting ~40% of the global population. These regions support some of the most extreme environments on Earth, characterized by extreme temperatures, low and variable rainfall, and low soil fertility. In these biomes, microorganisms provide vital ecosystem services and have evolved distinctive adaptation strategies to endure and flourish in the extreme. However, dryland microbiomes and the ecosystem services they provide are under threat due to intensifying desertification and climate change. In this review, we provide a synthesis of our current understanding of microbial life in drylands, emphasizing the remarkable diversity and adaptations of these communities. We then discuss anthropogenic threats, including the influence of climate change on dryland microbiomes and outline current knowledge gaps. Finally, we propose research priorities to address those gaps and safeguard the sustainability of these fragile biomes.

'Follow the Water': Microbial Water Acquisition in Desert Soils.

Water availability is the dominant driver of microbial community structure and function in desert soils. However, these habitats typically only receive very infrequent large-scale water inputs (e.g., from precipitation and/or run-off). In light of recent studies, the paradigm that desert soil microorganisms are largely dormant under xeric conditions is questionable. Gene expression profiling of microbial communities in desert soils suggests that many microbial taxa retain some metabolic functionality, even under severely xeric conditions. It, therefore, follows that other, less obvious sources of water may sustain the microbial cellular and community functionality in desert soil niches. Such sources include a range of precipitation and condensation processes, including rainfall, snow, dew, fog, and nocturnal distillation, all of which may vary quantitatively depending on the location and geomorphological characteristics of the desert ecosystem. Other more obscure sources of bioavailable water may include groundwater-derived water vapour, hydrated minerals, and metabolic hydro-genesis. Here, we explore the possible sources of bioavailable water in the context of microbial survival and function in xeric desert soils. With global climate change projected to have profound effects on both hot and cold deserts, we also explore the potential impacts of climate-induced changes in water availability on soil microbiomes in these extreme environments.

Iron acquisition and mineral transformation by cyanobacteria living in extreme environments.

Iron is an essential micronutrient for most living organisms, including cyanobacteria. These microorganisms have been found in Earth's driest polar and non-polar deserts, including the Atacama Desert, Chile. Iron-containing minerals were identified in colonized rock substrates from the Atacama Desert, however, the interactions between microorganisms and iron minerals remain unclear. In the current study, we determined that colonized gypsum rocks collected from the Atacama Desert contained both magnetite and hematite phases. A cyanobacteria isolate was cultured on substrates consisting of gypsum with embedded magnetite nanoparticles. Transmission electron microscopy imaging revealed a significant reduction in the size of magnetite nanoparticles due to their dissolution, which occurred around the microbial biofilms. Concurrently, hematite was detected, likely from the oxidation of the magnetite nanoparticles. Higher cell counts and production of siderophores were observed in cultures with magnetite nanoparticles suggesting that cyanobacteria were actively acquiring iron from the magnetite nanoparticles. Magnetite dissolution and iron acquisition by the cyanobacteria was further confirmed using large bulk magnetite crystals, uncovering a survival strategy of cyanobacteria in these extreme environments.

Adaptations of endolithic communities to abrupt environmental changes in a hyper-arid desert.

The adaptation mechanisms of microbial communities to natural perturbations remain unexplored, particularly in extreme environments. The extremophilic communities of halite (NaCl) nodules from the hyper-arid core of the Atacama Desert are self-sustained and represent a unique opportunity to study functional adaptations and community dynamics with changing environmental conditions. We transplanted halite nodules to different sites in the desert and investigated how their taxonomic, cellular, and biochemical changes correlated with water availability, using environmental data modeling and metagenomic analyses. Salt-in strategists, mainly represented by haloarchaea, significantly increased in relative abundance at sites characterized by extreme dryness, multiple wet/dry cycles, and colder conditions. The functional analysis of metagenome-assembled genomes (MAGs) revealed site-specific enrichments in archaeal MAGs encoding for the uptake of various compatible solutes and for glycerol utilization. These findings suggest that opportunistic salt-in strategists took over the halite communities at the driest sites. They most likely benefited from compounds newly released in the environment by the death of microorganisms least adapted to the new conditions. The observed changes were consistent with the need to maximize cellular bioenergetics when confronted with lower water availability and higher salinity, providing valuable information on microbial community adaptations and resilience to climate change.

