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Isolation and characterization of seven bacterial strains in Jeju Island§
Korean J. Microbiol. 2023;59(3):184-191
Published online September 30, 2023
© 2023 The Microbiological Society of Korea.

Minji Kim1, Ki-Eun Lee2, In-Tae Cha2, and Soo-Je Park1*

1Department of Biology, Jeju National University, Jeju 63243, Republic of Korea
2Species Diversity Research Division, National Institute of Biological Resource, Incheon 22689, Republic of Korea
Correspondence to: *E-mail: sjpark@jejunu.ac.kr; Tel.: +82-64-754-3524; Fax: +82-64-756-3541
§Supplemental material for this article may be found at http://www.kjom.org/main.html
Received May 23, 2023; Revised July 24, 2023; Accepted July 25, 2023.
Abstract
Microorganisms have diverse metabolisms or characteristics that allow them to adapt and survive in various environments. They are also considered critical players that contribute to biogeochemical cycles and recycling organic matter degradation in their habitats. In this study, we isolated 50 bacterial strains from marine and non-marine environments in Jeju Island. Finally, seven strains were selected based on the sequence similarity of the 16S ribosomal RNA gene and identified as undiscovered isolated strain (> 98.7%) in South Korea. Using phylogenetic analyses, the strains were classified into five genera from three phyla. We observed that the isolates have various metabolic characteristics, including denitrifying or degradation of polymers. In addition, some strains could be grown under acidic (pH 4.5) or saline (NaCl 10%, w/v) culture conditions. This present investigation into the characterization of the isolated strains may provide basal information on microorganisms.
Keywords : 16S rRNA gene, Jeju, sediment, undiscovered bacterial strain, wastewater
Body

Locey and Lennon (2016) claimed that the microorganisms diversity predicts to 1012 species on Earth and these microorganisms can live in a variety of environments, including a wide range of temperatures or salt concentrations. Microorganisms possess diverse characteristics (e.g., bactericide production, [in]organic matter degradation) for adaptation and survival in their habitats (Haruta and Kanno, 2015). Microorganisms contribute to the biogeochemical cycles, mainly that of carbon and nitrogen, even in the gut of animal (Arrigo, 2005; Lorenz and Lal, 2009; Engel and Moran, 2013). For example, cellulose and starch, which are the principal polysaccharides in plants, are the dominant carbohydrates in nature. As aforementioned, this vast majority of the global carbohydrate-biomass can be recycled by microbial degradation activities. In addition, microorganisms can degrade macromolecules produced by organisms (i.e., protein, chitin, or agar) and play a crucial role in the turnover of matter in extreme environments (Pérez Castro et al., 2021). Therefore, we applied these metabolic features to generate a number of biologically active compounds used in medicine, agriculture, and industry (Sanchez et al., 2012). Through these characteristics, microorganisms play a crucial role in the circulation of the ecosystem of the Earth, and they can also be utilized as microbial resources in a variety of industrial fields, including the biological industry. Consequently, the isolation and obtaining of microorganisms with diverse physio-biochemical and metabolic capabilities is an essential aspect of the Nagoya Protocol implementation agenda (Buck and Hamilton, 2011). In this study, we isolated microorganisms and confirmed their characteristics in diverse marine and non-marine environments in Jeju Island, South Korea.

