scholarly journals The Existence of Endophytic Actinobacteria from Rhododendron zoelerri Revealed by Culture-Dependent and Culture-Independent Approaches

2018 ◽  
Vol 25 (2) ◽  
pp. 54
Author(s):  
Yulin Lestari ◽  
Lia Aseptin Murdini ◽  
Dedy Duryadi Solihin
Keyword(s):  
Community Structure ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Morphological Characteristics ◽  
Rrna Gene ◽  
Culture Independent ◽  
Reference Strains ◽  
Culture Dependent ◽  
Endophytic Actinobacteria

Endophytic actinobacteria from medicinal plant may play a significant role in producing bioactive compounds. The information regarding their diversity is an important.  Rhododendron are traditionally used for treating human disorders. One of the selected Rhododendron used in this study was R.  zoelleri from Papua origin, which has been conserved and grown in Cibodas Botanical Garden, West Java, Indonesia. The aim of this study was to assess the existence of endophytic actinobacteria from R. zoelleri based on a culture-dependent and their community structure based on a culture-independent approach. Culturable actinobacteria were isolated and cultured on HV medium. Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis (PCR-DGGE) targeting the metagenomic 16S rRNA was used to analyse the structure of the actinobacterial community. Six culturable endophytic actinobacteria (200 cfu/g fresh weight) from R. zoelleri were successfully isolated, three isolates from leaf, and the other isolates were obtained from stem. The six culturable isolates were RZP 1.3, RZP 1.1, RZP 2.2, RZPB 1.1, RZPB 7.1, RZPB 4.1. Based on their morphological characteristics, the endophytes have Streptomyces characters. The existence of Streptomyces spp. were also confirmed with molecular analysis based on 16S rRNA gene. The phylogenetic analysis based on 16S rRNA gene to the reference strains available in EzTaxon-e database showed that six isolates were closely related to S. djakartensis strains of NBRC 15409ᵀ(99.19%), S. tritolerans strains of DAS 165T(99.90%), S. coelicoflavus strains of NBRC 15399T(99.59). However, they showed differences in morphological characteristics as compared with the reference strains. The metagenomic analysis of the DGGE profile based on 16S rRNA gene showed the community structure of endophytic actinobacteria from R. zoelleri which was represented by 13 DGGE bands. The bands were closely related to Agromyces, Gordonia, Microbacterium, Micromonospora, Propionibacterium, Saccharomonospora, Streptomyces which have 93.18%-100% similarity. Based on the data, it showed diversity of endophytic actinobacteria from R. zoelleri which may be further assess for their novelty and bioprospecting.

Culture-dependent and -independent approaches establish the complexity of a PAH-degrading microbial consortium

2005 ◽  
Vol 51 (11) ◽  
pp. 897-909 ◽  
Author(s):  
Marc Viñas ◽  
Jordi Sabaté ◽  
Caterina Guasp ◽  
Jorge Lalucat ◽  
Anna M Solanas
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Microbial Consortium ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Sole Source ◽  
Rrna Gene ◽  
Dominant Group ◽  
Clone Libraries ◽  
Variable Regions ◽  
Culture Dependent

A microbial consortium (AM) obtained by sequential enrichment in liquid culture with a polycyclic aromatic hydrocarbon (PAH) mixture of three- and four-ringed PAHs as a sole source of carbon and energy was examined using a triple-approach method based on various cultivation strategies, denaturing gradient gel electrophoresis (DGGE), and the screening of 16S and 18S rRNA gene clone libraries. Eleven different sequences by culture-dependent techniques and seven by both DGGE and clone libraries were obtained. The comparison of three variable regions (V3–V5) of the 16S rRNA gene between the sequences obtained yielded 19 different microbial components. Proteobacteria were the dominant group, representing 83% of the total, while the Cytophaga–Flexibacter–Bacteroides group (CFB) was 11% and the Ascomycota fungi 6%. β-Proteobacteria were predominant in the DGGE and clone library methods, whereas they were a minority in culturable strains. The highest diversity and number of noncoincident sequences were achieved by the cultivation method that showed members of the α-, β-, and γ-Proteobacteria; CFB bacterial group; and Asco mycota fungi. Only six of the 11 strains isolated showed PAH-degrading capability. The bacterial strain (AMS7) and the fungal strain (AMF1), which were similar to Sphingomonas sp. and Fusarium sp., respectively, achieved the greatest PAH depletion. The results indicate that polyphasic assessment is necessary for a proper understanding of the composition of a microbial consortium.Key words: microbial consortium, microbial diversity, PAHs, DGGE, 16S rRNA gene.

scholarly journals Influence of Particle Size on Bacterial Community Structure in Aquatic Sediments as Revealed by 16S rRNA Gene Sequence Analysis

