• Title/Summary/Keyword: Anaerobic culture

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Characterization of Microbial Diversity of Metal-Reducing Bacteria Enriched from Groundwater and Reduction/Biomineralization of Iron and Manganese (KURT 지하심부 지하수 내 토착 금속환원미생물의 종 다양성 및 철/망간의 환원과 생광물화작용)

  • Kim, Yumi;Oh, Jong-Min;Jung, Hea-Yeon;Lee, Seung Yeop;Roh, Yul
    • Economic and Environmental Geology
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    • v.47 no.4
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    • pp.431-439
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    • 2014
  • The purposes of this research were to investigate the enrichment of metal-reducing bacteria from KURT groundwater and the identification of the microbial diversity by 16S rRNA as well as to examine microbial Fe(III)/Mn(IV) reduction and to analyze morphological features of interactions between microbes and precipitates and their mineralogical composition. To cultivate metal-reducing bacteria from groundwater sampled at the KURT in S. Korea, different electron donors such as glucose, acetate, lactate, formate, pyruvate and Fe(III)-citrate as an electron accepter were added into growth media. The enriched culture was identified by 16S rRNA gene sequence analysis for the diversity of microbial species. The effect of electron donors (i.e., glucose, acetate, lactate, formate, pyruvate) and electron acceptors (i.e., akaganeite, manganese oxide) on microbial iron/manganese reduction and biomineralization were examined using the 1st enriched culture, respectively. SEM, EDX, and XRD analyses were used to determine morphological features, chemical composition of microbes and mineralogical characteristics of the iron and manganese minerals. Based on 16S rRNA gene analysis, the four species, Fusibacter, Desulfuromonas, Actinobacteria, Pseudomonas sp., from KURT groundwater were identified as anaerobic metal reducers and these microbes precipitated metals outside of cells in common. XRD and EDX analyses showed that Fe(III)-containing mineral, akaganeite (${\beta}$-FeOOH), reduced into Fe(II)/Fe(III)-containing magnetite ($Fe_3O_4$) and Mn(IV)-containing manganese oxide (${\lambda}-MnO_2$) into Mn(II)-containing rhodochrosite ($MnCO_3$) by the microbes. These results implicate that microbial metabolism and respiratory activities under anaerobic condition result in reduction and biomineralization of iron and manganese minerals. Therefore, the microbes cultivated from groundwater in KURT might play a major role to reduce various metals from highly toxic, mobile to less toxic, immobile.

Changes of Bacterial Population during the Decomposition Process of Red Tide Dinoflagellate, Cochiodinium polykrikoides in the Marine Sediment Addition of Yellow Loess (황토첨가 해양퇴적물에서 적조생물 Cochiodinium polykrikoides 분해중 세균군집의 변동)

  • PARK Young-Tae;LEE Won-Jae
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.31 no.6
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    • pp.920-926
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    • 1998
  • To investigate the effects of yellow loess on the microbial community after applying into C. polykrikoides as a red tide centrol method during decomposition process, we conducted this study using microcosm experiments, which consisted of sediment collected from Jinhae and Masan bay. The composition, number of bacteria and respiratory electron transport system activity (ETSA) were analyzed. The number of heterotrophic bacteria examined in the samples of both stations reached maximum value within 12 hrs with $10^7$ cells/dry g, independent with the yellow loess applied. In addition, a differenee in the variation of heterotrophic bacterial composition was not observed by adding the yellow loess, and Vibrio spp. always appeared during the culture periods, However, in day 8 culture, the sulfate reducing bacteria was $3.8\times10^7$ cells/dry g in Masan bay and $5.5\times10^6$ cells/dry g in Jinhae bay samples without yellow loess, and these were 120, 350 fold-and 160, 420 fold-increased when yellow loess was added (1 : 1, 1 : 2). The average ETSA was 6.8$\~$7.6 $\mu$g formazan $h^{-1}$ dry $g^{-1}$ independently with yellow loess in aerobic condition for both samples, but activity was decreased by addition of yellow loess in anaerobic. Thus the addition of yellow loess to marine sediment seems to have an effect to inhibit the anaerobic decomposition process and growth of sulfate reducing bacteria which lead to the bad condition of marine environments.

