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Microbial Population Differentials between Mucosal and Submucosal Intestinal Tissues in Advanced Crohn's Disease of the Ileum.

Chiodini RJ, Dowd SE, Chamberlin WM, Galandiuk S, Davis B, Glassing A - PLoS ONE (2015)

Bottom Line: A variety of organisms of the Phylum Firmicutes were increased in the subjacent submucosa as compared to the parallel mucosal tissue including Ruminococcus spp., Oscillospira spp., Pseudobutyrivibrio spp., and Tumebacillus spp.In addition, Propionibacterium spp. and Cloacibacterium spp. were increased as well as large increases in Proteobacteria including Parasutterella spp. and Methylobacterium spp.Our data demonstrate the existence of a distinct submucosal microbiome and ecosystem that is not well reflected in the mucosa and/or downstream fecal material.

View Article: PubMed Central - PubMed

Affiliation: St. Vincent Healthcare, Sisters of Charity of Leavenworth Health System, Billings, Montana, United States of America; Department of Biological and Physical Sciences, Montana State University-Billings, Billings, Montana, United States of America.

ABSTRACT
Since Crohn's disease is a transmural disease, we hypothesized that examination of deep submucosal tissues directly involved in the inflammatory disease process may provide unique insights into bacterial populations transgressing intestinal barriers and bacterial populations more representative of the causes and agents of the disease. We performed deep 16s microbiota sequencing on isolated ilea mucosal and submucosal tissues on 20 patients with Crohn's disease and 15 non-inflammatory bowel disease controls with a depth of coverage averaging 81,500 sequences in each of the 70 DNA samples yielding an overall resolution down to 0.0001% of the bacterial population. Of the 4,802,328 total sequences generated, 98.9% or 4,749,183 sequences aligned with the Kingdom Bacteria that clustered into 8545 unique sequences with <3% divergence or operational taxonomic units enabling the identification of 401 genera and 698 tentative bacterial species. There were significant differences in all taxonomic levels between the submucosal microbiota in Crohn's disease compared to controls, including organisms of the Order Desulfovibrionales that were present within the submucosal tissues of most Crohn's disease patients but absent in the control group. A variety of organisms of the Phylum Firmicutes were increased in the subjacent submucosa as compared to the parallel mucosal tissue including Ruminococcus spp., Oscillospira spp., Pseudobutyrivibrio spp., and Tumebacillus spp. In addition, Propionibacterium spp. and Cloacibacterium spp. were increased as well as large increases in Proteobacteria including Parasutterella spp. and Methylobacterium spp. This is the first study to examine the microbial populations within submucosal tissues of patients with Crohn's disease and to compare microbial communities found deep within the submucosal tissues with those present on mucosal surfaces. Our data demonstrate the existence of a distinct submucosal microbiome and ecosystem that is not well reflected in the mucosa and/or downstream fecal material.

No MeSH data available.


Related in: MedlinePlus

Intestinal tissues examined in present study.Mucosal tissues examined by other methods, such as snip biopsy (A), predominately examine microbes of the mucosal surface; Mucosal (B) and Submucosal (C) tissues examined by the present study examined bacterial populations deep within the diseased tissues.
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pone.0134382.g001: Intestinal tissues examined in present study.Mucosal tissues examined by other methods, such as snip biopsy (A), predominately examine microbes of the mucosal surface; Mucosal (B) and Submucosal (C) tissues examined by the present study examined bacterial populations deep within the diseased tissues.

Mentions: Methods were developed to completely separate the mucosal and submucosal layers (Fig 1) of intestinal tissues and obtain representative sections beyond that accessible by simple biopsy. Tissues were processed by a modification of methods previously described [7]. An appropriate section of intestinal tissue was cut from the sample, weighed, and then vortexed in ice-cold water for 15–20 seconds at high speed to remove surface contamination. Under magnification, the mucosal layer was manually excised from the tissue and the excised mucosal layer then transferred to a microcentrifuge vial suitable for bead-beating and 1.0 ml of ice-cold 1mM DL-dithiothreitol (DTT) added and agitated for 20–30 minutes at 3000 rpm on a pulsating vortex mixer. Undissolved tissue was removed and discarded, and the supernatant centrifuged at 10,000g for 3 minutes and DNA extracted from the sediment as described below.


