Pedobacter Lotistagni Sp. Nov. Isolated from Lotus Pond Water

Antonie van Leeuwenhoek (2015) 107:951–959 DOI 10.1007/s10482-015-0387-5

ORIGINAL PAPER

Pedobacter lotistagni sp. nov. isolated from lotus pond water

Hina Singh • Juan Du • Hien T. T. Ngo • Ki-Young Kim • Tae-Hoo Yi

Received: 27 November 2014 / Accepted: 13 January 2015 / Published online: 23 January 2015 Ó Springer International Publishing Switzerland 2015

Abstract A light-yellow coloured, Gram-stain neg- 23344 T. The DNA G?C content was determined to be ative, rod-shaped, aerobic, non-motile bacterium, 36.8 mol%. In DNA–DNA hybridization tests, the designated THG-DN6.8T, was isolated from a lotus DNA relatedness between strain THG-DN6.8T and its pond near Donghaksa temple in Daejeon, South closest phylogenetic neighbour P. koreensis was Korea. The strain was found to grow well on nutrient found to be below 10 %. The predominant isoprenoid agar, optimally at pH 6.0–7.5, at temperature quinone was identiﬁed as menaquinone MK-7 and the 25–28 °C and in the presence of 0.5 % NaCl. Based major polar lipid as phosphatidylethanolamine. The on 16S rRNA gene sequence analysis, strain THG- major fatty acids of strain THG-DN6.8T were identi- T DN6.8 was found to share the highest sequence ﬁed as iso-C15:0,C16:0,C18:0 and C16:1 x6c and/or T similarity with Pedobacter koreensis KCTC 12536 , C16:1 x7c (summed feature 3). On the basis of the followed by Pedobacter glacialis CCTCC AB phenotypic characteristics, genotypic analysis and 2012941T, Pedobacter kyungheensis KACC 16221T, chemotaxonomic characteristics, strain THG-DN6.8T Pedobacter caeni LMG 22862T, Pedobacter insulae is considered to represent a novel species of the genus KCTC 12820T and Pedobacter boryungensis KCTC Pedobacter, for which the name Pedobacter lotistagni sp. nov. is proposed. The type strain is THG-DN6.8T (= KCTC 42229T = JCM 30354T). Hina Singh and Juan Du have contributed equally to this work. Keywords Pedobacter lotistagni Gram-staining- Electronic supplementary material The online version of negative Menaquinone MK-7 16S rRNA this article (doi:10.1007/s10482-015-0387-5) contains supplementary material, which is available to authorized users.

H. Singh J. Du H. T. T. Ngo T.-H. Yi (&) Introduction Department of Oriental Medicinal Material & Processing, College of Life Science, Kyung Hee University Global The genus Pedobacter was first described by Steyn Campus, 1732 Deokyoungdaero, Giheung-gu, Yongin-si, et al. (1998) and belongs to the family Sphingobacte- Gyeonggi-do 446-701, Republic of Korea e-mail: [email protected] riaceae and order Sphingobacteriales. Mostly Pedob- acter species have been reported from soils (Yoon et al. K.-Y. Kim 2007; Luo et al. 2010; Zhou et al. 2012; Oh et al. 2013). Department of Genetic Engineering, College of Life Members of this genus are ubiquitous in nature and Science, Kyung Hee University Global Campus, 1732 Deokyoungdaero, Giheung-gu, Yongin-si, some species have been reported from various envi- Gyeonggi-do 446-701, Republic of Korea ronments such as a waterfall (Chung et al. 2014), 123 952 Antonie van Leeuwenhoek (2015) 107:951–959 glacial cryonite (Margesin et al. 2003), glacial water Suspended cells were placed on carbon- and formvar- (Qui et al. 2014), food (Derichs et al. 2014) and coated nickel grids for 30 s and grids were floated on compost (Lee et al. 2009). Currently, there are 45 one drop of 0.1 % (w/v) aqueous uranyl acetate, blotted species of the genus Pedobacter with validly published dry and then viewed with a transmission electron names (http://www.bacterio.net/pedobacter.html). microscope (Model JEM1010; JEOL) at 11,0009 Members of the genus Pedobacter are Gram-stain magnification under standard operating conditions. negative, rod-shaped bacteria, non-motile or motile by Gram-staining was determined using a bioMe´rieux gliding, with MK7 as the predominant isoprenoid Gram stain Kit according to the manufacturer’s quinone and G?C content of 36–45 mol% (Qui et al. instructions. Cells were grown in NB broth for 24 h 2014). More recently, the G?C content range of the at 28 °C and then tested for gliding motility by the genus has been emended by Kook et al. (2014)to hanging-drop technique (Skerman 1967). Growth on