2008 ◽  
Vol 74 (16) ◽  
pp. 5237-5240 ◽  
Author(s):  
Colin R. Jackson ◽  
Andrew Q. Weeks
Keyword(s):  
Particle Size ◽  
Community Structure ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Denaturing Gradient Gel Electrophoresis ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Gene Sequence Analysis ◽  
Gradient Gel Electrophoresis ◽  
Rrna Gene Sequencing

ABSTRACT Bacterial communities associated with sediment particles were examined using PCR-denaturing gradient gel electrophoresis and 16S rRNA gene sequencing. Particle size influenced community structure, with attached bacterial assemblages separating into 63- to 125-, 125- to 1,000-, and 1,000- to 2,000-μm fractions. Differences were particularly pronounced for the Verrucomicrobia-Planctomycetes, whose numbers were significantly reduced on coarser particles.

scholarly journals Taxonomic Structure and Stability of the Bacterial Community in Belgian Sourdough Ecosystems as Assessed by Culture and Population Fingerprinting

2008 ◽  
Vol 74 (8) ◽  
pp. 2414-2423 ◽  
Author(s):  
Ilse Scheirlinck ◽  
Roel Van der Meulen ◽  
Ann Van Schoor ◽  
Marc Vancanneyt ◽  
Luc De Vuyst ◽  
...  
Keyword(s):  
16S Rrna ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Taxonomic Structure ◽  
Rrna Gene ◽  
Type I ◽  
Technological Parameters ◽  
Structure And Stability ◽  
Two Samples ◽  
Culture Independent ◽  
Culture Dependent

ABSTRACT A total of 39 traditional sourdoughs were sampled at 11 bakeries located throughout Belgium which were visited twice with a 1-year interval. The taxonomic structure and stability of the bacterial communities occurring in these traditional sourdoughs were assessed using both culture-dependent and culture-independent methods. A total of 1,194 potential lactic acid bacterium (LAB) isolates were tentatively grouped and identified by repetitive element sequence-based PCR, followed by sequence-based identification using 16S rRNA and pheS genes from a selection of genotypically unique LAB isolates. In parallel, all samples were analyzed by denaturing gradient gel electrophoresis (DGGE) of V3-16S rRNA gene amplicons. In addition, extensive metabolite target analysis of more than 100 different compounds was performed. Both culturing and DGGE analysis showed that the species Lactobacillus sanfranciscensis, Lactobacillus paralimentarius, Lactobacillus plantarum, and Lactobacillus pontis dominated the LAB population of Belgian type I sourdoughs. In addition, DGGE band sequence analysis demonstrated the presence of Acetobacter sp. and a member of the Erwinia/Enterobacter/Pantoea group in some samples. Overall, the culture-dependent and culture-independent approaches each exhibited intrinsic limitations in assessing bacterial LAB diversity in Belgian sourdoughs. Irrespective of the LAB biodiversity, a large majority of the sugar and amino acid metabolites were detected in all sourdough samples. Principal component-based analysis of biodiversity and metabolic data revealed only little variation among the two samples of the sourdoughs produced at the same bakery. The rare cases of instability observed could generally be linked with variations in technological parameters or differences in detection capacity between culture-dependent and culture-independent approaches. Within a sampling interval of 1 year, this study reinforces previous observations that the bakery environment rather than the type or batch of flour largely determines the development of a stable LAB population in sourdoughs.

scholarly journals PENDEKATAN CULTURE INDEPENDENT UNTUK ANALISIS KOMUNITAS BAKTERI

OSEANA ◽  
2019 ◽  
Vol 42 (1) ◽  
pp. 9-17
Author(s):  
Nur Fitriah Afianti ◽  
Yeti Darmayati
Keyword(s):  
Bacterial Community ◽  
16S Rrna ◽  
Target Genes ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Single Strand Conformation Polymorphism ◽  
Molecular Techniques ◽  
Rrna Gene ◽  
Culturable Bacteria ◽  
Culture Independent ◽  
Culture Dependent

CULTURE INDEPENDENT APPROACH FOR BACTERIAL COMMUNITY ANALYSIS. Analysis of bacterial community can be through two approaches, through cultivation (culture dependent) and without cultivation (culture independent). Culture dependent approach is conventional method which only covered few bacteria because not all bacteria could be cultured. Culture independent approach with molecular techniques based on DNA communities can provide more information about the structure and diversity of bacteria in nature, both culturable bacteria and unculturable bacteria. 16S rRNA gene is commonly target gene used in bacterial communities analysis. Other specific target genes also being developed for specific groups of bacteria. Several methods are developed for the analysis of molecular markers 16S rRNA or other specific genes, including Denaturing Gradient Gel Electrophoresis (DGGE), Terminal Restriction Fragment Length Polymorphism (TRFLP), and Single Strand Conformation Polymorphism (SSCP).

