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Sphingomonas abietis sp. nov., an Endophytic Bacterium Isolated from Korean Fir
1Korean Collection for Type Cultures (KCTC), Biological Resource Center, Korea Research Institute of Bioscience and Biotechnology, Jeongeup 56212, Republic of Korea
2Department of Biosystem and Bioengineering, KRIBB School of Biotechnology, University of Science and Technology (UST), Daejeon 34113, Republic of Korea
3Present address: National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences, Shanghai 200032, P.R. China
J. Microbiol. Biotechnol. 2023; 33(10): 1292-1298
Published October 28, 2023 https://doi.org/10.4014/jmb.2303.03017
Copyright © The Korean Society for Microbiology and Biotechnology.
Abstract
Keywords
Graphical Abstract
Introduction
In this study, we isolated a new bacterial strain designated PAMB 00755T from the leaves of Korean fir (
Materials and Methods
Isolation and Ecology
Leaves were collected from the Korean fir growing on Hallasan Mountain (33°21'42''N, 126°31'45''E) on Jeju Island, South Korea. To prepare the sample, five grams of leaves were subjected to surface sterilization using 1%NaOCl solution. The leaves were rinsed five times with distilled water. The sterilized leaves were then homogenized using a blender with 10 ml of 1× phosphate-buffered saline (PBS). To remove any debris, the resulting mixture was filtered through four layers of sterile cheesecloth and then serially diluted using a standard dilution method with 1× PBS buffer. A volume of 100 μl from the sample solution was evenly spread onto Reasoner's 2A agar (R2A, Difco) plates, followed by incubation at 25°C for 1 week. Single colonies were then re-streaked onto fresh R2A medium. Strain PAMB 00755T, which formed circular, smooth, yellow colonies, was selected for the following study. The strain was stored at -80°C in 10% skimmed milk. It is currently accessible through the Korean Collection for Type Cultures (KCTC 92781T) and the Guangdong Microbial Culture Collection Center (GDMCC 1.3779T). Unless otherwise noted, bacterial cells were cultured on R2A for 3–4 days before performing subsequent tests.
16S rRNA Gene Sequence Analysis
The extraction of genomic DNA from the bacteria was performed to serve as a template for PCR amplification of the 16S rRNA gene. Universal bacterial primers, specifically 27F and 1492R, were used for the amplification process. The PCR products were sequenced by a commercial company (Macrogen Inc., Korea) using the primers 27F, 518F, 800R, and 1492R [24]. The almost full length of the 16S rRNA sequence was assembled using Vector NTI software (1.6.1). The obtained sequences were compared in the EzBioCloud and the NCBI database [25]. The alignment of multiple sequences was performed using BioEdit software. Subsequently, the construction of the phylogenetic tree was carried out using the Molecular Evolutionary Genetics Analysis (MEGA) software version 11.0. The neighbor-joining (NJ), maximum likelihood (ML), and minimum parsimony (MP) methods were used for this analysis, with 1,000 bootstrap iterations [26]. The outgroup was
Whole-Genome Characteristics
Genomic DNA was extracted as previously described [10]. For whole-genome sequencing, the PacBio RSII system (Pacific Biosciences, Inc., USA) as well as an Illumina sequencing platform by Macrogen, Inc. (Korea), were utilized. The obtained raw data were
Chemotaxonomic Characterization
Chemotaxonomic features, including cellular fatty acids, polyamines, isoprenoid quinones, and polar lipids, were investigated. For cellular fatty acid analysis, approximately 40 mg of fresh cells from the third quadrant of the streaked plates were collected. The cellular fatty acids were extracted as previously described [22] using a MIDI Sherlock Microbial Identification System (6.0). The samples were subjected to gas chromatography using a 6890N gas chromatography system (Agilent Technologies, USA), and the data were identified using the TSBA6 database of the Microbial Identification software package [31]. Polyamines were extracted from 0.1 g of freeze-dried cells and analyzed by comparison to spermine, putrescine, spermidine, and homospermidine standards in Ace Emzyme Inc. (Korea). Isoprenoid quinones were extracted using the method outlined by Collins
