Search Results (321)

The purpose of this work was to obtain specialized enrichment cultures from an original extreme acidophilic consortium of extremely acidophilic microorganisms and to study their microbial community composition and biotechnological potential. At temperatures of 25, 35, 40 and 50 °C, distinct enrichments of extremely acidophilic microorganisms used in the processes of bioleaching sulfide ores were obtained using nutrient media containing ferrous sulfate, elemental sulfur and a copper sulfide concentrate as nutrient inorganic substrates, with and without the addition of 0.02% yeast extract. The microbial community composition was studied using the sequencing of the V3–V4 hypervariable region of the 16S rRNA genes. The different growth conditions led to changes in the microbial composition and relative abundance of mesophilic and moderately thermophilic, strict autotrophic and mixotrophic microorganisms in members of the genera Acidithiobacillus, Sulfobacillus, Leptospirillum, Acidibacillus, Ferroplasma and Cuniculiplasma. The dynamics of the oxidation of ferrous iron, sulfur, and sulfide minerals (pyrite and chalcopyrite) by the enrichments was also studied in the temperature range of 25 to 50 °C. The study of enrichment cultures using the molecular biological method using the metabarcoding method of variable V3–24 V4 fragments of 16S rRNA genes showed that enrichment cultures obtained under different conditions differed in composition, which can be explained by differences in the physiological properties of the identified microorganisms. Regarding the dynamics of the oxidation of ferrous ions, sulfur, and sulfide minerals (pyrite and chalcopyrite), each enrichment culture was studied at a temperature range of 25 to 50 °C and indicated that all obtained enrichments were capable of oxidizing ferrous iron, sulfur and minerals at different rates. The obtained enrichment cultures may be used in further work to increase bioleaching by using the suitable inoculum for the temperature and process conditions. Full article

Tryptophan is synthesized in microorganisms via a five-step enzymatic pathway originating from chorismate, which is a product of the shikimate pathway. As a biosynthetic precursor to a wide range of high-value compounds such as indole-3-acetic acid, indigo, indirubin, and violacein, this pathway has been a central target for metabolic engineering to enhance microbial production. Anthranilate phosphoribosyltransferase (AnPRT) catalyzes the second step of the pathway by transferring a phosphoribosyl group from PRPP to anthranilate, forming phosphoribosyl anthranilate (PRA). AnPRT, the sole member of class IV phosphoribosyltransferases, adopts a unique fold and functions as a homodimer. While the structural basis of AnPRT activity has been elucidated in several organisms, thermostable variants remain underexplored despite their relevance for high-temperature bioprocessing. In this study, the crystal structure of AnPRT from the thermophilic archaeon Methanocaldococcus jannaschii (MjAnPRT) was determined at a 2.16 Å resolution. The enzyme exhibits a conserved dimeric architecture and key catalytic motifs. Comparative structural analysis with mesophilic and hyper thermophilic homologs revealed that MjAnPRT possesses enhanced local stability in catalytically important regions and strengthened inter-subunit interactions. These features likely contribute to its thermostability and provide a valuable framework for the rational design of robust AnPRTs for industrial and synthetic biology applications. Full article

Thermostable cellulases and xylanases have broad acceptance in food, feed, paper and pulp, and bioconversion of lignocellulosics. Thermophilic fungi serve as an excellent source of thermostable enzymes. This study characterized four endo-β-1,4-glucanases (two glycoside hydrolase (GH) family 5 and two GH7 members) and four endo-β-1,4-xylanases (two GH10 and two GH11 members) from thermophilic fungus Thielavia terrestris, along with one GH10 endo-β-1,4-xylanase each from thermophilic fungus Chaetomium thermophilum and mesophilic fungus Chaetomium globosum. Comparative analysis was conducted against three previously reported GH10 endoxylanases: two thermostable enzymes from the thermophilic fungus Humicola insolens and thermophilic bacterium Halalkalibacterium halodurans, and one mesophilic enzyme from model fungus Neurospora crassa. The GH10 xylanase TtXyn10C (Thite_2118148; UniProt G2R8T7) from T. terrestris demonstrated high thermostability and activity, with an optimal temperature of 80–85 °C. It retained over 60% of its activity after 2 h at 70 °C, maintained approximately 30% activity after 15 min at 80 °C, and showed nearly complete stability following 1 min of exposure to 95 °C. TtXyn10C exhibited specific activity toward beechwood xylan (1130 ± 15 U/mg) that exceeded xylanases from H. insolens and H. halodurans while being comparable to N. crassa xylanase activity. Furthermore, TtXyn10C maintained stability across a pH range of 3–9 and resisted trypsin digestion, indicating its broad applicability. The study expands understanding of enzymes from thermophilic fungi. The discovery of the TtXyn10C offers a new model for investigating the high activity-stability trade-off and structure-activity relationships critical for industrial enzymes. Full article