36.0–47.5 mol%. The major fatty acids are iso-C15:0, Reasoner’s 2A agar (R2A; Difco), tryptone soya agar C16:0, iso-C15:1 G, iso-C17:0 C16:1 x6c, C16:1 x7c and (TSA, Oxoid), NA, Luria–Bertani agar (LB; Oxoid), iso-C17:0 3-OH. Pedobacter species mostly contain Marine agar (MB; Difco) and MacConkey agar phosphatidylethanolamine as the major polar lipid (Oxoid) was also assessed at 28 °C for 7 days. Growth (Zhou et al. 2012). In this study, we characterized by at different temperatures (4, 10, 15, 18, 25, 28, 30, 35, means of a polyphasic approach that a new isolate from 37 and 42 °C) and pH conditions (pH 4.0–10.0, at a lotus pond near Donghaksa temple in Daejeon, South intervals of 0.5 pH units) were determined using NB Korea belongs to the genus Pedobacter. The pheno- after 4 days of incubation at 28 °C. The following pH typic and genotypic characterization of the novel buffers were used (final concentration, 100 mM): strain, THG-DN6.8T, are described in this report. acetate buffer was used for pH 4.0–6.5 and phosphate buffer was used for pH 7.0–10.0. The pH of NB was confirmed after autoclaving. The requirement of NaCl Materials and methods was tested using of 0–5 % (w/v) NaCl (at 0.5 % intervals) in NB after 4 days of incubation at 28 °C. Isolation of the bacterial strain Growth was estimated by monitoring the optical density at 600 nm. Anaerobic growth was tested in A water sample was collected from a lotus pond near serum bottles containing NB supplemented with Donghaksa temple in Daejeon in a sterile falcon tube and thioglycolate (0.1 %) and in which the air was transferred to the laboratory. The water sample was substituted with nitrogen gas. Production of flexiru- diluted in 0.85 % (w/v) saline solution, serially diluted up bin-type pigments was determined by the reversible to 10-6 and spread on different media including nutrient colour shift to red, purple or brown when yellow or agar (NA, Difco). The plates were incubated at 28 °Cfor orange colonies are covered with aqueous 20 % KOH 1 week. Single colonies were purified by subculturing solution (Fautz and Reichenbach 1980). Methyl red them on to new NA plates. Firstly, the isolate was and Voges-Proskauer reaction were tested in Clark- routinely cultured on NA at 28 °C and stored as glycerol Lubs’ medium (Scharlau). Catalase activity was suspension 25 % (v/v) at -80 °C. Strain THG-DN6.8T determined by the production of bubbles from 3 % has been deposited in the Korean Collection for Type (v/v) H2O2 solution mixed with freshly grown cells and Cultures (KCTC 42229T) and Japan Collection of oxidase activity was determined by using of 1 % (w/v) Microorganisms (JCM 30354T). For comparative studies, N,N,N,N-tetramethyl-p-phenylenediamine reagent six reference Pedobacter type strains (Table 1)were (Sigma) according to the manufacturer’s instructions. obtained from various culture collection centers. These Tests for hydrolysis were performed on NA containing: reference strains were cultured under the same optimum casein [2 % skim milk (Oxoid)], starch [1 % starch conditions as strain THG-DN6.8T. (Difco)], esculin [0.1 % esculin and 0.02 % ferric

