Analysis of the Intestinal Microbiota in Autistic Patients by Denaturing Gradient Gel Electrophoresis

2018 ◽  
Vol 10 (3) ◽  
pp. 440-446
Author(s):  
Na Li ◽  
Honglin Zhang ◽  
Renmin Zhang ◽  
Zhiyu Bai ◽  
Zhimao Bai ◽  
...  
Keyword(s):  
Intestinal Microbiota ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Study Groups ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Control Group ◽  
Healthy Children ◽  
Microbial Composition ◽  
Gradient Gel Electrophoresis

To analyze the differences in the composition of the intestinal microbiota between the autistic and healthy children, we selected 45 autistic and 20 control children aged 2 to 9 years to collect their fecal samples. The total microbial genome DNA of each fecal sample was extracted, and the V3 regions of microbial 16S rRNA genes were amplified. The intestinal microbial composition of both study groups was detected by PCR-based denaturing gradient gel electrophoresis. Quantity One and Biodap software were used to analyze the diversity and similarity of bacterial populations, and SPSS software was used for statistical analysis. The denaturant gradient gel electrophoresis profiles documented significant differences in the composition of intestinal microflora between the autism and control groups. Analysis of the excised bands demonstrated the abundance of bacteria species assigned to the genus Escherichia/Shigella in the gastrointestinal tract of the autism group but a low content in the control group. An opposite result was obtained for the Bacteroides genus. These data indicate that intestinal microbial composition may is correlated with the occurrence of the autism.

scholarly journals Effects of Lactococcus lactis on Composition of Intestinal Microbiota: Role of Nisin

2006 ◽  
Vol 72 (1) ◽  
pp. 239-244 ◽  
Author(s):  
Nete Bernbom ◽  
Tine Rask Licht ◽  
Carl-Henrik Brogren ◽  
Birthe Jelle ◽  
Anette H. Johansen ◽  
...  
Keyword(s):  
Lactococcus Lactis ◽  
Intestinal Microbiota ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Fecal Microbiota ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Enzyme Linked Immunosorbent Assay ◽  
Gradient Gel Electrophoresis ◽  
Intact Molecule ◽  
Human Flora

ABSTRACT This study examined the ability of (i) pure nisin, (ii) nisin-producing Lactococcus lactis strain CHCC5826, and (iii) the non-nisin-producing L. lactis strain CHCH2862 to affect the composition of the intestinal microbiota of human flora-associated rats. The presence of both the nisin-producing and the non-nisin-producing L. lactis strains significantly increased the number of Bifidobacterium cells in fecal samples during the first 8 days but decreased the number of enterococci/streptococci in duodenum, ileum, cecum, and colon samples as detected by selective cultivation. No significant changes in the rat fecal microbiota were observed after dosage with nisin. Pearson cluster analysis of denaturing gradient gel electrophoresis profiles of the 16S rRNA genes present in the fecal microbial population revealed that the microbiota of animals dosed with either of the two L. lactis strains were different from that of control animals dosed with saline. However, profiles of the microbiota from animals dosed with nisin did not differ from the controls. The concentrations of nisin estimated by competitive enzyme-linked immunosorbent assay (ELISA) were approximately 10-fold higher in the small intestine and 200-fold higher in feces than the corresponding concentrations estimated by a biological assay. This indicates that nisin was degraded or inactivated in the gastrointestinal tract, since fragments of this bacteriocin are detected by ELISA while an intact molecule is needed to retain biological activity.

scholarly journals Comparison of Different Denaturing Gradient Gel Electrophoresis Primer Sets for the Study of Marine Bacterioplankton Communities

2007 ◽  
Vol 73 (18) ◽  
pp. 5962-5967 ◽  
Author(s):  
Olga Sánchez ◽  
Josep M. Gasol ◽  
Ramon Massana ◽  
Jordi Mas ◽  
Carlos Pedrós-Alió
Keyword(s):  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Coastal System ◽  
Marine Bacterioplankton ◽  
Gradient Gel Electrophoresis ◽  
Primer Sets ◽  
Bacterial Groups ◽  
Plankton Communities

ABSTRACT An annual seasonal cycle of composition of a bacterioplankton community in an oligotrophic coastal system was studied by denaturing gradient gel electrophoresis (DGGE) using five different primer sets. Analysis of DGGE fingerprints showed that primer set 357fGC-907rM grouped samples according to seasons. Additionally, we used the set of 16S rRNA genes archived in the RDPII database to check the percentage of perfect matches of each primer for the most abundant bacterial groups inhabiting coastal plankton communities. Overall, primer set 357fGC-907rM was the most suitable for the routine use of PCR-DGGE analyses in this environment.

