Palaeococcus pacificus DY20341T, a hyperthermophilic archaeon from deep sea hydrothermal sediments, grows at temperature range from 50 to 90 ℃ (Topt = 80℃). To explore the molecular basis of its thermal adaptation, we identified six representative hyperthermostable proteins from its cytosolic fraction that remained soluble after a stringent in vitro heat treatment at 100 °C. MALDI-TOF mass spectrometry identified these as an inorganic pyrophosphatase (PPase), a 5’-methylthioadenosine phosphorylase (MTAP), and four putative transporter components (one TRAP and three ABC transporter solute-binding proteins). RT-qPCR analysis revealed that the transcription of all six corresponding genes was significantly upregulated when the growth temperature was elevated from 80 °C to 90 °C, suggesting a potential role in the cellular response to heat stress. Biochemical characterization of the recombinant PPase demonstrated remarkable thermostability and an optimal temperature of 90 °C. Kinetic analysis revealed high catalytic efficiency, with a maximal specific activity of 590 ± 30 U/mg and a strong substrate affinity ( K_m = 6.24 µM). Phylogenetic analysis of the identified thermostable MTAP (PAP_00250) revealed its evolutionary origin from hyperthermophilic archaea, distinguishing it clearly from a second MTAP homolog (PAP_07050) in the genome, which aligns with mesophilic bacteria. The structural stability of the identified ABC and TRAP transporter components hypothesizes that efficient nutrient scavenging remains critical at extreme temperatures. Together, these findings provide valuable insights into the stable molecular machinery and potential physiological strategies enabling P. pacificus to thrive in hydrothermal environments.
This study aimed to evaluate L-asparaginase (L-ASNase) production by fungi isolated from soils across different Antarctic islands. Out of 138 isolates, 33 filamentous fungi and 5 yeasts exhibited enzymatic activity in solid culture media. Filamentous fungi were identified by sequencing the Internal Transcribed Spacer (ITS) region, while yeasts were identified using the D1/D2 domains of the 26 S rDNA gene. The blue filamentous fungus Antarctomyces F13.CR.LASP exhibited a maximum L-ASNase productivity of 0.76 U/L. Regarding the yeasts, the isolate Vishniacozyma victoriae G.L11 reached a productivity of 0.17 U/L, while corresponding values for glutaminase and urease were 0.11 and 0.06 U/L, respectively. Furthermore, V. victoriae G.L11 was capable of producing the enzyme using different low-cost substrates, particularly pineapple peel waste, which yielded 5.17 U/L. In this scenario, future optimization could further enhance L-asparaginase production levels by these fungi, providing valuable insights into the biotechnological potential of Antarctic fungi.
Understanding the coordinated regulation of gene expression in response to environmental challenges is critical for unravelling the resistance strategies of the extremophiles that endure extreme habitats. Deinococcus radiodurans is a notable extremophile for rebuilding entire genome from the fragmented DNA, a property that enables it to withstand acute exposure to high levels of DNA-damaging agents, including gamma radiation, ultraviolet (UV) rays, desiccation, and chemical mutagens. In the context of DNA damage, a variety of DNA lesions from base damages to double strand breaks are acted upon by appropriate DNA repair pathways which operate either in tandem or in a sequential manner to efficiently restore the entire genome and restart normal growth. To achieve a high-regulatory specificity, a multi-factorial regulatory mechanism is required to activate or repress target pathway genes in vivo, in a dose and time dependent manner. Several studies have explored a role of cis-acting elements including genomic sequences like promoters, enhancers, or operators, structural elements, or trans-acting factors such as transcriptional regulators or non-coding RNAs (ncRNAs) in the regulation of DNA damage response. This review explores the intricate regulatory networks employed by D. radiodurans, offering a comprehensive overview of its DNA damage response mechanisms. These insights help shed light on the molecular strategies that enable this organism to thrive in extreme environments. The knowledge gaps which need the focus of future research are discussed.
