1. INTRODUCTION

The Yeongsan River is a prime domestic waterway that drains freshwater from an area spanning approximately 3,371 km2 and extending across 120 km in the southwestern part of Korea into the Yellow Sea. Additionally, it flaunts an elaborate estuary and extensive tidal flats (approximately 280 km2) owing to its macrotidal characteristics. Research focused on the rivers of Korea predominantly investigates the four principal rivers, including the Han and the Nakdong River, with surveys being conducted on the Geumgang, Yeongsan, and other river in recent times. The study of biological diversity or ecology in the Yeongsan River basin commenced in 1974 and has been constrained to a handful of instances until recently. The majority of research has emphasized the Yeongsan Lake (Son et al. 2018) and to the best of our knowledge only two studies have investigated phytoplankton and periphyton algae in the main stream of the Yeongsan River (Kim et al. 2003;Cho et al. 2010). As part of a research program orchestrated by the Nakdonggang National Institute of Biological Resources (NNIBR) in 2020-23, numerous bacterial strains were isolated from freshwater and sediment specimens sampled from the Yeongsan River basin. Among the aforementioned strains, a total of 45 bacterial strains were affiliated to unrecorded species in Korea thus far. These bacterial species belong to phyla Actinomycetota, Bacillota, Acidobacteriota, Bacteroidota, Verrucomicrobiota, and Pseudomonadota based on phylogenetic analysis individually.

2. MATERIALS AND METHODS

Freshwater and sediment samples were collected from the Yeongsan River basin individually (Table 1), and then inoculated on to Reasoner’s 2A agar (R2A; BD), tryptic soy agar (TSA; BD), 0.1 TSA, marine agar 2216 (MA; BD), 0.1 MA and nutrient agar using a standard dilution plating technique. The culture plates were incubated for 3 days at 25°C. All the strains were purified as single colonies and stored in a 20% glycerol suspension (v/v) at -80°C (Jeon et al. 2023). The designated strain IDs and culture media are summarized in Table 1. Following the attainment of the stationary phase by the bacterial cells, the colony morphologies of the strains were scrutinized on the agar plates using a magnifying glass. The cellular morphologies and sizes were further examined using a transmission electron microscope (H-7650; Hitachi, Japan), following which the strains were Gram stained using a Gram staining kit (BD Biosciences, Franklin Lakes, NJ, USA). The biochemical and enzymatic characteristics of the strains were assessed using the API 20NE and API ZYM (BioMérieux, Marcy-l’Étoile, France), respectively. Catalase and oxidase activity were examined using 3% hydrogen peroxide and oxidase reagent (BioMérieux), respectively. Phylogenetic positions were assigned to the Gram-negative and positive strains through 16S rRNA gene analysis. The 16S rRNA gene of individual strains was amplified using PCR and sequenced in adherence to a previously described procedure (Baek et al. 2020); these 16S rRNA gene sequences were analyzed using the EzBioCloud server per prior methodology (Yoon et al. 2017), and a cutoff value of 98.7% sequence similarity was applied to determine the bacterial species. For phylogenetic analyses, the 16S rRNA gene sequences were aligned using EzEditor (Jeon et al. 2014), and a phylogenetic tree was constructed via neighbor-joining (Saitou and Nei 1987) using MEGA version 11 (Tamura et al. 2021). The topology of the phylogenetic tree was evaluated using bootstrap analysis (Felsenstein 1985) with 1,000 replications.