Adequate treatment of the organic fraction of municipal solid waste (OFMSW) in co-digestion with sewage sludge (SS) through dark fermentation (DF) technologies has been widely studied and recognized. However, there is little experience with a high-solids approach, where practical and scalable conditions are established to lay the groundwork for further development of feasible industrial-scale projects. In this study, the biochemical hydrogen potential of OFMSW using a 7 L batch reactor at mesophilic conditions was evaluated. Parameters such as pH, redox potential, temperature, alkalinity, total solids, and substrate/inoculum ratio were adjusted and monitored. Biogas composition was analyzed by gas chromatography. The microbial characterization of SS and post-reaction percolate liquids was determined through metagenomics analyses. Results show a biohydrogen yield of 38.4 NmLH₂/gVS OFMSW, which forms ~60% of the produced biogas. Aeration was proven to be an efficient inoculum pretreatment method, mainly to decrease the levels of methanogenic archaea and metabolic competition, and at the same time maintain the required total solid (TS) contents for high-solids conditions. The microbial community analysis reveals that biohydrogen production was carried out by specific anaerobic and aerobic bacteria, predominantly dominated by the phylum Firmicutes, including the genus Bacillus (44.63% of the total microbial community), Clostridium, Romboutsia, and the phylum Proteobacteria, with the genus Proteus. Full article

The study of flora is crucial for conserving natural resources and assessing human impact on the environment. This paper explores floristic diversity, the role of plants, and the integration of technology in botanical research. In the studied area, 462 plant species were identified. Bioform analysis revealed a predominance of hemicryptophytes (45.45%) and therophytes (26.19%), suggesting a warm climate and significant zoo-anthropogenic influences. Other bioforms were present in lower percentages. Most plant species in Teleorman County are mesophilic (39.39%) and mesoxerophilic (30.95%), indicating adaptation to moderate or slightly dry environments. Regarding temperature affinity, the majority are micromesotherms (62.98%), suited for mild thermal conditions. Soil reaction analysis showed a preference for weakly acidic neutrophilic (39.82%) and euryionic (33.76%) soils, indicating tolerance for neutral to slightly acidic pH levels. The research evaluates the structure and diversity of flora in Teleorman County and emphasizes the influence of climate factors such as humidity, temperature, and soil pH on species distribution. Using the transect method and fractal analysis, this study concludes that temperature is the dominant climatic factor shaping local biodiversity. Full article

Tea has become one of the most popular drinks worldwide thanks to its pleasant sensory attributes and diverse health benefits. However, tannin-rich compositions have several negative effects and significantly impact the quality of tea beverages. Among various detannification methods, tannase treatment appears to be the most secure and environmentally friendly strategy. Although numerous microbial tannases have been identified and used in food processing, they are predominantly mesophilic with compromised heat tolerance, which limit their application in high-temperature tea extraction processing. Computer-assisted rational design and site-directed mutagenesis has emerged as a promising strategy in enzyme engineering to improve the thermostability of industrial enzymes. Nevertheless, relevant studies for tannase thermostability improvement remain lacking. In the present study, a novel thermophilic tannase called TanPL1 from marine fungus Penicillium longicatenatum strain SM102 was expressed in the food-grade host Yarrowia lipolytica. After purification and characterization, the thermostability of this enzyme was improved through site-directed mutagenesis guided by computer-aided rational design and molecular dynamics simulations. Then the thermostable mutant MuTanPL1 was applied in green tea processing for both polyphenol extraction and ester catechin hydrolysis. The tannase yield and specific activity values of 166.4 U/mL and 1059.3 U/mg, respectively, were achieved. The optimum pH and temperature of recombinant TanPL1 were determined to be 5.5 and 55 °C, respectively, and the enzyme exhibited high activity toward various gallic acid ester substrates. The site-directed mutagenesis method successfully generated a single-point mutant, MuTanPL1, with significantly enhanced thermostability and a higher optimum temperature of 60 °C. After 2 h of detannification by MuTanPL1, nearly all gallated catechins in green tea infusion were biotransformed. This resulted in a 202.4% and 12.1-fold increase in non-ester catechins and gallic acid levels, respectively. Meanwhile, the quality of the tea infusion was also markedly improved. Sensory evaluation and antioxidant activity assays revealed notable enhancements in these properties, while turbidity was reduced considerably. Additionally, the α-amylase inhibition activity of the tannase-treated tea infusion declined from 50.49% to 8.56%, revealing a significantly lower anti-nutritional effect. These findings suggest that the thermostable tannase MuTanPL1 holds strong application prospects in tea beverage processing. Full article