citrate (Difco)], Tween 80 [0.01 % CaCl22H2O and Morphological and physiological characterization 1 % Tween 80 (Sigma)], Tween 20 [0.01 % CaCl2- 2H2O and 1 % Tween 20 (Sigma)], chitin [1 % chitin After culturing for 3 days on NA at 28 °C, the from crab shell (Sigma)], L-tyrosine [0.5 % L-tyrosine morphology of the novel strain was examined. (Sigma)], carboxymethyl-cellulose (CMC) [0.1 % 123 Antonie van Leeuwenhoek (2015) 107:951–959 953

Table 1 Differential phenotypic characteristics of strain THG-DN6.8T and the reference type strains of the genus Pedobacter Characteristics 1 2 3 4 5 6 7

Nitrate reduction ---?--- Indole production ----?-- Motility -?----- Hydrolysis of: Tween 20 ?-?w ??? Tween 80 --?--?? L-tyrosine w ?????? Casein ----?-- Starch ???--?? DNA -?-???- Gelatin ----?-- Assimilation of (API 20NE) b-galactosidase (PNPG) ?????-- D-glucose ??-??w ? L-arabinose ? w -?-w - D-mannose ??????? D-mannitol -----w - N-acetyl-glucosamine w ?????w D-maltose ??-???w Enzyme activities (API ZYM) Esterase (C4) ???---- Esterase lipase (C8) ?????-- Lipase (C14) ??----? Leucine arylamidase ??????? Valine arylamidase ????-?? Cystine arylamidase ?-----? Trypsin w -?-?-- a-chymotrypsin ?-??-?- Naphtol-AS-BI-phosphohydrolase ????-?? a-galactosidase ?-??--? b-galactosidase ?????-- a-glucosidase ?-??-?? b-glucosidase ?-??--- N-Acetyl-b-glucosaminidase ????-?? a-Mannosidase w --?--- a-Fucosidase ---??-- DNA G?C Content (mol%) 38.8 38.0 39.4 42.4 42.7 39.4 38.5

All strains are positive for the following characteristics: hydrolysis of esculin and CMC; assimilation of D-Mannose and N-acetylglucosamine; oxidase, catalase, leucine arylamidase, alkaline phosphatase and acid phosphatase. All strains were negative for the following characteristics: hydrolysis of urea and chitin; assimilation of arginine dihydrolase, gluconate, caprate, adipate, malate, citrate and phenyl-acetate; glucose acidiﬁcation and b-glucuronidase Strains: 1, THG-DN6.8T;2,Pedobacter koreensis KCTC 12536T;3,Pedobacter glacialis CCTCC AB 2012941T;4,Pedobacter kyungheensis KACC 16221T;5,Pedobacter caeni LMG 22862T;6,Peobacter insulae KCTC 12820T;7,Pedobacter boryungensis KCTC 23344 T ?, Positive; -, negative; w, weakly positive