Genetic Diversity of the Biofilm CoveringMontacuta ferruginosa (Mollusca, Bivalvia) as Evaluated by Denaturing Gradient Gel Electrophoresis Analysis and Cloning of PCR-Amplified Gene Fragments Coding for 16S rRNA†

1998 ◽  
Vol 64 (9) ◽  
pp. 3464-3472 ◽  
Author(s):  
David C. Gillan ◽  
Arjen G. C. L. Speksnijder ◽  
Gabriel Zwart ◽  
Chantal De Ridder
Keyword(s):  
Genetic Diversity ◽  
16S Rrna ◽  
Dna Extraction ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Gradient Gel Electrophoresis ◽  
Biofilm Bacteria ◽  
Sequence Types

The shell of the bivalve Montacuta ferruginosa, a symbiont living in the burrow of an echinoid, is covered with a rust-colored biofilm. This biofilm includes different morphotypes of bacteria that are encrusted with a mineral rich in ferric ion and phosphate. The aim of this research was to determine the genetic diversity and phylogenetic affiliation of the biofilm bacteria. Also, the possible roles of the microorganisms in the processes of mineral deposition within the biofilm, as well as their impact on the biology of the bivalve, were assessed by phenotypic inference. The genetic diversity was determined by denaturing gradient gel electrophoresis (DGGE) analysis of short (193-bp) 16S ribosomal DNA PCR products obtained with primers specific for the domain Bacteria. This analysis revealed a diverse consortium; 11 to 25 sequence types were detected depending on the method of DNA extraction used. Individual biofilms analyzed by using the same DNA extraction protocol did not produce identical DGGE profiles. However, different biofilms shared common bands, suggesting that similar bacteria can be found in different biofilms. The phylogenetic affiliations of the sequence types were determined by cloning and sequencing the 16S rRNA genes. Close relatives of the genera Pseudoalteromonas,Colwellia, and Oceanospirillum (members of the γ-Proteobacteria lineage), as well as Flexibacter maritimus (a member of theCytophaga-Flavobacter-Bacteroides lineage), were found in the biofilms. We inferred from the results that some of the biofilm bacteria could play a role in the mineral formation processes.

scholarly journals Denaturing Gradient Gel Electrophoresis and Barcoded Pyrosequencing Reveal Unprecedented Archaeal Diversity in Mangrove Sediment and Rhizosphere Samples

2012 ◽  
Vol 78 (16) ◽  
pp. 5520-5528 ◽  
Author(s):  
Ana C. C. Pires ◽  
Daniel F. R. Cleary ◽  
Adelaide Almeida ◽  
Ângela Cunha ◽  
Simone Dealtry ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Mangrove Sediment ◽  
Bulk Sediment ◽  
Archaeal Diversity ◽  
Mangrove Species ◽  
Content Type ◽  
Gradient Gel Electrophoresis

ABSTRACTMangroves are complex ecosystems that regulate nutrient and sediment fluxes to the open sea. The importance of bacteria and fungi in regulating nutrient cycles has led to an interest in their diversity and composition in mangroves. However, very few studies have assessedArchaeain mangroves, and virtually nothing is known about whether mangrove rhizospheres affect archaeal diversity and composition. Here, we studied the diversity and composition ofArchaeain mangrove bulk sediment and the rhizospheres of two mangrove trees,Rhizophora mangleandLaguncularia racemosa, using denaturing gradient gel electrophoresis (DGGE) and pyrosequencing of archaeal 16S rRNA genes with a nested-amplification approach. DGGE profiles revealed significant structural differences between bulk sediment and rhizosphere samples, suggesting that roots of both mangrove species influence the sediment archaeal community. Nearly all of the detected sequences obtained with pyrosequencing were identified asArchaea, but most were unclassified at the level of phylum or below. Archaeal richness was, furthermore, the highest in theL. racemosarhizosphere, intermediate in bulk sediment, and the lowest in theR. manglerhizosphere. This study shows that rhizosphere microhabitats ofR. mangleandL. racemosa, common plants in subtropical mangroves located in Rio de Janeiro, Brazil, hosted distinct archaeal assemblages.