For this study, samples were collected from marine and terrestrial (i.e., non-marine) environments in Jeju Island, South Korea (see Table 1). To isolate bacterial strains, the samples were serially diluted to 10-4 using artificial media for freshwater (AFW) (Kim et al., 2020) or seawater (ASW) (Kim et al., 2021a). Final diluted aliquots were spread onto Reasoner’s 2A agar (R2A; Difco) or Marine 2216 agar (MA; Difco) and incubated at 30°C for two weeks. During cultivation, we carefully screened for fungal contamination, especially in soil samples. Then, to obtain a purified single colony, each colony was sub-cultivated on the same culture agar plate at least five times. Unless otherwise stated, all isolated strains were cultivated for three days. For further analysis, purified isolated-bacterial strains were preserved at -80°C in 30% (w/v) glycerol suspension. For phylogenetic analysis for each strain isolated in this study, we extracted genomic DNA (gDNA) using a Monarch® Nucleic Acid Purification kit (NEW England Biolabs). For 16S rRNA gene amplification, a universal bacterial 16S rRNA gene primer set (27F and 1492R) was used and polymerase chain reaction (PCR) was performed (Weisburg et al., 1991). The amplified 16S rRNA gene PCR products were sequenced by a certified service provider (Bioneer). DNA sequences were assembled using SeqMan (DNASTAR) and nearly full-length 16S rRNA sequences were obtained for all isolates. The sequences were edited using the BioEdit program and aligned using Clustal X (Thompson et al., 1997). The 16S rRNA gene sequences were initially compared to those of other bacterial strains with valid names using the EzBioCloud (https://www.ezbiocloud.net) and the NIBR database (https://species.nibr.go.kr/index.do). Phylogenetic trees were generated using the Kimura-two model (Kimura, 1989) and the neighbor-joining method (Saitou and Nei, 1987) in MEGA X (Kumar et al., 2018). Clade support was evaluated using 1,000 bootstrap replicates (Felsenstein, 1985). Cellular morphology was determined using a transmission electron microscopy (TEM) or scanning electron microscopy (SEM) (Supplementary data Fig. S1). Gram-staining was performed according to our previously published study (Kim et al., 2021b) for cells grown on R2A or MA at 30°C. Oxidase and catalase activities were tested using 1% (w/v) tetramethyl-p-phenylenediamine (Merck) and 3% (v/v) H2O2, respectively. Other physiological characteristics for growth range (optimal) of temperature, pH, and salt concentration were determined as described in our previous studies (Kim et al., 2020, 2021a, 2021b; Ryu et al., 2020). For pH range test, homopiperazine-1,4-bis-2-ethanesulfonic acid (pH 4.0–5.0), 2-(N-morpholino) ethanesulfonic acid (pH 5.5–6.5), 1,3-bis[tris(hydroxymethyl)methylamino]propane (pH 7.0–8.5), and 3-(cyclohexylamino)-1-propanesulfonic acid (pH 9.0–10.0) were used as a buffer (final conc. 10mM) (Ryu et al., 2020; Kim et al., 2021a). Biochemical characteristics were evaluated using API 20NE, API 32GN, and API ZYM strips (bioMérieux), according to the manufacturer’s instructions. The incubation time was 24 hours for API 20NE and 32GN, and 4 hours for API ZYM. During the API evaluation of marine isolates, we used a basal medium supplemented with NaCl (final 3% [w/v]), which was supplied by the API kit. To identify other metabolic capabilities, we estimated the denitrification and degradation activity of polymers such as tween 20, tween 80, starch or cellulose. For denitrification, each strain was cultivated under anaerobic culture condition on R2A or MA supplemented with nitrate (final conc. 5 mM). To estimate hydrolysis activity, we used R2A or MA added tween 20 and tween 80 (1% [v/v]), starch (1% [w/v]), and cellulose (1% [w/v]), and then incubated at 30°C for two weeks (Smibert and Krieg, 1994).

Summary of isolated strains from the phyla Pseudomonadota, Bacteroidota and Bacillota and their taxonomic affiliations.