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Longitudinal Study of the Subgingival Microbial Change after Tetracycline Topical Application (Tetracycline 국소도포가 치은연하 세균분포에 미치는 영향)

  • Choi, Kwang-Choon;Lee, Young-Hee;Lee, Jin-Yong;Chung, Chong-Pyoung;Son, Seong-Heui
    • The Journal of the Korean Society for Microbiology
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    • v.21 no.4
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    • pp.503-513
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    • 1986
  • Previous studies have developed the technique of topical application of tetracycline(TC) into the periodontal pockets and examined the change of clinical parameters and subgingival microbial morphotypes. The purpose of this study was to longitudinally examine the clinical and microbiological effects of topically applied TC in a double-blind and split-mouth design. Thirteen patients with moderate periodontitis, who were treated with or without TC application and scaling treatment, were examined. TC gel(3%) was used to apply into the selected periodontal pockets twice a week for 2 weeks. During the experiment, clinical parameters and subgingival microbial morphotypes were examined, and for isolation of black-pigmented Bacteroides(BPB) and streptococci, an anaerobic sample culturing was done at week 0, 2, and 7. In clinical observation the TC-scaled group exhibited a significant decrease of Gingival Inflammatory Index, Plaque Index, Sulcus Bleeding Index, pocket depth, and gingival crevicular fluid when compared to the TC-unsealed, placebo-scaled, and placebo-unsealed groups. The result of microbial morphotype observation showed a significant increase of coccal form and a decrease of spirochetes in the TC-scaled, TC-unscaled, and placebo-scaled groups. The culture study of streptococci revealed that TC with scaling treatment resulted in a significant increase of S. sanguis I at week 2, but its proportion had returned to the base line level. The anaerobic culture study showed that BPB was significantly reduced in the TC-scaled and TC-unsealed groups at week 7. Among BPB species, B. intermedius declined significantly with time treatment(week 2 and 7) in the TC-scaled and TC-unsealed groups. These results suggest that the settled pathogenic microflora can be succeeded by nonpathogenic microflora in periodontal pockets after TC treatment.

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Optimal conditions and effects of prebiotics for growth and antimicrobial substances production of Lactobacillus brevis BK11 (Lactobacillus brevis BK11의 증식과 항균물질 생산을 위한 최적 배양조건 및 prebiotics의 영향)

  • Lim, Eun-Seo
    • Korean Journal of Microbiology
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    • v.51 no.3
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    • pp.288-299
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    • 2015
  • Lactobacillus brevis BK11 obtained from Baikkimchi was selected to study the effects of culture medium, initial pH, atmosphere composition, incubation temperature and time, and prebiotics on growth and production of antimicrobial substances. Growth and antimicrobial substances production of L. brevis BK11 were significantly higher in MRS broth than in BHI or M17 broth. The production of cell mass, lactic acid, and bacteriocin by BK11 strain was at maximum in MRS broth adjusted to pH 6.0. Aerobic and microaerobic conditions were favored cell growth and antimicrobial substances production than anaerobic condition. Biomass and lactic acid production and antimicrobial substances activity of BK 11 were significantly better at 30 and $37^{\circ}C$ than at $25^{\circ}C$. Growth of the strain BK11 entered the stationary growth stage at 24 h after inoculation, and decreased after 36 h. Antimicrobial activities of cell-free culture supernatant and bacteriocin solution were highest when cultured in MRS broth with an initial pH 6.0 for 24-30 h at $37^{\circ}C$. In addition, the highest cell number and lactic acid and bacteriocin production were recorded in the presence of 1 and 2% (w/v) fructooligosaccharide (FOS), however, inulin and raffinose did not affect biological and physicochemical characteristics and antimicrobial activities of L. brevis BK11 cultures. According to these results, production of antimicrobial substances by L. brevis KB11 was closely associated with cell density. Under optimal conditions for antimicrobial substances production, L. brevis BK11 effectively inhibited the growth of Helicobacter pylori ATCC 43504.