The hyperthermophilic bacterium Thermotoga maritima possesses a unique peptidoglycan characterized by the presence of an unusual D-lysine. We have previously identified and characterized a lysine racemase involved in the D-lysine biosynthetic pathway in T. maritima. The lysine racemase possesses high racemase activity toward ornithine as well as lysine. However, the physiological significance of D-ornithine is unknown in T. maritima. In the present study, we characterized ornithine decarboxylase (TM1873), which catalyzes the first step of polyamine biosynthesis, to determine its substrate specificity for D-amino acids including D-ornithine. TM1873 displayed the highest decarboxylase activity toward L-ornithine, followed by L-lysine, and lowest activity toward L-arginine. Based on our kinetic analysis, the kcat/Km value for L-ornithine was higher than that for L-lysine. These results suggest that TM1873 has a dual specificity for L-ornithine and L-lysine. Decarboxylase activity of TM1873 was maintained under the physiological pH and growth temperature in T. maritima. Intriguingly, TM1873 also acted on D-ornithine and D-lysine, whereas these activities were lower than those toward corresponding L-amino acids. Additionally, diaminopimelate decarboxylase (TM1517) exhibited nominal activity toward D-ornithine and D-lysine. These results suggest that these two decarboxylases have the ability to metabolize basic D-amino acids, although their substrate preferences were oriented toward basic L-amino acids and meso- diaminopimelate, respectively.
Heavy metal (HM) contamination from mining poses a long-term threat to agricultural soils. This study assessed the impacts of long-term HM exposure on soil microbial communities by comparing contaminated and uncontaminated agricultural soils from northwestern China. Using physicochemical analyses and high-throughput amplicon sequencing of bacterial and fungal communities, we assessed impacts on microbial community composition, co-occurrence networks, and predicted functions. HM contamination significantly reduced fungal α-diversity and altered the composition of both bacterial and fungal communities. The relative abundances of Actinobacteria, Acidobacteria, and Chloroflexi increased significantly in contaminated soils. Fungal communities were markedly enriched in Mortierellomycota. Notably, HM contamination substantially reduced the complexity of microbial co-occurrence networks, as indicated by fewer nodes, edges, and keystone taxa. Predictive functional profiling suggested an increased predicted abundance of functions related to HM resistance and detoxification, alongside shifts in core nutrient cycling potentials. Specifically, carbon and nitrogen metabolic pathways were altered, and the proportion of saprotrophic fungi increased, suggesting changes in organic matter decomposition dynamics. In conclusion, long-term HM stress is associated with a shift toward a simplified state, characterized by taxonomic restructuring, less complex co-occurrence networks, and a functional shift toward stress resistance and altered nutrient cycling.
Glaciozyma antarctica is an Antarctic psychrophilic yeast providing an excellent eukaryotic model for dissecting molecular mechanisms of cold adaptation. Yet, its extracellular protein repertoire, the interface between the cell and extreme surroundings, remains unexplored. Here, we present the first integrated bioinformatics–proteomics secretome map of G. antarctica, linking computational prediction with experimental validation under varying temperature stress, yielding 9 high-confidence experimentally supported secreted proteins. Three complementary pipelines (Phobius, SignalP/TMHMM, and WoLF PSORT) predicted 278 high-confidence secreted proteins, predominantly glycosyl hydrolases, lipases, oxidoreductases, and stress-response factors. To validate these predictions, we performed LC–MS/MS analysis of extracellular fractions from cultures exposed to − 12 °C, 0 °C, and 12 °C. A total of 104 secreted proteins was identified, including 55 shared across all temperatures and multiple unique cold-induced candidates such as antifreeze proteins and glycosidases. Comparative analysis uncovered a temperature-dependent remodeling of the secretome and revealed several proteins lacking canonical signal peptides, highlighting potential unconventional secretion mechanisms or moonlighting roles in extracellular environments. These findings demonstrate that G. antarctica adapts to freezing environments by dynamically reshaping its extracellular proteome to maintain structural plasticity and metabolic efficiency. This integrative approach identifies promising cold-active enzymes for potential application in the low-temperature biocatalysis industry.