All strains were cultured at 30°C for three days

Strain ID NIBR IDa GenBank accession number Most closely related species Similarity (%)b Isolated mediumc Isolation source (GPS) Taxonomic affiliation (Phylum; Class; Order; Family)
A2 NIBRBAC000509118 OL825025 Pseudoalteromonas tetraodonis 99.66 MA Aquaculture wastewater (33°33’38”N 126°48’53”E) Pseudomonadota; Gammaproteobacteria; Alteromonadales; Pseudoalteromonadaceae
S2-11 NIBRBAC000509119 OL825026 Priestia endophytica 100 MA Intertidal (33°30’41”N 126°53’54”E) Bacillota; Bacilli; Bacillales; Bacillaceae
B1 NIBRBAC000509120 OL825027 Pseudoalteromonas carrageenovora 99.07 MA Aquaculture wastewater (33°33’38”N 126°48’53”E) Pseudomonadota; Gammaproteobacteria; Alteromonadales; Pseudoalteromonadaceae
Fe1 NIBRBAC000509121 OL825028 Rhodanobacter denitrificans 100 R2A Forest sediment (33°27’27”N 126°33’29”E) Pseudomonadota; Gammaproteobacteria; Lysobacterales; Rhodanobacteraceae
Y3-1 NIBRBAC000509398 OP185243 Flavobacterium branchiicola 98.99 R2A Artificial pond sediment (33°28’12”N 126°29’40”E) Bacteroidota; Flavobacteriia; Flavobacteriales; Flavobacteriaceae
G5-1 NIBRBAC000509399 OP185244 Novosphinogobium clariflavum 98.92 R2A Gotjawal sediment (33°17’26”N 126°16’04”E) Pseudomonadota; Alphaproteobacteria; Sphingomonadales; Sphingomonadaceae
G6-2-2 NIBRBAC000509400 OP185245 Flavobacterium bizetiae 99.49 R2A Gotjawal sediment (33°17’26”N 126°16’04”E) Bacteroidota; Flavobacteriia; Flavobacteriales; Flavobacteriaceae

a Cells deposited at National Institute of Biological Resource (NIBR).

b Estimated by 16S rRNA gene sequence.

c MA, Marine agar; R2A, Reasner’s 2A agar.



In the present study, 50 isolates were obtained from marine and non-marine environments. Based on the phylogenetic analysis of the 16S rRNA gene sequences, we excluded duplicate species and finally isolated seven strains (A2, S2-11, B1, Fe1, Y3-1, G5-1, and G6-2-2). The strains were > 98.7%, representing species previously undiscovered in South Korea, belonging to five genera in three phyla. For the identity of the strains isolated in this study, strain A2 (1,518 bp) was most closely related to Pseudoalteromonas tetraodonis KMM 458T (AF214729; 99.66% sequence similarity), strain B1 (1,420 bp) to Pseudoalteromonas carrageenovora NBRC 12985T (AB 680359; 99.07% sequence similarity), strain S2-11 (1,499 bp) to Priestia endophytica 2DT (AF295302; 100% sequence identity), strain Fe1 (1,469 bp) to Rhodanobacter denitrificans 2APBS1T (JF719060; 100% sequence identity), strain Y3-1 (1,447 bp) to Flavobacterium branchiicola 59B-3-09T (HE 612102; 98.99% sequence similarity), strain G5-1 (1,389 bp) to Novosphingobium clariflavum CICC 11035sT (KU530129; 98.92% sequence similarity), and strain G6-2-2 (1,416 bp) to Flavobacterium bizetiae CIP 105534T (MT117836; 99.49% sequence similarity). The phylogenetic relationships between the isolates and closely related species are presented in Fig. 1. In addition, the resultant phenotypic analysis (i.e., pH and salt) showed that three strains (A2, S2-11, and B1) and two strains (Fe1 and G-5-1) had slightly halophilic and acid-resistant properties, respectively. The morphological, physiological, and biochemical characteristics of the isolated strains are described in detail below.

Fig. 1. Neighbor-joining tree based on 16S rRNA gene sequences showing the phylogenetic relationships between the isolated strains and their relative strains.
The sequences of Galbibacter marinus ck-I2-15T (EU928746) (A), Salinicoccus sediminis SV-16T (KF701618) (B) and Staphylococcus felis DSM 7377T (MF678879) (C) were used as the outgroup in this study, respectively. The GenBank accession numbers are provided in parentheses. Bootstrap values (> 60%) are shown at each branch. Bar, 0.01 (A and B) and 0.02 (C) substitutions per nucleotide position.