Phenotypic Characteristics
After culturing strain PAMB 00755T on R2A medium for 3 days, the morphological features of the strain were examined using scanning electron microscopy (SEM) at the Korean Basic Science Institute in Chuncheon. To examine motility, a semi-solid R2A medium supplemented with 0.4% agar was used. The Gram staining procedure was conducted using a Gram staining kit (Difco), following the manufacturer’s instructions. Catalase activity was assessed by observing bubbles production upon addition of a 3% (v/v) hydrogen peroxide solution to fresh cells. Oxidase activity was determined by observing the development of a purple color using an oxidase reagent kit. The strain was grown on various media, comprising R2A, nutrient agar (NA), potato dextrose agar (PDA), trypticase soy agar (TSA), marine agar 2216 (MA), and Luria-Bertani (LB) agar to determine the optimum medium. The strain was grown on PDA at 4, 10, 15, 20, 25, 30, 37, 40, 45, 50 and 60°C for 7 days to determine the optimal temperature. Salt tolerance was tested by growing the strain in R2A broth with concentrations from 0 to 15% (w/v) in increments of 1% [33], and pH tolerance was tested by growing the strain at a pH from 3.0 to 12.0 in increments of 1. Other biochemical features were tested using API 20NE (bioMérieux; substrate utilization, France) or API ZYM (bioMérieux; enzyme activities; NaCl 0.85% medium) and API 50CH (bioMérieux; production from carbohydrates) following the manufacturer’s instructions.
Results and Discussion
Phenotypic, Physiological, and Biochemical Characteristics
Strain PAMB 00755T exhibited optimum (and robust) growth on both PDA and R2A media, limited growth on NA, and no growth on TSA, LB, or MA. The cells are Gram-negative, non-motile, and short rods or ovoid (0.3–1.0 × 0.4–1.2 μm) without flagella (Fig. S1). Colonies were yellowish and ranged from 2–4 mm in diameter after being grown for 3 days on PDA medium. The strain grew at pH 4.0–9.0 (optimum pH, 6.0), 0–1% of NaCl (optimum NaCl concentration, 0%), and 4–25°C (optimum temperature, 20°C). It tested positive for catalase and oxidase activity. Other characteristics that distinguished strain PAMB 00755T from closely related strains are shown in Table 1.
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Table 1 . Distinguishing physiological and biochemical characteristics of strain PAMB 00755T and its closely related type strains.
Characteristic 1 2 3 Isolation source Korea fir Garden soila Oil port waterb Colony color Yellow Light yellow Yellowb Colony shape Short rods or ovoid Rodsa Short rodsb Oxidase Positive Weaka Negativeb Colony size (mm) 2–4 0.6–1.2a 2–3b Cell size (µm) 0.3–1.0 × 0.4–1.2 0.3–0.5 × 1.3–2.2a 1.04–1.90 × 0.52–0.75b Growth at: Temperature (°C), range (optimum) 4–25 (20) 15–35 (30)a 20–35 (26)b Optimum media PDA and R2A NA and R2Aa LBb NaCl tolerance (%, w/v), range (optimum) 0–1 (0) 0–0.5 (0)a 0.5–3.0 (0.5)b pH, range (optimum) 4.0–9.0 (6) 5.0–7.0 (6.5)a 4.0–7.5 (7)b Assimilation (API 20NE) of: β-galactosidase + - - Glucose - - + Arabinose - - + API 50CH Arbutin w - w Esculin ferric citrate w - w Salicin + + + D-cellobiose w - Maltose w - w D-lactose w - w D-melezitose w - w API ZYM Trypsin w + w β-galactosidase + - - N-acetyl-β-glucosaminidase + - + Strains: 1,
Sphingomonas abietis PAMB 00755T; 2,S. chungangi KACC 19292T; 3,S. polyaromaticivorans KCTC 82794T; “+”, positive, “w”, weakly positive, “−“, negative. Unless stated otherwise, all data are from our study. aAkter and Huq, 2020, bLuo,et al ., 2012.
