Despite common perception, most of Earth is what is often referred to as an \'extreme environment.\' Yet to the organisms that call these places home, it is simply that (home). They have adapted to thrive in these environments and, in the process, have evolved many unique adaptations at the molecular- and \'omic-level. Scientists\' interest in these organisms has typically been in how they and their products can be harnessed for biotechnological applications and the environments where they are found, while the general public\'s veers more toward a fascination with their deviation from the \'norm\'. However, these organisms have so much more to tell us about Life and the myriad ways there are to perform \'simple\' biological processes.

Microorganisms that can withstand high pressure within an environment are termed piezophiles. These organisms are considered extremophiles and inhabit the deep marine or terrestrial subsurface. Because these microorganisms are not easily accessed and require expensive sampling methods and laboratory instruments, advancements in this field have been limited compared to other extremophiles. This review summarizes the current knowledge on piezophiles, notably the cellular and physiological adaptations that such microorganisms possess to withstand and grow in high-pressure environments. Based on existing studies, organisms from both the deep marine and terrestrial subsurface show similar adaptations to high pressure, including increased motility, an increase of unsaturated bonds within the cell membrane lipids, upregulation of heat shock proteins, and differential gene-regulation systems. Notably, more adaptations have been identified within the deep marine subsurface organisms due to the relative paucity of studies performed on deep terrestrial subsurface environments. Nevertheless, similar adaptations have been found within piezophiles from both systems, and therefore the microbial biogeography concepts used to assess microbial dispersal and explore if similar organisms can be found throughout deep terrestrial environments are also briefly discussed.

The deep-sea is characterized by extreme conditions, such as high hydrostatic pressure (HHP) and near-freezing temperature. Piezophiles, microorganisms adapted to high pressure, have developed key strategies to maintain the integrity of their lipid membrane at these conditions. The abundance of specific membrane lipids, such as those containing unsaturated and branched-chain fatty acids, rises with increasing HHP. Nevertheless, this strategy is not universal among piezophiles, highlighting the need to further understand the effects of HHP on microbial lipid membranes. Challenges in the study of lipid membrane adaptations by piezophiles also involve methodological developments, cross-adaptation studies, and insight into slow-growing piezophiles. Moreover, the effects of HHP on piezophiles are often difficult to disentangle from effects caused by low temperature that are often characteristic of the deep sea. Here, we review the knowledge of membrane lipid adaptation strategies of piezophiles, and put it into the perspective of marine systems, highlighting the future challenges of research studying the effects of HHP on the microbial lipid composition.

Protein adaptations to extreme environmental conditions are drivers in biotechnological process optimization and essential to unravel the molecular limits of life. Most proteins with such desirable adaptations are found in extremophilic organisms inhabiting extreme environments. The deep sea is such an environment and a promising resource that poses multiple extremes on its inhabitants. Conditions like high hydrostatic pressure and high or low temperature are prevalent and many deep-sea organisms tolerate multiple of these extremes. While molecular adaptations to high temperature are comparatively good described, adaptations to other extremes like high pressure are not well-understood yet. To fully unravel the molecular mechanisms of individual adaptations it is probably necessary to disentangle multifactorial adaptations. In this study, we evaluate differences of protein structures from deep-sea organisms and their respective related proteins from nondeep-sea organisms. We created a data collection of 1281 experimental protein structures from 25 deep-sea organisms and paired them with orthologous proteins. We exhaustively evaluate differences between the protein pairs with machine learning and Shapley values to determine characteristic differences in sequence and structure. The results show a reasonable discrimination of deep-sea and nondeep-sea proteins from which we distinguish correlations previously attributed to thermal stability from other signals potentially describing adaptions to high pressure. While some distinct correlations can be observed the overall picture appears intricate.

