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0INTERNATIONAL SCIENTIFIC AND PRACTICAL CONFERENCE "STATUS AND DEVELOPMENT PROSPECTS OF FUNDAMENTAL AND APPLIED MICROBIOLOGY: THE VIEWPOINT OF YOUNG SCIENTISTS" _25-26 SEPTEMBER, 2024_
BIOTECHNOLOGY POTENTIAL AND CHARACTERISTICS
OF THERMOPHILIC BACTERIA ISOLATED FROM THE KHOJA-OBI-GARM GEOTHERMAL SPRING (TAJIKISTAN)
1Dzhuraeva Munavvara, 2Birkeland Nils-Kaare, 3Bobodzhanova Khursheda
1Senior researcher, Center of Biotechnology of the TNU, Dushanbe, Tajikistan 2PhD, professor Department of Biological Sciences, University of Bergen, Bergen, Norway 3Candidat of biological science, associated professor, Director of the Center of Biotechnology of
the TNU, Dushanbe, Tajikistan https://doi.org/10.5281/zenodo.13846073
Abstract. To date, many strains of Thermus have been isolated from various geothermal environments such as hot springs and deep-sea hydrothermal vents. In an effort to recover thermophilic microorganisms and enzymes for industrial applications, a new thermophilic bacterium belonging to the Deinococcota phylum was isolatedfrom hot soil (93 oC; pH 8,5) in the Khodja-Obi-Garm field located on the Bank of the Varzob river, (Tajikistan). Draft genome sequencing yielded 2,169,573 bp of unique sequence data distributed into 124 contigs with an average GC content of67.76% and a genome completeness of 100%. Average nucleotide identity (ANI) and digital DNA - DNA hybridization (dDDH) values were 97.71 % and 68.6%, respectively, demonstrating that strain Thermus KhOG constitutes a separate and distinct genome species, and the first reported thermophile from Tajikistan.
Acknowledgments
This work was funded by the Eurasia Program of the Norwegian Directorate for Higher Education and Skills (HK-dir) (CPEA-LT-2017/10061).
Keywords: Tajikistan, thermostable enzymes, Thermus, geothermal spring
Extremophiles produce numerous extremozymes with different industrial applications, including agricultural, chemical, and medicinal applications. They are found in almost any habitat. Due to these advantages, extremozymes will be increasingly used in a wide range of consumer products. Extremophiles improve our understanding of macromolecular stability and physiochemical requirements for life. According to new research adaptations that enable survival under one stress, may also allow survival under other stress conditions. Finally, studying extreme organisms contributes greatly to biotechnology's ongoing development. Understanding how life can thrive on Earth may help biotechnology better understand and locate potential life in others [1].
The enzymes produced by extremophiles have significant applications in different industries like detergent, food, feed, starch, textile, leather, pulp and paper, and pharmaceuticals [2].
The impact of biotechnology on our lives is inescapable. Some of these impacts are well publicized, like the process of generating biofuels. However, there are numerous other applications that are not widely known outside of specialist circles that affect our daily life, such as food and drink (e.g. lactose-free milk [3] and bioinsecticides [4]), how we make and wash our clothes (e.g. cellulases to produce 'stone-washed' jeans [5], lipases [6], and proteases [7] in detergents), and the medications we take to remain healthy, just to name a few examples.
INTERNATIONAL SCIENTIFIC AND PRACTICAL CONFERENCE "STATUS AND DEVELOPMENT PROSPECTS OF FUNDAMENTAL AND APPLIED MICROBIOLOGY: THE VIEWPOINT OF YOUNG SCIENTISTS" _25-26 SEPTEMBER, 2024_
To date, few extremophiles/extremozymes have found their way into large-scale use in the field of biotechnology [8]; however, their potential is undeniable in many applications. Four success stories are the thermostable DNA polymerases used in the polymerase chain reaction (PCR) [9], various enzymes used in the process of making biofuels [10], organisms used in the mining process [11], and carotenoids used in the food and cosmetic industries [12]. Other potential applications include making lactose-free milk [3]; the production of antibiotics, anticancer, and antifungal drugs [13]; and the production of electricity or, more accurately, the leaching of electrons to generate current that can be used or stored [1]. Enzymes from thermophiles are of great interest for research and bioengineering due to their stability and efficiency [14].
In an effort to recover thermophilic microorganisms and enzymes for industrial applications, a new thermophilic bacterium belonging to the Deinococcota phylum was isolated from hot soil (93 oC; pH 8,5) in the Khodja-Obi-Garm field located on the Bank of the Varzob river, (Tajikistan), at 38°53'41.45" N, 68°47'15.64" E, at an elevation of 1835 m, with a conductivity of 4378.3 |S/cm [15, 16].
This bacterium was isolated from a hot water samples. The isolate was an obligately aerobic, Gram-negative, non-sporulating, and rod-shaped thermophilic bacteria, which formed yellow pigmented colonies on R2A agar plates. The isolate, designated strain Thermus KhOG, grew in the temperature range from 55 - 75C (optimum, 70°C), and at pH values ranging from 5 to 10 (optimum, pH 7.2). DNA was extracted from cells cultivated in R2A for 24 hours at 65°C with shaking using the GenElute bacterial genomic DNA kit (Sigma-Aldrich). Based on the 16S rRNA gene sequence, it was identified as a member of the Thermus genus, sharing 99.99% sequence identity with the species in this genus Thermusparvatiensis. For genome sequencing by Eurofins Genomics, a NEBNext Ultra II DNA preparation kit was used, and Illumina NovaSeq 6000 S2 paired-end genomic sequencing was performed. Reads with a maximum of 7 bases with a Phred score below 28 were discarded. Additional quality control was performed using the Trim Reads tool in the CLC Genomics Workbench v. 20.1. Assembly was performed using the CLC de novo assembly tool. Draft genome sequencing yielded 2,169,573 bp of unique sequence data distributed into 124 contigs with an average GC content of 67.76% and a genome completeness of 100%. Average nucleotide identity (ANI) and digital DNA - DNA hybridization (dDDH) values were 97.71 % and 68.6%, respectively, demonstrating that strain Thermus KhOG constitutes a separate and distinct genome species, and the first reported thermophile from Tajikistan. It actively degraded cellulose, casein, starch, and amylase, and yielded positive results for a large number of hydrolytic enzymes e.g., alkaline phosphatase, C-4 esterase, C-8 esterase lipase, C-14 lipase, chymotrypsin, acid phosphatase, phosphoamidase, a-glucosidase, at 65 oC, and thus represents a good source of potentially valuable industrial enzymes. This study demonstrated that diverse and novel thermophilic bacterial populations with biotechnological potentials thrive in high-altitude geothermal springs in Tajikistan. Most of the microbial isolates are still mostly uncharacterized and awaits further exploration. Environmental factors play an important role in structuring microbial communities, and hence these factors should be considered in future analyses.
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INTERNATIONAL SCIENTIFIC AND PRACTICAL CONFERENCE "STATUS AND DEVELOPMENT PROSPECTS OF FUNDAMENTAL AND APPLIED MICROBIOLOGY: THE VIEWPOINT OF YOUNG SCIENTISTS"
_25-26 SEPTEMBER, 2024_
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