Phylogenetic Analyses
The complete 16S rRNA gene amplicon (1443 nucleotides) of strain PAMB 00755T was obtained after assembly using Vector NTI software (1.6.1). Based on comparing the 16S rRNA sequence to the EzBioCloud and NCBI databases, strain PAMB 00755T is related to members of the genus
-
Fig. 1. Phylogenetic analysis of the 16S rRNA gene in strain PAMB 00755T and related species within the genus
Sphingomonas . The presented values indicate bootstrap values (>70%) obtained using the neighbor-joining (NJ), maximum likelihood (ML), and minimum parsimony (MP) algorithms. The scale bar represents 0.050 substitutions per nucleotide position.
Genomic and Phylogenomic Analysis
The complete genome of strain PAMB 00755T was sequenced, resulting in a complete circular chromosome of 4,429,509 bp after
-
Fig. 2. Phylogenomic tree of genus
Sphingomonas based on the up-to-date bacterial core gene set (UBCG) showing the position of strain PAMB 00755T. At the nodes, the gene support index (GSI) is presented on the left side, while bootstrap values are indicated on the right side. The scale bar corresponds to 0.20 substitutions per position.
Chemotaxonomic Features
Table 2 shows that C18:1
-
Table 2 . Comparisons of cellular fatty acid profiles between strain PAMB 00755T and closely related strains in the genus
Sphingomonas .1 2 3 C14:0 ND ND 0.8 C16:0 6.7 8.4 6.2 C17:0 1.1 ND ND C18:0 0.8 ND 0.5 C17:1 ω 6c 4.6 1.6 ND C18:1 ω 5c 1.9 ND 1.7 C19:0 cyclo ω8c ND ND 15.6 C12:0-2OH 1.3 ND 0.7 C13:0-2OH 0.7 ND ND C14:0-2OH 6.8 8.5 7.3 C15:0-2OH 0.6 ND ND Summed feature 3a 1.1 ND ND Summed feature 8a 72.1 81.5 67.2 Strains: 1,
Sphingomonas abietis PAMB 00755T; 2,S. chungangi KACC 19292T; 3,S. polyaromaticivorans KCTC 82794T; “ND” represents not detected. The major components (>10%) are highlighted in bold. All the data presented in this study were obtained from the present study. aSummed features refer to groups of two or three fatty acids that cannot be individually distinguished using gas chromatography with the MIDI System. Summed feature 3 includes C16:1ω 6c and/or C16:1ω 7c , while summed feature 8 includes C18:1ω 7c and/or C18:1ω 6c .
Collectively, the finding from phylogenetic, genomic, phenotypic, and chemotaxonomic analyses indicate that strain PAMB 00755T is a novel species within the genus
Description of Sphingomonas abietis
Strain PAMB 00755T is accessible from the Korea Collection for Type Culture (KCTC) with the designation KCTC 92781T and the Guangdong Microbial Culture Collection Center (GDMCC) with the designation GDMCC 1.3779T. The accession numbers for its 16S rRNA and whole-genome sequences are OP964609.1 and CP115174.1, respectively. Based on its distinct phenotypic, genotypic, and chemotaxonomic characteristics, strain PAMB 00755T was classified as a novel species within the genus
Data Availability
The 16S rRNA gene and whole-genome sequence of stain PAMB 00755T have been assigned GenBank accession numbers OP964609.1 and CP115174.1, respectively. This strain is accessible from the Korea Collection for Type Cultures (KCTC 92781T) and the Guangdong Microbial Culture Collection Center (GDMCC 1.3779T).
Author Contributions
JL: responsible for research design, project supervision, and manuscript revision. LJ: performed the experiments, analyzed the data, wrote the original manuscript draft, and revised the manuscript. HC: performed the experiments on the fatty acid profile. HC, YP, DJ, DC, YJ, and JHL: revised the manuscript. CYK: supervised the project. All authors have reviewed and approved the final manuscript.
