Hyun Mi Jin

Alteromonas as a key agent of polycyclic aromatic hydrocarbon biodegradation in crude oil-contaminated coastal sediment.

Following the 2007 oil spill in South Korean tidal flats, we sought to identify microbial players influencing the environmental fate of released polycyclic aromatic hydrocarbons (PAHs). Two years of monitoring showed that PAH concentrations in sediments declined substantially. Enrichment cultures were established using seawater and modified minimal media containing naphthalene as sole carbon source. The enriched microbial community was characterized by 16S rRNA-based DGGE profiling; sequencing selected bands indicated Alteromonas (among others) were active. Alteromonas sp. SN2 was isolated and was able to degrade naphthalene, phenanthrene, anthracene, and pyrene in laboratory-incubated microcosm assays. PCR-based analysis of DNA extracted from the sediments revealed naphthalene dioxygenase (NDO) genes of only two bacterial groups: Alteromonas and Cycloclasticus, having gentisate and catechol metabolic pathways, respectively. However, reverse transcriptase PCR-based analysis of field-fixed mRNA revealed in situ expression of only the Alteromonas-associated NDO genes; in laboratory microcosms these NDO genes were markedly induced by naphthalene addition. Analysis by GC/MS showed that naphthalene in tidal-flat samples was metabolized predominantly via the gentisate pathway; this signature metabolite was detected (0.04 μM) in contaminated field sediment. A quantitative PCR-based two-year data set monitoring Alteromonas-specific 16S rRNA genes and NDO transcripts in sea-tidal flat field samples showed that the abundance of bacteria related to strain SN2 during the winter season was 20-fold higher than in the summer season. Based on the above data, we conclude that strain SN2 and its relatives are site natives--key players in PAH degradation and adapted to winter conditions in these contaminated sea-tidal-flat sediments.

Metatranscriptomic Analysis of Lactic Acid Bacterial Gene Expression during Kimchi Fermentation

Barcode-based 16S rRNA gene pyrosequencing showed that the kimchi microbiome was dominated by six lactic acid bacteria (LAB), Leuconostoc (Lc.) mesenteroides, Lactobacillus (Lb.) sakei, Weissella (W.) koreensis, Lc. gelidum, Lc. carnosum, and Lc. gasicomitatum. Therefore, we used completed genome sequences of representatives of these bacteria to investigate metatranscriptomic gene-expression profiles during kimchi fermentation. Total mRNA was extracted from kimchi samples taken at five time points during a 29 day-fermentation. Nearly all (97.7%) of the metagenome sequences that were recruited on all LAB genomes of GenBank mapped onto the six LAB strains; this high coverage rate indicated that this approach for assessing processes carried out by the kimchi microbiome was valid. Expressed mRNA sequences (as cDNA) were determined using Illumina GA IIx. Assignment of mRNA sequences to metabolic genes using MG-RAST revealed the prevalence of carbohydrate metabolism and lactic acid fermentation. The mRNA sequencing reads were mapped onto genomes of the six LAB strains, which showed that Lc. mesenteroides was most active during the early-stage fermentation, whereas gene expression by Lb. sakei and W. koreensis was high during later stages. However, gene expression by Lb. sakei decreased rapidly at 25 days of fermentation, which was possibly caused by bacteriophage infection of the Lactobacillus species. Many genes related to carbohydrate transport and hydrolysis and lactate fermentation were actively expressed, which indicated typical heterolactic acid fermentation. Mannitol dehydrogenase-encoding genes (mdh) were identified from all Leuconostoc species and especially Lc. mesenteroides, which harbored three copies (two copies on chromosome and one copy on plasmid) of mdh with different expression patterns. These results contribute to knowledge of the active populations and gene expression in the LAB community responsible for an important fermentation process.

Microbial succession and metabolite changes during fermentation of dongchimi, traditional Korean watery kimchi.

Dongchimi, one of the most common types of watery kimchi in Korea, was prepared using radish and its pH values, microbial cell numbers, bacterial communities, and metabolites were monitored periodically to investigate the fermentation process of watery kimchi. The bacterial abundance increased quickly during the early fermentation period and the pH values concurrently decreased rapidly without any initial pH increase. After 15 days of fermentation, the bacterial abundance decreased rapidly with the increase of Saccharomyces abundance and then increased again with a decrease of Saccharomyces abundance after 40 days of fermentation, suggesting that bacteria and Saccharomyces have a direct antagonistic relationship. Finally, after 60 days of fermentation, a decrease in bacterial abundance and the growth of Candida were concurrently observed. Community analysis using pyrosequencing revealed that diverse genera such as Leuconostoc, Lactobacillus, Pseudomonas, Pantoea, and Weissella were present at initial fermentation (day 0), but Leuconostoc became predominant within only three days of fermentation and remained predominant until the end of fermentation (day 100). Metabolite analysis using (1)H NMR showed that the concentrations of free sugars (fructose and glucose) were very low during the early fermentation period, but their concentrations increased rapidly although lactate, mannitol, and acetate were produced. After 30 days of fermentation, quick consumption of free sugars and production of glycerol and ethanol were observed concurrently with the growth of Saccharomyces, levels of which might be considered for use as a potential indicator of dongchimi quality and fermentation time.

