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 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.

scholarly journals Phenotypic characterization of Astragalus glycyphyllos symbionts and their phylogeny based on the 16S rDNA sequences and RFLP of 16S rRNA gene

2014 ◽  
Vol 105 (6) ◽  
pp. 1033-1048 ◽  
Author(s):  
Sebastian Gnat ◽  
Magdalena Wójcik ◽  
Sylwia Wdowiak-Wróbel ◽  
Michał Kalita ◽  
Aneta Ptaszyńska ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Phenotypic Characterization ◽  
Rrna Gene ◽  
Rdna Sequences ◽  
16S Rdna Sequences ◽  
Astragalus Glycyphyllos Symbionts

scholarly journals Sulfate-Reducing Bacteria in Tubes Constructed by the Marine Infaunal Polychaete Diopatra cuprea

2004 ◽  
Vol 70 (12) ◽  
pp. 7053-7065 ◽  
Author(s):  
George Y. Matsui ◽  
David B. Ringelberg ◽  
Charles R. Lovell
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Microbial Communities ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Phospholipid Fatty Acid ◽  
Rrna Gene ◽  
Sandy Sediment ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Sulfate Reducing

ABSTRACT Marine infaunal burrows and tubes greatly enhance solute transport between sediments and the overlying water column and are sites of elevated microbial activity. Biotic and abiotic controls of the compositions and activities of burrow and tube microbial communities are poorly understood. The microbial communities in tubes of the marine infaunal polychaete Diopatria cuprea collected from two different sediment habitats were examined. The bacterial communities in the tubes from a sandy sediment differed from those in the tubes from a muddy sediment. The difference in community structure also extended to the sulfate-reducing bacterial (SRB) assemblage, although it was not as pronounced for this functional group of species. PCR-amplified 16S rRNA gene sequences recovered from Diopatra tube SRB by clonal library construction and screening were all related to the family Desulfobacteriaceae. This finding was supported by phospholipid fatty acid analysis and by hybridization of 16S rRNA probes specific for members of the genera Desulfosarcina, Desulfobacter, Desulfobacterium, Desulfobotulus, Desulfococcus, and Desulfovibrio and some members of the genera Desulfomonas, Desulfuromonas, and Desulfomicrobium with 16S rRNA gene sequences resolved by denaturing gradient gel electrophoresis. Two of six SRB clones from the clone library were not detected in tubes from the sandy sediment. The habitat in which the D. cuprea tubes were constructed had a strong influence on the tube bacterial community as a whole, as well as on the SRB assemblage.

scholarly journals Characterization of a Culturable Alphaproteobacterial Symbiont Common to Many Marine Sponges and Evidence for Vertical Transmission via Sponge Larvae

2006 ◽  
Vol 72 (5) ◽  
pp. 3724-3732 ◽  
Author(s):  
Julie J. Enticknap ◽  
Michelle Kelly ◽  
Olivier Peraud ◽  
Russell T. Hill
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Marine Sponges ◽  
Rrna Gene ◽  
Surrounding Water ◽  
Axinella Corrugata ◽  
Gradient Gel Electrophoresis ◽  
Related Group

ABSTRACT A closely related group of alphaproteobacteria were found to be present in seven genera of marine sponges from several locations and were shown to be transferred between sponge generations through the larvae in one of these sponges. Isolates of the alphaproteobacterium were cultured from the sponges Axinella corrugata, Mycale laxissima, Monanchora unguifera, and Niphates digitalis from Key Largo, Florida; Didiscus oxeata and Monanchora unguifera from Discovery Bay, Jamaica; an Acanthostronglyophora sp. from Manado, Indonesia; and Microciona prolifera from the Cheasapeake Bay in Maryland. Isolates were very similar to each other on the basis of 16S rRNA gene sequence (>99% identity) and are closely related to Pseudovibrio denitrificans. The bacterium was never isolated from surrounding water samples and was cultured from larvae of M. laxissima, indicating that it is a vertically transmitted symbiont in this sponge. Denaturing gradient gel electrophoresis, 16S rRNA gene clone library analysis, and fluorescent in situ hybridization with probes specific to the alphaproteobacterium confirmed the presence of this bacterium in the M. laxissima larvae. The alphaproteobacterium was densely associated with the larvae rather than being evenly distributed throughout the mesohyl. This is the first report of the successful culture of a bacterial symbiont of a sponge that is transferred through the gametes.

Impact of a probiotic, inulin, or their combination on the piglets’ microbiota at different intestinal locations

2015 ◽  
Vol 6 (4) ◽  
pp. 473-483 ◽  
Author(s):  
V.A. Sattler ◽  
K. Bayer ◽  
G. Schatzmayr ◽  
A.G. Haslberger ◽  
V. Klose
Keyword(s):  
16S Rrna ◽  
Gut Microbiota ◽  
16S Rrna Gene ◽  
Real Time ◽  
Relative Abundance ◽  
Real Time Pcr ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Feed Additives ◽  
Rrna Gene ◽  
16S Rrna Gene Pyrosequencing