Acknowledgments
We thank Dr. Gwang Joong Kim of Korea Basic Science Institute (KBI, Chuncheon Center) for kindly providing the FE-SEM image. Additionally, we would like to thank Dr. Aharon Oren from the Hebrew University of Jerusalem for his valuable comments and recommendations concerning nomenclature.
This work was funded by a grant from the Korea Research Institute of Bioscience and Biotechnology (KRIBB) Research Initiative (KGM5282331), and a grant from the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2020R111A2072308).
Conflict of Interest
The authors have no financial conflicts of interest to declare.
Abbreviations
KCTC Korean Collection for Type Cultures
KACC Korean Agricultural Culture Collection
GDMCC Guangdong Microbial Culture Collection
ANI average nucleotide identity
dDDH digital DNA-DNA hybridization
PGAP Prokaryotic Genome Annotation Pipeline
COG cluster of orthologous groups of proteins
GGDC genome-to-Genome distance calculator
DPG diphosphatidylglycerol
PE phosphatidylethanolamine
SGL sphingoglycolipid
PC phosphatidylcholine
PM phosphatidyl-N-methylethanolamine
APGL aminophosphoglycolipid
Supplemental Materials
References
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Related articles in JMB
Article
Research article
J. Microbiol. Biotechnol. 2023; 33(10): 1292-1298
Published online October 28, 2023 https://doi.org/10.4014/jmb.2303.03017
Copyright © The Korean Society for Microbiology and Biotechnology.
Sphingomonas abietis sp. nov., an Endophytic Bacterium Isolated from Korean Fir
Lingmin Jiang1,3, Hanna Choe1, Yuxin Peng1, Doeun Jeon1, Donghyun Cho1, Yue Jiang1, Ju Huck Lee1,2, Cha Young Kim1, and Jiyoung Lee1,2*
1Korean Collection for Type Cultures (KCTC), Biological Resource Center, Korea Research Institute of Bioscience and Biotechnology, Jeongeup 56212, Republic of Korea
2Department of Biosystem and Bioengineering, KRIBB School of Biotechnology, University of Science and Technology (UST), Daejeon 34113, Republic of Korea
3Present address: National Key Laboratory of Plant Molecular Genetics, CAS Center for Excellence in Molecular Plant Sciences, Chinese Academy of Sciences, Shanghai 200032, P.R. China
Correspondence to:Jiyoung Lee, jiyoung1@kribb.re.kr
Abstract
PAMB 00755T, a bacterial strain, was isolated from Korean fir leaves. The strain exhibits yellow colonies and consists of Gram-negative, non-motile, short rods or ovoid-shaped cells. It displays optimal growth conditions at 20°C, 0% NaCl, and pH 6.0. Results of 16S rRNA gene-based phylogenetic analyses showed that strain PAMB 00755T was most closely related to Sphingomonas chungangi MAH-6T (97.7%) and Sphingomonas polyaromaticivorans B2-7T (97.4%), and ≤96.5% sequence similarity to other members of the genus Sphingomonas. The values of average nucleotide identity (79.9–81.3%), average amino acid identity (73.3–75.9%), and digital DNA–DNA hybridization (73.3–75.9%) were significantly lower than the threshold values for species boundaries; these overall genome-related indexes (OGRI) analyses indicated that the strain represents a novel species. Genomic analysis revealed that the strain has a 4.4-Mbp genome encoding 4,083 functional genes, while the DNA G+C content of the whole genome is 66.1%. The genome of strain PAMB 00755T showed a putative carotenoid biosynthetic cluster responsible for its antioxidant activity. The respiratory quinone was identified as ubiquinone 10 (Q-10), while the major fatty acids in the profile were identified as C18:1ω7c and/or C18:1ω6c (summed feature 8). The major polar lipids of strain PAMB 00755T were diphosphatidylglycerol, phosphatidylethanolamine, sphingoglycolipid, and phosphatidylcholine. Based on a comprehensive analysis of genomic, phenotypic, and chemotaxonomic characteristics, we proposed the name Sphingomonas abietis sp. nov. for this novel species, with PAMB 00755T as the type strain (= KCTC 92781T = GDMCC 1.3779T).
