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Clinical and Diagnostic Laboratory Immunology logoLink to Clinical and Diagnostic Laboratory Immunology
. 1996 Jan;3(1):66–72. doi: 10.1128/cdli.3.1.66-72.1996

PCR detection of colonization by Helicobacter pylori in conventional, euthymic mice based on the 16S ribosomal gene sequence.

J G Smith 1, L Kong 1, G K Abruzzo 1, C J Gill 1, A M Flattery 1, P M Scott 1, D Bramhill 1, C Cioffe 1, C M Thompson 1, K Bartizal 1
PMCID: PMC170249  PMID: 8770506

Abstract

Many animal models of Helicobacter infection have been described, including infection in rhesus monkeys, ferrets, gnotobiotic piglets, and mice. These animal models utilize a combination of detection methods, including culture, urease testing, and histopathology, all of which may be unreliable, insensitive, or labor-intensive. Development of new animal models of Helicobacter pylori requires new methods of detection with increased sensitivity and specificity. We have developed sensitive and specific PCR primers based on the 16S ribosomal gene sequence of H. pylori. The primers detected single-copy 16S DNA representing 0.2 cell of pure H. pylori (2 cells in the presence of mouse stomach mucosal DNA) and did not cross-react with closely related bacteria. We were able to detect colonization by H. pylori in conventional, euthymic, outbred mice up to 4 weeks postinoculation with a high percentage of isolates tested. One isolate of H. pylori was detected by PCR in 100% of the mice at 6 months and 60% of the mice 1 year after inoculation. Approximately 10(3) to 10(4) H. pylori cells per stomach were detected by utilizing this PCR methodology semiquantitatively. These primers and PCR methodology have facilitated detection of H. pylori colonization in conventional, euthymic mice, colonization which may not have been detectable by other methods.

