Journal of Species Research 6(2):91-100, 2017

A report of 9 unrecorded radiation resistant bacterial species in Korea

Myung-Suk Kang1 and Sathiyaraj Srinivasan2,*

1Microorganism Resources Division, National Institute of Biological Resources, Incheon 22689, Republic of Korea 2Department of Bio & Environmental Technology, College of Natural Science, Seoul Women’s University, Seoul 01797, Republic of Korea

*Correspondent: [email protected]

Five bacterial strains, ES10-3-3-1, KKM10-2-2-1, Ant11, JM10-4-1-3, and KMS4-11 assigned to the genus were isolated from soil samples collected from Namyangju-si in Gyeonggi-do, Gangnam-gu and Dongdaemun-gu in Seoul, Korea. In addition, four bacterial strains, KKM10-2-7-2, JM10-2-5, JM10- 2-6-2, and KKM10-2-3 assigned to the genus Hymenobacter were isolated from soil samples collected from Gangnam-gu and Dongdaemun-gu in Seoul, in South Korea. The five Deinococcus species were Gram-stain positive, pink-pigmented, and short-rod or coccus shaped. The four Hymenobacter species were Gram-stain negative, red-pigmented, and short-rod shaped. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strains ES10-3-3-1, KKM10-2-2-1, Ant11, JM10-4-1-3, and KMS4-11 were most closely related to Deinococcus citri NCCP-154T (with 99.8% similarity), Deinococcus grandis DSM 12784T (99.0%), Deinococcus marmoris DSM 12784T (98.8%), Deinococcus claudionis PO-04- 19-125T (98.7%), and Deinococcus radioresistens 8AT (99.8%), respectively. KKM10-2-7-2, JM10-2-5, JM10-2-6-2, and KKM10-2-3 were most closely related to Hymenobacter algoricola VUG-A23aT (99.1% similarity), Hymenobacter elongatus VUG-A112T (99.1% similarity), Hymenobacter gelipurpurascens Txg1T (99.1% similarity), and Hymenobacter psychrotolerans Tibet-IIU11T (99.3% similarity), respectively. These nine species have never been reported in Korea; thus, five Deinococcus species are reported in the family Deinococcaceae, order Deinococcales, class Deinococci, phylum Deinococcus-Thermus and four Hymenobacter species are reported in the family Cytophagaceae, order Cytophagales, class Cytophagia, phylum Bacteroidetes. Keywords: 16S rRNA gene, Bacteroidetes, Deinococcus, Deinococcaceae, Deinococcus-Thermus, Hymenobacter, unrecorded species in Korea