Keywords: Polyphasic taxonomy, Korean fir, phylogeny, AntiSMASH, endophytic bacterium
Introduction
In this study, we isolated a new bacterial strain designated PAMB 00755T from the leaves of Korean fir (
Materials and Methods
Isolation and Ecology
Leaves were collected from the Korean fir growing on Hallasan Mountain (33°21'42''N, 126°31'45''E) on Jeju Island, South Korea. To prepare the sample, five grams of leaves were subjected to surface sterilization using 1%NaOCl solution. The leaves were rinsed five times with distilled water. The sterilized leaves were then homogenized using a blender with 10 ml of 1× phosphate-buffered saline (PBS). To remove any debris, the resulting mixture was filtered through four layers of sterile cheesecloth and then serially diluted using a standard dilution method with 1× PBS buffer. A volume of 100 μl from the sample solution was evenly spread onto Reasoner's 2A agar (R2A, Difco) plates, followed by incubation at 25°C for 1 week. Single colonies were then re-streaked onto fresh R2A medium. Strain PAMB 00755T, which formed circular, smooth, yellow colonies, was selected for the following study. The strain was stored at -80°C in 10% skimmed milk. It is currently accessible through the Korean Collection for Type Cultures (KCTC 92781T) and the Guangdong Microbial Culture Collection Center (GDMCC 1.3779T). Unless otherwise noted, bacterial cells were cultured on R2A for 3–4 days before performing subsequent tests.
16S rRNA Gene Sequence Analysis
The extraction of genomic DNA from the bacteria was performed to serve as a template for PCR amplification of the 16S rRNA gene. Universal bacterial primers, specifically 27F and 1492R, were used for the amplification process. The PCR products were sequenced by a commercial company (Macrogen Inc., Korea) using the primers 27F, 518F, 800R, and 1492R [24]. The almost full length of the 16S rRNA sequence was assembled using Vector NTI software (1.6.1). The obtained sequences were compared in the EzBioCloud and the NCBI database [25]. The alignment of multiple sequences was performed using BioEdit software. Subsequently, the construction of the phylogenetic tree was carried out using the Molecular Evolutionary Genetics Analysis (MEGA) software version 11.0. The neighbor-joining (NJ), maximum likelihood (ML), and minimum parsimony (MP) methods were used for this analysis, with 1,000 bootstrap iterations [26]. The outgroup was
Whole-Genome Characteristics
Genomic DNA was extracted as previously described [10]. For whole-genome sequencing, the PacBio RSII system (Pacific Biosciences, Inc., USA) as well as an Illumina sequencing platform by Macrogen, Inc. (Korea), were utilized. The obtained raw data were
Chemotaxonomic Characterization
Chemotaxonomic features, including cellular fatty acids, polyamines, isoprenoid quinones, and polar lipids, were investigated. For cellular fatty acid analysis, approximately 40 mg of fresh cells from the third quadrant of the streaked plates were collected. The cellular fatty acids were extracted as previously described [22] using a MIDI Sherlock Microbial Identification System (6.0). The samples were subjected to gas chromatography using a 6890N gas chromatography system (Agilent Technologies, USA), and the data were identified using the TSBA6 database of the Microbial Identification software package [31]. Polyamines were extracted from 0.1 g of freeze-dried cells and analyzed by comparison to spermine, putrescine, spermidine, and homospermidine standards in Ace Emzyme Inc. (Korea). Isoprenoid quinones were extracted using the method outlined by Collins