Microbial Population Differentials between Mucosal and Submucosal Intestinal Tissues in Advanced Crohn's Disease of the Ileum.

Chiodini RJ, Dowd SE, Chamberlin WM, Galandiuk S, Davis B, Glassing A - PLoS ONE (2015)

Intestinal tissues examined in present study.Mucosal tissues examined by other methods, such as snip biopsy (A), predominately examine microbes of the mucosal surface; Mucosal (B) and Submucosal (C) tissues examined by the present study examined bacterial populations deep within the diseased tissues.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4519195&req=5

pone.0134382.g001: Intestinal tissues examined in present study.Mucosal tissues examined by other methods, such as snip biopsy (A), predominately examine microbes of the mucosal surface; Mucosal (B) and Submucosal (C) tissues examined by the present study examined bacterial populations deep within the diseased tissues.
Mentions: Methods were developed to completely separate the mucosal and submucosal layers (Fig 1) of intestinal tissues and obtain representative sections beyond that accessible by simple biopsy. Tissues were processed by a modification of methods previously described [7]. An appropriate section of intestinal tissue was cut from the sample, weighed, and then vortexed in ice-cold water for 15–20 seconds at high speed to remove surface contamination. Under magnification, the mucosal layer was manually excised from the tissue and the excised mucosal layer then transferred to a microcentrifuge vial suitable for bead-beating and 1.0 ml of ice-cold 1mM DL-dithiothreitol (DTT) added and agitated for 20–30 minutes at 3000 rpm on a pulsating vortex mixer. Undissolved tissue was removed and discarded, and the supernatant centrifuged at 10,000g for 3 minutes and DNA extracted from the sediment as described below.

Bottom Line: A variety of organisms of the Phylum Firmicutes were increased in the subjacent submucosa as compared to the parallel mucosal tissue including Ruminococcus spp., Oscillospira spp., Pseudobutyrivibrio spp., and Tumebacillus spp.In addition, Propionibacterium spp. and Cloacibacterium spp. were increased as well as large increases in Proteobacteria including Parasutterella spp. and Methylobacterium spp.Our data demonstrate the existence of a distinct submucosal microbiome and ecosystem that is not well reflected in the mucosa and/or downstream fecal material.

View Article: PubMed Central - PubMed

Affiliation: St. Vincent Healthcare, Sisters of Charity of Leavenworth Health System, Billings, Montana, United States of America; Department of Biological and Physical Sciences, Montana State University-Billings, Billings, Montana, United States of America.

ABSTRACT
Since Crohn's disease is a transmural disease, we hypothesized that examination of deep submucosal tissues directly involved in the inflammatory disease process may provide unique insights into bacterial populations transgressing intestinal barriers and bacterial populations more representative of the causes and agents of the disease. We performed deep 16s microbiota sequencing on isolated ilea mucosal and submucosal tissues on 20 patients with Crohn's disease and 15 non-inflammatory bowel disease controls with a depth of coverage averaging 81,500 sequences in each of the 70 DNA samples yielding an overall resolution down to 0.0001% of the bacterial population. Of the 4,802,328 total sequences generated, 98.9% or 4,749,183 sequences aligned with the Kingdom Bacteria that clustered into 8545 unique sequences with <3% divergence or operational taxonomic units enabling the identification of 401 genera and 698 tentative bacterial species. There were significant differences in all taxonomic levels between the submucosal microbiota in Crohn's disease compared to controls, including organisms of the Order Desulfovibrionales that were present within the submucosal tissues of most Crohn's disease patients but absent in the control group. A variety of organisms of the Phylum Firmicutes were increased in the subjacent submucosa as compared to the parallel mucosal tissue including Ruminococcus spp., Oscillospira spp., Pseudobutyrivibrio spp., and Tumebacillus spp. In addition, Propionibacterium spp. and Cloacibacterium spp. were increased as well as large increases in Proteobacteria including Parasutterella spp. and Methylobacterium spp. This is the first study to examine the microbial populations within submucosal tissues of patients with Crohn's disease and to compare microbial communities found deep within the submucosal tissues with those present on mucosal surfaces. Our data demonstrate the existence of a distinct submucosal microbiome and ecosystem that is not well reflected in the mucosa and/or downstream fecal material.

No MeSH data available.


Related in: MedlinePlus