Ⓒ 2017 National Institute of Biological Resources DOI:10.12651/JSR.2017.6.2.091

Introduction water (http://www.bacterio.cict.fr/d/deinococcus.html). Members of the genus Deinococcus are Gram-positive In 2015, we collected diverse soil samples and isolat- (Brooks and Murray, 1981; Srinivasan et al., 2012a, ed unrecorded bacterial species in Korea. The identified 2012b), have red cell colors, and have L-ornithine as the bacterial species belonged to the phyla Deinococcus- di-amino acid in the cell-wall peptidoglycan. Thermus and Bacteroidetes. This report focuses on the iso­ The genus Hymenobacter was first proposed by Hirsch lation and description of unrecorded radiation-resistant (1998) and the type species Hymenobacter roseosalivarius species in the the genera Deinococcus and Hymenobacter. was isolated from continental Antarctic soils and sand- The genus Deinococcus was proposed by Brooks stone (Hirsch et al., 1998). Currently, there are 36 spe- and Murray (1981) and the type species Deinococcus cies of the genus Hymenobacter with validly published radiodurans was isolated from gamma-ray-irradiated names (http://www.bacterio.net/hymenobacter.html#r). food. Currently, the genus Deinococcus comprises 57 Members of the genus Hymenobacter are pink or red, species isolated from diverse environments such as air, rod-shaped, and Gram-negative. The Hymenobacter spe- hot springs, continental Antarctica, desert soil, fish, and cies inhabit various environments, such as soil, glaciers, 92 JOURNAL OF SPECIES RESEARCH Vol. 6, No. 2 marine and freshwater aquatic systems and air (Han et (Kim et al., 2012) and edited using the BioEdit program al., 2014). (Hall, 1999). Multiple alignments were performed with the MUSCLE program (Edgar, 2004). Evolutionary dis- tances were calculated using the two-parameter model Materials and Methods (Kimura, 1983). Phylogenetic trees were constructed us- ing the neighbor-joining method (Saitou and Nei, 1987) Various soil samples were suspended on distilled water in the MEGA5 program (Tamura, 2011) with bootstrap and serially diluted. The aliquot was inoculated onto values based on 1,000 replications (Felsenstein, 1985). R2A agar and incubated at 25°C for 3 days (Table 1). The designated strain IDs, sources, culture media, and incubation conditions are summarized in Table 1. All Results and Discussion strains were purified as single colonies and stored in 20% glycerol suspension at -80°C as well as lyophilized Based on the comparative 16S rRNA gene sequence ampoules. analyses and phylogenies, nine strains, designated ES10- Colony morphology and cell size of the strains were 3-3-1, KKM10-2-2-1, Ant11, JM10-4-1-3, KMS4-11, observed on R2A agar after cells were grown for 3 days KKM10-2-7-2, JM10-2-5, JM10-2-6-2, and KKM10- at 25°C by using transmission electron microscopy 2-3 were assigned to the species level. Morphology and (LIBRA 120, Carl Zeiss). Transmission electron micro- physiological characteristics are shown in the species graphs of the strains are shown in Fig. 1. Gram reaction description section. was performed according to the classic Gram procedure Strains ES10-3-3-1, KKM10-2-2-1, Ant11, JM10- described by Doetsch (1981). Biochemical characteris- 4-1-3, and KMS4-11 were most closely related to tics were tested by using Biolog Microstation with GEN Deinococcus citri NCCP-154T (AB558498; 99.85% 16S III microplate system. A single colony was selected and rRNA gene sequence similarity), Deinococcus grandis emulsified into ‘inoculating fluid A’ (Biolog) for subse- DSM 12784T (Y11329; 99.00% similarity), Deinococcus quent inoculation on to the MicroPlate test plate (Bio- marmoris DSM 12784T (JNIV01000230; 98.81% log). More fastidious organisms, including capnophilic similarity), Deinococcus claudionis PO-04-19-125T strains, were cultured on alternative media, according to (EF635406; 98.80% similarity), and Deinococcus the manufacturer’s instructions, and inocula prepared to radioresistens 8AT (KJ123751; 99.82% similarity), re- a specified transmittance using a turbidimeter, as spec- spectively (Table 1). ified in the user’s guide. For each isolate, 100 μL of the Strains KKM10-2-7-2, JM10-2-5, JM10-2-6-2, and cell suspension was inoculated into each well of the KKM10-2-3 were most closely related to Hymenobacter MicroPlate,­ using a multichannel pipette and incubated algoricola VUG-A23aT (EU155009; 99.18% 16S rRNA at 37°C for 24 h, according to growth characteristics. gene sequence similarity), Hymenobacter elongatus VUG- MicroPlates were read in a MicroStation semi-automated A112T (GQ454797; 99.10% similarity), Hymenobacter reader after 24 h and results interpreted by the identifica- gelipurpurascens Txg1T (Y18836; 99.13% similari- tion system’s software (GEN III database, version 5.2.1). ty), and Hymenobacter psychrotolerans Tibet-IIU11T The system indicated which isolates could not be identi- (DQ177475; 99.37% similarity), respectively (Table 2). fied after 20 h and required further incubation. Such iso- As expected from the high 16S rRNA gene sequence lates were re-incubated and re-read 3 to 6 h later. (Wragg similarities of the nine strains with their closest relatives, et al., 2014) Genomic DNA was extracted and 16S each strain formed a robust phylogenetic clade with the rRNA gene was amplified by PCR with 9F and 1492R most closely related species (Fig. 2). From the high 16S universal bacterial primers (Weisburg et al., 1991). The rRNA gene sequence similarity and robust formation of 16SrRNA gene sequences of the related taxa were ob- phylogenetic clades, it is concluded that strains ES10-3- tained from EzTaxon-e (http://eztaxon-e.ezbiocloud.net) 3-1, KKM10-2-2-1, Ant11, JM10-4-1-3, and KMS4-11

Table 1. 16S rRNA gene sequence similarity, isolation source, medium, and incubation conditions of unrecorded strains belonging to the genus Deinococcus. Strain ID Most closely related species Similarity (%) Isolation source Medium Incubation conditions ES10-3-3-1 Deinococcus citri 99.3 soil of Namyangju-si R2A 25°C, 3 d KKM10-2-2-1 Deinococcus grandis 99.0 soil of Gangnam-gu R2A 25°C, 3 d Ant11 Deinococcus marmoris 99.1 soil of Dongdaemun-gu R2A 25°C, 3 d JM10-4-1-3 Deinococcus claudionis 98.8 soil of Dongdaemun-gu R2A 25°C, 3 d KMS4-11 Deinococcus radioresistens 99.8 soil of Gangnam-gu R2A 25°C, 3 d June 2017 KANG AND SRINIVASAN-UNRECORDED RADIATION RESISTANT 93

(1) (2) (3)

(4) (5) (6)

(7) (8) (9)

Fig. 1. Transmission electron micrographs of the strains isolated in this study. Bar: 0.5 μm substitutions per cell size. Strains: 1, ES10-3-3- 10; 2, KKM10-2-2-1; 3, Ant11; 4, JM10-4-1-3; 5, KMS4-11; 6, KKM10-2-7-2; 7, JM10-2-5; 8, JM10-2-6-2; 9, KKM10-2-3.