3. RESULTS AND DISCUSSION

Total 45 bacterial strains were isolated (Table 1) and showed ≥98.7% 16S rRNA gene sequence similarities with unrecorded bacterial species in Korea. Phylogenetic analyses were conducted using the 16S rRNA gene sequences of the 45 strains (20YS10WR-12, R20SS-10, R20SS-19, 21YS07S-S28, 01TPW-2-14, 20CGW-82, 20CGW-69, YGJ-S-30, TM2-4, 20YS06W0N-35, 20YS 10WR-09, 21YS08-0M08, 21YS05S-0R27, 21YS03W- 49, 20YS06W0N-38, SJ2-62, 21YS05W-0N23, T10, 01MPW-2-28, 01TPW-2-35, YGS-S-32, SN-1, 21YS02S- 57, 20Ga-4, 21YS01W-91, 21YS16S-07, R20SW-21, YGS-S-16, 20YS10WR-01, 21YS07S-S12, 21YS10WISP05, 20DYS-68, SN-2, 21YS16S-51, NM40, 20YS 09W-41, 21YS14W-18, 20DYS-41, 20SRW-23, TM2-1, 21YS13S-68, 21YS07S-S44, 20DYW-34, 21YS21S-6, and R20SW-11) isolated from the Yeongsan River basin; contingent upon the results, the strains were assigned to the phyla Bacteroidota (four species), Pseudomonadota (26 species), Verrucomicrobiota (one species; Fig. 1), Actinomycetota (12 species), Acidobacteriota (one species), and Bacillota (one species; Fig. 2) and were categorized into nine classes, 18 orders, 25 families, and 39 genera. The taxonomic identities of the isolates have been documented in Table 1. Transmission electron micrographs of the isolates have been displayed in Fig. 3. The morphological, physiological, and biochemical properties of the isolated strains have been detailed in the subsequent subsections.

3.1. Description of Chitinophaga silvisoli 20YS10WR-12

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, spread, and yellow colored after incubation for 2-3 days at 20°C. Positive for Esculin hydrolyzed, gelatin hydrolyzed, β-galactosidase and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase and urease. L-arabinose (weakly) are utilized. It does not utilize D-glucose, D-mannose, D-mannitol, N-acetylglucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20YS10WR-12 (=NNIBR2021 641BA1529) was isolated from freshwater of the Yeongsan River, Yeongam-gun, Jeollanam-do, Korea. The Gen Bank accession number of 16S rRNA gene sequence is MZ976823.

3.2. Description of Flavobacterium lacunae R20SS-10

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are round, convex, and yellow colored after incubation for 2-3 days at 25°C. Positive for esculin hydrolysis (weakly), gelatin hydrolysis and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease and β-galactosidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain R20SS-10 (=NNIBR2020641BA613) was isolated from soil of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW020255.

3.3. Description of Flavobacterium saliperosum R20SS-19

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are round, convex, and yellow colored after incubation for 2-3 days at 25°C. Positive for gelatin hydrolysis. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, β-galactosidase and oxidase. Malic acid (Weakly) is utilized. It does not utilize D-glucose, L-Arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, Dmaltose, potassium gluconate, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain R20SS- 19 (=NNIBR2020641BA614) was isolated soil of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW020254.

3.4. Description of Flavobacterium urocaniciphilum 21YS07S-S28

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and yellow colored after incubation for 2-3 days at 25°C and pH 7.0. Positive for esculin hydrolysis and gelatin hydrolysis. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS07S-S28 (=NNIBR2021641BA1663) was isolated from soil of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OL677218.

3.5. Description of Rhodopseudomonas faecalis 01TPW-2-14

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped (vibrio or peanut shaped). Colonies grown on 0.1 TSA are circular, smooth, and ivory colored after incubation for 4 days at 25°C. Positive for arginine dihydrolase, urease, esculin hydrolysis and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, gelatin hydrolysis and β-galactosidase. Adipic acid and malic acid are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, trisodium citrate, and phenylacetic acid. Strain 01TPW-2-14 (=NNIBR2020641 BA786) was isolated from freshwater of the Yeongsan River, Gwansan-dong, Gwansan-gu, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW367778.

3.6. Description of Gemmobacter tilapiae 20CGW-82

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on 0.1 MA are circular, convex, smooth, and ivory colored after incubation for 7 days at 25°C. Positive for urease, esculin hydrolysis, β-galactosidase and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase and gelatin hydrolysis. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20CGW- 82 (=NNIBR2020641BA807) was isolated from freshwater of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW295520.