Phenotypic Characteristics
After culturing strain PAMB 00755T on R2A medium for 3 days, the morphological features of the strain were examined using scanning electron microscopy (SEM) at the Korean Basic Science Institute in Chuncheon. To examine motility, a semi-solid R2A medium supplemented with 0.4% agar was used. The Gram staining procedure was conducted using a Gram staining kit (Difco), following the manufacturer’s instructions. Catalase activity was assessed by observing bubbles production upon addition of a 3% (v/v) hydrogen peroxide solution to fresh cells. Oxidase activity was determined by observing the development of a purple color using an oxidase reagent kit. The strain was grown on various media, comprising R2A, nutrient agar (NA), potato dextrose agar (PDA), trypticase soy agar (TSA), marine agar 2216 (MA), and Luria-Bertani (LB) agar to determine the optimum medium. The strain was grown on PDA at 4, 10, 15, 20, 25, 30, 37, 40, 45, 50 and 60°C for 7 days to determine the optimal temperature. Salt tolerance was tested by growing the strain in R2A broth with concentrations from 0 to 15% (w/v) in increments of 1% [33], and pH tolerance was tested by growing the strain at a pH from 3.0 to 12.0 in increments of 1. Other biochemical features were tested using API 20NE (bioMérieux; substrate utilization, France) or API ZYM (bioMérieux; enzyme activities; NaCl 0.85% medium) and API 50CH (bioMérieux; production from carbohydrates) following the manufacturer’s instructions.
Results and Discussion
Phenotypic, Physiological, and Biochemical Characteristics
Strain PAMB 00755T exhibited optimum (and robust) growth on both PDA and R2A media, limited growth on NA, and no growth on TSA, LB, or MA. The cells are Gram-negative, non-motile, and short rods or ovoid (0.3–1.0 × 0.4–1.2 μm) without flagella (Fig. S1). Colonies were yellowish and ranged from 2–4 mm in diameter after being grown for 3 days on PDA medium. The strain grew at pH 4.0–9.0 (optimum pH, 6.0), 0–1% of NaCl (optimum NaCl concentration, 0%), and 4–25°C (optimum temperature, 20°C). It tested positive for catalase and oxidase activity. Other characteristics that distinguished strain PAMB 00755T from closely related strains are shown in Table 1.
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Table 1 . Distinguishing physiological and biochemical characteristics of strain PAMB 00755T and its closely related type strains..
Characteristic 1 2 3 Isolation source Korea fir Garden soila Oil port waterb Colony color Yellow Light yellow Yellowb Colony shape Short rods or ovoid Rodsa Short rodsb Oxidase Positive Weaka Negativeb Colony size (mm) 2–4 0.6–1.2a 2–3b Cell size (µm) 0.3–1.0 × 0.4–1.2 0.3–0.5 × 1.3–2.2a 1.04–1.90 × 0.52–0.75b Growth at: Temperature (°C), range (optimum) 4–25 (20) 15–35 (30)a 20–35 (26)b Optimum media PDA and R2A NA and R2Aa LBb NaCl tolerance (%, w/v), range (optimum) 0–1 (0) 0–0.5 (0)a 0.5–3.0 (0.5)b pH, range (optimum) 4.0–9.0 (6) 5.0–7.0 (6.5)a 4.0–7.5 (7)b Assimilation (API 20NE) of: β-galactosidase + - - Glucose - - + Arabinose - - + API 50CH Arbutin w - w Esculin ferric citrate w - w Salicin + + + D-cellobiose w - Maltose w - w D-lactose w - w D-melezitose w - w API ZYM Trypsin w + w β-galactosidase + - - N-acetyl-β-glucosaminidase + - + Strains: 1,
Sphingomonas abietis PAMB 00755T; 2,S. chungangi KACC 19292T; 3,S. polyaromaticivorans KCTC 82794T; “+”, positive, “w”, weakly positive, “−“, negative. Unless stated otherwise, all data are from our study. aAkter and Huq, 2020, bLuo,et al ., 2012..
