SCREENING AND CHARACTERIZATION OF PSYCHROPHYLIC XYLANOLYTIC BACTERIA

Temperature is the most significant parameter that plays a key role in the distribution of microorganisms on our planet. Psychrophilic microorganisms are the most prominent inhabitants of the cold ecosystems, producing cold-adapted enzymes, particularly useful due to their high catalytic activity at low temperatures, offering energy-saving advantages, this feature makes them more appealing and attractive in biotechnology [1]. Xylanolytic microorganisms, especially bacteria, have been reported from various extreme environments, such as marine habitats or Antarctic regions. They breakdown a xylan, the second most abundant naturally occurring renewable polysaccharide available on earth. The complete hydrolysis of xylan is challenging due to its heterogeneous structure, it requires a complex of several cooperatively acting enzymes collectively known as xylanases. The suitability of xylanases for its application in food and feed, paper and pulp, textile, pharmaceuticals, and lignocellulosic biorefinery has led to an increase in demand of xylanases globally. Although cold-adapted xylanases are not very common, some bacteria have been reported to produce them [2,3]. The objective of this study was to search and characterize psychrophylic xylanolytic bacteria, isolated from the compost sample in Vilnius. Ten isolated cultures were selected and genotyped using BOX-PCR. Our analysis showed that these 10 isolates belong to 10 different strains. 16S rRNA genes of these isolates were amplified and sequenced. BLAST analysis showed that the strains could be assigned to the genera Rhodococcus, Flavobacterium, Arthrobacter, Sporosarcina, Sphingobacterium and Pseudomonas. One of these strains (S31) was selected for further work due to its lowest 16S rRNA gene similarity (93.99%). The S31 strain was grown in a medium, where xylan was the only carbon source; cultivation was carried out in a cold room at 6 °C. The Congo red staining method was used to show the activity of xylanases. According to 16S rRNA gene sequence analysis as well as Oxford Nanopore PromethION and Illumina NovaSeq sequencing results, the S31 strain was identified as Pseudomonas bubulae S31 strain. The genome was submitted to the Genbank, acc. No. CP178394-CP178396. The genome consists of 3 contigs – chromosome and two plasmids. In the genome, 106 CAZY protein genes have been identified, 22 of which were supposed to be secreted. Three secreted protein genes have been identified that could potentially be involved in xylan degradation.