3.7. Description of Paracoccus luteus 20CGW-69

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex, smooth, and cream colored after incubation for 3 days at 25°C. Positive for urease, esculin hydrolysis and gelatin hydrolysis. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, β-galactosidase and oxidase. Larabinose, D-mannose, D-mannitol, potassium gluconate, and adipic acid are utilized. It does not utilize D-glucose, N-acetyl-glucosamine, D-maltose, capric acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20CGW-69 (=NNIBR2021641BA1593) was isolated from freshwater of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562842.

3.8. Description of Azospirillum agricola YGJ-S-30

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation at 25°C and pH 7.0. Positive for nitrate reduction, urease, esculin hydrolysis and β-galactosidase. But negative for indole production, glucose fermentation, arginine dihydrolase, gelatin hydrolysis and oxidase. L-arabinose, potassium gluconate, capric acid, malic acid, and trisodium citrate are utilized. It does not utilize D-glucose, D-mannose, D-mannitol, N-acetylglucosamine, D-maltose, adipic acid and phenylacetic acid. Strain YGJ-S-30 (=NNIBR2020641BA818) was isolated from soil of the Yeongsan River, Jiwon-dong, Dong-gu, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW293 922.

3.9. Description of Sphingomonas mesophila TM2-4

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are spread, and white colored after incubation for 1-2 days at 25°C and pH 7.0. Negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain TM2-4 (=NNIBR2020641BA828) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW34 5951.

3.10. Description of Novosphingobium piscinae 20YS06W0N-35

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire edge, and yellow colored after incubation for 1-2 days at 25°C and pH 7.0. Positive for esculin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. D-maltose are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, Nacetyl- glucosamine, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20YS06W0N-35 (=NNIBR2020641 BA829) was isolated from freshwater of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW34 5954.

3.11. Description of Sphingomonas laterariae 20YS10WR-09

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and ivory colored after incubation for 2-3 days at 20°C. Positive for esculin hydrolysis, β-galactosidase and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease and gelatin hydrolysis. L-arabinose (weakly) and N-acetyl-glucosamine (weakly) are utilized. It does not utilize D-glucose, D-mannose, D-mannitol, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20YS10WR-09 (=NNIBR 2021641BA1528) was isolated from freshwater of the Yeongsan River, Yeongam-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MZ976822.

3.12. Description of Sphingomonas lacunae 21YS08-0M08

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and yellow colored after incubation for 2-3 days at 20°C. Positive for oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis and β-galactosidase. Larabinose (weakly), and trisodium citrate (weakly) are utilized. It does not utilize D-glucose, D-mannose, Dmannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, and phenylacetic acid. Strain 21YS08-0M08 (=NNIBR2021641 BA1530) was isolated from freshwater of the Yeongsan River, Yeongam-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MZ976824.

3.13. Description of Sphingorhabdus planktonica 21YS05S-0R27

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex, smooth, opaque, and orange colored after incubation for 3 days at 25°C. Positive for arginine dihydrolase, urease, esculin hydrolysis, β-galactosidase and oxidase. But negative for nitrate reduction, indole production, glucose fermentation and gelatin hydrolysis. D-glucose, D-mannitol, D-maltose, potassium gluconate and adipic acid are utilized. It does not utilize L-arabinose, D-mannose, N-acetyl-glucosamine, capric acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS05S-0R27 (=NNIBR2021641BA1668) was isolated from soil of the Yeongsan River, Jiya-dong, Bukgu, Gwangju, Korea. The GenBank accession number of 16S rRNA gene sequence is MZ976831.