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CMC (Sigma)] and DNA [DNase agar, Scharlau system (Alliance 2690 system, Waters) as described (Spain); DNase activity revealed by flooding the plates previously (Mesbah et al. 1989) with reversed-phase with 1 N HCl]. Plates were evaluated after 4 days of column SunFireTM C18 (4.6 9 250 mm 9 5 lm), incubation at 28 °C. Basic chemical tests, carbon- flow rate of 1.0 ml/min, solvent mixture of 200 mM source assimilation and enzyme activities for the novel (NH4)H2PO4/acetonitrile (97:3, v/v) as mobile phase isolate and all reference strains were conducted using and wavelength at 270 nm. The genomic DNA of API 20NE (bioMe´rieux) and API ZYM (bioMe´rieux), Escherichia coli strain B (Sigma-Aldrich D4889) was according to the manufacturer’s instructions. API used as a standard. 20NE were recorded after incubation for 48 h, under the optimal conditions for each strain, while API ZYM DNA–DNA hybridization was recorded after incubation for 10 h. DNA–DNA hybridization was performed fluoromet- Molecular characterization and phylogenetic rically, according to the method developed by Ezaki construction et al. (1989) with modifications (Stabili et al. 2008), using photobiotin-labelled DNA probes and micro– Genomic DNA was extracted and purified using a dilution wells. DNA–DNA hybridization was carried commercial Genomic DNA extraction kit (Solgent, out to determine levels of relatedness of the novel Korea). The 16S rRNA gene was amplified with the strain THG-DN6.8T with its closest relative Pedob- universal bacterial primer pair 27F (50-TAC- acter koreensis KACC 15270T. The optimum rena- CAGGGTATCTAATCC-30) and 1492R (50- turation temperature (30 °C) was calculated as GGTTACCTTGTTACGACTT-30) (Weisburg et al. [(0.51 9 G?C content) ? 47] - 36 (Gillis et al. 1991) and the purified PCR products were sequenced 1970), where 36 °C is the correction for the presence by Solgent Co. Ltd (Daejeon, Korea). The 16S rRNA of 50 % formamide (McConaughy et al. 1969). gene sequences of related taxa were obtained from the Hybridization was performed with five replications GenBank database and EzTaxon e-server [http:// for each sample. The highest and lowest values eztaxon-e.ezbiocloud.net/; Kim et al. (2012)]. Seq- obtained for each sample were excluded and the Man software version 4.1 (DNASTAR, Inc.) was used means of the remaining three values were converted to to compile the nearly complete (1,433 bp) 16S rRNA percentage DNA–DNA relatedness values. sequence of strain THG-DN6.8T. The multiple align- ments were performed using the CLUSTAL_X pro- Chemotaxonomic characterization gram (Thompson et al. 1997) and gaps were edited using the BioEdit program (Hall 1999). The evolu- For quinone and polar lipids analysis, cells were tionary distances were calculated using the Kimura grown in NA broth at 28 °C, shaken at 160 rpm for two-parameter model (Kimura 1983). The phyloge- 2 day, centrifuged and freeze dried. Respiratory netic trees were constructed using the neighbour- quinones were extracted from 300 mg freeze-dried joining method (Saitou and Nei 1987), maximum- cells with chloroform:methanol (2:1, v/v), separated parsimony (Fitch 1971) and maximum-likelihood using hexane and eluted with hexane:diethyl ether method (Felsenstein 1981) in the MEGA5.2 Program (90:10, v/v), then eluent was evaporated by rotatory (Tamura et al. 2011), with bootstrap values based on evaporation and dissolved in acetone. Menaquinone 1,000 replications (Felsenstein 1985). purification was determined using a reverse-phase HPLC system (Waters, Alliance 2690 system) [wave- DNA G?C mol% content length 270 nm, solvent MeOH: isopropanol (7:5, v/v), flow rate; 1.0 ml/min] as previously described (Hirai- For determination of the DNA G?C content, genomic shi et al. 1996; Collins and Jones 1981; Tamaoka et al. DNA of strain THG-DN6.8T was extracted and puri- 1983). The polar lipids of strain THG-DN6.8T and its fied by the method of Moore and Dowhan (1995), then closest reference strain P. koreensis KACC 15270T degraded enzymatically into nucleosides using nucle- were analyzed as described by (Minnikin et al. 1984). ase P1 and alkaline phosphatase (Sigma). The nucleo- Two-dimensional thin layer chromatography (2D- sides were analyzed using a reverse-phase HPLC TLC) performed using TLC Kiesel gel 60 F254 plates 123 Antonie van Leeuwenhoek (2015) 107:951–959 955