Hadal snailfishes are the deepest-living fishes in the ocean, inhabiting trenches from depths of ∼6,000 to 8,000 m. While the microbial communities in trench environments have begun to be characterized, the microbes associated with hadal megafauna remain relatively unknown. Here, we describe the gut microbiomes of two hadal snailfishes, Pseudoliparis swirei (Mariana Trench) and Notoliparis kermadecensis (Kermadec Trench), using 16S rRNA gene amplicon sequencing. We contextualize these microbiomes with comparisons to the abyssal macrourid Coryphaenoides yaquinae and the continental shelf-dwelling snailfish Careproctus melanurus. The microbial communities of the hadal snailfishes were distinct from their shallower counterparts and were dominated by the same sequences related to the Mycoplasmataceae and Desulfovibrionaceae. These shared taxa indicate that symbiont lineages have remained similar to the ancestral symbiont since their geographic separation or that they are dispersed between geographically distant trenches and subsequently colonize specific hosts. The abyssal and hadal fishes contained sequences related to known, cultured piezophiles, microbes that grow optimally under high hydrostatic pressure, including Psychromonas, Moritella, and Shewanella. These taxa are adept at colonizing nutrient-rich environments present in the deep ocean, such as on particles and in the guts of hosts, and we hypothesize they could make a dietary contribution to deep-sea fishes by degrading chitin and producing fatty acids. We characterize the gut microbiota within some of the deepest fishes to provide new insight into the diversity and distribution of host-associated microbial taxa and the potential of these animals, and the microbes they harbor, for understanding adaptation to deep-sea habitats. IMPORTANCE Hadal trenches, characterized by high hydrostatic pressures and low temperatures, are one of the most extreme environments on our planet. By examining the microbiome of abyssal and hadal fishes, we provide insight into the diversity and distribution of host-associated life at great depth. Our findings show that there are similar microbial populations in fishes geographically separated by thousands of miles, reflecting strong selection for specific microbial lineages. Only a few psychropiezophilic taxa, which do not reflect the diversity of microbial life at great depth, have been successfully isolated in the laboratory. Our examination of deep-sea fish microbiomes shows that typical high-pressure culturing methodologies, which have largely remained unchanged since the pioneering work of Claude ZoBell in the 1950s, may simulate the chemical environment found in animal guts and helps explain why the same deep-sea genera are consistently isolated.

Deep-sea ecosystems share a common physical parameter, namely high hydrostatic pressure (HHP). Some of the microorganisms isolated at great depths have a high physiological plasticity to face pressure variations. The adaptive strategies by which deep-sea microorganisms cope with HHP variations remain to be elucidated, especially considering the extent of their biotopes on Earth. Herein, we investigated the gene expression patterns of Thermococcus piezophilus, a piezohyperthermophilic archaeon isolated from the deepest hydrothermal vent known to date, under sub-optimal, optimal and supra-optimal pressures (0.1, 50, and 90 MPa, respectively). At stressful pressures [sub-optimal (0.1 MPa) and supra-optimal (90 MPa) conditions], no classical stress response was observed. Instead, we observed an unexpected transcriptional modulation of more than a hundred gene clusters, under the putative control of the master transcriptional regulator SurR, some of which are described as being involved in energy metabolism. This suggests a fine-tuning effect of HHP on the SurR regulon. Pressure could act on gene regulation, in addition to modulating their expression.

A novel extreme thermophilic and piezophilic chemoorganoheterotrophic archaeon, strain EXT12cT, was isolated from a hydrothermal chimney sample collected at a depth of 2496 m at the East Pacific Rise 9° N. Cells were strictly anaerobic, motile cocci. The strain grew at NaCl concentrations ranging from 1 to 5 % (w/v; optimum, 2.0%), from pH 6.0 to 7.5 (optimum, pH 6.5-7.0), at temperatures between 60 and 95 °C (optimum, 80-85 °C), and at pressures from 0.1 to at least 50 MPa (optimum, 30 MPa). Strain EXT12cT grew chemoorganoheterotrophically on complex proteinaceous substrates. Its growth was highly stimulated by the presence of elemental sulphur or l-cystine, which were reduced to hydrogen sulfide. The DNA G+C content was 54.58 mol%. Phylogenetic analyses based on 16S rRNA gene sequences and concatenated ribosomal protein sequences showed that strain EXT12cT falls into the genus Thermococcus and is most closely related to Thermococcus nautili strain 30-1T. Overall genome relatedness index analyses (average nucleotide identity scores and in silico DNA-DNA hybridizations) showed a sufficient genomic distance between the new genome and the ones of the Thermococcus type strains for the delineation of a new species. On the basis of genotypic and phenotypic data, strain EXT12cT is considered to represent a novel species, for which the name Thermococcus henrietii sp. nov. is proposed, with the type strain EXT12cT (=UBOCC M-2417T=DSM 111004T).

Microorganisms living in deep-oil reservoirs face extreme conditions of elevated temperature and hydrostatic pressure. Within these microbial communities, members of the order Thermotogales are predominant. Among them, the genus Pseudothermotoga is widespread in oilfield-produced waters. The growth and cell phenotypes under hydrostatic pressures ranging from 0.1 to 50 MPa of two strains from the same species originating from subsurface, Pseudothermotoga elfii DSM9442 isolated from a deep African oil-producing well, and surface, P. elfii subsp. lettingae isolated from a thermophilic sulfate-reducing bioreactor, environments are reported for the first time. The data support evidence for the piezophilic nature of P. elfii DSM9442, with an optimal hydrostatic pressure for growth of 20 MPa and an upper limit of 40 MPa, and the piezotolerance of P. elfii subsp. lettingae with growth occurring up to 20 MPa only. Under the experimental conditions, both strains produce mostly acetate and propionate as volatile fatty acids with slight variations with respect to the hydrostatic pressure for P. elfii DSM9442. The data show that the metabolism of P. elfii DSM9442 is optimized when grown at 20 MPa, in agreement with its piezophilic nature. Both Pseudothermotoga strains form chained cells when the hydrostatic pressure increases, especially P. elfii DSM9442 for which 44% of cells is chained when grown at 40 MPa. The viability of the chained cells increases with the increase in the hydrostatic pressure, indicating that chain formation is a protective mechanism for P. elfii DSM9442.

