De-Chao Zhang

Characterization of the bacterial archaeal diversity in hydrocarbon-contaminated soil.

A polyphasic approach combining culture-based methods with molecular methods is useful to expand knowledge on microbial diversity in contaminated soil. Microbial diversity was examined in soil samples from a former industrial site in the European Alps (mainly used for aluminum production and heavily contaminated with petroleum hydrocarbons) by culture-dependent and culture-independent methods. The physiologically active eubacterial community, as revealed by fluorescence-in-situ-hybridization (FISH), accounted for 6.7% of the total (DAPI-stained) bacterial community. 4.4% and 2.0% of the DAPI-stained cells could be attributed to culturable, heterotrophic bacteria able to grow at 20°C and 10°C, respectively. The majority of culturable bacterial isolates (34/48) belonged to the Proteobacteria (with a predominance of Alphaproteobacteria and Gammaproteobacteria), while the remaining isolates were affiliated with the Actinobacteria, Cytophaga-Flavobacterium-Bacteroides and Firmicutes. A high fraction of the culturable, heterotrophic bacterial population was able to utilize hydrocarbons. Actinobacteria were the most versatile and efficient degraders of diesel oil, n-alkanes, phenol and PAHs. The bacterial 16S rRNA gene clone library contained 390 clones that grouped into 68 phylotypes related to the Proteobacteria, Bacteroidetes, Actinobacteria and Spirochaetes. The archaeal 16S rRNA gene library contained 202 clones and 15 phylotypes belonging to the phylum Euryarchaeota; sequences were closely related to those of methanogenic archaea of the orders Methanomicrobiales, Methanosarcinales, Methanobacteriales and Thermoplasmatales. A number of bacterial and archaeal phylotypes in the clone libraries shared high similarities with strains previously described to be involved in hydrocarbon biodegradation. Knowledge of the bacterial and archaeal diversity in the studied soil is important in order to get a better insight into the microbial structure of contaminated environments and to better exploit the bioremediation potential by identifying potential hydrocarbon degraders and consequently developing appropriate bioremediation strategies.

Dyadobacter psychrophilus sp. nov., a psychrophilic bacterium isolated from soil

A Gram-negative, aerobic bacterium, designated strain BZ26(T), was isolated from hydrocarbon-contaminated soil. The strain was psychrophilic, showing good growth over a temperature range of 1-20 degrees C. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain BZ26(T) was related to members of the genus Dyadobacter and had highest 16S rRNA gene sequence similarity to Dyadobacter alkalitolerans 12116(T) (98.1 %), Dyadobacter koreensis KCTC 12537(T) (97.5 %) and Dyadobacter ginsengisoli Gsoil 043(T) (97.2 %). Strain BZ26(T) had MK-7 as the major menaquinone and summed feature 3 (C(16 : 1)omega7c/iso-C(15 : 0) 2-OH), C(16 : 1)omega5c and iso-C(15 : 0) as major fatty acids. The genomic DNA G+C content of strain BZ26(T) was 48.9 mol%. On the basis of phenotypic characteristics and genotypic analysis, strain BZ26(T) is considered to represent a novel species of the genus Dyadobacter, for which the name Dyadobacter psychrophilus sp. nov. is proposed. The type strain is BZ26(T) (=DSM 22270(T) =CGMCC 1.8951(T)).

Nocardioides alpinus sp. nov., a psychrophilic actinomycete isolated from alpine glacier cryoconite.

A gram-positive, non-motile, rod-shaped, psychrophilic actinomycete, designated strain Cr7-14(T), was isolated from alpine glacier cryoconite. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain Cr7-14(T) was related to members of the genus Nocardioides and shared highest 16S rRNA gene sequence similarities with the type strains of Nocardioides furvisabuli (98.6 %), Nocardioides ganghwensis (98.2 %), Nocardioides oleivorans (98.1 %) and Nocardioides exalbidus (97.6 %). The predominant cellular fatty acids of strain Cr7-14(T) were C(17 : 1)ω8c (39.5 %) and iso-C(16 : 0) (32.4 %). The major menaquinone was MK-8(H(4)). The diagnostic diamino acid in the cell-wall peptidoglycan was ll-2,6-diaminopimelic acid. The predominant cell-wall sugars were galactose and rhamnose. The polar lipid pattern contained diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylinositol, four unknown glycolipids and two unknown polar lipids. The genomic DNA G+C content was 71.9 mol%. On the basis of phenotypic characteristics, phylogenetic analysis and DNA-DNA relatedness data, a novel species, Nocardioides alpinus sp. nov., is proposed, with Cr7-14(T) ( = DSM 23325(T) = LMG 26053(T) = CGMCC 1.10697(T)) as the type strain.