Research on the Diversity of Intestinal Microbial Communities of Red tilefish (Branchiostegus japonicus) by 16S rDNA Sequence Analysis (16S rDNA 염기서열 분석에 의한 옥돔(Branchiostegus japonicus)의 장내미생물 군집의 다양성 조사)

  • Kim, Min-Seon;Lee, Seung-Jong;Heo, Moon-Soo
    • Journal of Life Science
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    • v.28 no.3
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    • pp.361-368
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    • 2018
  • This study investigated the diversity of communities of intestinal microorganisms, separated from the intestinal organs of Red tilefish (Branchiostegus japonicas), collected on the Jeju Coast. First, in the isolation of 1.5% BHIA, MA, TSA and R2A Agar on the medium, there were most colonies in 1.5% BHIA. The results of aerobic culture and anaerobic culture were $1.7{\times}10^6CFU/g^{-1}$ and $1.1{\times}10^5cfu/g^{-1}$, respectively, on average, and 147 pure colonies were separated in total. In 16S rDNA sequencing, there were 58 genera and 74 species, showing 95-100% similarity with the basic strain. They were divided broadly into 5 phyla, and as the main phyletic group, Proteobacteria phylum comprised 50% with 9 families, 35 genera and 35 species of Moraxellaceae, Rhodobacteraceae, Shewanellae, Halomondaceae, Enterobacteriaceae, Vibrionaceae, Hahellaceae, Pseudomonadaceae, and Erythrobacteraceae, with the highest index of dominance. Actinobacteria phylum comprised 24% with 8 families, 11 genera and 17 species of Microbacteriaceae, Intrasporangiaceae, Dietziaceae, Dermabacteraceae, Dermacoccaceae, Nocardiodaceae, Brevibacteriaceae and Propionobacteriacea; Firmicutes phylum, 16% with 6 families, 8 genera and 17 species of Bacillaceae, Staphylcoccaceae, Planococcaceae, Streptococcaceae, Paenibacillaceae and Clostridiaceae; Bacteroidetes phylum, 6% with 2 families, 3 genera and 4 species of Cyclobacteriaceae and Flavobacteriaceae; and Deinococcus-Thermus phylum, 4% with 1 family, 1 genus and 1 species of Deinococcaceae.

Identification and Cultural Characterization of Antioxidant Producing Bacteria Isolated from the Jeju Coasts (제주 연안에서 분리한 항산화물질을 생산하는 균주의 동정 및 배양학적 특성)

  • Kim Man-Chul;Heo Moon-Soo
    • Journal of Life Science
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    • v.15 no.5 s.72
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    • pp.749-754
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    • 2005
  • An antioxidant- producing bacterium was isolated from sea water in Jeju island. The isolated strain, SC2-1 was gram-positive, catalase positive, facultatively anaerobic, oxidase negative, motile and small rods. The strain utilized sucrose, dextrose, fructose, mannitol and maltose as a sole carbon and energy source and sodium chloride was required for the bacteria growth. The radical scavenging activity of the culture supernatants was determined by DPPH (1,1-diphenyl-2-picrylhydrazyl) method. This bacterium was identified based on cellular fatty acids analysis and 16S rDNA sequencing, and then named Exiguobacterium sp. SC2-1. The modified optimal medium compositions required the addition of maltose $2.5\%(w/v)$, yeast extract $1.5\%(w/v)$ and $KH_{2} PO_{4} 0.05\%(w/v)$ in marine broth (Difco. Co. USA). Antioxidant activity of under optimal culture conditions were $93\%$.