Table 2. 16S rRNA gene sequence similarity, isolation source, medium, and incubation conditions of unrecorded strains belonging to the genus Hymenobacter. Strain ID Most closely related species Similarity (%) Isolation source Medium Incubation conditions KKM10-2-7-2 Hymenobacter algoricola 99.1 soil of Gangnam-gu R2A 25°C, 3 d JM10-2-5 Hymenobacter elongatus 99.2 soil of Dongdaemun-gu R2A 25°C, 3 d JM10-2-6-2 Hymenobacter gelipurpurascens 99.3 soil of Dongdaemun-gu R2A 25°C, 3 d KKM10-2-3 Hymenobacter psychrotolerans 98.8 soil of Gangnam-gu R2A 25°C, 3 d are members of the species Deinococcus citri (Ahmed 3 are members of the species Hymenobacter algoricola et al., 2014), Deinococcus grandis (Oyaizu et al., 1987), (Klassen and Foght, 2011), Hymenobacter elongatus Deinococcus marmoris (Hirsch et al., 2004), Deinococcus (Klassen and Foght, 2011), Hymenobacter gelipurpurascens claudionis (Callegan et al., 2008), and Deinococcus (Buczolits et al., 2006), and Hymenobacter psychrotolerans radioresistens (Srinivasan et al., 2015), respectively. (Zhang et al., 2008), respectively. KKM10-2-7-2, JM10-2-5, JM10-2-6-2, and KKM10-2- There are no reports of these nine species in the gen- 94 JOURNAL OF SPECIES RESEARCH Vol. 6, No. 2

(a)

Fig. 2. Neighbor-joining phylogenetic tree based on 16S rRNA gene sequences shows the relationship between the strains isolated in this study and their relatives of the genus Deinococcus (a) and Hymenobacter (b). Bootstrap values (>70%) are shown above nodes for the neighbor-joining methods. Bar: 0.02 substitutions per nucleotide position. June 2017 KANG AND SRINIVASAN-UNRECORDED RADIATION RESISTANT BACTERIA 95

(b)

Fig. 2. Continued. era Deinococcus or Hymenobacter having been isolated marmoris, Deinococcus claudionis, Deinococcus in Korea. The strains ES10-3-3-1, KKM10-2-2-1 Ant11, radioresistens, Hymenobacter algoricola, Hymenobacter JM10-4-1-3, KMS4-11 KKM10-2-7-2, JM10-2-5, elongatus, Hymenobacter gelipurpurascens, and JM10-2-6-2, and KKM10-2-3 are unreported strains of Hymenobacter psychrotolerans for. Deinococcus citri, Deinococcus grandis, Deinococcus 96 JOURNAL OF SPECIES RESEARCH Vol. 6, No. 2