scholarly journals Marinobacter gudaonensis sp. nov., isolated from an oil-polluted saline soil in a Chinese oilfield

2007 ◽  
Vol 57 (2) ◽  
pp. 250-254 ◽  
Author(s):  
Jun Gu ◽  
Hua Cai ◽  
Su-Lin Yu ◽  
Ri Qu ◽  
Bin Yin ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Saline Soil ◽  
Sequence Similarity ◽  
Eastern China ◽  
Rrna Gene ◽  
Respiratory Quinone ◽  
Shengli Oilfield ◽  
Major Respiratory Quinone ◽  
Reference Strains

Two novel strains, SL014B61AT and SL014B11A, were isolated from an oil-polluted saline soil from Gudao in the coastal Shengli Oilfield, eastern China. Cells of strains SL014B61AT and SL014B11A were motile, Gram-negative and rod-shaped. Growth occurred at NaCl concentrations of between 0 and 15 % and at temperatures of between 10 and 45 °C. Strain SL014B61AT had Q9 as the major respiratory quinone and C16 : 0 (21.2 %), C18 : 1ω9c (20.3 %), C16 : 1ω7c (7.3 %) and C16 : 1ω9c (6.4 %) as predominant fatty acids. The G+C content of the DNA was 57.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain SL014B61AT belonged to the genus Marinobacter in the class Gammaproteobacteria. Strain SL014B61AT showed the highest 16S rRNA gene sequence similarity with Marinobacter bryozoorum (97.9 %) and showed 97.8 % sequence similarity to Marinobacter lipolyticus. DNA–DNA relatedness to the reference strains Marinobacter bryozoorum and Marinobacter lipolyticus was 35.5 % and 33.8 %, respectively. On the basis of these data, it is proposed that strains SL014B61AT and SL014B11A represent a novel species, Marinobacter gudaonensis sp. nov. The type strain is strain SL014B61AT (=DSM 18066T=LMG 23509T=CGMCC 1.6294T).

scholarly journals Molecular Diversity Assessment of Plant Growth Promoting Rhizobacteria Using Denaturing Gradient Gel Electrophoresis (DGGE) of 16s rRNA Gene

2017 ◽  
Vol 5 (1) ◽  
pp. 72-80
Author(s):  
Umesh Prasad Shrivastava
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Uttar Pradesh ◽  
Plant Growth Promoting Rhizobacteria ◽  
Melting Behavior ◽  
Rrna Gene ◽  
Dgge Profile ◽  
Diversity Assessment

The rhizobacteria were isolated from rhizosphere of rice plant of different fields of 4 districts of Nepal and 5 districts of Bihar and Uttar Pradesh, adjoining states of India with Nepal. The DGGE analysis was performed for diversity analysis. For the construction of dendrogram, 16S rRNA gene was amplified by two different sets of primers. The DGGE ladder consisting of PCR amplified products of nine pure bacterial cultures were obtained. The first DGGE ladder was prepared by 400 bp fragment of 16S rDNA with GC clamp and the second DGGE ladder was prepared with 200 bp fragment of 16S rDNA with GC clamp. The perpendicular DGGE of these amplicons based on their melting behavior clearly demonstrated separation of different isolates. The 16S rDNA fragment amplified with primer set of V2-V3 regions with GC clamp showed separation between 40-60% of denaturant. The DGGE profile based on primer set F352T and 519r for various bacteria present in soil samples of 5 districts of India and 4 districts of Nepal revealed that the number of bands which might be specific for diazotrophic isolates varied from 2 to 11. The dendrogram constructed based on DGGE profile of various samples of 5 districts of India and 4 districts of Nepal showed that all the samples could be clustered in nine groups with 58-96% similarity to each other. Among all these 37 samples, only Var-4 and Var-5 showed 100% similarity, no other samples from any site showed 100% similarity. Int. J. Appl. Sci. Biotechnol. Vol 5(1): 72-80

scholarly journals Amplification of 16S rRNA gene sequences to differentiate two highly related bradyrhizobia species

2007 ◽  
Vol 42 (9) ◽  
pp. 1361-1364 ◽  
Author(s):  
Adriana Giongo ◽  
Adriana Ambrosini ◽  
João Ruy Jardim Freire ◽  
Maria Helena Bodanese Zanettini ◽  
Luciane Maria Pereira Passaglia
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Bradyrhizobium Japonicum ◽  
Diagnostic Method ◽  
Molecular Diagnostic ◽  
Rrna Gene ◽  
16S Rrna Gene Sequences ◽  
Closely Related Species ◽  
Reference Strains ◽  
Commercial Inoculants