Chitin is the second most abundant polysaccharide and can serve as a carbon and nitrogen source for microorganisms in various ecosystems, including hypersaline environments. However, chitin metabolism in extremely halophilic archaea of the class Halobacteria has not been systematically analyzed, and the pathways of N-acetylglucosamine (GlcNAc) utilization in these organisms remain poorly understood. In this study, we performed a large-scale comparative genomic analysis of the class Halobacteria to assess prevalence and organization of genes, presumably involved in chitin utilization. Potential chitinolytic species are unevenly distributed across the class Halobacteria and tend to cluster within only 7 out of its 117 recognized genera. Glycoside hydrolases from GH18 and GH3 families were found as predominant endochitinases and β-hexosaminidases, respectively. Most putative chitinolytic haloarchaea lacked the genes involved in GlcNAc catabolism in bacteria and hyperthermophilic archaeon Thermococcus kodakaraensis. However, many of them harbor enzymes of pathways previously proposed for Natrarchaeobius and nanohaloarchaeon Ca. “Nanohalobium constans”. Analysis of gene neighborhoods demonstrated a conserved organization of the gene clusters associated with chitin metabolism in some genomes, resembling the clusters previously described for Natrarchaeobius species. Overall, our results indicate that Halomicrobium, Natrarchaeobius, Natrialba, Saliphagus, Halocatena, Haladaptatus, and Salinarchaeum genera are enriched in species with chitinolytic potential.
With the growing challenges of resource shortages and environmental pollution, bioleaching has attracted increasing attention for its high potential in processing refractory and low-grade ores. However, the slow growth rate and low biomass production of autotrophic bacteria commonly used in bioleaching limit their industrial application. Here, based on high-throughput sequencing analysis, which revealed a strong positive correlation between Sulfobacillus and Alicyclobacillus in mining environments, we developed a mixed-bacterial system composed of Sulfobacillus thermosulfidooxidans and Alicyclobacillus ferrooxydans to enhance the efficiency of bioleaching. The mixed-bacterial system achieved a pyrite leaching rate up to 91.14%, compared to 67.54% by S. thermosulfidooxidans and 17.63% by A. ferrooxydans. The improved leaching performance was attributed to the increased production of total extracellular polymeric substances (EPS), higher cell density and stronger redox capacity in the bioleaching solution. This study contributes to the development of a greener and more sustainable metallurgical industry.
Hypersaline environments exhibit extreme physiochemical conditions yet support diverse microbial communities. These communities are not only ecologically important but also possess substantial potential for biotechnological exploitation. In this study, we employed a comparative metagenomic approach to assess microbial diversity using two distinct methodologies: (1) direct DNA extraction from raw sediment, and (2) DNA extraction following halophilic enrichment in selective media. Sediment samples were collected from multiple sites and pooled together within the Rann of Kachchh and close-by saltpans and were analysed using 16S rRNA sequencing coupled with bioinformatics pipelines. The results revealed pronounced differences in microbial community composition between the two approaches. Raw sediment samples exhibited significantly higher alpha diversity, with dominant taxa including Halobacterota, Cyanobacteria, and Desulfobacterota, with a substantial proportion of unclassified genera. In contrast, enriched samples were dominated by fast-growing, culturable genera such as Halobacterium, Alkalibacillus, and Candidatus haloredivivus. Principal Coordinate Analysis (PCoA) of beta diversity demonstrated distinct clustering between raw and enriched communities, even within samples from the same sites, underscoring the selective bias introduced by enrichment procedures. These findings emphasise that the methodological choice strongly influences the observed microbial diversity. The aim of this study was to compare microbial community composition in raw hypersaline sediments and enrichment cultures using metagenomic sequencing, to evaluate how enrichment selectively favours specific halophilic taxa. This comparative approach allows identification of the microbial groups that rapidly proliferate under controlled hypersaline conditions, thereby complementing direct environmental sequencing. By integrating both direct and enrichment-based metagenomic approaches, a more comprehensive understanding of microbial community structure in hypersaline environments can be achieved.
Chitin is the second most abundant polysaccharide in nature and constitutes a key structural component in the cell wall of fungi and invertebrate exoskeletons. Chitinases produced by Antarctic organisms have demonstrated high catalytic efficiency, being promising tools for sustainable biotechnological processes. Therefore, the objective of this study was to identify the genes responsible for chitin degradation in the genomes of two Antarctic Arthrobacter strains, as well as to evaluate their biotechnological potential. Whole-genome sequencing followed by phylogenomic analysis revealed that both strains 492 and 285 belong to the species Arthrobacter psychrochitiniphilus. Genome assembly and annotation confirmed the genetic potential for chitinase production, showing that both bacteria possess the gene encoding type C chitinase (Chi C) from the GH18 family. Genomic comparative analysis revealed that they possess a total of eleven genes related to glycoside hydrolases (GH) and carbohydrate-binding modules (CBM), highlighting GH13 and GH65. Arthrobacter psychrochitiniphilus 492 was classified as psychrophilic, while A. psychrochitiniphilus 285 was classified as mesophilic-psychrotolerant. The enzyme production by A. psychrochitiniphilus 285 was 19.9 U/L, in 96 h at 15 °C, and the chitinase showed activity against the Aspergillus sp. series nigri CBMAI 1846, indicating promising potential for the formulation of bioinoculants and sustainable agriculture.