This study examines biodegradability (BD) and optimum conditions for the solid-state anaerobic digestion (SS-AD) of organic solid poultry waste (organs, intestines, offal, and unprocessed meat) to maximize biomethane production. Three main parameters, substrate-to-inoculum (S/I) ratio, pH, and temperature, were evaluated for the SS-AD of organic solid poultry waste. pH was evaluated at non-adjusted pH, initially adjusted pH, and controlled pH conditions at a constant S/I ratio of 0.5 and temperature of 35 ± 1 °C. The S/I ratios were examined at (0.3, 0.5, 1, and 2) at a controlled pH of ≈7.9 and temperature of 35 ± 1 °C. The temperature was assessed at mesophilic (35 ± 1 °C) and thermophilic (55 ± 1 °C) conditions with a constant S/I ratio of 0.5 and controlled pH of ≈7.9. The results demonstrate that the highest biomethane production and BD were achieved with a controlled pH of ≈7.9 (689 ± 10 mg/L, 97.5 ± 1.4%). The initially adjusted pH (688 ± 14 mg/L, 97.3 ± 1.9%) and an S/I ratio of 0.3 (685 ± 8 mg/L, 96.8 ± 1.2%) had approximately equivalent outcomes. The thermophilic conditions yielded 78% lower biomethane yield than mesophilic conditions. The challenge of lower biomethane yield under thermophilic conditions will be resolved in future studies by determining the rate-limiting step. These observations highlight that SS-AD is a promising technology for biomethane production from solid organic poultry waste. Full article

The increasing demand for sustainable protein sources highlights Hermetia illucens (Black Soldier Fly, BSF) as a promising alternative. However, microbiological safety remains a key concern. This study investigated the microbial diversity of BSF larvae, comparing two processing methods: (1) boiling followed by drying and (2) drying alone. Microbial diversity was assessed via 16S rRNA sequencing, while bacterial loads were quantified using culture-based methods on samples from a French company. A systematic review complemented this analysis by synthesizing the existing knowledge on BSF microbiota. The rearing conditions varied, with substrate pH ranging from 4.1 to 9.0 and ambient temperatures between 24.6 °C and 42.7 °C. Mesophilic bacteria, spores, and lactic acid bacteria reached up to 8.6, 7.7, and 8.5 log CFU/g in the substrates and larvae, while yeasts, molds, and sulfite-reducing bacteria remained below 4.8 log CFU/g. Boiling reduced most loads below detection thresholds, particularly for yeasts, molds, and ASR. Salmonella, Listeria monocytogenes, Cronobacter sp., and coagulase-positive staphylococci were absent, whereas Clostridium perfringens and Escherichia coli were variably detected. Metabarcoding showed shifts in composition, with Proteobacteria, Bacteroidota, Actinobacteriota, and Firmicutes (Bacillota and Clostridiota) dominating. Process 1 more effectively reduced the bacterial loads, though Bacillus and Clostridium remained. Campylobacter sp. detection in powders raises food safety concerns. Full article

Small heat shock proteins (sHsps) are ubiquitous low-molecular-weight chaperones that prevent protein aggregation under cellular stress conditions. In the absence of stress, they assemble into large oligomers. In response to stress, such as elevated temperatures, they undergo conformational changes that expose hydrophobic surfaces, allowing them to interact with denatured proteins. At heat shock temperatures in bacteria, large sHsp oligomers disassemble into smaller oligomeric forms. Methanogens are a diverse group of microorganisms, ranging from thermophilic to psychrophilic and halophilic species. Accordingly, their sHsps exhibit markedly different temperature dependencies based on their optimal growth temperatures. In this study, we characterized sHsps from both hyperthermophilic and mesophilic methanogens to investigate the mechanisms underlying their temperature-dependent behavior. Using analytical ultracentrifugation, we observed the dissociation of sHsps from a mesophilic methanogen into dimers. The dissociation equilibrium of these oligomers was found to be dependent not only on temperature but also on protein concentration. Furthermore, by generating various mutants, we identified the specific amino acid residues responsible for the temperature dependency observed. The C-terminal region containing the IXI/V motif and the α-crystallin domain were found to be the primary determinants of oligomer stability and its temperature dependence. Full article