Description of Deinococcus citri ES10-3-3-1 stachyose, and sucrose are utilized as sole carbon source. But, acetoacetic acid, N-acetyl-D-galactosamine, N-ace- Cell is Gram-stain-positive, non-flagellated, and coc- tyl-D-mannosamine, N-acetyl-neuraminic acid, N-ace- cus-shaped. Colonies are pale pinkish-colored after 3 tyl-β-D-glucosamine, bromo-succinic acid, citric acid, days of incubation on R2A at 25°C. In the GN3 micro- dextrin, formic acid, L-fucose, gentiobiose, α-hydroxy- plates, L-, D-fructose, D-fructose-6-phosphate, butric acid, β-hydroxy-D,L-butyric acid, p-hydroxy- D-fucose, L-fucose, L-galactonic acid lactone, D-galac­ phenylacetic acid, inosine, α-keto-butyric acid, L-lactic turonic acid, D-gluconic acid, α-D-glucose, glucuron­ acid, α-D-lactose, D-malic acid, β-methyl-D-glucoside, amide, D-glucuronic acid, glycerol, D-mannitol, D- methyl pyruvate, mucic acid, myo-inositol, propionic mannose, D-melibiose, 3-methyl glucose, pectin, glycyl- acid, D-raffinose, D-saccharic acid, D-salicin, D-treha- L-protine, D-, D-sorbitol, sucrose, and D-turanose lose, D-turanose, and Tween 40 are not utilized. are utilized as sole carbon source. But acetic acid, aceto- In sensitivity tests, the tetrazolium redox dye is re- acetic acid, N-acetyl-D-galactosamine, N-acetyl-D-man- duced at pH 6 but not at pH 5; the dye is reduced in the nosamine, N-acetyl-neuraminic acid, N-acetyl-β-D-glu- presence of minocycline, tetrazolium blue, and tetra- cosamine, γ-amino-butryric acid, D-arabitol, L-arginine, zolium violet, but not in the presence of 1% NaCl, 1% D-aspartic acid, L-aspartic acid, bromo-succinic acid, sodium lactate, 4% NaCl, 8% NaCl, aztreonam, fusidic D-cellobiose, citric acid, dextrin, formic acid, D-galac­ acid, guanidine HCl, lincomycin, lithium chloride, mi- tose, gelatin, gentiobiose, D-glucose-6-phosphate, L- nocycline, nalidixic acid, niaproof 4, potassium tellu- , L-, α-hydroxybutric acid, β-hy- rite, D-Serine, sodium bromate, sodium butyrate, tro- droxy-D,L-butyric acid, p-hydroxy-phenylacetic acid, leandomycin, and vancomycin. Strain KKM10-2-2-1 inosine, α-keto-butyric acid, α-keto-glutaric acid, L-lac- (=NIBRBAC000003991) has been isolated from a soil tic acid, D-lactic acid methyl ester, α-D-lactose, D-malic sample, Gangnam-gu in Seoul, South Korea. acid, L-malic acid, D-maltose, β-methyl-D-glucoside, methyl pyruvate, mucic acid, myo-inositol, propionic acid, L-pyroglutamic acid, quinic acid, D-raffinose, L- Description of Deinococcus marmoris Ant11 rhamnose, D-saccharic acid, D-salicin, L-serine, stachy- Cells are Gram-stain-positive, and short rod or coc- ose, D-trehalose, and Tween 40 are not utilized. ccus-shaped. Colonies are circular and pale red-colored In sensitivity tests, the tetrazolium redox dye is re- after 3 days of incubation on R2A at 25°C. In the GN3 duced at pH 6 but not at pH 5; the dye is reduced in the microplates, acetoacetic acid, D-fructose-6-phosphate, presence of 1% NaCl, aztreonam, nalidixic acid, potas- D-galactose, glucuronamide, D-glucuronic acid, and sium tellurite, rifamycin SV, D-serine, tetrazolium blue, L-histidine are utilized as sole carbon source. But acetic and tetrazolium violet; but not in the presence of 1% acid, N-acetyl-D-galactosamine, N-acetyl-D-mannos­ sodium lactate, 4% NaCl, 8% NaCl, fusidic acid, guan- amine, N-acetyl-neuraminic acid, N-acetyl-β-D-glucos- idine HCl, lincomycin, lithium chloride, minocycline, amine, L-alanine, γ-amino-butryric acid, D-arabitol, L- niaproof 4, sodium bromate, sodium butyrate, troleando­ arginine, D-aspartic acid, L-aspartic acid, bromo-succinic mycin, and vancomycin. Strain ES10-3-3-1 (=NIBRBAC acid, D-cellobiose, citric acid, dextrin, formic acid, D- 000003990) was isolated from a soil sample, Namyangju- fructose, D-fucose, L-fucose, L-galactonic acid lactone, si in Gyeonggi-do, Korea. D-galacturonic acid, gelatin, gentiobiose, D-gluconic acid, α-D-glucose, D-glucose-6-phosphate, L-glutamic Description of Deinococcus grandis KKM10-2-2-1 acid, glycerol, α-hydroxybutric acid, β-hydroxy-D, L- Cells are Gram-stain-positive, non-flagellated, and butyric acid, p-hydroxy-phenylacetic acid, inosine, α- short rod-shaped. Colonies are pinkish-colored after 3 keto-butyric acid, α-keto-glutaric acid, L-lactic acid, D- days of incubation on R2A at 25°C. In the GN3 micro- lactic acid methyl ester, α-D-lactose, D-malic acid, L- plates, acetic acid, L-alanine, γ-amino-butryric acid, malic acid, D-maltose, D-mannitol, D-mannose, D-mel- D-arabitol, L-arginine, D-aspartic acid, L-aspartic acid, ibiose, β-methyl-D-glucoside, 3-methyl glucose, methyl D-cellobiose, D-fructose, D-fructose-6-phosphate, D-fu- pyruvate, mucic acid, myo-inositol, pectin, propionic cose, L-galactonic acid lactone, D-galactose, D-galac- acid, Glycyl-L-protine, L-pyroglutamic acid, quinic turonic acid, gelatin, D-gluconic acid, α-D-glucose, D- acid, D-raffinose, L-rhamnose, D-saccharic acid, D-sal- glucose-6-phosphate, glucuronamide, D-glucuronic