• Title/Summary/Keyword: 16S rRNA gene-based sequencing

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Exploring the role and characterization of Burkholderia cepacia CD2: a promising eco-friendly microbial fertilizer isolated from long-term chemical fertilizer-free soil

  • HyunWoo Son;Justina Klingaite;Sihyun Park;Jae-Ho Shin
    • Journal of Applied Biological Chemistry
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    • v.66
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    • pp.394-403
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    • 2023
  • In the pursuit of sustainable and environmentally-friendly agricultural practices, we conducted an extensive study on the rhizosphere bacteria inhabiting soils that have been devoid of chemical fertilizers for an extended period exceeding 40 years. Through this investigation, we isolated a total of 80 species of plant growth-promoting rhizosphere bacteria and assessed their potential to enhance plant growth. Among these isolates, Burkholderia cepacia CD2 displayed remarkable plant growth-promoting activity, making it an optimal candidate for further analysis. Burkholderia cepacia CD2 exhibited a range of beneficial characteristics conducive to plant growth, including phosphate solubilization, siderophore production, denitrification, nitrate utilization, and urease activity. These attributes are well-known to positively influence the growth and development of plants. To validate the taxonomic classification of the strain, 16S rRNA gene sequencing confirmed its placement within the Burkholderia genus, providing further insights into its phylogenetic relationship. To delve deeper into the potential mechanisms underlying its plant growth-promoting properties, we sought to confirm the presence of specific genes associated with plant growth promotion in CD2. To achieve this, whole genome sequencing (WGS) was performed by Plasmidsaurus Inc. (USA) utilizing Oxford Nanopore technology (Abingdon, UK). The WGS analysis of the genome of CD2 revealed the existence of a subsystem function, which is thought to be a pivotal factor contributing to improved plant growth. Based on these findings, it can be concluded that Burkholderia cepacia CD2 has the potential to serve as a microbial fertilizer, offering a sustainable alternative to chemical fertilizers.

First Report of Pectobacterium aroidearum Causing Soft Rot on Ficus carica in Korea

  • Kyoung-Taek Park;Leonid N. Ten;Soo-Min Hong;Song-Woon Nam;Chang-Gi Back;Seung-Yeol Lee;Hee-Young Jung
    • Research in Plant Disease
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    • v.30 no.1
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    • pp.88-94
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    • 2024
  • In July 2021, symptoms of soft rot were observed on the stems of Ficus carica in Yeongam, Jeollanamdo, Korea. To accurately diagnose the cause, infected stem was collected and bacterial strain was isolated. Among these, the pathogenic strain KNUB-08-21 was identified as Pectobacterium aroidearum through 16S rRNA gene sequencing and phylogenetic analysis based on the concatenated sequences of the dnaX, leuS, and recA genes. The affiliation of the isolate with this bacterial species was also confirmed by its biochemical characteristics obtained using API ID 32 GN system. Artificial inoculation confirmed the strain's pathogenicity in figs, causing significant damage to both stems and fruits. To our knowledge, this is the first report of P. aroidearum causing soft rot disease in F. carica in Korea.

High-throughput sequencing-based metagenomic and transcriptomic analysis of intestine in piglets infected with salmonella

  • KyeongHye, Won;Dohyun, Kim;Donghyun, Shin;Jin, Hur;Hak-Kyo, Lee;Jaeyoung, Heo;Jae-Don, Oh
    • Journal of Animal Science and Technology
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    • v.64 no.6
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    • pp.1144-1172
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    • 2022
  • Salmonella enterica serovar Typhimurium isolate HJL777 is a virulent bacterial strain in pigs. The high rate of salmonella infection are at high risk of non-typhoidal salmonella gastroenteritis development. Salmonellosis is most common in young pigs. We investigated changes in gut microbiota and biological function in piglets infected with salmonella via analysis of rectal fecal metagenome and intestinal transcriptome using 16S rRNA and RNA sequencing. We identified a decrease in Bacteroides and increase in harmful bacteria such as Spirochaetes and Proteobacteria by microbial community analysis. We predicted that reduction of Bacteroides by salmonella infection causes proliferation of salmonella and harmful bacteria that can cause an intestinal inflammatory response. Functional profiling of microbial communities in piglets with salmonella infection showed increasing lipid metabolism associated with proliferation of harmful bacteria and inflammatory responses. Transcriptome analysis identified 31 differentially expressed genes. Using gene ontology and Innate Immune Database analysis, we identified that BGN, DCN, ZFPM2 and BPI genes were involved in extracellular and immune mechanisms, specifically salmonella adhesion to host cells and inflammatory responses during infection. We confirmed alterations in gut microbiota and biological function during salmonella infection in piglets. Our findings will help prevent disease and improve productivity in the swine industry.