Conversion of Ribosome-Protected mRNA to DNA and Deep Sequencing for Ribosome Profiling in Haloferax volcanii.

The translation of messenger RNA (mRNA) into protein is an essential process for all forms of life. The ability to monitor this process in a quantitative way by ribosome profiling-based approaches has revolutionized our ability to monitor protein synthesis in vivo and to explore and model complex cellular processes. Ribosome profiling is a high-throughput technique that globally analyzes the full set of ribosomes engaged in translation, providing insights into important aspects of the mechanism of protein synthesis and its regulation. This protocol covers the construction of a ribosome profiling library from culture harvesting, footprint isolation via ultracentrifugation, gel-based size fractionation, and footprint sequencing for a model halophilic archaeon, Haloferax volcanii. This approach has revealed the first global view of translation in the archaea.

Small RNA-Sequencing Library Preparation for the Halophilic Archaeon Haloferax volcanii.

Posttranscriptional regulation actuated by small RNAs (sRNAs) plays essential roles in a wide variety of cellular processes, especially in stress responses and environmental signaling. Hundreds of sRNAs have recently been discovered in archaea using genome-wide approaches but the molecular mechanisms of only a few have been characterized experimentally. Here, we describe how to build sRNA sequencing libraries using size-selected total RNA in the model archaeon, Haloferax volcanii , to provide a tool to further characterize sRNAs in archaea.

Viruses Ubiquity and Diversity in Atacama Desert Endolithic Communities.

Viruses are key players in the environment, and recent metagenomic studies have revealed their diversity and genetic complexity. Despite progress in understanding the ecology of viruses in extreme environments, viruses' dynamics and functional roles in dryland ecosystems, which cover about 45% of the Earth's land surfaces, remain largely unexplored. This study characterizes virus sequences in the metagenomes of endolithic (within rock) microbial communities ubiquitously found in hyper-arid deserts. Taxonomic classification and network construction revealed the presence of novel and diverse viruses in communities inhabiting calcite, gypsum, and ignimbrite rocks. Viral genome maps show a high level of protein diversity within and across endolithic communities and the presence of virus-encoded auxiliary metabolic genes. Phage-host relationships were predicted by matching tRNA, CRISPR spacer, and protein sequences in the viral and microbial metagenomes. Primary producers and heterotrophic bacteria were found to be putative hosts to some viruses. Intriguingly, viral diversity was not correlated with microbial diversity across rock substrates.

Adaptation of Cyanobacteria to the Endolithic Light Spectrum in Hyper-Arid Deserts.

In hyper-arid deserts, endolithic microbial communities survive in the pore spaces and cracks of rocks, an environment that enhances water retention and filters UV radiation. The rock colonization zone is enriched in far-red light (FRL) and depleted in visible light. This poses a challenge to cyanobacteria, which are the primary producers of endolithic communities. Many species of cyanobacteria are capable of Far-Red-Light Photoacclimation (FaRLiP), a process in which FRL induces the synthesis of specialized chlorophylls and remodeling of the photosynthetic apparatus, providing the ability to grow in FRL. While FaRLiP has been reported in cyanobacteria from various low-light environments, our understanding of light adaptations for endolithic cyanobacteria remains limited. Here, we demonstrated that endolithic Chroococcidiopsis isolates from deserts around the world synthesize chlorophyll f, an FRL-specialized chlorophyll when FRL is the sole light source. The metagenome-assembled genomes of these isolates encoded chlorophyll f synthase and all the genes required to implement the FaRLiP response. We also present evidence of FRL-induced changes to the major light-harvesting complexes of a Chroococcidiopsis isolate. These findings indicate that endolithic cyanobacteria from hyper-arid deserts use FRL photoacclimation as an adaptation to the unique light transmission spectrum of their rocky habitat.

