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. 1997 Nov;41(5):646–650. doi: 10.1136/gut.41.5.646

Search for Mycobacterium paratuberculosis DNA in orofacial granulomatosis and oral Crohn's disease tissue by polymerase chain reaction

M Riggio 1, J Gibson 1, A Lennon 1, D Wray 1, D MacDonald 1
PMCID: PMC1891551  PMID: 9414972

Abstract

Background—Although intestinal Crohn's disease has long been suspected to have a mycobacterial cause, possible mycobacterial involvement in orofacial granulomatosis (OFG) and oral lesions of Crohn's disease has not yet been investigated.
Aims—As the slow growing Mycobacterium paratuberculosis has been implicated in the aetiology of intestinal Crohn's disease, the potential involvement of this mycobacterial species in OFG and oral lesions of Crohn's disease was investigated.
Patients—To attempt detection of the organism in OFG and oral Crohn's disease tissue samples, a polymerase chain reaction (PCR) assay was used on archival formalin fixed, paraffin wax embedded oral tissue sections from 30 patients with OFG, seven with Crohn's disease, and 12 normal controls.
Methods—The PCR assay used was based on primers targeting the 5' region of the multicopy IS900 DNA insertion element of the M paratuberculosis genome. In order to achieve maximum sensitivity, two rounds of PCR were carried out and amplicons confirmed by Southern blot hybridisation to a digoxigenin labelled IS900 DNA probe.
Results—None of the OFG and oral lesions of Crohn's disease samples were positive for M paratuberculosis and all normal controls were also negative. 
Conclusions—These results suggest that M paratuberculosis may not be a major aetiological agent in OFG or oral Crohn's disease lesions, although the use of paraffin wax embedded tissue as opposed to fresh tissue as a sample source could underestimate the true prevalence of the organism. 



Keywords: oral Crohn's disease; Mycobacterium paratuberculosis; orofacial granulomatosis; polymerase chain reaction

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Figure 1 .

Figure 1

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: Agarose gel electrophoresis of PCR products (20 µl) obtained in the PCR sensitivity assay following two rounds of 40 cycles of amplification with M paratuberculosis IS900 primers P90+ and P91+. Extracted tisssue DNA from PCR negative OFG samples was spiked with serial 10-fold dilutions of M paratuberculosis DNA. Lanes 1-6, M paratuberculosis DNA at 100 pg (lane 1), 10 pg (lane 2), 1 pg (lane 3), 100 fg (lane 4), 10 fg (lane 5), 1 fg (lane 6); lane 7, 100 bp DNA ladder. 


Figure 2 .

Figure 2

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: (A) 2% agarose gel electrophoresis of selected PCR products (20 µl) obtained from tissue DNA samples following two rounds of 40 cycles of amplification with M paratuberculosis IS900 primers P90+ and P91+. Lane 1, 100 bp DNA ladder; lanes 2-8, OFG samples; lanes 9-11, oral Crohn's disease samples; lanes 12-14, normal samples; lane 15, negative PCR control; lane 16, positive PCR control. The single PCR product obtained from the samples analysed, which is slightly smaller in size than the positive control PCR product, is shown in lane 6. (B) Corresponding Southern blot hybridisation. For orientation of the membrane, lanes 1 and 16 correspond to the 100 bp DNA ladder and PCR positive control lanes, respectively. The PCR product in lane 6 did not hybridise to the probe. 


Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Asquith P., Thompson R. A., Cooke W. T. Oral manifestations of Crohn's disease. Gut. 1975 Apr;16(4):249–254. doi: 10.1136/gut.16.4.249. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Chiodini R. J., Van Kruiningen H. J., Merkal R. S. Ruminant paratuberculosis (Johne's disease): the current status and future prospects. Cornell Vet. 1984 Jul;74(3):218–262. [PubMed] [Google Scholar]
  3. Cook S. M., Bartos R. E., Pierson C. L., Frank T. S. Detection and characterization of atypical mycobacteria by the polymerase chain reaction. Diagn Mol Pathol. 1994 Mar;3(1):53–58. doi: 10.1097/00019606-199403010-00009. [DOI] [PubMed] [Google Scholar]
  4. Croft C. B., Wilkinson A. R. Ulceration of the mouth, pharynx, and larynx in Crohn's disease of the intestine. Br J Surg. 1972 Apr;59(4):249–252. doi: 10.1002/bjs.1800590402. [DOI] [PubMed] [Google Scholar]
  5. Dell'Isola B., Poyart C., Goulet O., Mougenot J. F., Sadoun-Journo E., Brousse N., Schmitz J., Ricour C., Berche P. Detection of Mycobacterium paratuberculosis by polymerase chain reaction in children with Crohn's disease. J Infect Dis. 1994 Feb;169(2):449–451. doi: 10.1093/infdis/169.2.449. [DOI] [PubMed] [Google Scholar]
  6. Dudeney T. P. Crohn's disease of the mouth. Proc R Soc Med. 1969 Dec 12;62(12):1237–1237. [PMC free article] [PubMed] [Google Scholar]
  7. Dumonceau J. M., Van Gossum A., Adler M., Fonteyne P. A., Van Vooren J. P., Deviere J., Portaels F. No Mycobacterium paratuberculosis found in Crohn's disease using polymerase chain reaction. Dig Dis Sci. 1996 Feb;41(2):421–426. doi: 10.1007/BF02093838. [DOI] [PubMed] [Google Scholar]
  8. Fidler H. M., Thurrell W., Johnson N. M., Rook G. A., McFadden J. J. Specific detection of Mycobacterium paratuberculosis DNA associated with granulomatous tissue in Crohn's disease. Gut. 1994 Apr;35(4):506–510. doi: 10.1136/gut.35.4.506. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Frank T. S., Cook S. M. Analysis of paraffin sections of Crohn's disease for Mycobacterium paratuberculosis using polymerase chain reaction. Mod Pathol. 1996 Jan;9(1):32–35. [PubMed] [Google Scholar]
  10. Frank T. S., Cook S. M., Del Buono E. A., Wilson M. D. A simplified method for detecting cytomegalovirus by polymerase chain reaction from histologic sections of small biopsies. Mod Pathol. 1992 Jul;5(4):449–454. [PubMed] [Google Scholar]
  11. Ghandour K., Issa M. Oral Crohn's disease with late intestinal manifestations. Oral Surg Oral Med Oral Pathol. 1991 Nov;72(5):565–567. doi: 10.1016/0030-4220(91)90495-x. [DOI] [PubMed] [Google Scholar]
  12. Green E. P., Tizard M. L., Moss M. T., Thompson J., Winterbourne D. J., McFadden J. J., Hermon-Taylor J. Sequence and characteristics of IS900, an insertion element identified in a human Crohn's disease isolate of Mycobacterium paratuberculosis. Nucleic Acids Res. 1989 Nov 25;17(22):9063–9073. doi: 10.1093/nar/17.22.9063. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lisby G., Andersen J., Engbaek K., Binder V. Mycobacterium paratuberculosis in intestinal tissue from patients with Crohn's disease demonstrated by a nested primer polymerase chain reaction. Scand J Gastroenterol. 1994 Oct;29(10):923–929. doi: 10.3109/00365529409094864. [DOI] [PubMed] [Google Scholar]
  14. Lisby G., Milman N., Jacobsen G. K. Search for Mycobacterium paratuberculosis DNA in tissue from patients with sarcoidosis by enzymatic gene amplification. APMIS. 1993 Nov;101(11):876–878. doi: 10.1111/j.1699-0463.1993.tb00195.x. [DOI] [PubMed] [Google Scholar]
  15. Millar D. S., Withey S. J., Tizard M. L., Ford J. G., Hermon-Taylor J. Solid-phase hybridization capture of low-abundance target DNA sequences: application to the polymerase chain reaction detection of Mycobacterium paratuberculosis and Mycobacterium avium subsp. silvaticum. Anal Biochem. 1995 Apr 10;226(2):325–330. doi: 10.1006/abio.1995.1232. [DOI] [PubMed] [Google Scholar]
  16. Mitchell I. C., Turk J. L., Mitchell D. N. Detection of mycobacterial rRNA in sarcoidosis with liquid-phase hybridisation. Lancet. 1992 Apr 25;339(8800):1015–1017. doi: 10.1016/0140-6736(92)90536-c. [DOI] [PubMed] [Google Scholar]
  17. Rowbotham D. S., Mapstone N. P., Trejdosiewicz L. K., Howdle P. D., Quirke P. Mycobacterium paratuberculosis DNA not detected in Crohn's disease tissue by fluorescent polymerase chain reaction. Gut. 1995 Nov;37(5):660–667. doi: 10.1136/gut.37.5.660. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Saboor S. A., Johnson N. M., McFadden J. Detection of mycobacterial DNA in sarcoidosis and tuberculosis with polymerase chain reaction. Lancet. 1992 Apr 25;339(8800):1012–1015. doi: 10.1016/0140-6736(92)90535-b. [DOI] [PubMed] [Google Scholar]
  19. Sanderson J. D., Moss M. T., Tizard M. L., Hermon-Taylor J. Mycobacterium paratuberculosis DNA in Crohn's disease tissue. Gut. 1992 Jul;33(7):890–896. doi: 10.1136/gut.33.7.890. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Schiller K. F., Golding P. L., Peebles R. A., Whitehead R. Crohn's disease of the mouth and lips. Gut. 1971 Oct;12(10):864–865. [PubMed] [Google Scholar]
  21. Vary P. H., Andersen P. R., Green E., Hermon-Taylor J., McFadden J. J. Use of highly specific DNA probes and the polymerase chain reaction to detect Mycobacterium paratuberculosis in Johne's disease. J Clin Microbiol. 1990 May;28(5):933–937. doi: 10.1128/jcm.28.5.933-937.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wiesenfeld D., Ferguson M. M., Mitchell D. N., MacDonald D. G., Scully C., Cochran K., Russell R. I. Oro-facial granulomatosis--a clinical and pathological analysis. Q J Med. 1985 Jan;54(213):101–113. [PubMed] [Google Scholar]

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