acid, icin, D-serine, L-serine, D-sorbitol, stachyose, sucrose, L-glutamic acid, glycerol, L-histidine, α-keto-glutaric D-trehalose, D-turanose, and Tween 40 are not utilized. acid, D-lactic acid methyl ester, L-malic acid, D-malt- In sensitivity tests, the tetrazolium redox dye is re- ose, D-mannitol, D-mannose, D-melibiose, 3-methyl duced at pH 5 and pH 6; the dye is reduced in the pres- glucose, pectin, glycyl-L-protine, L-pyroglutamic acid, ence of minocycline, tetrazolium blue, and tetrazolium quinic acid, L-rhamnose, D-serine, L-serine, D-sorbitol, violet, but not in the presence of 1% NaCl, 1% sodium June 2017 KANG AND SRINIVASAN-UNRECORDED RADIATION RESISTANT BACTERIA 97 lactate, 4% NaCl, 8% NaCl, aztreonam, fusidic acid, acetic acid, acetoacetic acid, N-acetyl-D-galactosamine, guanidine HCl, lincomycin, lithium chloride, nalidixic N-acetyl-D-mannosamine, N-acetyl-neuraminic acid, acid, niaproof 4, potassium tellurite, rifamycin SV, D- N-acetyl-β-D-glucosamine, L-alanine, γ-amino-butryric Serine, sodium bromate, sodium butyrate, troleando­ acid, D-arabitol, L-arginine, D-aspartic acid, L-aspar- mycin, and vancomycin. Strain Ant11 ( =NIBRBAC tic acid, bromo-succinic acid, D-cellobiose, citric acid, 000003985) has been isolated from a soil sample, Dong- dextrin, formic acid, D-fructose, D-fucose, L-fucose, daemun-gu in Seoul, South Korea. L-galactonic acid lactone, D-galactose, D-galacturonic acid, gelatin, gentiobiose, D-gluconic acid, α-D-glucose, Description of Deinococcus claudionis JM10-4-1-3 D-glucose-6-phosphate, D-glucuronic acid, L-glutamic acid, glycerol, L-histidine, α-hydroxybutric acid, β-hy- Cells are Gram-stain-positive, non-flagellated, and droxy-D, L-butyric acid, p-hydroxy-phenylacetic acid, short rod-shaped. Colonies are pale pinkish-colored after inosine, α-keto-butyric acid, α-keto-glutaric acid, L-lac- 3 days of incubation on R2A at 25°C. In the GN3 micro­ tic acid, D-lactic acid methyl ester, α-D-lactose, D-malic plates, L-alanine, D-fructose, D-fructose-6-phosphate, acid, L-malic acid, D-maltose, D-mannitol, D-mannose, D-fucose, L-fucose, L-galactonic acid lactone, D-galac­ D-melibiose, β-methyl-D-glucoside, 3-methyl glucose, turonic acid, D-gluconic acid, α-D-glucose, glucuron­ methyl pyruvate, mucic acid, myo-inositol, pectin, pro- amide, D-glucuronic acid, glycerol, D-mannitol, D- pionic acid, Glycyl-L-protine, L-pyroglutamic acid, mannose, D-melibiose, 3-methyl glucose, pectin, glycyl- quinic acid, D-raffinose, L-rhamnose, D-saccharic acid, L-protine, D-serine, D-sorbitol, sucrose, and D-turano­ D-salicin, D-serine, L-serine, D-sorbitol, stachyose, su- seare utilized as sole carbon source. But acetic acid, ac- crose, D-trehalose, D-turanose, and Tween 40 are not etoacetic acid, N-acetyl-D-galactosamine, N-acetyl-D- utilized. mannosamine, N-acetyl-neuraminic acid, N-acetyl-β-D- In sensitivity tests, the tetrazolium redox dye is not re- glucosamine, γ-amino-butryric acid, D-arabitol, L-ar- duced at pH 5 and pH 6; the dye is reduced in the pres- ginine, D-aspartic acid, L-aspartic acid, bromo-succinic ence of minocycline, sodium butyrate, tetrazolium blue, acid, D-cellobiose, citric acid, dextrin, formic acid, D-ga- and tetrazolium violet; but not reduced in the presence of lactose, gelatin, gentiobiose, D-glucose-6-phosphate, 1% NaCl, 1% sodium lactate, 4% NaCl, 8% NaCl, aztre- L-glutamic acid, L-histidine, α-hydroxybutric acid, β- onam, fusidic acid, guanidine HCl, lincomycin, lithium hydroxy-D, L-butyric acid, p-hydroxy-phenylacetic chloride, nalidixic acid, niaproof 4, potassium tellurite, acid, inosine, α-keto-butyric acid, α-keto-glutaric acid, rifamycin SV, D-serine, sodium bromate, troleandomy- L-lactic acid, D-lactic acid methyl ester, α-D-lactose, cin, and vancomycin. Strain KMS4-11 ( =NIBRBAC D-malic acid, L-malic acid, D-maltose, β-methyl-D- 000003983) has been isolated from a soil sample, Gang­ glucoside, methyl pyruvate, mucic acid, myo-inositol, nam-gu in Seoul, South Korea. propionic acid, L-pyroglutamic acid, quinic acid, D- raffinose, L-rhamnose, D-saccharic acid, D-salicin, L- serine, stachyose, D-trehalose, and Tween 40 are not uti- Description of Hymenobacter algoricola KKM10-2-7-2 lized. Cells are Gram-stain-negative, non-flagellated, and In sensitivity tests, the tetrazolium redox dye is re- short rod-shaped. Colonies are circularand pale pinkish- duced at pH 6 but not at pH 5; the dye is reduced at 1% colored after 3 days of incubation on R2A at 25°C. In NaCl, aztreonam, nalidixic acid, potassium tellurite, ri- the GN3 microplates, acetoacetic acid, D-fructose, D- famycin SV, D-serine, tetrazolium blue, and tetrazolium fructose-6-phosphate, L-fucose, L-galactonic acid lac- violet; but not in the presence of 1% sodium lactate, 4% tone, D-galactose, glucuronamide, D-glucuronic acid, NaCl, 8% NaCl, fusidic acid, guanidine HCl, lincomy- L-histidine, and D-raffinose are utilized as sole carbon cin, lithium chloride, minocycline, niaproof 4, sodium source. But acetic