Description of Pseudoalteromonas tetraodonis A2

Strain A2 was isolated from aquaculture wastewater from Gujwa-eup, South Korea. A2 cells are Gram-stain-negative, rod-shaped (1.5 × 2.5 µm) and aerobic. The colonies are swarming and ivory colored after 3 days of incubation on MA at 30°C. Growth occurs at 4–37°C (optimum, 25°C), pH 6.0–9.0 (optimum, pH 7.0–7.5), and with 0.5–10% (w/v) NaCl (optimum, 3–4%). The cells could hydrolyze tween 80 variably but not starch, cellulose and tween 20. Cells are positive for oxidase, catalase, and glucose acidification, and negative for the reduction of nitrates (NO3-) to nitrite (NO2-), reduction of nitrates (NO3-) to nitrogen (N2), and indole production. In addition, the cells are positive for the utilization of D-glucose, D-mannitol, D-maltose, D-melibiose, L-histidine, L-proline, D-sucrose, acetate, L-alanine, glycogen, and L-serine, and weakly positive for propionate, but cells are negative for the utilization of L-arabinose, D-mannose, N-acetyl-D-glucosamine, gluconate, caprate, adipate, malate, citrate, phenyl-acetate, salicin, L-fucose, D-sorbitol, valerate, citrate, 2-ketogluconate, 3-hydroxy-butyrate, 4-hydroxy-benzoate, L-rhamnose, N-acetyl-D-glucosamine, D-ribose, inositol, itaconate, suberate, malonate, lactate, 5-ketogluconate and 3-hydroxy-benzoate as energy and carbon source. Moreover, cells show positive activity for β-glucosidase (esculin hydrolysis), protease (gelatin hydrolysis), β-galactosidase (PNPG), alkaline phosphatase, leucine arylamidase, acid phosphatase and naphtol-AS-BI-phosphohydrolase, and negative activity for arginine dihydrolase, urease, lipase (C14), cystine arylamidase, trypsin, α-chymotrypsin, α-galactosidase, β-galactosidase (ONPG), β-glucuronidase, α-glucosidase, β-glucosidase (6-Br-2-naphthyl-β-D-glucopyranoside), N-acetyl-β-glucosaminidase, α-mannosidase and α-fucosidase. Lastly, the cells have low activity for esterase (C4), esterase lipase (C8) and valine arylamidase.

Description of Priestia endophytica S2-11

Strain S2-11 was isolated from the intertidal region of Hado-ri in South Korea. S2-11 cells are Gram-stain-negative, rod-shaped (2.0 × 3.5 µm) and strictly aerobic. The colonies are circular, entire, and yellow colored after 3 days of incubation on MA at 30°C. Growth occurs at 10–45°C (optimum, 30°C), pH 6.0–8.5 (optimum, pH 7.0) and with 0.0–10% (w/v) NaCl (optimum, 0–0.5%). The cells could hydrolyze starch but not cellulose, tween 20 and 80. Cells are positive for oxidase and catalase, and negative for the reduction of nitrates (NO3-) to nitrite (NO2-), reduction of nitrates (NO3-) to nitrogen (N2), indole production, and glucose acidification. Additionally, the cells are positive for utilization of D-glucose, L-arabinose, D-mannose, N-acetyl-D-glucosamine, D-maltose, gluconate, malate, D-mannitol, salicin, D-melibiose, D-sorbitol, L-histidine, 2-ketogluconate, L-proline, L-rhamnose, D-ribose, inositol, D-sucrose and glycogen, and weakly positive for lactate and L-alanine, but the cells are negative for the utilization of caprate, adipate, citrate, phenyl-acetate, propionate, valerate, 3-hydroxy-butyrate, 4-hydroxy-benzoate, itaconate, suberate, malonate, acetate, 5-ketogluconate, 3-hydroxy-benzoate and L-serine as energy and carbon source. Moreover, the cells are positive activity for β-glucosidase (esculin hydrolysis), alkaline phosphatase, esterase (C4), leucine arylamidase and naphtol-AS-BI-phosphohydrolase, and negative activity for arginine dihydrolase, urease, protease (gelatin hydrolysis), β-galactosidase (PNPG), esterase lipase (C8), lipase (C14), valine arylamidase, cystine arylamidase, trypsin, α-chymotrypsin, acid phosphatase, α-galactosidase, β-galactosidase (ONPG), β-glucuronidase, α-glucosidase, β-glucosidase (6-Br-2-naphthyl-β-D-glucopyranoside), N-acetyl-β-glucosaminidase, α-mannosidase, and α-fucosidase.