Comparative Genomics Reveals Adaptation by Alteromonas sp. SN2 to Marine Tidal-Flat Conditions: Cold Tolerance and Aromatic Hydrocarbon Metabolism

Alteromonas species are globally distributed copiotrophic bacteria in marine habitats. Among these, sea-tidal flats are distinctive: undergoing seasonal temperature and oxygen-tension changes, plus periodic exposure to petroleum hydrocarbons. Strain SN2 of the genus Alteromonas was isolated from hydrocarbon-contaminated sea-tidal flat sediment and has been shown to metabolize aromatic hydrocarbons there. Strain SN2s genomic features were analyzed bioinformatically and compared to those of Alteromonas macleodii ecotypes: AltDE and ATCC 27126. Strain SN2s genome differs from that of the other two strains in: size, average nucleotide identity value, tRNA genes, noncoding RNAs, dioxygenase gene content, signal transduction genes, and the degree to which genes collected during the Global Ocean Sampling project are represented. Patterns in genetic characteristics (e.g., GC content, GC skew, Karlin signature, CRISPR gene homology) indicate that strain SN2s genome architecture has been altered via horizontal gene transfer (HGT). Experiments proved that strain SN2 was far more cold tolerant, especially at 5°C, than the other two strains. Consistent with the HGT hypothesis, a total of 15 genomic islands in strain SN2 likely confer ecological fitness traits (especially membrane transport, aromatic hydrocarbon metabolism, and fatty acid biosynthesis) specific to the adaptation of strain SN2 to its seasonally cold sea-tidal flat habitat.

Isolation of a BTEX-degrading bacterium, Janibacter sp. SB2, from a sea-tidal flat and optimization of biodegradation conditions.

An enrichment culture was established using seawater containing BTEX (benzene, toluene, ethylbenzene and xylene) compounds to isolate a BTEX-degrading bacterium from contaminated sea-tidal flat. The enriched microbial communities were characterized by 16S rRNA-based DGGE profiling, which indicated that a Janibacter species was dominant during the enrichment. Strain SB2, corresponding to the major band and able to degrade all BTEX compounds, was isolated and characterized. NH4Cl, NaH2PO4, cell mass and BTEX concentrations were used as independent variables to optimize the degradation of BTEX by strain SB2 in a tidal flat and a statistically significant (R(2)=0.8933, p<0.0001) quadratic polynomial mathematical model was suggested. For the initial concentration of 240 mg/L BTEX in a slurry system containing 3.0×10(7) cells/L, 45.5% BTEX removal was observed under the optimum condition of NH4Cl and NaH2PO4, while 32.2% BTEX removal was observed under the untreated condition of NH4Cl and NaH2PO4.

Altererythrobacter gangjinensis sp. nov., a marine bacterium isolated from a tidal flat.

A Gram-stain-negative, ochre-pigmented, strictly aerobic bacterium, designated strain KJ7(T), was isolated from a tidal flat of the Gangjin bay in South Korea. Cells were halotolerant, non-motile, catalase- and oxidase-positive rods. Growth of strain KJ7(T) was observed at 5-35 °C (optimum, 25 °C), at pH 6.0-9.5 (optimum, pH 6.5-7.0) and in the presence of 0-9 % (w/v) NaCl (optimum, 2 %). The major cellular fatty acids were C18 : 1ω7c, C17 : 1ω6c, summed feature 3 (comprising C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0. The polar lipid pattern indicated the presence of phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, a sphingoglycolipid, an unidentified phospholipid and two unidentified lipids. The G+C content of the genomic DNA was 60.2±0.9 mol% and the predominant respiratory quinone was Q-10. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain KJ7(T) formed a phyletic lineage distinct from other members of the genus Altererythrobacter and was most closely related to Altererythrobacter luteolus SW-109(T) and Altererythrobacter namhicola KYW48(T) (95.6 and 95.0 % 16S rRNA gene sequence similarity, respectively). On the basis of phenotypic, chemotaxonomic and molecular features, strain KJ7(T) represents a novel species of the genus Altererythrobacter, for which the name Altererythrobacter gangjinensis sp. nov. is proposed. The type strain is KJ7(T) ( = KACC 16190(T) = JCM 17802(T)).