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Selected References

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  1. Andersen L. P., Holck S., Povlsen C. O., Elsborg L., Justesen T. Campylobacter pyloridis in peptic ulcer disease. I. Gastric and duodenal infection caused by C. pyloridis: histopathologic and microbiologic findings. Scand J Gastroenterol. 1987 Mar;22(2):219–224. doi: 10.3109/00365528708991883. [DOI] [PubMed] [Google Scholar]
  2. Baskerville A., Newell D. G. Naturally occurring chronic gastritis and C pylori infection in the rhesus monkey: a potential model for gastritis in man. Gut. 1988 Apr;29(4):465–472. doi: 10.1136/gut.29.4.465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bhatia S. J., Kochar N., Abraham P., Nair N. G., Mehta A. P. Lactobacillus acidophilus inhibits growth of Campylobacter pylori in vitro. J Clin Microbiol. 1989 Oct;27(10):2328–2330. doi: 10.1128/jcm.27.10.2328-2330.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Blomberg L., Henriksson A., Conway P. L. Inhibition of adhesion of Escherichia coli K88 to piglet ileal mucus by Lactobacillus spp. Appl Environ Microbiol. 1993 Jan;59(1):34–39. doi: 10.1128/aem.59.1.34-39.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Booth L., Holdstock G., MacBride H., Hawtin P., Gibson J. R., Ireland A., Bamforth J., DuBoulay C. E., Lloyd R. S., Pearson A. D. Clinical importance of Campylobacter pyloridis and associated serum IgG and IgA antibody responses in patients undergoing upper gastrointestinal endoscopy. J Clin Pathol. 1986 Feb;39(2):215–219. doi: 10.1136/jcp.39.2.215. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Buck G. E. Campylobacter pylori and gastroduodenal disease. Clin Microbiol Rev. 1990 Jan;3(1):1–12. doi: 10.1128/cmr.3.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Buck G. E., Gourley W. K., Lee W. K., Subramanyam K., Latimer J. M., DiNuzzo A. R. Relation of Campylobacter pyloridis to gastritis and peptic ulcer. J Infect Dis. 1986 Apr;153(4):664–669. doi: 10.1093/infdis/153.4.664. [DOI] [PubMed] [Google Scholar]
  8. Cantorna M. T., Balish E. Inability of human clinical strains of Helicobacter pylori to colonize the alimentary tract of germfree rodents. Can J Microbiol. 1990 Apr;36(4):237–241. doi: 10.1139/m90-041. [DOI] [PubMed] [Google Scholar]
  9. Chen M., Lee A., Hazell S. Immunisation against gastric helicobacter infection in a mouse/Helicobacter felis model. Lancet. 1992 May 2;339(8801):1120–1121. doi: 10.1016/0140-6736(92)90720-n. [DOI] [PubMed] [Google Scholar]
  10. Dick-Hegedus E., Lee A. Use of a mouse model to examine anti-Helicobacter pylori agents. Scand J Gastroenterol. 1991 Sep;26(9):909–915. doi: 10.3109/00365529108996241. [DOI] [PubMed] [Google Scholar]
  11. Doidge C., Crust I., Lee A., Buck F., Hazell S., Manne U. Therapeutic immunisation against Helicobacter infection. Lancet. 1994 Apr 9;343(8902):914–915. doi: 10.1016/s0140-6736(94)90032-9. [DOI] [PubMed] [Google Scholar]
  12. Drazek E. S., Dubois A., Holmes R. K. Characterization and presumptive identification of Helicobacter pylori isolates from rhesus monkeys. J Clin Microbiol. 1994 Jul;32(7):1799–1804. doi: 10.1128/jcm.32.7.1799-1804.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dubois A., Fiala N., Heman-Ackah L. M., Drazek E. S., Tarnawski A., Fishbein W. N., Perez-Perez G. I., Blaser M. J. Natural gastric infection with Helicobacter pylori in monkeys: a model for spiral bacteria infection in humans. Gastroenterology. 1994 Jun;106(6):1405–1417. doi: 10.1016/0016-5085(94)90392-1. [DOI] [PubMed] [Google Scholar]
  14. Eaton K. A., Morgan D. R., Krakowka S. Persistence of Helicobacter pylori in conventionalized piglets. J Infect Dis. 1990 Jun;161(6):1299–1301. doi: 10.1093/infdis/161.6.1299. [DOI] [PubMed] [Google Scholar]
  15. Eckloff B. W., Podzorski R. P., Kline B. C., Cockerill F. R., 3rd A comparison of 16S ribosomal DNA sequences from five isolates of Helicobacter pylori. Int J Syst Bacteriol. 1994 Apr;44(2):320–323. doi: 10.1099/00207713-44-2-320. [DOI] [PubMed] [Google Scholar]
  16. Fox J. G., Correa P., Taylor N. S., Lee A., Otto G., Murphy J. C., Rose R. Helicobacter mustelae-associated gastritis in ferrets. An animal model of Helicobacter pylori gastritis in humans. Gastroenterology. 1990 Aug;99(2):352–361. doi: 10.1016/0016-5085(90)91016-y. [DOI] [PubMed] [Google Scholar]