This study provides the first culture-based inventory of haloarchaeal diversity in Moroccan Atlantic solar salterns (Oualidia and Khenifiss), using sediment samples collected across a gradient of moderate to high salinity (16.7–29.2
Leptolyngbya JSC-1 is a thermophilic and siderophilic cyanobacterium inhabiting iron-rich hot springs. Response surface method (RSM) is being reported for the first time for optimizing growth conditions of this thermophilic and siderophilic cyanobacterium. Using response surface quadratic model of Box-Behnken design, optimal culture conditions (A: temperature, 45 °C; B: Fe concentration, 42 µM; and C: light intensity, 2000 lx which is equivalent to 27 µmol photons m⁻² s⁻¹ intensity of cool white fluorescent lamp) were determined. The significant model terms were found to be B, AB, A2, B2, and C2. The model R2 value (coefficient of determination) was 0.939, suggesting that the fitted model could explain 93.9
Astrobiology assesses the habitability of planetary bodies and the potential for extraterrestrial life. Analog environments on Earth serve as sites for studying extreme environments that resemble extraterrestrial conditions, aiding in validating life-detection methods, mission instrumentation, and biosignature preservation. These environments function as a source of model microorganisms and communities that define the habitability and biochemistry of such extraterrestrial environments. Well-known analog environments include the Atacama Desert (Chile) for space mission validation, the McMurdo Dry Valleys (Antarctica) for Mars analog studies, and Rio Tinto (Spain) for extreme acidic environments. Although significant research has been conducted on these sites, various alternative environments may also offer valuable opportunities for astrobiological studies. Saudi Arabia encompasses a variety of pristine (or with minimal anthropic influence) extreme environments with conditions analogous to extraterrestrial settings (e.g., deserts and salt flats as analogs to Mars, and terrestrial and marine volcanic fields as analogs to icy moons), yet their potential remains largely unexplored. Recent studies have identified a volcanic crater with sodium phosphates and chlorates that mimics Enceladus’s ocean chemistry, and researchers have cultured Halalkalibacterium halodurans strains with adaptations to survive these conditions, offering valuable biological models. Additionally, complex metabolic landscapes with implications for icy moon habitability have been observed in Red Sea systems, which could be employed as valuable natural laboratories in astrobiological research. Furthermore, these findings underscore the potential of the Saudi Arabian extremophilic microbiome for space-related research. This review explores the microbial diversity of extreme environments in Saudi Arabia, emphasizing their potential as new terrestrial analogs to Mars and icy moons and the role of their microbiomes as terrestrial proxies for extraterrestrial life.
Cellulose-hydrolyzing enzyme, cellulase, has a wide range of importance in many industries such as bioethanol, textiles, and food production. A novel isolate, Bacillus thuringiensis TS-04, from the salterns of Tuticorin, India, was identified through 16 S rDNA sequencing and selected for its superior cellulolytic activity (41.49 IU/mL) compared to other isolates. Optimization using Box-Behnken Design and Response Surface Methodology evaluated the effects of pH, incubation time, and inoculum percentage. The optimized conditions (pH 6.0, 72 h, 2.5
Photosynthetic life is based on absorbing sunlight and turning it into biologically usable energy. In many cases however, canopy-like structures and cavern-like habitats in terrestrial environments can limit the intensity and alter the spectra of light. One acclimation to use filtered light in the near infrared range, typically between 700 and 800 nm is named far-red light photoacclimation or FaRLiP as in recent studies of cyanobacteria. Here we report the common capacity for FaRLiP in the dominant cyanobacterial genera in a canyon hot spring microbial mat ecosystem. We identified FaRLiP in the genomes of cyanobacterial isolates and the metagenomes of mat samples. We show using absorption spectroscopy and HPLC that under far red-light specific isolates show an increase in far red-light absorption and the presence of Chl f. Springs in narrow canyons are a microniche where FaRLiP seems highly ecologically advantageous.