(10 9 10 cm; Merck). The plates were first developed of 4–37 °C (optimum, 25–28 °C), at pH 6.0–8.5 with chloroform:methanol:water (65:25:4, v/v/v) fol- (optimum, 6.0–7.0) and at 0.5–2.0 % NaCl (optimum, lowed by a second development with chloroform: 0.5 %). Phenotypic analysis showed that strain THG- methanol:acetic acid:water (80:12:15:4, v/v/v/v). For DN6.8T cells are Gram-strain negative, non-motile, the presence of total and specific lipids, the plates were aerobic and rod shaped with size range approximately developed using sprays of 5 % molybdatophosphoric 0.3–0.5 9 1.3–2.0 lm (Supplementary Fig. S2). Tests acid (total lipids, Sigma), 0.2 % ninhydrin (aminoli- for oxidase and catalase activity were found to be pids, Sigma) and 2.5 % a-naphthol-sulfuric acid positive. Nitrate reduction and indole production were (glycolipids, Sigma) and drying at 120 °C for found to be negative. Strain THG-DN6.8T was found 5–10 min. TLC plates were also sprayed with molyb- to be unable to hydrolyze Tween 80, casein, chitin, denum blue reagent (Sigma) for detecting phospho- gelatin, urea and DNA but able to hydrolyze L- lipids. No heating step is needed for this reagent. tyrosine, starch, Tween 20, CMC and esculin. The For fatty acid analysis, strains THG-DN6.8T and all biochemical and physiological characteristics of strain the reference strains were cultured on NA at 28 °C for THG-DN6.8T and the most closely related Pedobacter 48 h. Cells in exponential growth phase were used. type strains are given in Table 1. The results suggest Fatty acids were extracted, methylated and saponified that the novel isolate represents a novel species of the by the methods described for the Sherlock Microbial genus Pedobacter. Identification system (MIDI) and analyzed by capil- Strain THG-DN6.8T was found to contain MK-7 as lary GLC (Hewlet Packard 6890) using the TSBA the predominant isoprenoid quinone, in line with all library version 6.1 (Sasser 1990). other members of the genus Pedobacter. As shown in Supplementary Fig. S3, the major polar lipid was identified as phosphatidylethanolamine, which is similar to the polar lipid profile of the reference strain P. Results and discussion koreensis KACC 15270T. In addition, some unidentified aminophospholipids (APL1-3) and unidentified The 16S rRNA sequence of strain THG-DN6.8T polar lipids (L1-2) were also detected in both the (NCBI Genbank accession number KM035962) strains, whilst an additional unidentified polar lipid revealed that strain THG-DN6.8T belongs to the genus (L3) was observed in the reference strain only. An Pedobacter within the family Sphingobacteriaceae. unidentified aminolipid (AL) was detected in the novel The highest sequence similarity was found with isolate but not in P. koreensis KACC 15270T. The species of the genus Pedobacter including P. koreen- previously reported polar lipid profile of Pedobacter sis KACC 15270T (97.3 %), Pedobacter glacialis jejuensis KACC 17172T (Kook et al. 2014) is similar in KACC 11491T (96.7 %), Pedobacter kyungheensis comparison with that of strain THG-DN6.8T as both NBRC 106274T (96.2 %), Pedobacter caeni (95.6 %), strains contain phosphatidylethanolamine as the major Pedobacter insulae (95.7 %) and Pedobacter bor- polar lipid and significant amounts of unidentified yungensis (95.6 %). A neighbour-joining phyloge- aminophospholipids. Thus, the polar lipid profile of netic tree based on 16S rRNA gene sequences showed novel isolate is in line with those of members of the the position of strain THG-DN6.8T clustered within genus Pedobacter. The major fatty acids of strain T the genus Pedobacter (Fig. 1). A phylogenetic tree THG-DN6.8 were identified as iso-C15:0 (17.4 %), built using the maximum-likelihood method is pre- C16:0 (16.6 %), C18:0 (10.2 %) and C16:1 x6c and/or sented as Supplementary Fig. S1. These results C16:1 x7c (11.5 %) as was seen in the closest related suggest that strain THG-DN6.8T represents a novel reference type strains (Table 2). The proportions of species within the genus Pedobacter. C16:0 and C18:0 were found to be notably different from Colonies of strain THG-DN6.8T on NA agar were those in the closely related species. observed to be light-yellow, round, sticky and raised The DNA G?C content of novel isolate was with approximate diameter 2–3 mm. Strain THG- determined to be 36.8 mol%, which lies within the DN6.8T was found to grow on LB, R2A and NA but range (36.0–47.5 mol%) for members of the genus not to grow on TSA, MA and MacConkey agar. The Pedobacter (Kook et al. 2014). The DNA–DNA strain was found to grows best on NA at temperatures hybridization value of strain THG-DN6.8T with its 123 956 Antonie van Leeuwenhoek (2015) 107:951–959