3.14. Description of Erythrobacter sanguineus 21YS03W-49

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex, smooth, and orange colored after incubation for 3 days at 25°C. Positive for esculin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. D-glucose, D-mannose, D-maltose, adipic acid and malic acid are utilized. It does not utilize L-arabinose, D-mannitol, N-acetyl-glucosamine, potassium gluconate, capric acid, trisodium citrate, and phenylacetic acid. Strain 21YS03W-49 (=NNIBR2021641BA1588) was isolated from freshwater of the Yeongsan River, Damyang-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562844.

3.15. Description of Bordetella hinzii 20YS06W0N-38

Cells are Gram-stain-negative, aerobic, non-flagellated, and short rod-shaped. Colonies grown on R2A are convex with entire edge, and light pink colored after incubation for 1-2 days at 25°C and pH 7.0. Positive for esculin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20YS06W0N-38 (=NNIBR2020641 BA830) was isolated from freshwater of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW34 5955.

3.16. Description of Pararobbsia alpina SJ2-62

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation for 1-2 days at 25°C and pH 7.0. Positive for urease and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, esculin hydrolysis, gelatin hydrolysis and oxidase. D-glucose, D-Mannose, D-Mannitol and N-acetyl-glucosamine are utilized. It does not utilize L-arabinose, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain SJ2-62 (=NNIBR2020641BA822) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW34 5944.

3.17. Description of Polynucleobacter yangtzensis 21YS05W-0N23

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex, smooth, translucent, and cream colored after incubation for 3 days at 25°C. Positive for arginine dihydrolase, urease and esculin hydrolysis. But negative for nitrate reduction, indole production, glucose fermentation, gelatin hydrolysis, β-galactosidase and oxidase. Potassium gluconate, and trisodium citrate are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, capric acid, adipic acid, malic acid and phenylacetic acid. Strain 21YS05W-0N23 (=NNIBR2021 641BA1667) was isolated from freshwater of the Yeongsan River, Jiya-dong, Buk-gu, Gwangju, Korea. The GenBank accession number of 16S rRNA gene sequence is MZ976830.

3.18. Description of Rhodoferax bucti T10

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation for 1-2 days at 25°C and pH 7.0. Positive for urease, esculin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, gelatin hydrolysis and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, Dmaltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain T10 (=NNIBR2020641BA417) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam- do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW345948.

3.19. Description of Aquabacterium pictum 01MPW-2-28

Cells are Gram-stain-negative, aerobic, non-flagellated, and oval-shaped. Colonies grown on 0.1 MA are circular, convex with smooth, and ivory colored after incubation for 3 days at 25°C. Positive for urease, esculin hydrolysis and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, gelatin hydrolysis and β-galactosidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 01MPW- 2-28 (=NNIBR2020641BA784) was isolated from freshwater of the Yeongsan River, Gwangsan-dong, Gwangsan- gu, Gwangju, Korea. The GenBank accession number of 16S rRNA gene sequence is MW367779.

3.20. Description of Malikia spinosa 01TPW-2-35

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on 0.1 TSA are mucoid, irregular, convex, opaque, and white colored after incubation for 3 days at 25°C. Positive for esculin hydrolysis and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and β-galactosidase. D-glucose and D-mannitol are utilized. It does not utilize L-arabinose, D-mannose, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 01TPW-2-35 (=NNIBR2020641BA 785) was isolated from freshwater of the Yeongsan River, Gwangsan-dong, Gwangsan-gu, Gwangju, Korea. The GenBank accession number of 16S rRNA gene sequence is MW397777.

3.21. Description of Comamonas kerstersii YGS-S-32

Cells are Gram-stain-negative, aerobic, non-flagellated, and short rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation at 25°C and pH 7.0. Positive for nitrate reduction and oxidase. But negative for indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis and β-galactosidase. D-maltose are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetylglucosamine, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain YGS-S-32 (=NNIBR2020641BA821) was isolated from soil of the Yeongsan River, Hwasun-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW293925.