Description of Pseudoalteromonas carrageenovora B1

Strain B1 was isolated from aquaculture wastewater from Gujwa-eup, South Korea. B1 cells are Gram-stain-negative, rod-shaped (1.5 × 2.5 µm) and strictly aerobic. The colonies are irregular and ivory colored after 3 days of incubation on MA at 30°C. Growth occurs at 10–37°C (optimum, 25°C), pH 5.5–9.0 (optimum, pH 7.0–7.5), and with 0.5–10% (w/v) NaCl (optimum, 3–4%). The cells could hydrolyze tween 80 variably but not starch, cellulose and tween 20. Cells are positive for oxidase, catalase, and glucose acidification, and negative for the reduction of nitrates (NO3-) to nitrite (NO2-), reduction of nitrates (NO3-) to nitrogen (N2), and indole production. The cells are also positive for utilization of D-glucose, D-mannose, D-mannitol, D-maltose, malate, propionate, valerate, L-histidine, L-proline, D-sucrose, acetate, L-alanine and L-serine, and weakly positive for gluconate and glycogen, but the cells are negative for utilization of L-arabinose, N-acetyl-D-glucosamine, caprate, adipate, citrate, phenyl-acetate, salicin, D-melibiose, L-fucose, D-sorbitol, 2-ketogluconate, 3-hydroxy-butyrate, 4-hydroxy-benzoate, L-rhamnose, N-acetyl-D-glucosamine, D-ribose, inositol, itaconate, suberate, malonate, lactate, 5-ketogluconate, and 3-hydroxy-benzoate as energy and carbon source. Moreover, cells are positive activity for β-glucosidase (esculin hydrolysis), β-galactosidase (PNPG), alkaline phosphatase, leucine arylamidase, acid phosphatase, and naphthol-AS-BI-phosphohydrolase, and weak activity for esterase (C4), esterase lipase (C8), valine arylamidase and α-chymotrypsin. The cells are negative activity for arginine dihydrolase, urease, protease (gelatin hydrolysis), lipase (C14), cystine arylamidase, trypsin, α-glalactosidase, β-galactosidase (ONPG), β-glucuronidase, α-glucosidase, β-glucosidase (6-Br-2-naphthyl- β-D-glucopyranoside), N-acetyl-β-glucosaminidase, α-mannosidase, and α-fucosidase.

Description of Rhodanobacter denitrificans Fe1

Strain Fe1 was isolated from pine forest sediments in Jeju-si, South Korea. Fe1 cells are Gram-stain-negative, rod-shaped (0.5 × 1.3 µm) and facultatively anaerobic. The colonies are circular and yellow colored after 3 days of incubation on R2A at 30°C. Growth occurs at 10–37°C (optimum, 30°C), pH 4.5–8.0 (optimum, pH 6.5) and with 0.0–2.0% (w/v) NaCl (optimum, 0.0–0.5%). The cells could not hydrolyze starch, cellulose, tween 20 and 80. Cells are positive for oxidase, catalase, and the reduction of nitrates (NO3-) to nitrogen (N2), and negative for the reduction of nitrates (NO3-) to nitrite (NO2-), indole production, and glucose acidification. Additionally, the cells are positive for the utilization of D-glucose, N-acetyl-D- glucosamine, D-maltose, 3-hydroxy-butyrate and acetate, and negative for the utilization of L-arabinose, D-mannose, D- mannitol, gluconate, caprate, adipate, malate, citrate, phenyl- acetate, salicin, D-melibiose, L-fucose, D-sorbitol, propionate, valerate, L-histidine, 2-ketogluconate, 4-hydroxy-benzoate, L- proline, L-rhamnose, D-ribose, inositol, D-sucrose, itaconate, suberate, malonate, lactate, L-alanine, 5-ketogluconate, glycogen, 3-hydroxy-benzoate and L-serine as energy and carbon source. Moreover, the cells are positive activity for β-glucosidase (esculin hydrolysis), alkaline phosphatase, esterase (C4), esterase lipase (C8), leucine arylamidase, valine arylamidase, cystine arylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase, α-glucosidase, β-glucosidase (6-Br-2-naphthyl-β- D-glucopyranoside) and N-acetyl-β-glucosaminidase, and weak activity for α-chymotrypsin. But the cells are negative activity for arginine dihydrolase, urease, protease (gelatin hydrolysis), β-galactosidase (PNPG), lipase (C14), trypsin, α-galactosidase, β-galactosidase (ONPG), β-glucuronidase, α-mannosidase and α-fucosidase.