L-asparaginase (L-ASNase) is a key industrial enzyme significant for cancer therapy and the food industry for reducing dietary acrylamide. The hyperthermophilic L-ASNase from Thermococcus sibiricus (TsAI) was previously shown to exhibit high activity and thermostability and is promising for biotechnology. To gain insights into structure-functional relationships of TsAI, determination of the substrate specificity, kinetic parameters, structural characterization, and molecular docking were performed. TsAI characteristics were compared with the TsAI^D54G/T56Q mutant, which exhibited increased activity after a double mutation in the substrate-binding region. TsAI and TsAI^D54G/T56Q were found to display high activity towards D-asparagine—62% and 21% of L-asparaginase activity, respectively—and low L-glutaminase coactivity of ~5%. Restoring the mesophilic-like triad GSQ in the mutant resulted in a two-fold increase in activity towards L-asparagine compared with TsAI. Crystal structures of TsAI forms solved at 1.9 Å resolution revealed that double mesophilic-like mutation increased the flexibility of the loop M51-L57, located in close proximity to the active site. Structural superposition and mutational analysis indicate that mobility of this loop is essential for the activity of thermo-ASNases. Molecular docking, without taking into account the temperature factor, showed that, in contrast to L-asparagine interaction, D-asparagine orientation in the TsAI and TsAI^D54G/T56Q active sites is similar and not optimal for catalysis. Under real conditions, high temperatures can induce structural changes that reduce L-ASNase discrimination towards D-asparagine. Overall, the obtained structural and biochemical data provide a basis for a more detailed understanding of thermo-ASNase functioning and possibilities to engineer improved variants for future biotechnological application. Full article

Often thought of as a mesic paradise, forest ecosystems are a mosaic of microhabitats with temporal oscillations that cause significant environmental stresses, providing habitats for extremophilic and extremotolerant fungi. Adapted to survive and thrive under conditions lethal to most mesophiles (e.g., extreme temperatures, pH, water potential, radiation, salinity, nutrient scarcity, and pollutants), these species are increasingly recognized as vital yet underappreciated elements of forest biodiversity and function. This review examines the current understanding of the roles of extremophilic fungi in forests, scrutinizing their presence in these ecosystems with a critical eye. Particularly under severe environmental conditions, extremophilic fungi play a crucial role in forest ecosystems, as they significantly enhance decomposition and nutrient cycling, and foster mutualistic interactions with plants that increase stress resilience. This helps to maintain ecosystem stability. We examine the definition of “extreme” within forest settings, survey the known diversity and distribution of these fungi across various forest stress niches (cold climates, fire-affected areas, acidic soils, canopy surfaces, polluted sites), and delve into their possible ecological functions, including decomposition of recalcitrant matter, nutrient cycling under stress, interactions with plants (pathogenesis, endophytism, perhaps mycorrhizae), bioremediation, and contributions to soil formation. However, the review stresses significant methodological difficulties, information gaps, and field-based natural biases. We recommend overcoming cultural constraints, enhancing the functional annotation of “omics” data, and planning investigations that clarify the specific activities and interactions of these cryptic creatures within the forest matrix to further advance the field. Here, we demonstrate that moving beyond simple identification to a deeper understanding of function will enable us to more fully appreciate the value of extremophilic fungi in forest ecosystems, particularly in relation to environmental disturbances and climate change. Full article

The production and utilization of conventional plastics have raised concerns regarding plastic waste management and environmental safety. In response, the emergence of biodegradable bioplastics presents a possible solution for sustainability. On the other hand, the efficacy of biodegradation is strictly dependent on both the bioplastic type and the conditions in which the biodegradation occurs. This review offers a comprehensive overview of the biodegradation behavior of several bioplastics under a managed (industrialized or controlled) environment, such as industrial composting and anaerobic digestion (at either mesophilic or thermophilic temperature), as well as under less studied unmanaged (natural or open) environments, including soil, seawater, and freshwater. Although the biodegradation trend of some bioplastics is well known, further investigation should be pursued for others in order to clearly have the knowledge and the ability to choose the most viable bioplastic for a specific application and future end-of-life. Full article