Glaciimonas immobilis gen. nov., sp. nov., a member of the family Oxalobacteraceae isolated from alpine glacier cryoconite

Strains Cr9-30(T) and Cr9-12 were isolated from alpine glacier cryoconite. Both strains were Gram-negative-staining, non-motile, rod-shaped and psychrophilic, showing good growth over the temperature range 1-20 °C. Phylogenetic analysis of 16S rRNA gene sequences revealed that the two strains formed a distinct branch within the family Oxalobacteraceae and were most closely related to members of the genus Collimonas. The 16S rRNA gene sequence similarity between strains Cr9-30(T) and Cr9-12 was 99.0 %. The two strains showed highest 16S rRNA gene sequence pairwise similarity with Collimonas pratensis LMG 23965(T) (96.6 and 96.1 % for strains Cr9-30(T) and Cr9-12, respectively), Collimonas arenae LMG 23964(T) (96.5 and 96.3 %, respectively) and Collimonas fungivorans LMG 21973(T) (96.4 and 96.2 %, respectively). The predominant cellular fatty acids were summed feature 3 (C₁₆:₁ω7c and/or iso-C₁₅:₀ 2-OH), C₁₆:₀ and C₁₈:₁ω7c. The DNA G+C content of strain Cr9-30(T) was 51.0 mol%. On the basis of phenotypic characteristics and phylogenetic analysis, strains Cr9-30(T) and Cr9-12 represent a novel species in a new genus of the family Oxalobacteraceae, for which the name Glaciimonas immobilis gen. nov., sp. nov. is proposed. The type strain of Glaciimonas immobilis is Cr9-30(T) ( = DSM 23240(T) = LMG 25547(T)).

Luteimonas terricola sp. nov., a psychrophilic bacterium isolated from soil.

Strain BZ92r(T) was isolated from hydrocarbon-contaminated soil. Cells were Gram-negative, aerobic, rod-shaped and cold-adapted (growth at 1-25 degrees C). The major fatty acids were iso-C(15 : 0) (25.6 %), iso-C(17 : 1)omega9c (24.9 %), iso-C(11 : 0) (18.4 %) and iso-C(11 : 0) 3-OH (16.2 %). The predominant ubiquinone was ubiquinone-8. The genomic DNA G+C content was 72.0 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain BZ92r(T) was a member of the genus Luteimonas (94.5-95.2 % 16S rRNA gene sequence similarity). On the basis of phenotypic, chemotaxonomic and phylogenetic distinctiveness, strain BZ92r(T) was considered to represent a novel species of the genus Luteimonas. The name Luteimonas terricola sp. nov. is proposed, with BZ92r(T) (=DSM 22344(T) =CGMCC 1.8985(T)) as the type strain.

Pedobacter bauzanensis sp. nov., isolated from soil

A Gram-negative, aerobic, rod-shaped, non-motile bacterium, designated BZ42(T), was isolated from the soil of an industrial site. Strain BZ42(T) was able to grow at 5-25 °C. The major fatty acids were iso-C(15 : 0) (46.2 %), C(16 : 1)ω7c and/or iso-C(15 : 0) 2-OH (23.2 %) and iso-C(17 : 0) 3-OH (9.1 %). The predominant menaquinone was MK-7. The genomic DNA G+C content was 36.5 mol% (HPLC). 16S rRNA gene sequence phylogenetic analysis revealed that strain BZ42(T) was a member of the genus Pedobacter, family Sphingobacteriaceae, and 16S rRNA gene sequence similarities between strain BZ42(T) and the type strains of species of the genus Pedobacter with validly published names were 90.4-93.2 %. On the basis of phenotypic, chemotaxonomic and phylogenetic distinctiveness, strain BZ42(T) was considered to represent a novel species of the genus Pedobacter, for which the name Pedobacter bauzanensis sp. nov. is proposed. The type strain is BZ42(T) (=DSM 22554(T) =CGMCC 1.10187(T) =CIP 110134(T)).

Colwellia polaris sp. nov., a psychrotolerant bacterium isolated from Arctic sea ice

A novel psychrotolerant, Gram-negative, aerobic bacterium, designated strain 537T, was isolated from sea-ice samples from the Arctic. Strain 537T was able to grow at 4-26 degrees C, with optimum growth occurring at 20-21 degrees C. Strain 537T had Q-8 as the major respiratory quinone and contained iso-C15:0 2-OH and/or C16:1 omega7c (22.95 %), C15:1 (17.64 %) and C17:1 omega8c (13.74 %) as the predominant cellular fatty acids. The genomic DNA G+C content was 38.9 mol%. A phylogenetic analysis based on 16S rRNA gene sequences indicated that strain 537T formed a coherent cluster within the genus Colwellia. The highest level of 16S rRNA gene sequence similarity (97.5 %) exhibited by strain 537T was obtained with respect to the type strain of Colwellia aestuarii. On the basis of phenotypic, chemotaxonomic and phylogenetic properties and DNA-DNA relatedness data, strain 537T represents a novel species of the genus Colwellia, for which the name Colwellia polaris sp. nov. is proposed. The type strain is 537T (=CGMCC 1.6132T =JCM 13952T).