Draft Metagenomes of Endolithic Cyanobacteria and Cohabitants from Hyper-Arid Deserts.

Cyanobacteria are essential to microbial communities inhabiting translucent rocks in hyper-arid deserts. Metagenomic studies revealed unique adaptations of these cyanobacteria, but validation of the corresponding metabolic pathways remained challenging without access to isolates. Here, we present high-quality metagenome-assembled genomes for cyanobacteria, and their heterotrophic companions, isolated from endolithic substrates.

Post-transcriptional regulation of redox homeostasis by the small RNA SHOxi in haloarchaea.

While haloarchaea are highly resistant to oxidative stress, a comprehensive understanding of the processes regulating this remarkable response is lacking. Oxidative stress-responsive small non-coding RNAs (sRNAs) have been reported in the model archaeon, Haloferax volc anii, but targets and mechanisms have not been elucidated. Using a combination of high throughput and reverse molecular genetic approaches, we elucidated the functional role of the most up-regulated intergenic sRNA during oxidative stress in H. volcanii, named Small RNA in Haloferax Oxidative Stress (SHOxi). SHOxi was predicted to form a stable secondary structure with a conserved stem-loop region as the potential binding site for trans-targets. NAD-dependent malic enzyme mRNA, identified as a putative target of SHOxi, interacted directly with a putative 'seed' region within the predicted stem loop of SHOxi. Malic enzyme catalyzes the oxidative decarboxylation of malate into pyruvate using NAD+ as a cofactor. The destabilization of malic enzyme mRNA, and the decrease in the NAD+/NADH ratio, resulting from the direct RNA-RNA interaction between SHOxi and its trans-target was essential for the survival of H. volcanii to oxidative stress. These findings indicate that SHOxi likely regulates redox homoeostasis during oxidative stress by the post-transcriptional destabilization of malic enzyme mRNA. SHOxi-mediated regulation provides evidence that the fine-tuning of metabolic cofactors could be a core strategy to mitigate damage from oxidative stress and confer resistance. This study is the first to establish the regulatory effects of sRNAs on mRNAs during the oxidative stress response in Archaea.

Cellular life from the three domains and viruses are transcriptionally active in a hypersaline desert community.

Microbial communities play essential roles in the biosphere and understanding the mechanisms underlying their functional adaptations to environmental conditions is critical for predicting their behaviour. This aspect of microbiome function has not been well characterized in natural high-salt environments. To address this knowledge gap, and to build a general framework relating the genomic and transcriptomic components in a microbiome, we performed a meta-omic survey of extremophile communities inhabiting halite (salt) nodules in the Atacama Desert. We found that the major phyla of this halophilic community have different levels of total transcriptional activity, at the selected time-points, and that different metabolic pathways were activated in their transcriptomes. We report that a novel Dolichomastix alga-the only eukaryote found in this system-was the most active community member. It produced the vast majority of the community's photosynthetic transcripts despite being outnumbered by Cyanobacteria. The divergence in the transcriptional landscapes of these segregated communities, compared with the relatively stable metagenomic functional potential, suggests that microbiomes in each salt nodule undergo unique transcriptional adjustments to adapt to local conditions. We also report the characterization of several previously unknown halophilic viruses, many of which exhibit transcriptional activity indicative of host infection.

Rock structure drives the taxonomic and functional diversity of endolithic microbial communities in extreme environments.

Endolithic (rock-dwelling) microbial communities are ubiquitous in hyper-arid deserts around the world and the last resort for life under extreme aridity. These communities are excellent models to explore biotic and abiotic drivers of diversity because they are of low complexity. Using high-throughput amplicon and metagenome sequencing, combined with X-ray computed tomography, we investigated how water availability and substrate architecture modulated the taxonomic and functional composition of gypsum endolithic communities in the Atacama Desert, Chile. We found that communities inhabiting gypsum rocks with a more fragmented substrate architecture had higher taxonomic and functional diversity, despite having less water available. This effect was tightly linked with community connectedness and likely the result of niche differentiation. Gypsum communities were functionally similar, yet adapted to their unique micro-habitats by modulating their carbon and energy acquisition strategies and their growth modalities. Reconstructed population genomes showed that these endolithic microbial populations encoded potential pathways for anoxygenic phototrophy and atmospheric hydrogen oxidation as supplemental energy sources.