Comparative Genomic Analysis and Benzene, Toluene, Ethylbenzene, and o-, m-, and p-Xylene (BTEX) Degradation Pathways of Pseudoxanthomonas spadix BD-a59

ABSTRACT Pseudoxanthomonas spadix BD-a59, isolated from gasoline-contaminated soil, has the ability to degrade all six BTEX (benzene, toluene, ethylbenzene, and o-, m-, and p-xylene) compounds. The genomic features of strain BD-a59 were analyzed bioinformatically and compared with those of another fully sequenced Pseudoxanthomonas strain, P. suwonensis 11-1, which was isolated from cotton waste compost. The genome of strain BD-a59 differed from that of strain 11-1 in many characteristics, including the number of rRNA operons, dioxygenases, monooxygenases, genomic islands (GIs), and heavy metal resistance genes. A high abundance of phage integrases and GIs and the patterns in several other genetic measures (e.g., GC content, GC skew, Karlin signature, and clustered regularly interspaced short palindromic repeat [CRISPR] gene homology) indicated that strain BD-a59s genomic architecture may have been altered through horizontal gene transfers (HGT), phage attack, and genetic reshuffling during its evolutionary history. The genes for benzene/toluene, ethylbenzene, and xylene degradations were encoded on GI-9, -13, and -21, respectively, which suggests that they may have been acquired by HGT. We used bioinformatics to predict the biodegradation pathways of the six BTEX compounds, and these pathways were proved experimentally through the analysis of the intermediates of each BTEX compound using a gas chromatograph and mass spectrometry (GC-MS). The elevated abundances of dioxygenases, monooxygenases, and rRNA operons in strain BD-a59 (relative to strain 11-1), as well as other genomic characteristics, likely confer traits that enhance ecological fitness by enabling strain BD-a59 to degrade hydrocarbons in the soil environment.

Litorimonas taeanensis gen. nov., sp. nov., isolated from a sandy beach

A heterotrophic, gram-negative, prosthecate bacterium, designated strain G5(T), was isolated from a sandy beach of Taean in South Korea. Cells of strain G5(T) were aerobic, catalase- and oxidase-positive, straight to slightly curved motile rods with a single flagellum and formed yellow-orange colonies on agar. Growth occurred at 15-40 °C (optimum 25-30 °C) and pH 6-9 (optimum pH 7-8). The major cellular fatty acids were C(18 : 1)ω7c, C(17 : 0), C(16 : 0), 11-methyl C(18 : 1)ω7c, C(17 : 1)ω8c and C(17 : 1)ω6c. The polar lipid pattern indicated the presence of phosphatidylglycerol, monoglycosyldiglyceride, glucuronopyranosyldiglyceride and two unidentified glycolipids. The G+C content of the genomic DNA was 63.6 mol% and the major quinone was Q-10. Comparative 16S rRNA gene sequence analysis showed that strain G5(T) belonged to the branch containing the genera Hellea, Robiginitomaculum and Hypomonas within the family Hyphomonadaceae. Within this group, strain G5(T) was most closely related to Hellea balneolensis 26III/A02/215(T) with 95.8 % 16S rRNA gene sequence similarity. Based on its phylogenetic position and its phenotypic, chemotaxonomic and molecular properties, strain G5(T) represents a novel species of a novel genus of the family Hyphomonadaceae, for which the name Litorimonas taeanensis gen. nov., sp. nov. is proposed. The type strain is G5(T) ( = KACC 13701(T)  = DSM 22008(T)).

Complete Genome Sequence of the Polycyclic Aromatic Hydrocarbon-Degrading Bacterium Alteromonas sp. Strain SN2

Alteromonas sp. strain SN2, able to metabolize polycyclic aromatic hydrocarbons, was isolated from a crude oil-contaminated sea-tidal flat. Here we report the complete 4.97-Mb genome sequence and annotation of strain SN2. These will advance the understanding of strain SN2s adaptation to the sea-tidal flat ecosystem and its pollutant metabolic versatility.

Aestuariibaculum suncheonense gen. nov., sp. nov., a marine bacterium of the family Flavobacteriaceae isolated from a tidal flat and emended descriptions of the genera Gaetbulibacter and Tamlana

A Gram-staining-negative, yellow-pigmented, strictly aerobic bacterium, designated strain SC17(T), was isolated from sediment of a tidal flat of Suncheon bay in South Korea. Cells were halotolerant, catalase- and oxidase-positive and non-motile rods. Growth of strain SC17(T) was observed at 5-40 °C (optimum, 25-30 °C), at pH 6.0-8.5 (optimum, pH 7.0) and in the presence of 1-8 % (w/v) NaCl (optimum, 1-2 %). The major cellular fatty acids were iso-C(15 : 0), summed feature 3 (comprising C(16 : 1)ω7c and/or C(16 : 1)ω6c and/or iso-C(15 : 0) 2-OH), iso-C(17 : 0) 3-OH, iso-C(15 : 1) G and anteiso-C(15 : 0.) The polar lipid content consisted of phosphatidylethanolamine and unidentified amino lipids and lipids. The G+C content of the genomic DNA was 46.4 mol% and the only respiratory quinone detected was menaquinone-6 (MK-6). Phylogenetic inference based on 16S rRNA gene sequences showed that strain SC17(T) formed a distinct phyletic lineage within the family Flavobacteriaceae and was most closely related to members of the genera Gaetbulibacter and Tamlana with 95.0-95.8 % sequence similarity. On the basis of phenotypic and molecular features, strain SC17(T) represents a novel genus of the family Flavobacteriaceae, for which the name Aestuariibaculum suncheonense gen. nov., sp. nov. is proposed. The type strain is SC17(T) (= KACC 16186(T) = JCM 17789(T)). Emended descriptions of the genera Gaetbulibacter and Tamlana are also proposed.