Sphingomonas glacialis sp. nov., a psychrophilic bacterium isolated from alpine glacier cryoconite.

A non-motile, rod-shaped, yellow bacterium, designated C16y(T), was isolated from alpine glacier cryoconite. Cells behaved Gram-positively, were aerobic and psychrophilic (good growth at 1-25 °C). Phylogenetic analysis based on 16S rRNA gene sequences showed that strain C16y(T) was related to the genus Sphingomonas and had highest 16S rRNA gene sequence similarity with Sphingomonas oligophenolica JCM 12082(T) (97.6  %) and Sphingomonas echinoides DSM 1805(T) (97.2  %). DNA-DNA hybridization demonstrated that strain C16y(T) could not be considered as a member of either Sphingomonas oligophenolica or Sphingomonas echinoides. Strain C16y(T) contained Q-10 as the predominant ubiquinone and C₁₈:₁ and C₁₆:₀ were the dominant fatty acids. The polar lipid profile contained phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, sphingoglycolipid, five unidentified glycolipids, two unidentified aminophospholipids and two unidentified lipids. The major polyamines were the triamines sym-homospermidine and spermidine. The G+C content was 67.9 mol%. Combined data from phenotypic, phylogenetic and DNA-DNA relatedness studies demonstrated that strain C16y(T) is a representative of a novel species of the genus Sphingomonas, for which we propose the name Sphingomonas glacialis sp. nov. The type strain is C16y(T) (=DSM 22294(T) =CGMCC 1.8957(T) =CIP 110131(T) [corrected]).

Devosia psychrophila sp. nov. and Devosia glacialis sp. nov., from alpine glacier cryoconite, and an emended description of the genus Devosia

Two psychrophilic strains, Cr7-05(T) and Cr4-44(T), isolated from alpine glacier cryoconite, were characterized by using a polyphasic approach. Both strains were psychrophilic, showing good growth over a temperature range of 1-20 °C. The chemotaxonomic characteristics of these isolates included the presence of C(18:1)ω7c and summed feature 3 (C(16:1)ω7c and/or C(16:1)ω6c) as the major cellular fatty acids, Q-10 as the predominant ubiquinone and diphosphatidylglycerol, phosphatidylglycerol and unknown glycolipids as major polar lipids. The DNA G+C contents of strains Cr7-05(T) and Cr4-44(T) were 61.4 and 63.6 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the two isolates belong to the genus Devosia. The 16S rRNA gene sequence similarity between the two strains was 98.6%, but DNA-DNA hybridization indicated 54% relatedness. Strains Cr7-05(T) and Cr4-44(T) exhibited 16S rRNA gene sequence similarity of 94.7-97.2 and 94.9-96.9%, respectively, to the type strains of recognized Devosia species. On the basis of phenotypic characteristics, phylogenetic analysis and DNA-DNA relatedness data, strains Cr7-05(T) and Cr4-44(T) represent two novel species within the genus Devosia, for which the names Devosia psychrophila sp. nov. (type strain Cr7-05(T) =DSM 22950(T) =CGMCC 1.10210(T) =CIP 110130(T)) and Devosia glacialis sp. nov. (type strain Cr4-44(T) =CGMCC 1.10691(T) =LMG 26051(T)) are proposed. An emended description of the genus Devosia is also provided.

Isolation and Characterization of Bacteria from Ancient Siberian Permafrost Sediment

In this study, we isolated and characterized bacterial strains from ancient (Neogene) permafrost sediment that was permanently frozen for 3.5 million years. The sampling site was located at Mammoth Mountain in the Aldan river valley in Central Yakutia in Eastern Siberia. Analysis of phospolipid fatty acids (PLFA) demonstrated the dominance of bacteria over fungi; the analysis of fatty acids specific for Gram-positive and Gram-negative bacteria revealed an approximately twofold higher amount of Gram-negative bacteria compared to Gram-positive bacteria. Direct microbial counts after natural permafrost enrichment showed the presence of (4.7 ± 1.5) × 108 cells g−1 sediment dry mass. Viable heterotrophic bacteria were found at 0 °C, 10 °C and 25 °C, but not at 37 °C. Spore-forming bacteria were not detected. Numbers of viable fungi were low and were only detected at 0 °C and 10 °C. Selected culturable bacterial isolates were identified as representatives of Arthrobacter phenanthrenivorans, Subtercola frigoramans and Glaciimonas immobilis. Representatives of each of these species were characterized with regard to their growth temperature range, their ability to grow on different media, to produce enzymes, to grow in the presence of NaCl, antibiotics, and heavy metals, and to degrade hydrocarbons. All strains could grow at −5 °C; the upper temperature limit for growth in liquid culture was 25 °C or 30 °C. Sensitivity to rich media, antibiotics, heavy metals, and salt increased when temperature decreased (20 °C > 10 °C > 1 °C). In spite of the ligninolytic activity of some strains, no biodegradation activity was detected.