Sweet potatoes play an important role in the global food supply, as they are rich in bioactive components and have numerous health benefits. Their minimally processed, ready-to-eat form is increasingly popular among consumers; however, discoloration and microbiological problems threaten the safety of these products. The aim of this study is to investigate the shelf life of cleaned, cut, ready-to-eat, vacuum-packed, and refrigerated Bonita (white) and Covington (orange) varieties of sweet potatoes after soaking in apple and chokeberry pomace extracts and treatment with citric and ascorbic acids. A series of microbiological and analytical tests was conducted during the storage period. The microbiological tests included the enumeration of cells of mesophilic aerobic and facultative anaerobic microbes, as well as lactobacilli, lactococci, Enterobacteriaceae, yeasts, and moulds. The analytical tests encompassed the determination of the total phenolic content, antioxidant capacity, water-soluble solid content, and pH value. The prevalent microbial groups detected in the examined sweet potato varieties were lactic acid bacteria, which were present in both fresh samples and following storage. This study established that low-temperature refrigeration (5 °C), vacuum packaging, and organic acid treatment can effectively control lactic acid bacteria, which are pivotal to spoilage. The combination of preservation steps is of particular significance for ready-to-cook sweet potatoes, as this approach effectively extends the shelf life of these products. Full article

Fruits, such as apricots, cherries, raspberries, and strawberries, are very often processed into jams to extend their shelf life. Jams are highly appreciated by consumers, and their storage conditions are important for preserving their quality. This study investigates the impact of storage on the microbiological quality and physicochemical parameters of five commercial fruit (apricot, sour cherry, white cherry, raspberry, and strawberry) jams. The pH, titratable acidity, moisture, sugar content, viscosity, and color were evaluated immediately after opening the jam containers and during storage at 2–4 °C and 20 °C. The total number of mesophilic aerobic germs (TMAG) and the total number of yeasts and molds (TYM) were also determined. The samples were analyzed during storage at 14, 42, and 56 days after opening the jam jars. The pH of the fruit jam samples varied from 2.91 (strawberry jam) to 3.47 (sour cherry jam) and decreased during storage, while the titratable acidity (TA) ranged from 0.46 (cherry and raspberry jams) to 0.52% malic acid (apricot jam) and increased during storage, regardless of the storage temperature. The moisture content was between 15.78% (apricot jam) and 21.82% (raspberry jam), and decreased in all the jam samples during storage. The sugar content (30.40–44.59 g per 100 g of jam) was typical for low-sweetened jams and increased during storage. Also, the viscosity of the fruit jam samples increased during storage (more in the samples stored at room temperature). Under the storage conditions, all the jam samples lost their specific color. Immediately after opening the jam jars, no yeasts or molds were found in the apricot and sour cherry jam samples. The highest number of yeasts and molds was detected in the white cherry jam samples (4.25 log₁₀ CFU/g). The TYM increased during storage, as did the TMAG. The time–temperature interaction factor influenced the physicochemical and microbiological properties of the jam samples. Full article

Antarctica, with its harsh environmental conditions, is home to a wide variety of microorganisms, including filamentous fungi. The survival of Antarctic mycoflora has led to increasing interest in their adaptation. Extreme low temperatures, low water availability, frequent freeze–thaw cycles, strong winds, ultraviolet radiation, etc., are all prerequisites for accelerated production of reactive oxygen species (ROS) and the occurrence of oxidative stress (OS). Antarctic fungi have effective mechanisms to cope with the damaging effects of ROS. While the distribution, morphology, physiology, and biotechnological potential of these fungi are well documented, the role of oxidative stress in their adaptation is poorly understood. This has been one of the main research topics of our team for a long time, and a number of papers on our results have been published. This review summarizes the existing literature on the induction of oxidative stress in Antarctic psychrophilic, psychrotolerant, and mesophilic fungi by extreme conditions. The most recent and relevant studies reporting on the relationship between cold stress and OS biomarkers and the damaging effects of ROS on key intracellular biomolecules are presented. The contribution of both enzymatic and non-enzymatic antioxidant defenses to the fungal cell response is discussed. Full article