acid, N-acetyl-D-galactosamine, N- bromate, sodium butyrate, troleandomycin, and vanco- acetyl-D-mannosamine, N-acetyl-neuraminic acid, N- mycin. Strain JM10-4-1-3 ( =NIBRBAC000003992) acetyl-β-D-glucosamine, L-alanine, γ-amino-butryric has been isolated from a soil sample, Dongdaemun-gu acid, D-arabitol, L-arginine, D-aspartic acid, L-aspartic in Seoul, South Korea. acid, bromo-succinic acid, D-cellobiose, citric acid, dex- trin, formic acid, D-fucose, D-galacturonic acid, gelatin, Description of Deinococcus radioresistens KMS4-11 gentiobiose, D-gluconic acid, α-D-glucose, D-glucose- Cells are Gram-stain-positive, non-flagellated, and 6-phosphate, L-glutamic acid, glycerol, α-hydroxybutric short rod or coccus-shaped. Colonies are circular and acid, β-hydroxy-D, L-butyric acid, p-hydroxy-phenyl­ pink-colored after 3 days of incubation on R2A at 25°C. acetic acid, inosine, α-keto-butyric acid, α-keto-glutaric In the GN3 microplates, D-fructose-6-phosphate, and acid, L-lactic acid, D-lactic acid methyl ester, α-D-lac- glucuronamide are utilized as sole carbon source. But tose, D-malic acid, L-malic acid, D-maltose, D-manni- 98 JOURNAL OF SPECIES RESEARCH Vol. 6, No. 2 tol, D-mannose, D-melibiose, β-methyl-D-glucoside, 4, rifamycin SV, sodium bromate, troleandomycin, and 3-methyl glucose, methyl pyruvate, mucic acid, myo- vancomycin. Strain JM10-2-5 (=NIBRBAC000003987) inositol, pectin, propionic acid, Glycyl-L-protine, L- has been isolated from a soil sample, Dongdaemun-gu pyroglutamic acid, quinic acid, L-rhamnose, D-saccharic in Seoul, South Korea. acid, D-salicin, D-serine, L-serine, D-sorbitol, stachyose, sucrose, D-trehalose, D-turanose, and Tween 40 are not Description of Hymenobacter gelipurpurascens utilized. JM10-2-6-2 In sensitivity tests, the tetrazolium redox dye is not re- Cells are Gram-stain-negative, non-flagellated, and duced at pH 5, pH 6; the dye is reduced in the presence short rod-shaped. Colonies are circular and pale red-col- of minocycline, potassium tellurite, tetrazolium blue, and ored after 3 days of incubation on R2A at 25°C. In the tetrazolium violet; but not reduced in the presence of 1% GN3 microplates, N-acetyl-D-galactosamine, N-acetyl- NaCl, 1% sodium lactate, 4% NaCl, 8% NaCl, aztreonam, D-mannosamine, γ-amino-butryric acid, D-arabitol, fusidic acid, guanidine HCl, lincomycin, lithium chloride, bromo-succinic acid, D-fructose-6-phosphate, D-fucose, nalidixic acid, niaproof 4, rifamycin SV, D-serine, sodium L-fucose, L-galactonic acid lactone, D-galactose, gela- bromate, sodium butyrate, troleandomycin, and vanco- tin, gentiobiose, α-D-glucose, glucuronamide, D-glucu- mycin. Strain KKM10-2-7-2 (=NIBRBAC000003986) ronic acid, glycerol, L-histidine, β-hydroxy-D, L-butyric has been isolated from a soil sample, Gangnam-gu in acid, α-D-lactose, D-maltose, D-mannose, D-melibiose, Seoul, South Korea. β-methyl-D-glucoside, methyl pyruvate, myo-inositol, Glycyl-L-protine, D-salicin, sucrose, D-trehalose, and Description of Hymenobacter elongatus JM10-2-5 D-turanose are utilized as sole carbon source. But acetic acid, acetoacetic acid, N-acetyl-neuraminic acid, N-ace- Cells are Gram-stain-negative, non-flagellated, and tyl-β-D-glucosamine, L-alanine, L-arginine, D-aspartic short rod-shaped. Colonies are circular and pale pinkish- acid, L-aspartic acid, D-cellobiose, citric acid, dextrin, colored after 3 days of incubation on R2A at 25°C. In formic acid, D-fructose, D-galacturonic acid, D-glucon- the GN3 microplates, acetoacetic acid, D-fructose-6- ic acid, D-glucose-6-phosphate, L-glutamic acid, α-hy- phosphate, glucuronamide, D-glucuronic acid, L-histidine, droxybutric acid, p-hydroxy-phenylacetic acid, inosine, and L-malic acid are utilized as sole carbon source. But α-keto-butyric acid, α-keto-glutaric acid, L-lactic acid, acetic acid, N-acetyl-D-galactosamine, N-acetyl-D-man- D-lactic acid methyl ester, D-malic acid, L-malic acid, nosamine, N-acetyl-neuraminic acid, N-acetyl-β-D-glu- D-mannitol, 3-methyl glucose, mucic acid, pectin, propi- cosamine, L-alanine, γ-amino-butryric acid, D-arabitol, onic acid, L-pyroglutamic acid, quinic acid, D-raffinose, L-arginine, D-aspartic acid, L-aspartic acid, bromo-suc- L-rhamnose, D-saccharic acid, D-serine, L-serine, D- cinic acid, D-cellobiose, citric acid, dextrin, formic acid, sorbitol, stachyose, and Tween 40 are not utilized. D-fructose, D-fucose, L-fucose, L-galactonic acid lac- In sensitivity tests, the tetrazolium redox dye is re- tone, D-galactose, D-galacturonic acid, gelatin, gentio- duced at pH 5 and pH 6; the dye is reduced in the pres- biose, D-gluconic acid, α-D-glucose, D-glucose-6-phos- ence of aztreonam, lincomycin, minocycline, nalidixic phate, L-glutamic acid, glycerol, α-hydroxybutric acid, acid, rifamycin SV, D-serine, sodium butyrate, tetrazoli- β-hydroxy-D, L-butyric acid, p-hydroxy-phenylacetic um blue, and tetrazolium violet; but not in the presence acid, inosine, α-keto-butyric acid, α-keto-glutaric acid, of 1% NaCl, 1% sodium lactate, 