Haloarchaea thrive in extreme environments where hypersalinity, DNA damage, and nutrient scarcity pose significant stresses. Here, we provide insight into how defined phosphate levels, UV stress, and Orc1/Cdc6 homolog type influence growth and genome dynamics (ploidy) in the model haloarchaeon Haloferax volcanii. During phosphate limitation, cells exhibited reduced growth, heightened UV sensitivity and dynamic changes in chromosome ploidy, with an early log-phase increase followed by a reduction at later growth stages. To uncover underlying mechanisms, we examined the sixteen Orc1/Cdc6 homologs of H. volcanii using bioinformatics, AI-based structural predictions, genetics, site-directed mutagenesis, and biochemical assays. When constitutively expressed, Orc10 and Orc14 were found to increase ploidy, while Orc1 and Orc10 impaired growth in a manner exacerbated by low phosphate and UV stress. Substitutions at the lysine acetylation site within the Walker A motif (Orc1 K165) and conserved DNA-binding residues (R498 in Orc1 and R304 in Orc14) were found to affect Orc/Cdc6 homolog functions. Moreover, a ∆pat2 lysine acetyltransferase mutant was found to exhibit reduced ploidy and a striking phosphate-dependent flocking phenotype during transition to stationary phase. These results highlight the interplay between phosphate availability, chromosomal regulation, and stress adaptation. Together, this study provides insight into mechanisms that promote H. volcanii survival in conditions that induce DNA damage and phosphate limitation.
In this study, Sphingomonas sp. PAMC26617 and Sphingomonas sp. PAMC26621 isolated from rocks of an Arctic lichen, Umbillicaria sp. and Cetraria sp. respectively, were evaluated for their tolerance to abiotic stress of heavy metal(loid)s (HMMs), salt, and osmotic (drought-like) stress at two different temperatures. HMMs resistance protein (CzcC, CzcD, CopB, CopC, CopD, ArsH, and CorC), and osmoprotectant enzymes such as gamma-glutamyl phosphate reductase (ProA), glutamate 5-kinase (ProB), and pyrroline-5-carboxylate reductase (ProC) were identified in both strains during genome analysis. Heavy metal (HM) resistance protein (CorA) was only identified in Sphingomonas sp. PAMC26621. In addition, both strains showed tolerance to CuSO4·5H2O (0.1 mM at 15 °C and 0.25 mM at 25 °C), CoCl2·6H2O (0.25 mM at 15 °C and 0.1 mM at 25 °C), ZnSO4·7H2O (0.1 mM at 15 °C and 25 °C) and Na2HAsO4·7H2O (10 mM at 15 °C and 25 °C), except for Cd(NO3)2·4H2O. Sphingomonas sp. PAMC26617 showed 0.05 mM Cd(NO3)2·4H2O at 15 °C and 0.01 mM at 25 °C and Sphingomonas sp. PAMC26621 showed 0.01 mM Cd(NO3)2·4H2O at 15 °C and 25 °C, respectively. Furthermore, both strains showed tolerance up to 100 mM NaCl and 100 mM D-mannitol at both temperatures. This study is very informative and helpful in understanding the diversity and adaptability of lichen-associated bacteria in harsh environments.
The bacterial phylum Caldisericota has a wide geographical and environmental distribution, but cultivated representatives of this phylum have so far been limited to a single strain. We have isolated a novel anaerobic, thermophilic, fermentative bacterium, strain AR60T from a thermal spring in Sakhalin Island, Russia. Cells of strain AR60T were long rods and tended to form aggregates. Growth was observed between 50 and 70 °C (optimum at 60 °C) and between pH 6.0 and 7.5 (optimum pH 6.5). The isolate was capable for fermentative growth on yeast extract, tryptone and peptone. Thiosulfate, S0, DMSO stimulated growth. The strain contained iso-C17:0 as the major fatty acid. Genome size of strain AR60T was 1.58 Mb; G + C content of the genomic DNA was 34.47
Two extremely halophilic archaeal strains, HT40T and LN261T, were isolated from salt crystal samples collected from the Dingyuan Salt Mine in Anhui, China. Strain HT40T was found to possess two copies of the 16 S rRNA gene (rrnA and rrnB), while only one copy was detected in strain LN261T. Phylogenetic analysis based on the 16 S rRNA gene sequences revealed that the rrnA and rrnB genes of HT40T showed high sequence similarities with Halovenus aranensis EB27T (94.73