Characterization and Antimicrobial Activity against Gardnerella vaginalis of Vaginal Lactobacillus spp. Isolated from Korean Women (한국 여성의 질에서 분리한 유산균의 Gardnerella vaginalis에 대한 항균효과 및 특성 규명)

  • Kim, YongGyeong;Kang, Chang-Ho;Shin, YuJin;Paek, Nam-Soo;So, Jae-Seong
    • KSBB Journal
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    • v.30 no.5
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    • pp.239-244
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    • 2015
  • Bacterial vaginosis (BV) is caused by microbial imbalance of the vaginal ecosystem and overgrowth of anaerobic bacteria. The antibiotic treatment often results in very high recurrence of BV because it disturbs the vaginal ecosystem. The high recurrence rates suggest a need for alternative therapeutic methods and probiotics are being recognized as alternative or additional treatment method for BV. The purpose of this study was to investigate how human vaginal isolates of Lactobacillus spp. inhibit the BV-associated pathogen Gardnerella vaginalis. Results show that selected strains significantly reduced the viability of G. vaginalis. Among these selected strains KLB410 and KLB416 were further selected based on acid/bile tolerance and identified through 16S rRNA gene sequencing being Lactobacillus plantarum. Further studies are underway to demonstrate that the selected strain can be applied as potential probiotics for recovering vaginal ecosystem.

Impact of a Glyphosate-Tolerant Soybean Line on the Rhizobacteria, Revealed by Illumina MiSeq

  • Lu, Gui-Hua;Zhu, Yin-Ling;Kong, Ling-Ru;Cheng, Jing;Tang, Cheng-Yi;Hua, Xiao-Mei;Meng, Fan-Fan;Pang, Yan-Jun;Yang, Rong-Wu;Qi, Jin-Liang;Yang, Yong-Hua
    • Journal of Microbiology and Biotechnology
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    • v.27 no.3
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    • pp.561-572
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    • 2017
  • The global commercial cultivation of transgenic crops, including glyphosate-tolerant soybean, has increased widely in recent decades with potential impact on the environment. The bulk of previous studies showed different results on the effects of the release of transgenic plants on the soil microbial community, especially rhizosphere bacteria. In this study, comparative analyses of the bacterial communities in the rhizosphere soils and surrounding soils were performed between the glyphosate-tolerant soybean line NZL06-698 (or simply N698), containing a glyphosate-insensitive EPSPS gene, and its control cultivar Mengdou12 (or simply MD12), by a 16S ribosomal RNA gene (16S rDNA) amplicon sequencing-based Illumina MiSeq platform. No statistically significant difference was found in the overall alpha diversity of the rhizosphere bacterial communities, although the species richness and evenness of the bacteria increased in the rhizosphere of N698 compared with that of MD12. Some influence on phylogenetic diversity of the rhizosphere bacterial communities was found between N698 and MD12 by beta diversity analysis based on weighted UniFrac distance. Furthermore, the relative abundances of part rhizosphere bacterial phyla and genera, which included some nitrogen-fixing bacteria, were significantly different between N698 and MD12. Our present results indicate some impact of the glyphosate-tolerant soybean line N698 on the phylogenetic diversity of rhizosphere bacterial communities together with a significant difference in the relative abundances of part rhizosphere bacteria at different classification levels as compared with its control cultivar MD12, when a comparative analysis of surrounding soils between N698 and MD12 was used as a systematic contrast study.

Identification of Lactobacillus spp. associated with nematodes in peach farm soil (복숭아 농장 토양에서 Nematodes와 연관된 Lactobacillus spp.의 분리 및 동정)

  • Lee, Woo-Hyun;Choi, Jae Im;Lee, Jin Il;Lee, Won-Pyo;Yoon, Sung-Sik
    • Korean Journal of Microbiology
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    • v.53 no.3
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    • pp.163-169
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    • 2017
  • Strains D4 and D5 were isolated from peach-rotten soil during the peach harvest season. The isolates were identified based on morphological and biochemical characterization, and identification was determined by 16S rRNA gene sequencing. Results showed that D4 has high similarity to Lactobacillus plantarum ATCC $14917^T$ and Lactobacillus pentosus ATCC $8041^T$ at 99.05% and 98.98%, respectively. D5 was also similar to Lactobacillus pentosus ATCC $8041^T$ and Lactobacillus plantarum ATCC $14917^T$ at 98.71% and 98.64%, respectively. In contrast, isolates showed differences in carbohydrate utilization in comparison to Lactobacillus plantarum ATCC $14917^T$ and Lactobacillus pentosus ATCC $8041^T$. In view of this we performed VITEK MS matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis, multiplex PCR fingerprinting, and random amplified polymorphic DNA (RAPD)-PCR to further confirm the identification of D4 and D5. The results of these analyses showed that both strains were most similar to Lactobacillus plantarum.