Description of Flavobacterium branchiicola Y3-1

Strain Y3-1 was isolated from artificial pond sediment in Jeju-si, South Korea. Y3-1 cells are Gram-stain-negative, rod-shaped (1.0 × 0.3 µm) and aerobic. The colonies are circular and yellow colored after 3 days of incubation on R2A at 30°C. Growth occurs at 18–30°C (optimum, 25°C), pH 6.0–8.0 (optimum, pH 7.0) and with 0.0–1.0% (w/v) NaCl (optimum, 0.0%). The cells could not hydrolyze starch, cellulose, tween 20 and 80. Cells are positive for oxidase, catalase, and reduction of nitrates (NO3-) to nitrite (NO2-), and negative for indole production and glucose acidification. In addition, the cells are positive for the utilization of D-glucose, L-arabinose, D-mannose, N-acetyl-D-glucosamine, D-maltose, D-melibiose, D-sucrose and glycogen, and weakly positive for L-proline and acetate, but cells are negative for the utilization of D-mannitol, gluconate, caprate, adipate, malate, citrate, phenyl-acetate, salicin, L-fucose, D-sorbitol, propionate, caprate, valerate, citrate, L-histidine, 2-ketogluconate, 3-hydroxy-butyrate, 4- hydroxy-benzoate, L-rhamnose, D-ribose, inositol, itaconate, suberate, malonate, lactate, L-alanine, 5-ketogluconate, 3- hydroxy-benzoate and L-serine as energy and carbon source. Moreover, cells are positive activity for β-glucosidase (esculin hydrolysis), alkaline phosphatase, esterase (C4), esterase lipase (C8), leucine arylamidase, valine arylamidase, cystine arylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase, α-galactosidase, α-glucosidase, β-glucosidase (6-Br-2-naphthyl- β-D-glucopyranoside) and N-acetyl-β-glucosaminidase, and negative activity for arginine dihydrolase, urease, protease (gelatin hydrolysis), β-galactosidase (PNPG), lipase (C14), trypsin, α-chymotrypsin, β-galactosidase (ONPG), β-glucuronidase, α-mannosidase and α-fucosidase.

Description of Novosphingobium clariflavum G5-1

Strain G5-1 was isolated from sediments in Gotjawal, South Korea. G5-1 cells are Gram-stain-negative, rod-shaped (0.9 × 0.3 µm) and strictly aerobic. The colonies are circular, entire, and yellow colored after 3 days of incubation on R2A at 30°C. The cells are hydrolyzed with tween 20. Growth occurs at 18–37°C (optimum, 25°C), pH 4.5–8.5 (optimum, pH 7.0) and with 0.0–2.0% (w/v) NaCl (optimum, 0.0%). The cells could hydrolyze tween 20 variably but not starch, cellulose and tween 80. Cells are positive for oxidase and catalase activity, and negative for the reduction of nitrates (NO3-) to nitrite (NO2-), reduction of nitrates (NO3-) to nitrogen (N2), indole production and glucose acidification. In addition, cells are positive for utilization of D-glucose, D-maltose, 3-hydroxy-butyrate and L-proline, and weakly positive for acetate, but cells are negative for the utilization of L-arabinose, D-mannose, D- mannitol, N-acetyl-D-glucosamine, gluconate, caprate, adipate, malate, citrate, phenyl-acetate, salicin, D-melibiose, L-fucose, D-sorbitol, propionate, valerate, L-histidine, 2-ketogluconate, 4-hydroxy-benzoate, L-rhamnose, N-acetyl-D-glucosamine, D-ribose, inositol, itaconate, suberate, malonate, lactate, L- alanine, 5-ketogluconate, glycogen, 3-hydroxy-benzoate and L-serine as energy and carbon source. Moreover, cells are positive activity for alkaline phosphatase, esterase (C4), esterase lipase (C8), leucine arylamidase, valine arylamidase, acid phosphatase, naphtol-AS-BI-phosphohydrolase and α- glucosidase, and negative activity for arginine dihydrolase, urease, β-glucosidase (esculin hydrolysis), protease (gelatin hydrolysis), β-galactosidase (PNPG), lipase (C14), cystine arylamidase, trypsin, α-chymotrypsin, α-galactosidase, β- galactosidase (ONPG), β-glucuronidase, β-glucosidase (6-Br- 2-naphthyl-β-D-glucopyranoside), N-acetyl-β-glucosaminidase, α-mannosidase, and α-fucosidase.