Environmental Factors Driving Spatial Heterogeneity in Desert Halophile Microbial Communities.

Spatial heterogeneity in microbial communities is observed in all natural ecosystems and can stem from both adaptations to local environmental conditions as well as stochastic processes. Extremophile microbial communities inhabiting evaporitic halite nodules (salt rocks) in the Atacama Desert, Chile, are a good model ecosystem for investigating factors leading to microbiome heterogeneity, due to their diverse taxonomic composition and the spatial segregation of individual nodules. We investigated the abiotic factors governing microbiome composition across different spatial scales, allowing for insight into the factors that govern halite colonization from regional desert-wide scales to micro-scales within individual nodules. We found that water availability and community drift account for microbiome assembly differently at different distance scales, with higher rates of cell dispersion at the smaller scales resulting in a more homogenous composition. This trend likely applies to other endoliths, and to non-desert communities, where dispersion between communities is limited. At the intra-nodule scales, a light availability gradient was most important in determining the distribution of microbial taxa despite intermixing by water displacement via capillary action.

Mechanism of water extraction from gypsum rock by desert colonizing microorganisms.

Microorganisms, in the most hyperarid deserts around the world, inhabit the inside of rocks as a survival strategy. Water is essential for life, and the ability of a rock substrate to retain water is essential for its habitability. Here we report the mechanism by which gypsum rocks from the Atacama Desert, Chile, provide water for its colonizing microorganisms. We show that the microorganisms can extract water of crystallization (i.e., structurally ordered) from the rock, inducing a phase transformation from gypsum (CaSO4·2H2O) to anhydrite (CaSO4). To investigate and validate the water extraction and phase transformation mechanisms found in the natural geological environment, we cultivated a cyanobacterium isolate on gypsum rock samples under controlled conditions. We found that the cyanobacteria attached onto high surface energy crystal planes ({011}) of gypsum samples generate a thin biofilm that induced mineral dissolution accompanied by water extraction. This process led to a phase transformation to an anhydrous calcium sulfate, anhydrite, which was formed via reprecipitation and subsequent attachment and alignment of nanocrystals. Results in this work not only shed light on how microorganisms can obtain water under severe xeric conditions but also provide insights into potential life in even more extreme environments, such as Mars, as well as offering strategies for advanced water storage methods.

Ribosome profiling in archaea reveals leaderless translation, novel translational initiation sites, and ribosome pausing at single codon resolution.

High-throughput methods, such as ribosome profiling, have revealed the complexity of translation regulation in Bacteria and Eukarya with large-scale effects on cellular functions. In contrast, the translational landscape in Archaea remains mostly unexplored. Here, we developed ribosome profiling in a model archaeon, Haloferax volcanii, elucidating, for the first time, the translational landscape of a representative of the third domain of life. We determined the ribosome footprint of H. volcanii to be comparable in size to that of the Eukarya. We linked footprint lengths to initiating and elongating states of the ribosome on leadered transcripts, operons, and on leaderless transcripts, the latter representing 70% of H. volcanii transcriptome. We manipulated ribosome activity with translation inhibitors to reveal ribosome pausing at specific codons. Lastly, we found that the drug harringtonine arrested ribosomes at initiation sites in this archaeon. This drug treatment allowed us to confirm known translation initiation sites and also reveal putative novel initiation sites in intergenic regions and within genes. Ribosome profiling revealed an uncharacterized complexity of translation in this archaeon with bacteria-like, eukarya-like, and potentially novel translation mechanisms. These mechanisms are likely to be functionally essential and to contribute to an expanded proteome with regulatory roles in gene expression.

SnapShot: Microbial Extremophiles.