Bacterial Community Dynamics during Composting of Food Wastes (음식물 쓰레기 퇴비화 과정에 따른 세균군집 구조의 변화)

  • Shin, Ji-Hye;Lee, Jin-Woo;Nam, Ji-Hyun;Park, Se-Yong;Lee, Dong-Hun
    • Korean Journal of Microbiology
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    • v.45 no.2
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    • pp.148-154
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    • 2009
  • Composting is a biological process converting solid organic waste into valuable materials such as fertilizer. The change of bacterial populations in a composting reactor of food waste was investigated for 2 months. Based on shifts in temperature profile, the composting process could be divided into the first phase ($2^{\circ}C\sim55^{\circ}C$), the second phase ($55^{\circ}C\sim97^{\circ}C$), and the third phase ($50^{\circ}C\sim89^{\circ}C$). The number of total bacteria was $1.66\times10^{11}$ cell/g, $0.29\times10^{11}$ cell/g, and $0.28\times10^{11}$ cell/g in the first, second, and third stages, respectively. The proportions of thermophiles increased from 33% to 89% in the second stage. T-RFLP analysis and nucleotide sequencing of 16S rRNA gene demonstrated that the change of bacterial community structure was coupled with shifts in composting stages. The structure of bacterial community in the ultra-thermophilic second stage reflected that of seeding starter. The major decomposers driving the ultra-thermophilic composting were identified as phylotypes related to Bacillus and Pseudomonas.

Acidophilic Bacterial Communities of Soil and Enrichment Cultures from Two Abandoned Mine Sites of the Korean Peninsula

  • Mishra, Debaraj;Lee, Sun-Hee;Kim, Jae-Hee;Kim, Dong-Jin;Rhee, Young-Ha
    • Korean Journal of Environmental Biology
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    • v.29 no.4
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    • pp.265-273
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    • 2011
  • Bacterial diversity based on the denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rRNA gene sequences was determined for soil samples from two abandoned mine sites and the corresponding enrichment cultures using soil sample as key inoculum. Sequencing analysis of DGGE bands obtained from both the soil samples matched mostly with sequences of uncultured and newly described organisms, or organisms recently associated with the acid mine drainage environment. However, the enrichment of soil samples in ferrous sulfate and elemental sulfur media yielded sequences that were consistent with well-known iron- and sulfur-oxidizing acidophilic bacteria. Analysis of enrichment cultures of soil samples from Dalsung mine revealed abundant ${\gamma}$-$Proteobacteria$, whereas that of Gubong mine sample displayed acidophilic groups of ${\gamma}$-$Proteobacteria$, ${\alpha}$-$Proteobacteria$, $Actinobacteria$ and $Firmicutes$. Chemical elemental analysis of the mine samples indicated that the Dalsung site contained more iron and sulfate along with other toxic components as compared with those of the Gubong site. Biogeochemistry was believed to be the primary control on the acidophilic bacterial group in the enrichment samples.

Association of Salivary Microbiota with Dental Caries Incidence with Dentine Involvement after 4 Years

  • Kim, Bong-Soo;Han, Dong-Hun;Lee, Ho;Oh, Bumjo
    • Journal of Microbiology and Biotechnology
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    • v.28 no.3
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    • pp.454-464
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    • 2018
  • Salivary microbiota alterations can correlate with dental caries development in children, and mechanisms mediating this association need to be studied in further detail. Our study explored salivary microbiota shifts in children and their association with the incidence of dental caries with dentine involvement. Salivary samples were collected from children with caries and their subsequently matched caries-free controls before and after caries development. The microbiota was analyzed by 16S rRNA gene-based high-throughput sequencing. The salivary microbiota was more diverse in caries-free subjects than in those with dental caries with dentine involvement (DC). Although both groups exhibited similar shifts in microbiota composition, an association with caries was found by function prediction. Analysis of potential microbiome functions revealed that Granulicatella, Streptococcus, Bulleidia, and Staphylococcus in the DC group could be associated with the bacterial invasion of epithelial cells, phosphotransferase system, and ${\text\tiny{D}}-alanine$ metabolism, whereas Neisseria, Lautropia, and Leptotrichia in caries-free subjects could be associated with bacterial motility protein genes, linoleic acid metabolism, and flavonoid biosynthesis, suggesting that functional differences in the salivary microbiota may be associated with caries formation. These results expand the current understanding of the functional significance of the salivary microbiome in caries development, and may facilitate the identification of novel biomarkers and treatment targets.

Cultivation-Dependent and -Independent Characterization of Microbial Community Producing Polyhydroxyalkanoates from Raw Glycerol

  • Ciesielski, Slawomir;Pokoj, Tomasz;Klimiuk, Ewa
    • Journal of Microbiology and Biotechnology
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    • v.20 no.5
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    • pp.853-861
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    • 2010
  • High substrate costs decrease the profitability of polyhydroxyalkanoates (PHAs) production, and thus low-cost carbon substrates coming from agricultural and industrial residuals are tested for the production of these biopolymers. Among them, crude glycerol, formed as a by-product during biodiesel production, seems to be the most promising source of carbon. The object of this study was to characterize the mixed population responsible for the conversion of crude glycerol into PHAs by cultivation-dependent and -independent methods. Enrichment of the microbial community was monitored by applying the Ribosomal Intergenic Spacer Analysis (RISA), and the identification of community members was based on 16S rRNA gene sequencing of cultivable species. Molecular analysis revealed that mixed populations consisted of microorganisms affiliated with four bacterial lineages: ${\alpha}$, ${\gamma}$-Proteobacteria, Actinobacteria, and Bacteroides. Among these, three Pseudomonas strains and Rhodobacter sp. possessed genes coding for polyhydroxyalkanoates synthase. Comparative analysis revealed that most of the microorganisms detected by direct molecular analysis were obtained by the traditional culturing method.