Natural feed additives are used to maintain health and to promote performance of pigs without antibiotics. Effects of a probiotic, inulin, and their combination (synbiotic), on the microbial diversity and composition at different intestinal locations were analysed using denaturing gradient gel electrophoresis (DGGE), real-time PCR, and 16S rRNA gene pyrosequencing. Bacterial diversity assessed by DGGE and/or pyrosequencing was increased by inulin in all three gut locations and by the synbiotic in the caecum and colon. In contrast, the probiotic did only affect the microbiota diversity in the ileum. Shifts in the DGGE microbiota profiles of the caecum and colon were detected for the pro- and synbiotic fed animals, whereas inulin profiles were more similar to the ones of the control. 16S rRNA gene pyrosequencing revealed that all three additives could reduce Escherichia species in each gut location, indicating a potential beneficial effect on the gut microbiota. An increase of relative abundance of Clostridiaceae in the large intestine was found in the inulin group and of Enterococcaceae in the ileum of probiotic fed pigs. Furthermore, real-time PCR results showed that the probiotic and synbiotic increased bifidobacterial numbers in the ileum, which was supported by sequencing results. The probiotic and inulin, to different extents, changed the diversity, relative abundance of phylotypes, and community profiles of the porcine microbiota. However, alterations of the bacterial community were not uniformly between gut locations, demonstrating that functionality of feed additives is site specific. Therefore, gut sampling from various locations is crucial when investigations aim to identify the composition of a healthy gut microbiota after its manipulation through feed additives.

scholarly journals Cryptic haplotypes of "Candidatus Liberibacter africanus"

2015 ◽  
Author(s):  
Warrick Nelson ◽  
Sandrine Eveillard ◽  
Marie-Pierre Dubrana ◽  
Joseph Bové
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Rdna Sequences ◽  
New Information ◽  
Candidatus Liberibacter ◽  
Citrus Leaves

“Candidatus Liberibacter africanus” (Laf) has long been recognised as a causal agent of the devastating citrus disease huanglongbing (HLB) or citrus greening. This species is currently restricted to Africa, the Arabian Peninsula and some Indian Ocean islands and vectored by the African citrus psyllid, Trioza erytreae. Blotchy mottle on citrus leaves is characteristic of the disease. Somewhat similar symptoms in the Rutaceous tree Calodendrum capensis (Cape Chestnut) resulted in the discovery of Laf outside commercial citrus crops in South Africa. This was classed as a subspecies of Laf (capensis, hence LafC). In subsequent surveys of both commercial citrus crops and Calodendrum, both natural and ornamental specimens, LafC was not found in the citrus crop, nor has Laf been found in C. capensis. HLB was reported from Madagascar in 1968 but no sequences from this source have so far been published. Until fairly recently, only the reference 16S rRNA gene sequences of Laf (L22533) and LafC (AF137368) had been deposited in GenBank. Both of these reference sequences contain a number of unresolved nucleotides. Resolving these nucleotide positions by aligning against more recently available sequences, it becomes evident that these unresolved positions represent one percentage point difference in similarity between Laf and LafC. The originally reported 97.4% similarity is therefore incorrect based on this new information. Recalculating the similarity on the full length 16S rDNA sequence results in 99.54% similarity, a value too high to justify a subspecies status. LafC should therefore be reduced to that of a haplotype of Laf. Further, the six 16S rRNA gene sequences currently available in GenBank identified as the species Laf separate into 2 haplotype groups. The 3 haplotypes of Laf are therefore LafA designated as the first accession sequenced (L22533), LafC for the former capensis subspecies and to recognise the prior use of this term, and LafB for the third haplotype not previously recognised. Thus the cryptic presence of 3 haplotypes is revealed by this review of the Laf 16S rDNA sequences.

scholarly journals Differentiation of Phylogenetically Related Slowly Growing Mycobacteria Based on 16S-23S rRNA Gene Internal Transcribed Spacer Sequences

1998 ◽  
Vol 36 (1) ◽  
pp. 139-147 ◽  
Author(s):  
Andreas Roth ◽  
Marga Fischer ◽  
Mohamed E. Hamid ◽  
Sabine Michalke ◽  
Wolfgang Ludwig ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Internal Transcribed Spacer ◽  
Sequence Divergence ◽  
Comparative Sequence Analysis ◽  
Its Sequences ◽  
23S Rrna ◽  
Rrna Gene ◽  
Its Sequencing

Interspecific polymorphisms of the 16S rRNA gene (rDNA) are widely used for species identification of mycobacteria. 16S rDNA sequences, however, do not vary greatly within a species, and they are either indistinguishable in some species, for example, in Mycobacterium kansasii and M. gastri, or highly similar, for example, in M. malmoense and M. szulgai. We determined 16S-23S rDNA internal transcribed spacer (ITS) sequences of 60 strains in the genus Mycobacterium representing 13 species (M. avium, M. conspicuum, M. gastri, M. genavense, M. kansasii,M. malmoense, M. marinum, M. shimoidei, M. simiae, M. szulgai,M. triplex, M. ulcerans, and M. xenopi). An alignment of these sequences together with additional sequences available in the EMBL database (for M. intracellulare, M. phlei, M. smegmatis, and M. tuberculosis) was established according to primary- and secondary-structure similarities. Comparative sequence analysis applying different treeing methods grouped the strains into species-specific clusters with low sequence divergence between strains belonging to the same species (0 to 2%). The ITS-based tree topology only partially correlated to that based on 16S rDNA, but the main branching orders were preserved, notably, the division of fast-growing from slowly growing mycobacteria, separate branching for M. simiae, M. genavense, and M. triplex, and distinct branches for M. xenopi and M. shimoidei. Comparisons of M. gastri with M. kansasii and M. malmoense with M. szulgairevealed ITS sequence similarities of 93 and 88%, respectively.M. marinum and M. ulcerans possessed identical ITS sequences. Our results show that ITS sequencing represents a supplement to 16S rRNA gene sequences for the differentiation of closely related species. Slowly growing mycobacteria show a high sequence variation in the ITS; this variation has the potential to be used for the development of probes as a rapid approach to mycobacterial identification.

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