A 16S rRNA gene PCR-based assay was developed aiming at a fast molecular diagnostic method to differentiate the two phylogenetically closely related species Bradyrhizobium japonicum and B. elkanii, isolated from soybean nodules, in order to identify those more competitive and comprising greater nitrogen fixation ability for use in the formulation of commercial inoculants. The assay used was able to discriminate ten reference strains belonging to these two Bradyrhizobium species, as well as to efficiently identify 37 strains isolated from fields cultivated with soybean.

scholarly journals Mucosa-Associated Faecalibacterium prausnitzii Phylotype Richness Is Reduced in Patients with Inflammatory Bowel Disease

2015 ◽  
Vol 81 (21) ◽  
pp. 7582-7592 ◽  
Author(s):  
Mireia Lopez-Siles ◽  
Margarita Martinez-Medina ◽  
Carles Abellà ◽  
David Busquets ◽  
Miriam Sabat-Mir ◽  
...  
Keyword(s):  
Inflammatory Bowel Disease ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Bowel Disease ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Gastrointestinal Disease ◽  
Rrna Gene ◽  
Content Type ◽  
Faecalibacterium Prausnitzii ◽  
Inflammatory Bowel

ABSTRACTFaecalibacterium prausnitziidepletion in intestinal diseases has been extensively reported, but little is known about intraspecies variability. This work aims to determine if subjects with gastrointestinal disease host mucosa-associatedF. prausnitziipopulations different from those hosted by healthy individuals. A new species-specific PCR-denaturing gradient gel electrophoresis (PCR-DGGE) method targeting the 16S rRNA gene was developed to fingerprintF. prausnitziipopulations in biopsy specimens from 31 healthy control (H) subjects and 36 Crohn's disease (CD), 23 ulcerative colitis (UC), 6 irritable bowel syndrome (IBS), and 22 colorectal cancer (CRC) patients. The richness ofF. prausnitziisubtypes was lower in inflammatory bowel disease (IBD) patients than in H subjects. The most prevalent operational taxonomic units (OTUs) consisted of four phylotypes (OTUs with a 99% 16S rRNA gene sequence similarity [OTU99]), which were shared by all groups of patients. Their distribution and the presence of some disease-specificF. prausnitziiphylotypes allowed us to differentiate the populations in IBD and CRC patients from that in H subjects. At the level of a minimum similarity of 97% (OTU97), two phylogroups accounted for 98% of the sequences. Phylogroup I was found in 87% of H subjects but in under 50% of IBD patients (P= 0.003). In contrast, phylogroup II was detected in >75% of IBD patients and in only 52% of H subjects (P= 0.005). This study reveals that even though the main members of theF. prausnitziipopulation are present in both H subjects and individuals with gut diseases, richness is reduced in the latter and an altered phylotype distribution exists between diseases. This approach may serve as a basis for addressing the suitability ofF. prausnitziiphylotypes to be quantified as a putative biomarker of disease and depicting the importance of the loss of these subtypes in disease pathogenesis.

Characterization of bacterial community structure dynamics in a rat burn wound model using 16S rRNA gene sequencing

2022 ◽  
Author(s):  
Chen Zheng-li ◽  
Peng Yu ◽  
Wu Guo-sheng ◽  
Hong Xu-Dong ◽  
Fan Hao ◽  
...  
Keyword(s):  
Community Structure ◽  
Bacterial Community ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Bacterial Community Structure ◽  
Bacterial Species ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Sprague Dawley ◽  
Rrna Gene Sequencing

Abstract Burns destroy the skin barrier and alter the resident bacterial community, thereby facilitating bacterial infection. To treat a wound infection, it is necessary to understand the changes in the wound bacterial community structure. However, traditional bacterial cultures allow the identification of only readily growing or purposely cultured bacterial species and lack the capacity to detect changes in the bacterial community. In this study, 16S rRNA gene sequencing was used to detect alterations in the bacterial community structure in deep partial-thickness burn wounds on the back of Sprague-Dawley rats. These results were then compared with those obtained from the bacterial culture. Bacterial samples were collected prior to wounding and 1, 7, 14, and 21 days after wounding. The 16S rRNA gene sequence analysis showed that the number of resident bacterial species decreased after the burn. Both resident bacterial richness and diversity, which were significantly reduced after the burn, recovered following wound healing. The dominant resident strains also changed, but the inhibition of bacterial community structure was in a non-volatile equilibrium state, even in the early stage after healing. Furthermore, the correlation between wound and environmental bacteria increased with the occurrence of burns. Hence, the 16S rRNA gene sequence analysis reflected the bacterial condition of the wounds better than the bacterial culture. 16S rRNA sequencing in the Sprague-Dawley rat burn model can provide more information for the prevention and treatment of burn infections in clinical settings and promote further development in this field.

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