Microbiology and Antibiotic Selection for Diabetic Foot Infections (감염된 당뇨병성 족부 병변의 균주 조사 및 항생제 선택)

  • Park, Se-Jin;Jung, Haw-Jae;Shin, Hun-Kyu;Kim, Eu-Gene;Lim, Jong-Jun;Yoon, Ji-Woong
    • Journal of Korean Foot and Ankle Society
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    • v.13 no.2
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    • pp.150-155
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    • 2009
  • Purpose: To help the empirical antibiotics selection in diabetic foot infection patients, we investigated prevalence of microorganisms and their antibiotics sensitivity results. Materials and Methods: Patients who came to our clinics to treat diabetic foot infections with deep ulceration and were followed up more than 6 months until complete recovery were adopted. From March 2006 to June 2009, there were 140 patients who corresponded with such a inclusion criteria. Wound cultures were done by deep tissue or bone debris at first visit to our clinics. Microorganisms which was documented by wound culture and most susceptible antibiotics by minimum inhibitory concentrations were surveyed retrospectively. Results: Microorganisms were confirmed in 113 cases (80.7%). In the other 27 cases (19.3%), there were no cultured microorganisms. In bacterial growth group, there were 72 cases (63.7%) of gram-positive bacteria and 41 cases (36.3%) of gram-negative bacteria. All of them were aerobic microorganisms and there were no anaerobic microorganisms. Methicillin-sensitive staphylococcus aureus was the most common pathogen and accounted for 35 cases (31.0%). As other common pathogens, there were Enterobacter cloacae (11 cases, 9.7%), pseudomonas aeruginosa (10 cases, 8.8%), Methicillin-resistant staphylococcus aureus (10 cases, 8.8%) and enterococcus faecalis (6 cases, 5.3%), and so on. Common susceptible antibiotics in gram positive microorganism were vancomycin (60 cases, 83.3%), teicoplanin (60 cases, 83.3%), nitrofurantoin (60 cases, 83.3%) and ciprofloxacin (53 cases, 73.6%). In gram negative ones, common susceptible antibiotics were imipenem (35 cases, 85.3%), piperacillin/tazobactam (33 cases, 80.5%) and gentamicin (31 cases, 75.6%). Conclusion: Methicillin-sensitive Staphylococcus aureus in gram positive and enterobacter cloacae in gram negative was the most common pathogen in each group. Ciprofloxacin and gentamicin might be adaptable as a first-line empirical antibiotics in infected diabetic foot patients.

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Evaluation of different types of mixed microbial culture for biomethanation of CO2 (식종슬러지 종류에 따른 이산화탄소 이용 바이오메탄 생산 비교)

  • Kim, Tae-Hoon;Lim, Byung-Seo;Yi, Sung-Ju;Yun, Gwang-Sue;Ahn, Byung-Kyu;Enkhtsog, Michidmaa;Yun, Yeo-Myeong
    • Journal of the Korea Organic Resources Recycling Association
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    • v.28 no.1
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    • pp.65-72
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    • 2020
  • The aims of this study were to compare the biomethanation of CO2 through specific methanogenic activity (SMA) test which was inoculated with four different types of mixed microbial culture obtained from full-scale anaerobic digestion (AD) plants. The experimental results showed that CH4 conversion was the highest in the samples inoculated by seed sludge taken from ADs of food waste and brewery; under this condition, the produced biomethane contains 89.3-91.9% of CH4. Meanwhile, the lowest level was obtained in the sample from sewage sludge. The measured ratio of CH4 production rate to CO2 consumption rate in all reactors was higher than the theoretical value (1) in the middle of the period and soon dropped to 0.7-0.8. It might be due to changed metabolic pathways in the reactor by the degradation of residual organic matter and the increased activity of homoacetogenic bacteria.

IDENTIFICATION OF PORPHYROMONAS ENDODONTALIS USING POLYMERASE CHAIN REACTION(RCR) (중합효소연쇄반응(Polymerase Chain Reaction)을 이용한 Porphyromonas endodontalis의 동정에 대한 연구)