3.22. Description of Ideonella paludis SN-1

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are convex with entire edge, and white colored after incubation for 1-2 days at 25°C and pH 7.0. Positive for nitrate reduction. But negative for indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, Dmaltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain SN-1 (=NNIBR2020641BA824) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW345946.

3.23. Description of Paucibacter toxinivorans 21YS02S-57

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, smooth, convex, and ivory colored after incubation for 3 days at 25°C. Positive for nitrate reduction, esculin hydrolysis and β-galactosidase. But negative for indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. D-maltose and potassium gluconate are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS02S-57 (=NNIBR2021641BA1587) was isolated from soil of the Yeongsan River, Damyang-gun, Jeollanam- do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562843.

3.24. Description of Undibacterium amnicola 20Ga-4

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire edge, and cream white colored after incubation for 3 days at 25°C. Positive for oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis and β-galactosidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20Ga-4 (=NNIBR2020641BA781) was isolated from freshwater of the Yeongsan River, Damyang-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MT893381.

3.25. Description of Cupriavidus agavae 21YS01W-91

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, rough, opaque, and ivory colored after incubation for 3 days at 25°C. Positive for nitrate reduction, arginine dihydrolase, urease and esculin hydrolysis. But negative for indole production, glucose fermentation, gelatin hydrolysis, β-galactosidase and oxidase. Potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, Dmannitol, N-acetyl-glucosamine and D-maltose. Strain 21YS01W-91 (=NNIBR2021641BA1586) was isolated from freshwater of the Yeongsan River, Damyang-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562839.

3.26. Description of Thauera terpenica 21YS16S-07

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on MA are circular, and ivory colored after incubation at 25°C. Positive for nitrate reduction (weakly), indole production (weakly) and oxidase. But negative for glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis and β-galactosidase. Malic acid is utilized. It does not utilize D-glucose, L-arabinose, Dmannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain 21YS 16S-07 (=NNIBR2022641BA2315) was isolated from soil of the Yeongsan River, Samdo-dong, Naju-si, Jeollanam- do, Korea. The GenBank accession number of 16S rRNA gene sequence is OQ073742.

3.27. Description of Pectobacterium punjabense R20SW-21

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex, smooth, and white colored after incubation for 2-3 days at 25°C. Positive for nitrate reduction, glucose fermentation, esculin hydrolysis and β-galactosidase. But negative for indole production, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. Dglucose, L-arabinose, D-mannose, D-mannitol, N-acetyl- glucosamine, malic acid, and trisodium citrate are utilized. It does not utilize D-maltose, potassium gluconate, capric acid, adipic acid, and phenylacetic acid. Strain R20SW-21 (=NNIBR2020641BA616) was isolated from freshwater of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW020252.

3.28. Description of Arenimonas malthae YGS-S-16

Cells are Gram-stain-negative, aerobic, non-flagellated, and short rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation at 25°C and pH 7.0. Positive for esculin hydrolysis and oxidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and β-galactosidase. Malic acid is utilized. It does not utilize D-glucose, Larabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain YGS-S-16 (=NNIBR2020641BA820) was isolated from soil of the Yeongsan River, Hwasun-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW293924.

3.29. Description of Lysobacter xinjiangensis 20YS10WR-01

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and yellow colored after incubation for 2-3 days at 20°C. Positive for nitrate reduction, gelatin hydrolysis and oxidase. But negative for indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis and β-galactosidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20YS10 WR-01 (=NNIBR2021641BA1527) was isolated from freshwater of the Yeongsan River, Yeongam-gun, Jeollanam- do, Korea. The GenBank accession number of 16S rRNA gene sequence is MZ976821.

3.30. Description of Luteimonas notoginsengisoli 21YS07S-S12

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and yellow colored after incubation at 25°C. Positive for esculin hydrolysis, gelatin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS07S-S12 (=NNIBR 2021641BA1662) was isolated from soil of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OL677 217.