Phylogenetic Analyses
The complete 16S rRNA gene amplicon (1443 nucleotides) of strain PAMB 00755T was obtained after assembly using Vector NTI software (1.6.1). Based on comparing the 16S rRNA sequence to the EzBioCloud and NCBI databases, strain PAMB 00755T is related to members of the genus
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Figure 1. Phylogenetic analysis of the 16S rRNA gene in strain PAMB 00755T and related species within the genus
Sphingomonas . The presented values indicate bootstrap values (>70%) obtained using the neighbor-joining (NJ), maximum likelihood (ML), and minimum parsimony (MP) algorithms. The scale bar represents 0.050 substitutions per nucleotide position.
Genomic and Phylogenomic Analysis
The complete genome of strain PAMB 00755T was sequenced, resulting in a complete circular chromosome of 4,429,509 bp after
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Figure 2. Phylogenomic tree of genus
Sphingomonas based on the up-to-date bacterial core gene set (UBCG) showing the position of strain PAMB 00755T. At the nodes, the gene support index (GSI) is presented on the left side, while bootstrap values are indicated on the right side. The scale bar corresponds to 0.20 substitutions per position.
Chemotaxonomic Features
Table 2 shows that C18:1
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Table 2 . Comparisons of cellular fatty acid profiles between strain PAMB 00755T and closely related strains in the genus
Sphingomonas ..1 2 3 C14:0 ND ND 0.8 C16:0 6.7 8.4 6.2 C17:0 1.1 ND ND C18:0 0.8 ND 0.5 C17:1 ω 6c 4.6 1.6 ND C18:1 ω 5c 1.9 ND 1.7 C19:0 cyclo ω8c ND ND 15.6 C12:0-2OH 1.3 ND 0.7 C13:0-2OH 0.7 ND ND C14:0-2OH 6.8 8.5 7.3 C15:0-2OH 0.6 ND ND Summed feature 3a 1.1 ND ND Summed feature 8a 72.1 81.5 67.2 Strains: 1,
Sphingomonas abietis PAMB 00755T; 2,S. chungangi KACC 19292T; 3,S. polyaromaticivorans KCTC 82794T; “ND” represents not detected. The major components (>10%) are highlighted in bold. All the data presented in this study were obtained from the present study. aSummed features refer to groups of two or three fatty acids that cannot be individually distinguished using gas chromatography with the MIDI System. Summed feature 3 includes C16:1ω 6c and/or C16:1ω 7c , while summed feature 8 includes C18:1ω 7c and/or C18:1ω 6c ..
Collectively, the finding from phylogenetic, genomic, phenotypic, and chemotaxonomic analyses indicate that strain PAMB 00755T is a novel species within the genus
Description of Sphingomonas abietis
Strain PAMB 00755T is accessible from the Korea Collection for Type Culture (KCTC) with the designation KCTC 92781T and the Guangdong Microbial Culture Collection Center (GDMCC) with the designation GDMCC 1.3779T. The accession numbers for its 16S rRNA and whole-genome sequences are OP964609.1 and CP115174.1, respectively. Based on its distinct phenotypic, genotypic, and chemotaxonomic characteristics, strain PAMB 00755T was classified as a novel species within the genus
Data Availability
The 16S rRNA gene and whole-genome sequence of stain PAMB 00755T have been assigned GenBank accession numbers OP964609.1 and CP115174.1, respectively. This strain is accessible from the Korea Collection for Type Cultures (KCTC 92781T) and the Guangdong Microbial Culture Collection Center (GDMCC 1.3779T).
Author Contributions
JL: responsible for research design, project supervision, and manuscript revision. LJ: performed the experiments, analyzed the data, wrote the original manuscript draft, and revised the manuscript. HC: performed the experiments on the fatty acid profile. HC, YP, DJ, DC, YJ, and JHL: revised the manuscript. CYK: supervised the project. All authors have reviewed and approved the final manuscript.
Acknowledgments
We thank Dr. Gwang Joong Kim of Korea Basic Science Institute (KBI, Chuncheon Center) for kindly providing the FE-SEM image. Additionally, we would like to thank Dr. Aharon Oren from the Hebrew University of Jerusalem for his valuable comments and recommendations concerning nomenclature.