Description of Flavobacterium bizetiae G6-2-2

Strain G6-2-2 was isolated from sediments in Gotjawal, South Korea. G6-2-2 cells are Gram-stain-negative, rod shaped (1.4 × 0.3 µm) and aerobic. The colonies are irregular and yellow colored after 3 days of incubation on R2A at 30°C. Growth occurs at 10–30°C (optimum, 25°C), pH 6.0–8.0 (optimum, pH 7.0) and with 0.0–1.0% (w/v) NaCl (optimum, 0.0%). The cells are positive for oxidase, catalase, and reduction of nitrates (NO3-) to nitrite (NO2-). In addition, the cells are positive for the utilization of D-glucose, L-arabinose, D-mannose, N-acetyl-D-glucosamine, D-maltose, salicin, L- rhamnose and glycogen, but cells are negative for the utilization of D-mannitol, gluconate, caprate, adipate, malate, citrate, phenyl-acetate, D-melibiose, L-fucose, D-sorbitol, propionate, valerate, L-histidine, 2-ketogluconate, 3-hydroxy- butyrate, 4-hydroxy-benzoate, L-proline, D-ribose, inositol, D-sucrose, itaconate, suberate, malonate, acetate, lactate, L-alanine, 5-ketogluconate, 3-hydroxy-benzoate and L-serine as energy and carbon source. Moreover, the cells are positive activity for β-glucosidase (esculin hydrolysis), alkaline phosphatase, esterase (C4), esterase lipase (C8), leucine arylamidase, valine arylamidase, acid phosphatase, naphtol- AS-BI-phosphohydrolase, α-glucosidase and β-glucosidase (6-Br-2-naphthyl-β-D-glucopyranoside), and negative activity for arginine dihydrolase, urease, protease (gelatin hydrolysis), β-galactosidase (PNPG), lipase (C14), cystine arylamidase, trypsin, α-chymotrypsin, α-galactosidase, β-galactosidase (ONPG), β-glucuronidase, N-acetyl-β-glucosaminidase, α-mannosidase and α-fucosidase.

적 요

미생물은 다양한 환경에 적응하고 생존하기 위하여 다양한 대사나 특성을 가지고 있으며, 생지화학적 순환과 유기물 분해 등에서 중요한 역할을 수행하고 있다. 본 연구에서는 제주도의 토양 및 해양 환경에서 50 여 종의 미생물을 발굴하였고, 분자계통학적 분석을 통하여 최종적으로 7종의 미생물을 분리하였다. 이들 미생물은 3문(Phylum) 5속(Genus)로 분류되었으며, 모두 국내에서 발견되지 않은 분리주로 확인되었다. 생리화학적 분석 결과, 분리된 균주들은 탈질화나 항생제 내성 또는 고분자 물질 분해능을 포함한 다양한 대사적 특성을 가지고 있음이 관찰되었다. 또한 일부 미생물들은 산성(pH 4.5) 또는 염(NaCl 10%, w/v) 배양 조건에서 성장할 수 있었다. 본 연구 결과는, 국내 미생물의 특성 분석에 대한 기초적인 정보를 제공할 것으로 기대된다.

Acknowledgments

This work was supported by grants from the National Institute of Biological Resources funded by the Ministry of Environment (No. NIBR202203112).

Conflict of Interest

Soo-Je Park is Editor of KJM. He was not involved in the review process of this article. Also, Authors have no conflicts of interest to report.