Fig. 1 Neighbour-joining tree based on 16S rRNA gene Numbers at nodes indicate bootstrap percentages (based on sequence analysis, showing relationships between strain THG- 1,000 resampled datasets). Bootstrap values less than 80 % are DN6.8T and members of the genus Pedobacter. Filled circles not indicated. Mucilaginibacter auburnensis JM-1070T was indicate that the corresponding nodes were also recovered in the used as an out group. Scale bar, 0.01 substitutions per nucleotide tree generated with the maximum-parsimony algorithm. position closest reference strain P. koreensis KACC 15270T considered to represent a novel species of the genus was determined to be 9.5 ± 1.5. This low DNA–DNA Pedobacter, for which name Pedobacter lotistagni sp. relatedness suggests that strain THG-DN6.8T is a nov. is proposed. novel Pedobacter species, following the recommen- dations of Stackebrandt and Goebel (1994). Description of Pedobacter lotistagni sp. nov The characteristics of the novel isolate are consis- tent with the description of the genus Pedobacter with Pedobacter lotistagni (lo.ti.stag’ni. L. fem. n. lotos regard to morphological, biochemical and chemotax- lotus; L. neut. n. stagnum pond; N.L. gen. n. lotistagni onomic properties. On the basis of the data from this of a lotus pond). polyphasic study, including analysis of 16S rRNA Cells are Gram-staining-negative, rod-shaped, aer- gene sequences, phylogenetic, phenotypic, biochem- obic and non-motile. Cell size is approximately ical and chemotaxonomic properties, strain THG- 0.3–0.5 lm in width and 1.3–2.0 lm in length. DN6.8T (= KCTC 42229T = JCM 30354T)is Colonies are light-yellow, round, sticky, raised and 123 Antonie van Leeuwenhoek (2015) 107:951–959 957

Table 2 Fatty acid proﬁles of strain THG-DN6.8T and reference type strains of the genus Pedobacter Fatty acid 1 2 3 4 5 6 7