Analyses of microbial activity in biomass-recycle reactors using denaturing gradient gel electrophoresis of 16S rDNA and 16S rRNA PCR products

2002 ◽  
Vol 48 (4) ◽  
pp. 333-341 ◽  
Author(s):  
Christine A Morgan ◽  
Andre Hudson ◽  
Allan Konopka ◽  
Cindy H Nakatsu
Keyword(s):  
Community Structure ◽  
16S Rrna ◽  
16S Rdna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Community Response ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Specific Content ◽  
Gradient Gel Electrophoresis

The relationship between mixed microbial community structure and physiology when grown under substrate-limited conditions was investigated using continuous-flow bioreactors with 100% biomass recycle. Community structure was analyzed by denaturing gradient gel electrophoresis (DGGE) of the PCR and RT-PCR amplified V3 region of 16S rDNA and 16S rRNA templates, respectively. Comparisons were made of communities exposed to different types of transient conditions (e.g., long- and short-term starvation, increasing nutrients). With progressively more stringent substrate limitation over time, the specific content of community RNA declined by more than 10-fold and closely followed the decline in specific growth rate. In contrast, the DNA content was variable (up to 3-fold differences) and did not follow the same trend. Cluster analysis of the presence or absence of individual bands indicated that the fingerprints generated by the two templates were different, and community response was first observed in the rRNA fraction. However, both the rDNA and rRNA fingerprints provided a picture of temporal population dynamics. Dice similarity coefficients gave a quantitative measure of the differences and changes between the communities. In comparison, standard cultivation techniques yielded only a quarter of the phylotypes detected by DGGE, but included the most dominant population based on rRNA. Nucleotide-sequence analyses of the almost complete 16S rRNA genes of these isolates place them in the same group of organisms that is typically cultivated from environmental samples: α, β, and γ Proteobacteria and the high GC and the low GC Gram-positive divisions.Key words: 16S rRNA, DGGE, community analysis, biomass-recycle reactor.

scholarly journals Development and Application of a Selective PCR-Denaturing Gradient Gel Electrophoresis Approach To Detect a Recently Cultivated Bacillus Group Predominant in Soil

2004 ◽  
Vol 70 (10) ◽  
pp. 5801-5809 ◽  
Author(s):  
Vesela A. Tzeneva ◽  
Youguo Li ◽  
Andreas D. M. Felske ◽  
Willem M. de Vos ◽  
Antoon D. L. Akkermans ◽  
...  
Keyword(s):  
16S Rrna ◽  
The Netherlands ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Physiological State ◽  
Soil Samples ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Gradient Gel Electrophoresis

ABSTRACT The worldwide presence of a hitherto-nondescribed group of predominant soil microorganisms related to Bacillus benzoevorans was analyzed after development of two sets of selective primers targeting 16S rRNA genes in combination with denaturing gradient gel electrophoresis (DGGE). The high abundance and cultivability of at least some of these microorganisms makes them an appropriate subject for studies on their biogeographical dissemination and diversity. Since cultivability can vary significantly with the physiological state and even between closely related strains, we developed a culture-independent 16S rRNA gene-targeted DGGE fingerprinting protocol for the detection of these bacteria from soil samples. The composition of the B. benzoevorans relatives in the soil samples from The Netherlands, Bulgaria, Russia, Pakistan, and Portugal showed remarkable differences between the different countries. Differences in the DGGE profiles of these communities in archived soil samples from the Dutch Wieringermeer polder were observed over time during which a shift from anaerobic to aerobic and from saline to freshwater conditions occurred. To complement the molecular methods, we additionally cultivated B. benzoevorans-related strains from all of the soil samples. The highest number of B. benzoevorans relatives was found in the soils from the northern part of The Netherlands. The present study contributes to our knowledge of the diversity and abundance of this interesting group of microbes in soils throughout the world.

scholarly journals Evaluations of Different Hypervariable Regions of Archaeal 16S rRNA Genes in Profiling of Methanogens by Archaea-Specific PCR and Denaturing Gradient Gel Electrophoresis

2007 ◽  
Vol 74 (3) ◽  
pp. 889-893 ◽  
Author(s):  
Zhongtang Yu ◽  
Rubén García-González ◽  
Floyd L. Schanbacher ◽  
Mark Morrison
Keyword(s):  
16S Rrna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Specific Pcr ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
V Region

ABSTRACT Different hypervariable (V) regions of the archaeal 16S rRNA gene (rrs) were compared systematically to establish a preferred V region(s) for use in Archaea-specific PCR-denaturing gradient gel electrophoresis (DGGE). The PCR products of the V3 region produced the most informative DGGE profiles and permitted identification of common methanogens from rumen samples from sheep. This study also showed that different methanogens might be detected when different V regions are targeted by PCR-DGGE. Dietary fat appeared to transiently stimulate Methanosphaera stadtmanae but inhibit Methanobrevibacter sp. strain AbM4 in rumen samples.

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