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In the 1980s, Escherichia coli was the preferred host for heterologous protein expression owing to its capacity for rapid growth in complex media; well-studied genetics; rapid and direct transformation with foreign DNA; and easily scalable fermentation. Despite the relative ease of use of E. coli for achieving the high expression of many recombinant proteins, for some proteins, e.g., membrane proteins or proteins of eukaryotic origin, this approach can be rather ineffective. Another microorganism long-used and popular as an expression system is baker's yeast, Saccharomyces cerevisiae . In spite of a number of obvious advantages of these yeasts as host cells, there are some limitations on their use as expression systems, for example, inefficient secretion, misfolding, hyperglycosylation, and aberrant proteolytic processing of proteins. Over the past decade, nontraditional yeast species have been adapted to the role of alternative hosts for the production of recombinant proteins, e.g., Komagataella phaffii , Yarrowia lipolytica , and Schizosaccharomyces pombe . These yeast species' several physiological characteristics (that are different from those of S. cerevisiae ), such as faster growth on cheap carbon sources and higher secretion capacity, make them practical alternative hosts for biotechnological purposes. Currently, the K. phaffii -based expression system is one of the most popular for the production of heterologous proteins. Along with the low secretion of endogenous proteins, K. phaffii efficiently produces and secretes heterologous proteins in high yields, thereby reducing the cost of purifying the latter. This review will discuss practical approaches and technological solutions for the efficient expression of recombinant proteins in K. phaffii , mainly based on the example of enzymes used for the feed industry.

Xylanases (EC 3.2.1.8) hydrolyze the hemicellulose of plant cell walls. Xylanases are used in the food and paper industries and for bioconversion of lignocellulose to biofuel. In this work, the producer-strain with four copies of the xAor xylanase gene was organized in two tandem copies for optimal expression in Komagataella phaffii T07 yeast. The secreted 35 kDa xylanase was purified from culture medium by gel filtration on Sephadex G-25 and anion exchange chromatography on DEAE-Sepharose 6HF. Tryptic peptides of the recombinant enzyme were analyzed by liquid chromatography-tandem mass spectrometry where the amino acid sequence corresponded to Protein Accession # O94163 for Endo-1,4-beta-xylanase from Aspergillus oryzae RIB40. The recombinant xylanase was produced in a bioreactor where the secreted enzyme hydrolyzed oat xylane with an activity of 258240 IU/mL. High activity in the culture medium suggested xylanase could be used for industrial applications without being purified or concentrated. The pH optimum for xylanase xAor was 7.5, though the enzyme was active from pH 2.5 to pH 10. Xylanase was active at temperatures from 35 °C to 85 °C with a maximum at 60 °C. In conclusion, this protocol yields soluble, secreted xylanase suitable for industrial scale production.

In the south of western Siberia (Russia), there are many unique and unexplored soda, saline, and freshwater lakes. In this study, the results are presented on microbial diversity, its metabolic potential, and their relation with a set of geochemical parameters for a hypersaline lake ecosystem in the Novosibirsk region (Oblast). The metagenomic approach used in this work allowed us to determine the composition and structure of a floating microbial community, the upper layer of silt, and the strata of bottom sediments in a natural saline lake via two bioinformatic approaches, whose results are in good agreement with each other. In the floating microbial community and in the upper layers of the bottom sediment, bacteria of the Proteobacteria (Gammaproteobacteria), Cyanobacteria, and Bacteroidetes phyla were found to predominate. The lower layers were dominated by Proteobacteria (mainly Deltaproteobacteria), Gemmatimonadetes, Firmicutes, and Archaea. Metabolic pathways were reconstructed to investigate the metabolic potential of the microbial communities and other hypothetical roles of the microbial communities in the biogeochemical cycle. Relations between different taxa of microorganisms were identified, as was their potential role in biogeochemical transformations of C, N, and S in a comparative structural analysis that included various ecological niches.

This is data on the microbial diversity in the floating cyanobacterial community and sediment samples from the lake Solenoe (Novosibirsk region, Russia) obtained by metagenomic methods. Such a detailed data of the microbial diversity of the Novosibirsk oblast lake ecosystem was carried out for the first time. The purpose of our work was to reveal microbial taxonomic diversity and abundance, metabolic pathways and new enzyme findings the studied lake ecosystem using the next-generation sequencing (NGS) technology and metagenomic analysis. The data was obtained using metagenomics DNA whole genome sequencing (WGS) on Illumina NextSeq and NovaSeq. The raw sequence data used for analysis is available in NCBI under the Sequence Read Archive (SRA) with the BioProjects and SRA accession numbers: PRJNA493912 (SRR7943696), PRJNA493952 (SRR7943839) and PRJNA661775 (SRR12601635, SRR12601634, SRR12601633) corresponding to floating cyanobacterial community and sediment layers samples, respectively., Competing Interests: The authors declare that they have not known competing financial interests or personal relationships which have, or could be perceived to have, influenced the work reported in this article., (© 2021 The Authors.)