4% NaCl, 8% NaCl, L-lactic acid, D-lactic acid methyl ester, α-D-lactose, fusidic acid, guanidine HCl, lithium chloride, niaproof D-malic acid, D-maltose, D-mannitol, D-mannose, D- 4, potassium tellurite, sodium bromate, troleandomy- melibiose, β-methyl-D-glucoside, 3-methyl glucose, cin, and vancomycin. Strain JM10-2-6-2 (=NIBRBAC methyl pyruvate, mucic acid, myo-inositol, pectin, pro- 000003988) has been isolated from a soil sample, Dong- pionic acid, glycyl-L-protine, L-pyroglutamic acid, daemun-gu in Seoul, South Korea. quinic acid, D-raffinose, L-rhamnose, D-saccharic acid, D-salicin, D-serine, L-serine, D-sorbitol, stachyose, su- Description of Hymenobacter psychrotolerans crose, D-trehalose, D-turanose, and Tween 40 are not KKM10-2-3 utilized. In sensitivity tests, the tetrazolium redox dye is re- Cells are Gram-stain-negative, non-flagellated, and duced at pH 5 and pH 6; the dye is reduced in the pres- short rod-shaped. Colonies are circular and pale red- ence of minocycline, potassium tellurite, D-serine, sodium colored after 3 days of incubation on R2A at 25°C. In butyrate, tetrazolium blue, and tetrazolium violet; but the GN3 microplates, N-acetyl-D-galactosamine, N- not in the presence of 1% NaCl, 1% sodium lactate, 4% acetyl-D-mannosamine, N-acetyl-neuraminic acid, N- NaCl, 8% NaCl, aztreonam, fusidic acid, guanidine HCl, acetyl-β-D-glucosamine, bromo-succinic acid, citric acid, lincomycin, lithium chloride, nalidixic acid, niaproof dextrin, formic acid, L-fucose, gentiobiose, p-hydroxy- June 2017 KANG AND SRINIVASAN-UNRECORDED RADIATION RESISTANT BACTERIA 99 phenylacetic acid, inosine, α-D-lactose, β-methyl-D- classification of ‘Taxeobacter ocellatus’, ‘Taxeobacter glucoside, 3-methyl glucose, methyl pyruvate, myo-ino- gelupurpurascens’ and ‘Taxeobacter chitinovorans’ as sitol, D-raffinose, D-trehalose, and D-turanose are uti- Hymenobacter ocellatus sp. nov., Hymenobacter lized as sole carbon source. But acetic acid, acetoacetic gelipurpurascens sp. nov. and Hymenobacter chitinivorans acid, L-alanine, γ-amino-butryric acid, D-arabitol, L- sp. nov., respectively, and emended description of the arginine, D-aspartic acid, L-aspartic acid, D-cellobiose, genus Hymenobacter Hirsch et al. 1999. Int J Syst Evol D-fructose, D-fructose-6-phosphate, D-fucose, L-galac­ Microbiol 56:2071-2078. tonic acid lactone, D-galactose, D-galacturonic acid, Callegan, R.P., M.F. Nobre, P.M. McTernan, J.R. Battista, R. gelatin, D-gluconic acid, α-D-glucose, D-glucose-6- Navarro-Gonzalez, C.P. McKay, M.S. da Costa and F.A. phosphate, glucuronamide, D-glucuronic acid, L-glu- Rainey. 2008. Description of four novel psychrophilic, tamic acid, glycerol, L-histidine, α-hydroxybutric acid, ionizing radiation-sensitive Deinococcus species from β-hydroxy-D, L-butyric acid, α-keto-butyric acid, α- alpine environments. Int J Syst Evol Microbiol 58:1252- keto-glutaric acid, L-lactic acid, D-lactic acid methyl 1258 ester, D-malic acid, L-malic acid, D-maltose, D-manni- Doetsch, R.N. 1981. Determinative methods of light micros- tol, D-mannose, D-melibiose, mucic acid, pectin, propi- copy. Manual of Methods for General Bacteriology, pp. 21-33. In: P. Gerhardt, R.G.E. Murray, R.N. Costilow, onic acid, glycyl-L-protine, L-pyroglutamic acid, quinic acid, L-rhamnose, D-saccharic acid, D-salicin, D-serine, E.W. Nester, W.A. Wood, N.R. Krieg and G.H. Phillips L-serine, D-sorbitol, stachyose, sucrose, and Tween 40 (eds.), American Society for Microbiology. Washington, are not utilized. DC, USA. In sensitivity tests, the tetrazolium redox dye is re- Edgar, R.C. 2004. MUSCLE: multiple sequence alignment duced at pH 6 but not at pH 5; the dye is reduced in the with high accuracy and high throughput Nucleic Acids presence of aztreonam, lincomycin, minocycline, nali- Res. 32:1792-1797. dixic acid, potassium tellurite, rifamycin SV, D-serine, Felsenstein, J. 1985. Confidence limit on phylogenies: an approach using the bootstrap. Evolution 39:783-791. tetrazolium violet, and troleandomycin; but not in the Hall, T.A. 1999. BioEdit: a user-friendly biological sequence presence of 1% NaCl, 1% sodium lactate, 4% NaCl, 8% alignment editor and analysis program for Windows 95/ NaCl, fusidic acid, guanidine HCl, lithium chloride, nia­ 98/NT. Nucleic Acids Symp Ser 41:95-98. proof 4, sodium bromate, sodium butyrate, tetrazolium Han, L., S.-J. Wu, C.-Y. Qin, Y.-H. Zhu, Z.-Q. Lu, B. Xie and blue, and vancomycin. Strain KKM10-2-3 (=NIBRBAC J. Lv. 2014. Hymenobacter qilianensis sp. nov., isolated 000003989) has been isolated from a soil sample, Gang­ from a subsurface sandstone sediment in the permafrost nam-gu in Seoul, South Korea. region of Qilian Mountains, China and emended descrip- tion of the genus Hymenobacter. Antonie van Leeuwen- hoek 105:971-978 Acknowledgements Hirsch P., C.A. Gallikowski, J. Siebert, K. Peissl, R. Krop- penstedt, P. Schumann, E. Stackebrandt and R. Ander- This work was supported by a research grant from son. 2004. sp. nov., Deinococcus