References
  1. Arrigo KR. 2005. Marine microorganisms and global nutrient cycles. Nature 437, 349-355.
    Pubmed CrossRef
  2. Buck M and Hamilton C. 2011. The Nagoya protocol on access to genetic resources and the fair and equitable sharing of benefits arising from their utilization to the convention on biological diversity. RECIEL 20, 47-61.
    CrossRef
  3. Engel P and Moran NA. 2013. The gut microbiota of insects - diversity in structure and function. FEMS Microbiol. Rev. 37, 699-735.
    Pubmed CrossRef
  4. Felsenstein J. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39, 783-791.
    Pubmed CrossRef
  5. Haruta S and Kanno N. 2015. Survivability of microbes in natural environments and their ecological impacts. Microbes Environ. 30, 123-125.
    Pubmed KoreaMed CrossRef
  6. Kim M, Cha IT, Lee KE, Lee BH, and Park SJ. 2021a. Kineobactrum salinum sp. nov., isolated from marine sediment. Int. J. Syst. Evol. Microbiol. 71, 004586.
    CrossRef
  7. Kim M, Hur M, Jung YJ, and Park SJ. 2020. Isolation and characterization of methylamine-degrading bacteria from soil. Korean J. Microbiol. 56, 140-145.
  8. Kim M, Lee KE, Cha IT, and Park SJ. 2021b. Draconibacterium halophilum sp. nov., a halophilic bacterium isolated from marine sediment. Curr. Microbiol. 78, 2440-2446.
    Pubmed CrossRef
  9. Kimura M. 1989. The neutral theory of molecular evolution and the world view of the neutralists. Genome 31, 24-31.
    Pubmed CrossRef
  10. Kumar S, Stecher G, Li M, Knyaz C, and Tamura K. 2018. MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol. Biol. Evol. 35, 1547-1549.
    Pubmed KoreaMed CrossRef
  11. Locey KJ and Lennon JT. 2016. Scaling laws predict global microbial diversity. Proc. Natl. Acad. Sci. USA 113, 5970-5975.
    Pubmed KoreaMed CrossRef
  12. Lorenz K and Lal R. 2009. Biogeochemical C and N cycles in urban soils. Environ. Int. 35, 1-8.
    CrossRef
  13. Pérez Castro S, Borton MA, Regan K, Hrabe de Angelis I, Wrighton KC, Teske AP, Strous M, and Ruff SE. 2021. Degradation of biological macromolecules supports uncultured microbial populations in guaymas basin hydrothermal sediments. ISME J. 15, 3480-3497.
    Pubmed KoreaMed CrossRef
  14. Ryu D, Kim M, Han B, Lee KE, Lee BH, Lee EY, Jung GY, Kim SJ, and Park SJ. 2020. Ferrovibrio terrae sp. nov., isolated from soil. Int. J. Syst. Evol. Microbiol. 70, 1042-1047.
    Pubmed CrossRef
  15. Saitou N and Nei M. 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol. Biol. Evol. 4, 406-425.
    CrossRef
  16. Sanchez S, Guzmán-Trampe S, Ávalos M, Ruiz B, Rodríguez-Sanoja R, and Jiménez-Estrada M. 2012. Microbial natural products, pp. 65-108. In Civjan N (ed.). Natural products in chemical biology, John Wiley & Sons, Inc., Hoboken, New Jersey, USA.
    KoreaMed CrossRef
  17. Smibert RM and Krieg NR. 1994. Phenotypic characterization, pp. 607-654. In Gerhardt P, Murray RGE, Wood WA, and Krieg NR (eds.). Methods for General and Molecular Bacteriology, American Society for Microbiology, Washington DC, USA.
  18. Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, and Higgins DG. 1997. The CLUSTAL_X Windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res. 25, 4876-4882.
    Pubmed KoreaMed CrossRef
  19. Weisburg WG, Barns SM, Pelletier DA, and Lane DJ. 1991. 16S ribosomal DNA amplification for phylogenetic study. J. Bacteriol. 173, 697-703.
    Pubmed KoreaMed CrossRef


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