Identification of microorganisms by MALDI-TOF mass spectrometry is a very efficient method with high throughput, speed, and accuracy. However, it is significantly limited by the absence of a universal database of reference mass spectra. This problem can be solved by creating an Internet platform for open databases of protein spectra of microorganisms. Choosing the optimal mathematical apparatus is the pivotal issue for this task. In our previous study we proposed the geometric approach for processing mass spectrometry data, which represented a mass spectrum as a vector in a multidimensional Euclidean space. This algorithm was implemented in a Jacob4 stand-alone package. We demonstrated its efficiency in delimiting two closely related species of the Bacillus pumilus group. In this study, the geometric approach was realized as R scripts which allowed us to design a Web-based application. We also studied the possibility of using full spectra analysis (FSA) without calculating mass peaks (PPA), which is the logical development of the method. We used 74 microbial strains from the collections of ICiG SB RAS, UNIQEM, IEGM, KMM, and VGM as the models. We demonstrated that the algorithms based on peak-picking and analysis of complete data have accuracy no less than that of Biotyper 3.1 software. We proposed a method for calculating cut-off thresholds based on averaged intraspecific distances. The resulting database, raw data, and the set of R scripts are available online at https://icg-test.mydisk.nsc.ru/s/qj6cfZg57g6qwzN., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2020 Starostin, Demidov, Ershov, Bryanskaya, Efimov, Shlyakhtun and Peltek.)

Background: The Uzon Caldera is one of the places on our planet with unique geological, ecological, and microbiological characteristics. Uzon oil is the youngest on Earth. Uzon oil has unique composition, with low proportion of heavy fractions and relatively high content of saturated hydrocarbons. Microbial communities of the «oil site» have a diverse composition and live at high temperatures (up to 97 °C), significant oscillations of Eh and pH, and high content of sulfur, sulfides, arsenic, antimony, and mercury in water and rocks., Results: The study analyzed the composition, structure and unique genetics characteristics of the microbial communities of the oil site, analyzed the metabolic pathways in the communities. Metabolic pathways of hydrocarbon degradation by microorganisms have been found. The study found statistically significant relationships between geochemical parameters, taxonomic composition and the completeness of metabolic pathways. It was demonstrated that geochemical parameters determine the structure and metabolic potential of microbial communities., Conclusions: There were statistically significant relationships between geochemical parameters, taxonomic composition, and the completeness of metabolic pathways. It was demonstrated that geochemical parameters define the structure and metabolic potential of microbial communities. Metabolic pathways of hydrocarbon oxidation was found to prevail in the studied communities, which corroborates the hypothesis on abiogenic synthesis of Uzon hydrothermal petroleum.

For accurate species-level identification of microorganisms, researchers today increasingly use a combination of standard microbiological cultivation and visual observation methods with molecular biological and genetic techniques that help distinguish between species and strains of microorganisms at the level of DNA or RNA molecules. The aim of this work was to identify microorganisms from the ICG SB RAS Collection using an integrated approach that involves a combination of various phenotypic and genotypic characteristics. Key molecular-genetic and phenotypic characteristics were determined for 93 microbial strains from the ICG SB RAS Collection. The strains were characterized by means of morphological, physiological, moleculargenetic, and mass-spectrometric parameters. Specific features of the growth of the strains on different media were determined, and cell morphology was evaluated. The strains were tested for the ability to utilize various substrates. The strains studied were found to significantly differ in their biochemical characteristics. Physiological characteristics of the strains from the collection were identified too, e.g., the relationship with oxygen, type of nutrition, suitable temperature and pH ranges, and NaCl tolerance. In this work, the microorganisms analyzed were combined into separate groups based on the similarities of their phenotypic characteristics. This categorization, after further refinement and expansion of the spectrum of taxa and their metabolic maps, may serve as the basis for the creation of an "artificial" classification that can be used as a key for simplified and quicker identification and recognition of microorganisms within both the ICG SB RAS Collection and other collections., (Copyright © AUTHORS, 2018.)

The Halomonas sp. isolate SL48-SHIP-3 genome was obtained from metagenomics sequencing of the microbial mat of Salt Lake Number 48 (54.201806N, 78.179194E; Novosibirsk region, Russia). The sequenced and annotated genome is 2,575,909 bp and encodes 2,368 genes., (Copyright © 2019 Shipova et al.)

The Phormidium sp. strain SL48-SHIP genome was obtained from metagenomics sequencing of the microbial mat of Salt Lake No. 48 (54.201806 N, 78.179194 E; Novosibirsk Region, Russia). The sequenced and annotated genome is 4,384,607 bp and encodes 3,807 genes., (Copyright © 2019 Rozanov et al.)

