Discovery of two novel Flavobacterium species with potential for complex polysaccharide degradation

Abstract

Polysaccharides are recognized for their extensive biological functions, holding significant promise for applications in both medicine and food industries. However, their utilization is frequently constrained by challenges such as high molecular weights and indistinct sugar chain structures. Recently, two novel bacterial strains, N6^T and J3^T, were isolated from the Nakdong River in Korea. These strains, which belong to the phylum Bacteroidota, are Gram-stain-negative, non-motile, aerobic, rod-shaped bacteria and have shown polysaccharide-degrading capabilities. Through comprehensive analyses, including 16S rRNA gene sequencing, whole-genome sequencing, and detailed morphological, physiological, and chemotaxonomic characterizations, these strains have been identified as new species within the genus Flavobacterium. KEGG pathway analysis further confirmed their robust capabilities for carbohydrate utilization. Additional investigations using the dbCAN and dbCAN-PUL databases identified the presence of carbohydrate-hydrolyzing enzymes (CAZymes) and polysaccharide utilization loci (PULs) within these strains, suggesting their potential to degrade various polysaccharides. Subsequent in vitro growth experiments demonstrated that strains N6^T and J3^T can degrade chitin, β-glucan, κ-carrageenan, and cellulose. Given their diverse polysaccharide degradation abilities, these strains are formally proposed to be named Flavobacterium polysaccharolyticum sp. nov. and Flavobacterium aureirubrum sp. nov. The type strains are designated as N6^T (= KCTC 102173^T = GDMCC 1.4609^T) and J3^T (= KCTC 102172^T = GDMCC 1.4608^T), respectively.

Keywords: Bacteroidota; Flavobacterium; Polyphasic taxonomy; Polysaccharide.

Fig. 1

Maximum likelihood (ML) tree based on 16S rRNA gene sequences showing the position of strains N6^T and J3^T closely related taxa within the genus Flavobacterium. The sequences of Gramella echinicola, Christiangramia aestuariivivens, Gramella marina, Gramella marina, Christiangramia forsetii were used as an outgroup. Bootstrap values are shown at branching points. In the phylogenetic tree, the outgroup is indicated by a green branch, and strains N6^T and J3^T are marked with red font. The outer circle includes purple and cyan bar charts representing the similarity of 16S rRNA gene sequences between Flavobacterium strains and strains N6^T and J3^T, respectively.

Fig. 2

Scanning electron microscope image exhibiting cell morphology of the strains N6^T (A-C) and J3^T (D-F). The magnifications are: A and D, ×22,000; B, C, and E, ×35,000; F, ×30,000. Scale bars represent 400 nm.

Fig. 3

Maximum likelihood (ML) tree showing the phylogenetic position of Flavobacterium. The sequences of Christiangramia forsetii and Christiangramia echinicolawere used as an outgroup. Taxon names are followed by strain names and GenBank accession numbers. Bootstrap values are shown at branching points. In the phylogenetic tree, the outgroup is indicated by a red branch, and strains N6^T and J3^T are marked with red font. The inner circle uses square colors to represent different sources of the strains, while the outer circle uses five colored shapes to indicate the presence of CAZymes enriched based on PULs. Solid shapes indicate the presence of these CAZymes, while hollow shapes indicate their absence.

Fig. 4

The functional features between strains N6^T and J3^T with closely related strains of the genus Flavobacterium. (A). Pairwise comparisons of average nucleotide identity (ANI). The color indicates the value of ANI, the value range is 70–100 with color turning from blue to red. (B). Heatmap of discriminated metabolic pathways within the genome of the two novel strains and the reference strains in genus Flavobacterium, The scale bar indicates that the intensity of color change reflects the gene number of pathways.

Fig. 5

The carbohydrate-active enzyme (from dbCAN2 meta server) compositions of the strains N6^T and J3^T. The number of cazymes: GH: Glycoside Hydrolases; GT: Glycosyl Transferases; PL: Polysaccharide Lyases; CE: Carbohydrate Esterases; AA: Auxiliary Activities; CBM: Carbohydrate-Binding Modules.

Fig. 6

Growth assays of the strains of N6^T and J3^T cultured in basal medium supplemented with different polysaccharides.