3.31. Description of Luteolibacter luojiensis 21YS10W-ISP05

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are convex with entire edge, and white colored after incubation for 2-3 days at 20°C. Positive for β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS10WISP05 (=NNIBR2021641BA1531) was isolated from freshwater of the Yeongsan River, Yeongam-gun, Jeollanam- do, Korea. The GenBank accession number of 16S rRNA gene sequence is MZ976825.

3.32. Description of Mycolicibacterium vaccae 20DYS-68

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on MA are circular, convex and yellow colored after incubation for 7 days at 25°C. Positive for urease. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, esculin hydrolysis, gelatin hydrolysis, β-galactosidase and oxidase. D-mannitol, potassium gluconate and malic acid are utilized. It does not utilize D-glucose, L-arabinose, D-mannose, N-acetyl- glucosamine, D-maltose, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain 20DYS- 68 (=NNIBR2020641BA809) was isolated from soil of the Yeongsan River, Damyang-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW295522.

3.33. Description of Angustibacter luteus SN-2

Cells are Gram-stain-negative, aerobic, non-flagellated, and long rod-shaped. Colonies grown on R2A are circular, convex with entire, and ivory colored after incubation for 1-2 days at 25°C. Negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain SN-2 (=NNIBR2020 641BA825) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW345947.

3.34. Description of Ornithinibacter aureus 21YS16S-51

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, and bright yellow colored after incubation for 7 days at 25°C. Positive for indole production. But negative for nitrate reduction, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetylglucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS16S-51 (=NNIBR2022641BA 2316) was isolated from soil of the Yeongsan River, Samdo- dong, Naju-si, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OQ073 743.

3.35. Description of Janibacter cremeus NM40

Cells are Gram-stain-negative, aerobic, non-flagellated, and cocci-shaped. Colonies grown on MA are circular, convex with entire, and white colored after incubation for 1-2 days at 25°C. Positive for β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis, gelatin hydrolysis and oxidase. L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate and malic acid are utilized. It does not utilize D-glucose, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain NM40 (=NNIBR2020641BA703) was isolated from freshwater of the Yeongsan River, Suncheon-si, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW345953.

3.36. Description of Aurantimicrobium minutum 21YS14W-18

Cells are Gram-stain-negative, aerobic, non-flagellated, and cocci-shaped. Colonies grown on R2A are circular, convex, smooth, and yellow colored after incubation at 25°C and pH 7.0. Positive for esculin hydrolysis. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS14W-18 (=NNIBR 2021641BA1659) was isolated from freshwater of the Yeongsan River, Hampyeong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OL677214.

3.37. Description of Luedemannella flava 20DYS-41

Cells are Gram-stain-negative, aerobic, non-flagellated, and branching hyphae-shaped. Colonies grown on R2A are circular, dry and yellowish ivory colored after incubation for 7 days at 25°C. Positive for nitrate reduction, urease, esculin hydrolysis, gelatin hydrolysis and β-galactosidase. But negative for indole production, glucose fermentation, arginine dihydrolase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20DYS-41 (=NNIBR2020641BA808) was isolated from soil of the Yeongsan River, Damyang-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW295521.

3.38. Description of Nocardioides iriomotensis 20SRW-23

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on MA are circular, flat, smooth, and beige colored after incubation for 7 days at 25°C. Positive for nitrate reduction, indole production, esculin hydrolysis, gelatin hydrolysis, β-galactosidase and oxidase. But negative for glucose fermentation, arginine dihydrolase and urease. D-glucose (weakly), D-mannose, D-mannitol, D-maltose, potassium gluconate and malic acid are utilized. It does not utilize L-arabinose, N-acetyl-glucosamine, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain 20SRW-23 (=NNIBR2020641BA806) was isolated from freshwater of the Yeongsan River, Suncheon- si, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW295519.