Straight

C16:0 16.6 5.4 5.0 7.1 3.1 7.1 3.0

C18:0 10.2 3.2 tr 2.3 tr 4.6 tr Branched

iso-C15:0 17.5 14.1 7 21.8 19.2 18.6 29.3

iso-C16:0 2.0 2.2 tr tr tr 1.3 tr

anteiso-C15:0 1.4 2.2 6.8 tr 2.5 1.9 2.1

iso-C16:1 H 1.4 2.2 tr tr tr 3.1 tr Unsaturated

C15:1 x6c 2.3 3.6 6.5 tr ND 4.8 4.2

anteiso-C17:1 x9c 6.1 1.6 6.9 tr ND 1.9 1.2

C18:3 x6c 2.7 tr ND tr ND tr ND Hydroxy

iso-C15:0 3OH 2.1 1.8 1.5 3.1 3 2.5 2.5

iso-C16:0 3OH 4.1 9.6 tr 1.4 1.8 8.0 5.4

iso-C17:0 3OH 7.1 4.9 6.5 15.2 2.1 8.6 6.8 Summed feature 3* 11.5 27.6 24.2 29.9 45.2 19.7 17.9 For fatty acid analysis, novel isolate and all reference strains were cultured on NA at 28 °C for 48 h and cells at exponential growth phase were used for analysis. Summed feature 3* could not be separated by the Microbial Identiﬁcation System (MIDI). Fatty acids amounting to less than 1.0 % in all strains are not listed. Summed feature 3* consists of C16:1 x6c and/or C16:1 x7c Strains: 1, THG-DN6.8T;2,Pedobacter koreensis KCTC 12536T;3,Pedobacter glacialis CCTCC AB 2012941T;4,Pedobacter kyungheensis KACC 16221T;5,Pedobacter caeni LMG 22862T;6,Peobacter insulae KCTC 12820T;7,Pedobacter boryungensis KCTC 23344T ND, not detected; tr, trace amount (\0.5 %) have a diameter of 2–3 mm. Positive for oxidase and a-chymotrypsin, acid phosphatase, Naphtol-AS-BI- catalase tests. Grows on R2A, NA and LB but does not phosphohydrolase, a-galactosidase, a-glucosidase, b- grow on TSA, MA and MacConkey agar. Growth glucosidase and N-acetyl-b-glucosaminidase; weakly occurs at 4–37 °C and the optimum temperature is positive for leucine arylamidase, trypsin, b-galactosi- 25–28 °C. Growth occurs at pH 6.0–8.5 and optimum dase, a-mannosidase; and negative for b-glucuronidase pH 6.0–7.0. Grows well in the presence of 0.5–2.0 % and a-fucosidase. MK-7 menaquinone is the predomi- NaCl (w/v) and the optimum is 0.5 %. Unable to nant isoprenoid quinone. The major polar lipid is hydrolyze Tween 80, casein, chitin, gelatin, urea and phosphatidylethanolamine. The major fatty acids are

DNA but able to hydrolyze L-tyrosine, starch, Tween 20, iso-C15:0,C16:0,C18:0 and C16:1 x6c and/or C16:1 CMC and esculin. No ﬂexirubin-type pigments. Nega- x7c (summed feature 3 as deﬁned by MIDI). The tive in the Methyl Red-Voges-Proskauer test, for nitrate DNA G?C content of the type strain is 36.8 mol%. reduction and indole production. According to API The type strain, THG-DN6.8T (= KCTC T T 20NE testing, positive for assimilation of D-glucose, L- 42229 = JCM 30354 ), was isolated from a lotus arabinose, D-mannose, D-maltose, N-acetylglucosamine pond in Daejeon, South Korea. The NCBI GenBank and D-maltose but negative for assimilation of D- accession number for the 16S rRNA gene sequence of mannitol, gluconate, caprate, adipate, malate, citrate strain THG-DN6.8T is KM035962. and phenyl-acetate. According to API ZYM tests, positive for the following enzyme activities: alkaline Acknowledgments This work was conducted under the phosphatase, esterase (C4), esterase lipase (C8), lipase industrial infrastructure program (No. N0000888) for fundamental technologies which is funded by the Ministry of (C14), valine arylamidase, cystine arylamidase, Trade, Industry & Energy (MOTIE, Korea).

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Zhou Z, Jiang F, Wang S, Peng F, Dai J, Li W, Fang C (2012) daechungensis, Pedobacter terricola, Pedobacter gluco- Pedobacter arcticus sp. nov., a facultative psychrophile sidilyticus and Pedobacter lentus. Int J Syst Evol Microbiol isolated from Arctic soil, and emended descriptions of the 62:1963–1969 genus Pedobacter, Pedobacter heparinus, Pedobacter

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