Halorubrum sp. strain 48-1-W was isolated from a water sample from a saline lake (Novosibirsk Region, Russia, 54°14'N 78°13'E). The sequenced and annotated genome is 3,584,929 bp and contains 3,506 genes.

The Bacillus altitudinis strain KL4 was isolated from bottom sediments in Lake Krotovaya Lyaga (Novosibirsk Region, Russia, 53.7°N, 77.9°E). The sequenced and annotated genome is 3,738,419 bp long and carries 3,909 genes., (Copyright © 2018 Rozanov et al.)

Bacillus altitudinis strain KU-skv2(2) was isolated from a microbial mat on an anthropogenic pipe from Caldera Uzon (Kamchatka, Russia, 54°30'0.23″N, 160°0'15.18″E). The sequenced and annotated genome is 3,739,340 bp in size and encodes 3,929 genes., (Copyright © 2018 Rozanov et al.)

Background: Microbial mats are a good model system for ecological and evolutionary analysis of microbial communities. There are more than 20 alkaline hot springs on the banks of the Barguzin river inflows. Water temperature reaches 75 °C and pH is usually 8.0-9.0. The formation of microbial mats is observed in all hot springs. Microbial communities of hot springs of the Baikal rift zone are poorly studied. Garga is the biggest hot spring in this area., Results: In this study, we investigated bacterial and archaeal diversity of the Garga hot spring (Baikal rift zone, Russia) using 16S rRNA metagenomic sequencing. We studied two types of microbial communities: (i) small white biofilms on rocks in the points with the highest temperature (75 °C) and (ii) continuous thick phototrophic microbial mats observed at temperatures below 70 °C. Archaea (mainly Crenarchaeota; 19.8% of the total sequences) were detected only in the small biofilms. The high abundance of Archaea in the sample from hot springs of the Baikal rift zone supplemented our knowledge of the distribution of Archaea. Most archaeal sequences had low similarity to known Archaea. In the microbial mats, primary products were formed by cyanobacteria of the genus Leptolyngbya. Heterotrophic microorganisms were mostly represented by Actinobacteria and Proteobacteria in all studied samples of the microbial mats. Planctomycetes, Chloroflexi, and Chlorobi were abundant in the middle layer of the microbial mats, while heterotrophic microorganisms represented mostly by Firmicutes (Clostridia, strict anaerobes) dominated in the bottom part. Besides prokaryotes, we detect some species of Algae with help of detection their chloroplasts 16 s rRNA., Conclusions: High abundance of Archaea in samples from hot springs of the Baikal rift zone supplemented our knowledge of the distribution of Archaea. Most archaeal sequences had low similarity to known Archaea. Metagenomic analysis of microbial communities of the microbial mat of Garga hot spring showed that the three studied points sampled at 70 °C, 55 °C, and 45 °C had similar species composition. Cyanobacteria of the genus Leptolyngbya dominated in the upper layer of the microbial mat. Chloroflexi and Chlorobi were less abundant and were mostly observed in the middle part of the microbial mat. We detected domains of heterotrophic organisms in high abundance (Proteobacteria, Firmicutes, Verrucomicrobia, Planctomicetes, Bacteroidetes, Actinobacteria, Thermi), according to metabolic properties of known relatives, which can form complete cycles of carbon, sulphur, and nitrogen in the microbial mat. The studied microbial mats evolved in early stages of biosphere formation. They can live autonomously, providing full cycles of substances and preventing live activity products poisoning.

The Anoxybacillus flavithermus KU2-6-11 was isolated from sediments of a nameless hot spring. The hot spring is located in Uzon caldera (Kamchatka, Russia). The sequenced and annotated genome is 2,646,305 bp and encodes 2787genes. The draft genome sequence of the Anoxybacillus flavithermus KU2-6-11 has been deposited at DDBJ/EMBL/GenBank under the accession PEDM01000000 and the sequences could be found at the site https://www.ncbi.nlm.nih.gov/nuccore/PEDM01000000.

The Thermoactinomyces sp. strain Gus2-1 was isolated from hot-spring sediments sample from the hot-spring Gusikha in Bargusin Valley (Baikal Rift Zone, Russia). The sequenced and annotated genome is 2,623,309 bp and encodes 2513 genes. The draft genome sequence of the Thermoactinomyces sp. strain Gus2-1 has been deposited at DDBJ/EMBL/GenBank under the accession JPZM01000000 and the sequences could be found at the site https://www.ncbi.nlm.nih.gov/nuccore/JPZM01000000.