  • Lee, Sang-Yup;Yoon, Soo-Han
    • Restorative Dentistry and Endodontics
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    • v.23 no.1
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    • pp.328-338
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    • 1998
  • Porphyromonas endodontalis, an anaerobic Gram negative cocobacillus which was known to be associated with the infected root canals and periapical lesions, is very difficult to culture and to detect by the traditional method in that it requires much time to induce the specific black pigmentation, and it is very sensitive to oxygen and the antibiotics added in the culture medium. In this study, the nucleotide sequences of the 'probe h' (0.73kb), one of the specific DNA probes top. endodontalis (ATCC 35406) which had been developed by our department, was determined and then a pair of primers for PCR amplification was fabricated to identify P. endodontalis. The plasmids containing 'probe h' were purified by $Wizard^{TM}$ Midipreps DNA Purification System (Promega Corp.), and the nucleotide sequences of the 'probe h' were determined by the dideoxy chain termination method using TaqTrack Sequencing System (Promega Corp.) and detected by fluorescent labelling method. The sense/antisense PCR primers were designed with computer software (Lasergene, DNASTAR Ind. PCR was done with a programmable GeneAmp PCR System 2400 (Perkin Elmer-Cetus Co.). Each sample containing the whole genomic DNA of P. endodontalis and other black-pigmented Bacteroides was itailly denatured at $94^{\circ}C$ for 5 min and then subjected to 30 cycles, each of them consisting of 60s at $94^{\circ}C$, 60s at $60^{\circ}C$, and 90s. at $72^{\circ}C$. The amplified DNA was resolved electrophoretically in a 1.0 % agarose gel in 1X TAE buffer, stained with EtBr, and photographed on a UV transilluminator. The results were as follows : 1. The nucleotide sequences of 'probe h' (743 base pairs) were obtained by dideoxy chain termination method, and from that results the specific primers to P. endodontalis (ATCC 35406), 'Primer H1/ Primer H2', were designed. 2. It has been found that 'Primer H1/H2' could detect P. endodontalis (ATCC 35406) using PCR. 3. The PCR system with this primers may be a powerful technique to amplify the specific sequences of 'probe h' of P. endodontalis (ATCC 35406) that produce distinct identification of it from other black-pigmented Bacteroides, and this could help us to determine the nature of periapical disease.

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Effects of Increasing Inclusion Levels of Rumen Cellulolytic Bacteria Culture on In vivo Ruminal Fermentation Patterns in Hanwoo Heifers (반추위 섬유소분해 박테리아 배양액의 투여 수준에 따른 한우 반추위 발효에 미치는 영향)

  • Park, Joong-Kook;Jeong, Chan-Sung;Park, Do-Yeun;Kim, Hyun-Cheol;Lee, Seung-Cheol;Kim, Chang-Hyun
    • Journal of Animal Science and Technology
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    • v.51 no.1
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    • pp.45-52
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    • 2009
  • This experiment was conducted to observe the effects of anaerobic cellulolytic bacteria culture (Ruminococcus flavefaciens H-20 and Fibrobactor succinogenes H-23) on in vivo ruminal fermentation characteristics in Hanwoo heifers. Four ruminally cannulated Hanwoo heifers ($221\pm7.5kg$) receiving a basal diet containing 3 kg of mixture hay (tall fescue and ochardgrass) and 2 kg of concentrate per day were in a $4\times4$ Latin square with 21-day periods. Treatments were the basal diet without the culture additive (control), the basal diet plus 50 ml/day of bacteria culture of H-20 and H-23 (1%), 150 ml/day of H-20 and H-23 (3%), and 250 ml/day of H-20 and H-23 (5%). In the whole experimental periods, ruminal pH did not differ between treatments. However, the concentration of ruminal ammonia-N was increased in the 3% treatment relative to control and the 1% treatment at 1 hr post-feeding (p<0.05). Avicelase and CMCase (carboxymethyl cellulase) activities in rumen fluid showed no significant difference among treatments. However, xylanase activity was higher in the 5% (119.49, xylose ${\mu}mol$/ml/min) than the 3% treatment (71.02, xylose ${\mu}mol$/ml/min) at 0 hr post-feeding (p<0.05). Concentrations of ruminal total VFA, acetate, propionate and valerate were unaffected by treatments, while butyrate was higher in the 3% treatment (24.48 mM) than control (15.71 mM) at 1 hr post-feeding (p<0.05). Results indicate that minimum 3% inclusion of cellulolytic bacteria cultures improved ruminal fermentation, especially ammonia-N concentration and butyric acid production.