3.39. Description of Nocardioides szechwanensis TM2-1

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation for 1-2 days at 25°C and pH 7.0. Positive for nitrate reduction, gelatin hydrolysis and β-galactosidase. But negative for indole production, glucose fermentation, arginine dihydrolase, urease, esculin hydrolysis and oxidase. It does not utilize D-glucose, L-arabinose, Dmannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain TM2-1 (=NNIBR2020641BA826) was isolated freshwater of the Yeongsan River, Jangseong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW345949.

3.40. Description of Schumannella luteola 20YS09W-41

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, smooth, convex, and yellow colored after incubation for 3 days at 25°C. Positive for esculin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. D-glucose, L-arabinose, D-mannose, D-mannitol, D-maltose and malic acid are utilized. It does not utilize N-acetylglucosamine, potassium gluconate, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain 20YS09W-41 (=NNIBR2021641BA1592) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562840.

3.41. Description of Propionicimonas paludicola 20DYW-34

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, smooth, convex, and ivory colored after incubation for 3 days at 25°C. Positive for glucose fermentation, esculin hydrolysis and β-galactosidase. But negative for nitrate reduction, indole production, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. Larabinose, D-mannose, D-mannitol and N-acetyl-glucosamine are utilized. It does not utilize D-glucose, Dmaltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 20DYW-34 (=NNIBR2021641BA1594) was isolated from freshwater of the Yeongsan River, Damyanggun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562841.

3.42. Description of Nocardioides aquaticus 21YS13S-68

Cells are Gram-stain-negative, aerobic, non-flagellated, and cocci-shaped. Colonies grown on R2A are irregular, dry, undulate, and yellow colored after incubation at 25°C and pH 7.0. Positive for esculin hydrolysis and gelatin hydrolysis. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, β-galactosidase and oxidase. D-glucose, D-mannitol, potassium gluconate and malic acid are utilized. It does not utilize L-arabinose, D-mannose, N-acetyl-glucosamine, D-maltose, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain 21YS13S-68 (=NNIBR2021641BA1658) was isolated from soil of the Yeongsan River, Hampyeong-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OL677213.

3.43. Description of Nocardioides simplex 21YS07S-S44

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, convex with entire, and white colored after incubation at 25°C. Positive for nitrate reduction, esculin hydrolysis, gelatin hydrolysis and β-galactosidase. But negative for indole production, glucose fermentation, arginine dihydrolase, urease and oxidase. D-glucose, Dmaltose, potassium gluconate and malic acid are utilized. It does not utilize L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, capric acid, adipic acid, trisodium citrate, and phenylacetic acid. Strain 21YS07SS44 (=NNIBR2021641BA1664) was isolated from soil of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is OL677219.

3.44. Description of Geothrix fermentans 21YS21S-6

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are circular, smooth, convex, and yellow colored after incubation for 3 days at 25°C. Positive for nitrate reduction and esculin hydrolysis. But negative for indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis, β-galactosidase and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain 21YS21S-6 (=NNIBR2021641BA1591) was isolated from freshwater of the Yeongsan River, Jangseong-gun, Jeollanam- do, Korea. The GenBank accession number of 16S rRNA gene sequence is OK562847.

3.45. Description of Lactococcus hircilactis R20SW-11

Cells are Gram-stain-negative, aerobic, non-flagellated, and rod-shaped. Colonies grown on R2A are round, convex, and cream colored after incubation for 2-3 days at 25°C. Positive for esculin hydrolysis (weakly) and β-galactosidase. But negative for nitrate reduction, indole production, glucose fermentation, arginine dihydrolase, urease, gelatin hydrolysis and oxidase. It does not utilize D-glucose, L-arabinose, D-mannose, D-mannitol, N-acetyl-glucosamine, D-maltose, potassium gluconate, capric acid, adipic acid, malic acid, trisodium citrate, and phenylacetic acid. Strain R20SW-11 (=NNIBR2020641BA615) was isolated from freshwater of the Yeongsan River, Muan-gun, Jeollanam-do, Korea. The GenBank accession number of 16S rRNA gene sequence is MW020253.