This work was funded by a grant from the Korea Research Institute of Bioscience and Biotechnology (KRIBB) Research Initiative (KGM5282331), and a grant from the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2020R111A2072308).
Conflict of Interest
The authors have no financial conflicts of interest to declare.
Abbreviations
KCTC Korean Collection for Type Cultures
KACC Korean Agricultural Culture Collection
GDMCC Guangdong Microbial Culture Collection
ANI average nucleotide identity
dDDH digital DNA-DNA hybridization
PGAP Prokaryotic Genome Annotation Pipeline
COG cluster of orthologous groups of proteins
GGDC genome-to-Genome distance calculator
DPG diphosphatidylglycerol
PE phosphatidylethanolamine
SGL sphingoglycolipid
PC phosphatidylcholine
PM phosphatidyl-N-methylethanolamine
APGL aminophosphoglycolipid
Supplemental Materials
Fig 1.
Fig 2.
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Table 1 . Distinguishing physiological and biochemical characteristics of strain PAMB 00755T and its closely related type strains..
Characteristic 1 2 3 Isolation source Korea fir Garden soila Oil port waterb Colony color Yellow Light yellow Yellowb Colony shape Short rods or ovoid Rodsa Short rodsb Oxidase Positive Weaka Negativeb Colony size (mm) 2–4 0.6–1.2a 2–3b Cell size (µm) 0.3–1.0 × 0.4–1.2 0.3–0.5 × 1.3–2.2a 1.04–1.90 × 0.52–0.75b Growth at: Temperature (°C), range (optimum) 4–25 (20) 15–35 (30)a 20–35 (26)b Optimum media PDA and R2A NA and R2Aa LBb NaCl tolerance (%, w/v), range (optimum) 0–1 (0) 0–0.5 (0)a 0.5–3.0 (0.5)b pH, range (optimum) 4.0–9.0 (6) 5.0–7.0 (6.5)a 4.0–7.5 (7)b Assimilation (API 20NE) of: β-galactosidase + - - Glucose - - + Arabinose - - + API 50CH Arbutin w - w Esculin ferric citrate w - w Salicin + + + D-cellobiose w - Maltose w - w D-lactose w - w D-melezitose w - w API ZYM Trypsin w + w β-galactosidase + - - N-acetyl-β-glucosaminidase + - + Strains: 1,
Sphingomonas abietis PAMB 00755T; 2,S. chungangi KACC 19292T; 3,S. polyaromaticivorans KCTC 82794T; “+”, positive, “w”, weakly positive, “−“, negative. Unless stated otherwise, all data are from our study. aAkter and Huq, 2020, bLuo,et al ., 2012..
-
Table 2 . Comparisons of cellular fatty acid profiles between strain PAMB 00755T and closely related strains in the genus
Sphingomonas ..1 2 3 C14:0 ND ND 0.8 C16:0 6.7 8.4 6.2 C17:0 1.1 ND ND C18:0 0.8 ND 0.5 C17:1 ω 6c 4.6 1.6 ND C18:1 ω 5c 1.9 ND 1.7 C19:0 cyclo ω8c ND ND 15.6 C12:0-2OH 1.3 ND 0.7 C13:0-2OH 0.7 ND ND C14:0-2OH 6.8 8.5 7.3 C15:0-2OH 0.6 ND ND Summed feature 3a 1.1 ND ND Summed feature 8a 72.1 81.5 67.2 Strains: 1,
Sphingomonas abietis PAMB 00755T; 2,S. chungangi KACC 19292T; 3,S. polyaromaticivorans KCTC 82794T; “ND” represents not detected. The major components (>10%) are highlighted in bold. All the data presented in this study were obtained from the present study. aSummed features refer to groups of two or three fatty acids that cannot be individually distinguished using gas chromatography with the MIDI System. Summed feature 3 includes C16:1ω 6c and/or C16:1ω 7c , while summed feature 8 includes C18:1ω 7c and/or C18:1ω 6c ..
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