Identification of a novel bacterial sequence associated with Crohn's disease

doi:10.1053/gast.2000.8519

Gastroenterology

Volume 119, Issue 1, July 2000, Pages 23-31

https://doi.org/10.1053/gast.2000.8519 Get rights and content

Abstract

Background & Aims: Enteric microorganisms are implicated in the pathogenesis of Crohn's disease (CD), but no clear bacterial or viral species has been identified. In this study, representational difference analysis (RDA) was used to isolate DNA segments preferentially abundant in lamina propria mononuclear cells of lesional mucosa vs. adjacent uninvolved mucosa. Methods: Two RDA-derived microbial sequences were isolated (I1 and I2) and identified as novel homologues of the ptxR and tetR bacterial transcription-factor families. Results: Quantitative competitive polymerase chain reaction of paraffin-embedded intestinal specimens from 212 patients showed that I2 DNA was present in many CD colonic lesions (43%), but was infrequent in other colonic specimens (9% of ulcerative colitis lesions and 5% of non–inflammatory bowel disease diseases; P < 0.0001). I2 was prevalent in ileal specimens, regardless of disease status (43%-54%). Enzyme-linked immunosorbent assay analysis of 150 individuals with an I2 glutathione-S-transferase fusion protein showed frequent immunoglobulin A seroreactivity in CD (54% of patients), but infrequent seroreactivity in patients with ulcerative colitis, other inflammatory enteric diseases, or normals (10%, 19%, and 4%, respectively; P < 0.001 to 0.00001). Conclusions: These findings relate CD to a novel lesion-localized and immunologically associated bacterial sequence, suggesting that the microorganism expressing the I2 gene product may be related to CD pathogenesis.

GASTROENTEROLOGY 2000;119:23-31

Section snippets

Patients

Lamina propria mononuclear cells were produced from a therapeutic colectomy specimen of a single Cedars-Sinai Medical Center patient with CD. Endoscopic biopsy specimens were obtained from 7 patients with CD at UCLA Medical Center. Paraffin blocks of ileal and colonic resection specimens were obtained from the surgical pathology archives of the UCLA and Cedar-Sinai Medical Centers. All available CD cases between 1993 and 1997 were evaluated, yielding 121 (61 colonic and 60 ileal), which

Cloning of gene segments associated with histologically involved CD mucosa

RDA was performed between lamina propria mononuclear cells derived from involved (lesional) and uninvolved (nonlesional) mucosa of a single CD colectomy specimen. After 3 cycles of RDA, 4 major DNA bands were obtained from the involved mucosa (designated I1 to I4), and 2 bands from the uninvolved mucosa (designated U1 and U2) (Figure 1A).

. Representational difference analysis of involved and uninvolved CD mucosa. (A) Lamina propria mononuclear cells were prepared from involved and uninvolved

Discussion

This study describes a novel bacterial sequence, I2, isolated by subtractive cloning in lamina propria mononuclear cells from colonic mucosa histologically involved with CD. PCR analysis indicated that I2 DNA was commonly detectable in involved CD colonic mucosa, but was uncommon in control inflammatory and noninflammatory mucosa. Serum antibodies to an I2-encoded peptide were also observed to be disease associated.

Acknowledgements

The authors thank Christopher Denny for advice on RDA; Sydney Finegold for advice and efforts in bacterial culture; Gunther Harth, Diana Lester, and Marcus Horwitz for mycobacterial isolates and cultures; and Karin Reimann and Hallie Sandusky for technical assistance.

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^☆: Address requests for reprints to: Jonathan Braun, M.D., Ph.D., Department of Pathology and Laboratory Medicine, UCLA School of Medicine, CHS 13-222, Los Angeles, California 90095-1732. e-mail: [email protected]; fax: (310) 825-5674.

^☆☆: Supported by National Institutes of Health grant DK46763, DK43026, the UCLA Clinical and Fundamental Immunology Training Grant (to C. L. S. and H. D., AI 07126-23), the Crohn's and Colitis Foundation of America, the Jonnson Comprehensive Cancer Center, and the Feintech Family Chair of Inflammatory Bowel Disease.

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Alimentary TractIdentification of a novel bacterial sequence associated with Crohn's disease☆,☆☆

Abstract

Section snippets

Patients

Cloning of gene segments associated with histologically involved CD mucosa

Discussion

Acknowledgements

Gastroenterology

Gastroenterology

Gastroenterology

Res Immunol

Gastroenterology

Am J Hum Genet

Gastroenterology

Curr Opin Immunol

J Mol Biol

Biochem Biophys Res Commun

Am J Clin Nutr

Anal Biochem

J Microbiol Methods

Emerging infectious diseases: public health issues for the 21st century

Science

Crohn's disease: Occurrence, course and prognosis. An epidemiologic cohort-study

Dan Med Bull

Antibodies to Mycobacterium paratuberculosis–specific protein antigens in Crohn's disease

Clin Exp Immunol

Occurrence, in Crohn's disease, of antibodies directed against a species-specific recombinant polypeptide of Mycobacterium paratuberculosis

Clin Diag Lab Immunol

Immunoglobulin A (IgA) and IgG serum antibodies to mycobacterial antigens in Crohn's disease patients and their relatives

J Clin Microbiol

Normal luminal bacteria, especially Bacteroides species, mediate chronic colitis, gastritis, and arthritis in HLA-B27/human β2 microglobulin transgenic rats

J Clin Invest

T cell receptor-alpha,beta-deficient mice fail to develop colitis in the absence of a microbial environment

Am J Pathol

The role of the fecal stream in Crohn's disease: an historical and analytic review

Inflamm Bowel Dis

An in-depth study of Crohn's disease in two French families

Gastroenterology

Two stage genomewide search in inflammatory bowel disease provides evidence for susceptibility loci on chromosomes 3, 7 and 12

Nat Genet

Alimentary Tract
Identification of a novel bacterial sequence associated with Crohn's disease☆,☆☆