Seoul Women’s University (2017) and a grant from the saxicola sp. nov., and Deinococcus marmoris sp. nov.,

National Institute of Biological Resources (NIBR) fund- low temperature and draught-tolerating, UV-resistant ed by the Ministry of Environment (MOE) of the Repub- bacteria from continental Antarctica. Syst Appl Microbi- lic of Korea (NIBR 201501113). ol 27:636-645. Hirsch, P., W. Ludwig, C. Hethke, M. Sittig, B. Hoffmann and C.A. Gallikowski. 1998. Hymenobacter roseosalivarius References gen. nov., sp. nov. from continental Antarctic soils and sandstone: bacteria of the Cytophaga/Flavobacterium/ Ahmed, I., S. Abbas, T. Kudo, M. Iqbal, T. Fujiwara and M. Bacteroides line of phylogenetic descent. Syst Appl Mi- Ohkua. 2014. Deinococcus citri sp. nov., isolated from crobiol 21:374-383. citrus leaf canker lesions. Int J Syst Evol Microbiol 64: Kampfer, P., N. Lodders, B. Huber, E. Falsen and H.-J. Busse. 4134-4140. 2008. Deinococcus aquatilis sp. nov., isolated from water. Brooks, B.W. and R.G.E. Murray. 1981. Nomenclature for Int J Syst Evol Microbiol 58:2803-2806. “Micrococcus radiodurans” and other radiation-resistant Kim, O.S., Y.J. Cho, K. Lee, S.H. Yoon, M. Kim, H. Na, S.C. cocci: Deinococcaceae fam. nov. and Deinococcus gen. Park, Y.S. Jeon, J.H. Lee, H. Yi, S. Won and J. Chun. nov., including five species. Int J Syst Bacteriol 19:353- 2012. Introducing EzTaxon-e: a prokaryotic 16S rRNA 360. gene sequence database with phylotypes that represent Buczolits, S., E.B.M. Denner, P. Kämpfer and H.J. Busse. uncultured species. Int J Syst Evol Microbiol 62:716-721. 2006. Proposal of Hymenobacter norwichensis sp. nov., Klassen, J.L. and J.M. Foght. 2011. Characterization of 100 JOURNAL OF SPECIES RESEARCH Vol. 6, No. 2

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