BACKGROUND: The deep ocean is characterized by low temperatures, high hydrostatic pressures, and low concentrations of organic matter. While these conditions likely select for distinct genomic characteristics within prokaryotes, the attributes facilitating adaptation to the deep ocean are relatively unexplored. In this study, we compared the genomes of seven strains within the genus Colwellia, including some of the most piezophilic microbes known, to identify genomic features that enable life in the deep sea.
RESULTS: Significant differences were found to exist between piezophilic and non-piezophilic strains of Colwellia. Piezophilic Colwellia have a more basic and hydrophobic proteome. The piezophilic abyssal and hadal isolates have more genes involved in replication/recombination/repair, cell wall/membrane biogenesis, and cell motility. The characteristics of respiration, pilus generation, and membrane fluidity adjustment vary between the strains, with operons for a nuo dehydrogenase and a tad pilus only present in the piezophiles. In contrast, the piezosensitive members are unique in having the capacity for dissimilatory nitrite and TMAO reduction. A number of genes exist only within deep-sea adapted species, such as those encoding d-alanine-d-alanine ligase for peptidoglycan formation, alanine dehydrogenase for NADH/NAD+ homeostasis, and a SAM methyltransferase for tRNA modification. Many of these piezophile-specific genes are in variable regions of the genome near genomic islands, transposases, and toxin-antitoxin systems.
CONCLUSIONS: We identified a number of adaptations that may facilitate deep-sea radiation in members of the genus Colwellia, as well as in other piezophilic bacteria. An enrichment in more basic and hydrophobic amino acids could help piezophiles stabilize and limit water intrusion into proteins as a result of high pressure. Variations in genes associated with the membrane, including those involved in unsaturated fatty acid production and respiration, indicate that membrane-based adaptations are critical for coping with high pressure. The presence of many piezophile-specific genes near genomic islands highlights that adaptation to the deep ocean may be facilitated by horizontal gene transfer through transposases or other mobile elements. Some of these genes are amenable to further study in genetically tractable piezophilic and piezotolerant deep-sea microorganisms.

Hyperthermophiles, living in environments above 80°C and usually coupling with multi-extreme environmental stresses, have drawn great attention due to their application potential in biotechnology and being the primitive extant forms of life. Studies on their survival and adaptation mechanisms have extended our understanding on how lives thrive under extreme conditions. During these studies, the \"cross-stress\" behavior in various organisms has been observed between the extreme high temperature and other environmental stresses. Despite the broad observation, the global view of the cross-stress behavior remains unclear in hyperthermophiles, leaving a knowledge gap in our understanding of extreme adaptation. In this study, we performed a global quantitative proteomic analysis under extreme temperatures, pH, hydrostatic pressure (HP), and salinity on an archaeal strain, Thermococcus eurythermalis A501, which has outstanding growth capability on a wide range of temperatures (50-100°C), pH (4-9), and HPs (0.1-70 MPa), but a narrow range of NaCl (1.0-5.0 %, w/v). The proteomic analysis (79.8% genome coverage) demonstrated that approximately 61.5% of the significant differentially expressed proteins (DEPs) responded to multiple stresses. The responses to most of the tested stresses were closely correlated, except the responses to high salinity and low temperature. The top three enriched universal responding processes include the biosynthesis and protection of macromolecules, biosynthesis and metabolism of amino acids, ion transport, and binding activities. In addition, this study also revealed that the specific dual-stress responding processes, such as the membrane lipids for both cold and HP stresses and the signal transduction for both hyperosmotic and heat stresses, as well as the sodium-dependent energetic processes might be the limiting factor of the growth range in salinity. The present study is the first to examine the global cross-stress responses in a piezophilic hyperthermophile at the proteomic level. Our findings provide direct evidences of the cross-stress adaptation strategy (33.5% of coding-genes) to multiple stresses and highlight the specific and unique responding processes (0.22-0.63% of coding genes for each) to extreme temperature, pH, salinity, and pressure, which are highly relevant to the fields of evolutionary biology as well as next generation industrial biotechnology (NGIB).