Background: Nothing is currently known about microbial composition of saline lakes of the Novosibirsk region and its dependence on physical-chemical parameters of waters. We studied the structure of microbial communities of saline lakes of the Novosibirsk region and the effect of physical-chemical parameters of waters on microbial communities of these lakes., Results: According to the ion content, the lakes were classified either as chloride or chloride-sulfate types. Water salinity ranges from 4.3 to 290 g L(-1). Many diverse microbial communities were found. Filamentous and colonial Cyanobacteria of the genera Scytonema, Aphanocapsa, and/or filamentous Algae dominated in littoral communities. Spatial and temporal organization of planktonic microbial communities and the quantities of Archaea and Bacteria were investigated using fluorescent in situ hybridization. We have found that the dominant planktonic component is represented by Archaea, or, less frequently, by Bacteria. Various phylogenetic groups (Bacteria, Archaea, Algae, and Cyanobacteria) are nonuniformly distributed. The principal component analysis was used to detect environmental factors that affect microorganism abundance. We found the principal components responsible for 71.1 % of the observed variation. It was demonstrated that two-block partial least squares was a better method than principal component analysis for analysis of the data. We observed general relationships between microbial abundance and water salinity., Conclusions: We have performed the first-ever study of the structure of the microbial communities of eleven saline lakes in the Novosibirsk region along with their physical-chemical parameters of waters. Our study demonstrates that saline lakes in the Novosibirsk region contain a unique microbial communities that may become a prolific source of microorganisms for fundamental and applied studies in various fields of ecology, microbiology, geochemistry, and biotechnology, and deserve further metagenomic investigation.

Microorganism identification by MALDI TOF mass-spectrometry is based on the comparison of the mass spectrum of the studied organism with those of reference strains. It is a rapid and reliable method. However, commercial databases and programs are mostly designed for identification of clinically important strains and can be used only for particular mass spectrometer models. The need for open platforms and reference databases is obvious. In this study we describe a geometric approach for microorganism identification by mass spectra and demonstrate its capabilities by analyzing 24 strains belonging to the Bacillus pumilus group. This method is based on representing mass spectra as points on a multidimensional space, which allows us to use geometric distances to compare the spectra. Delimitation of microorganisms performed by geometric approach correlates well with the results of molecular phylogenetic analysis and clustering using Biotyper 3.1. All three methods used allowed us to reliably divide the strains into two groups corresponding to closely related species, Bacillus pumilus and Bacillus altitudinis. The method developed by us will be implemented in a Web interface designed for using open reference databases for microorganism identification. The data is available at http://www.bionet.nsc.ru/mbl/database/database.html.

A Halorubrum H3 strain was isolated from a water and silt sample from Burlinskoye Lake (Altai Krai, Russia, 53°8'19″N 78°24'27″E). According to 16S rRNA sequences, this strain is most closely related to Halorubrum saccharovorum. The completely sequenced and annotated genome is 3,282,373 bp and contains 3,237 genes., (Copyright © 2015 Rozanov et al.)

Anoxybacillus flavithermus strain 25 was isolated from a sediment sample from the Garga hot spring in the Barguzin Valley, Baikal Region, Russian Federation (54°19'3.72″N, 110°59'38.4″E). The sequenced and annotated genome is 2,838,680 bp and encodes 3,009 genes., (Copyright © 2014 Rozanov et al.)

The Geobacillus icigianus G1w1(T) strain was isolated from sludge samples of unnamed vaporing hydrothermal (97°С) outlets situated in a geyser in the Troinoy region (Valley of Geysers, Kronotsky Nature Reserve, Kamchatka, Russian Federation; 54°25'51.40″N, 160°7'41.40″E). The sequenced and annotated genome is 3,457,810 bp and encodes 3,342 genes., (Copyright © 2014 Bryanskaya et al.)

Background: Geothermal areas are of great interest for the study of microbial communities. The results of such investigations can be used in a variety of fields (ecology, microbiology, medicine) to answer fundamental questions, as well as those with practical benefits. Uzon caldera is located in the Uzon-Geyser depression that is situated in the centre of the Karym-Semyachin region of the East Kamchatka graben-synclinorium. The microbial communities of Zavarzin spring are well studied; however, its benthic microbial mat has not been previously described., Results: Pyrosequencing of the V3 region of the 16S rRNA gene was used to study the benthic microbial community of the Zavarzin thermal spring (Uzon Caldera, Kamchatka). The community is dominated by bacteria (>95% of all sequences), including thermophilic, chemoorganotrophic Caldiserica (33.0%) and Dictyoglomi (24.8%). The benthic community and the previously examined planktonic community of Zavarzin spring have qualitatively similar, but quantitatively different, compositions., Conclusions: In this study, we performed a metagenomic analysis of the benthic microbial mat of Zavarzin spring. We compared this benthic community to microbial communities found in the water and of an integral probe consisting of water and bottom sediments. Various phylogenetic groups of microorganisms, including potentially new ones, represent the full-fledged trophic system of Zavarzin. A thorough geochemical study of the spring was performed.