Extremophiles are remarkable examples of life's resilience, thriving in hot springs at boiling temperatures, in brine lakes saturated with salt, and in the driest deserts. We review the biogeography, currently known limits of life, and molecular adaptations to extremes. See the online interactive map for additional detail on biogeography, environmental microbiology, and exemplary species. To view this SnapShot, open or download the PDF.

Regulatory Noncoding Small RNAs Are Diverse and Abundant in an Extremophilic Microbial Community.

Regulatory small RNAs (sRNAs) play large-scale and essential roles in many cellular processes across all domains of life. Microbial sRNAs have been extensively studied in model organisms, but very little is known about the dynamics of sRNA synthesis and their roles in the natural environment. In this study, we discovered hundreds of intergenic (itsRNAs) and antisense (asRNAs) sRNAs expressed in an extremophilic microbial community inhabiting halite nodules (salt rocks) in the Atacama Desert. For this, we built SnapT, a new sRNA annotation pipeline that can be applied to any microbial community. We found asRNAs with expression levels negatively correlated with that of their overlapping putative target and itsRNAs that were conserved and significantly differentially expressed between 2 sampling time points. We demonstrated that we could perform target prediction and correlate expression levels between sRNAs and predicted target mRNAs at the community level. Functions of putative mRNA targets reflected the environmental challenges members of the halite communities were subjected to, including osmotic adjustments to a major rain event and competition for nutrients.IMPORTANCE Microorganisms in the natural world are found in communities, communicating and interacting with each other; therefore, it is essential that microbial regulatory mechanisms, such as gene regulation affected by small RNAs (sRNAs), be investigated at the community level. This work demonstrates that metatranscriptomic field experiments can link environmental variation with changes in RNA pools and have the potential to provide new insights into environmental sensing and responses in natural microbial communities through noncoding RNA-mediated gene regulation.

Current State of and Future Opportunities for Prediction in Microbiome Research: Report from the Mid-Atlantic Microbiome Meet-up in Baltimore on 9 January 2019.

Accurate predictions across multiple fields of microbiome research have far-reaching benefits to society, but there are few widely accepted quantitative tools to make accurate predictions about microbial communities and their functions. More discussion is needed about the current state of microbiome analysis and the tools required to overcome the hurdles preventing development and implementation of predictive analyses. We summarize the ideas generated by participants of the Mid-Atlantic Microbiome Meet-up in January 2019. While it was clear from the presentations that most fields have advanced beyond simple associative and descriptive analyses, most fields lack essential elements needed for the development and application of accurate microbiome predictions. Participants stressed the need for standardization, reproducibility, and accessibility of quantitative tools as key to advancing predictions in microbiome analysis. We highlight hurdles that participants identified and propose directions for future efforts that will advance the use of prediction in microbiome research.

Halophilic microbial community compositional shift after a rare rainfall in the Atacama Desert.

Understanding the mechanisms underlying microbial resistance and resilience to perturbations is essential to predict the impact of climate change on Earth's ecosystems. However, the resilience and adaptation mechanisms of microbial communities to natural perturbations remain relatively unexplored, particularly in extreme environments. The response of an extremophile community inhabiting halite (salt rocks) in the Atacama Desert to a catastrophic rainfall provided the opportunity to characterize and de-convolute the temporal response of a highly specialized community to a major disturbance. With shotgun metagenomic sequencing, we investigated the halite microbiome taxonomic composition and functional potential over a 4-year longitudinal study, uncovering the dynamics of the initial response and of the recovery of the community after a rainfall event. The observed changes can be recapitulated by two general modes of community shifts-a rapid Type 1 shift and a more gradual Type 2 adjustment. In the initial response, the community entered an unstable intermediate state after stochastic niche re-colonization, resulting in broad predicted protein adaptations to increased water availability. In contrast, during recovery, the community returned to its former functional potential by a gradual shift in abundances of the newly acquired taxa. The general characterization and proposed quantitation of these two modes of community response could potentially be applied to other ecosystems, providing a theoretical framework for prediction of taxonomic and functional flux following environmental changes.