  17. Goodwin C. S., Blincow E., Peterson G., Sanderson C., Cheng W., Marshall B., Warren J. R., McCulloch R. Enzyme-linked immunosorbent assay for Campylobacter pyloridis: correlation with presence of C. pyloridis in the gastric mucosa. J Infect Dis. 1987 Mar;155(3):488–494. doi: 10.1093/infdis/155.3.488. [DOI] [PubMed] [Google Scholar]
  18. Hazell S. L., Eichberg J. W., Lee D. R., Alpert L., Evans D. G., Evans D. J., Jr, Graham D. Y. Selection of the chimpanzee over the baboon as a model for Helicobacter pylori infection. Gastroenterology. 1992 Sep;103(3):848–854. doi: 10.1016/0016-5085(92)90016-r. [DOI] [PubMed] [Google Scholar]
  19. Ho S. A., Hoyle J. A., Lewis F. A., Secker A. D., Cross D., Mapstone N. P., Dixon M. F., Wyatt J. I., Tompkins D. S., Taylor G. R. Direct polymerase chain reaction test for detection of Helicobacter pylori in humans and animals. J Clin Microbiol. 1991 Nov;29(11):2543–2549. doi: 10.1128/jcm.29.11.2543-2549.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Karita M., Li Q., Cantero D., Okita K. Establishment of a small animal model for human Helicobacter pylori infection using germ-free mouse. Am J Gastroenterol. 1994 Feb;89(2):208–213. [PubMed] [Google Scholar]
  21. Karita M., Li Q., Okita K. Evaluation of new therapy for eradication of H. pylori infection in nude mouse model. Am J Gastroenterol. 1993 Sep;88(9):1366–1372. [PubMed] [Google Scholar]
  22. Krakowka S., Morgan D. R., Kraft W. G., Leunk R. D. Establishment of gastric Campylobacter pylori infection in the neonatal gnotobiotic piglet. Infect Immun. 1987 Nov;55(11):2789–2796. doi: 10.1128/iai.55.11.2789-2796.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lee A., Fox J. G., Otto G., Murphy J. A small animal model of human Helicobacter pylori active chronic gastritis. Gastroenterology. 1990 Nov;99(5):1315–1323. doi: 10.1016/0016-5085(90)91156-z. [DOI] [PubMed] [Google Scholar]
  24. Linton D., Moreno M., Owen R. J., Stanley J. 16S rrn gene copy number in Helicobacter pylori and its application to molecular typing. J Appl Bacteriol. 1992 Dec;73(6):501–506. doi: 10.1111/j.1365-2672.1992.tb05012.x. [DOI] [PubMed] [Google Scholar]
  25. Marchetti M., Aricò B., Burroni D., Figura N., Rappuoli R., Ghiara P. Development of a mouse model of Helicobacter pylori infection that mimics human disease. Science. 1995 Mar 17;267(5204):1655–1658. doi: 10.1126/science.7886456. [DOI] [PubMed] [Google Scholar]
  26. Marshall B. J., Warren J. R. Unidentified curved bacilli in the stomach of patients with gastritis and peptic ulceration. Lancet. 1984 Jun 16;1(8390):1311–1315. doi: 10.1016/s0140-6736(84)91816-6. [DOI] [PubMed] [Google Scholar]
  27. Morgan D. D., Owen R. J. Use of DNA restriction endonuclease digest and ribosomal RNA gene probe patterns to fingerprint Helicobacter pylori and Helicobacter mustelae isolated from human and animal hosts. Mol Cell Probes. 1990 Aug;4(4):321–334. doi: 10.1016/0890-8508(90)90023-s. [DOI] [PubMed] [Google Scholar]
  28. Nader de Macías M. E., Apella M. C., Romero N. C., González S. N., Oliver G. Inhibition of Shigella sonnei by Lactobacillus casei and Lact. acidophilus. J Appl Bacteriol. 1992 Nov;73(5):407–411. doi: 10.1111/j.1365-2672.1992.tb04996.x. [DOI] [PubMed] [Google Scholar]
  29. Rauws E. A., Royen E. A., Langenberg W., Woensel J. V., Vrij A. A., Tytgat G. N. 14C-urea breath test in C pylori gastritis. Gut. 1989 Jun;30(6):798–803. doi: 10.1136/gut.30.6.798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Reid G., Cook R. L., Bruce A. W. Examination of strains of lactobacilli for properties that may influence bacterial interference in the urinary tract. J Urol. 1987 Aug;138(2):330–335. doi: 10.1016/s0022-5347(17)43137-5. [DOI] [PubMed] [Google Scholar]
  31. Rudmann D. G., Eaton K. A., Krakowka S. Ultrastructural study of Helicobacter pylori adherence properties in gnotobiotic piglets. Infect Immun. 1992 May;60(5):2121–2124. doi: 10.1128/iai.60.5.2121-2124.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Syed S. A., Abrams G. D., Freter R. Efficiency of various intestinal bacteria in assuming normal functions of enteric flora after association with germ-free mice. Infect Immun. 1970 Oct;2(4):376–386. doi: 10.1128/iai.2.4.376-386.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Taylor D. E., Eaton M., Chang N., Salama S. M. Construction of a Helicobacter pylori genome map and demonstration of diversity at the genome level. J Bacteriol. 1992 Nov;174(21):6800–6806. doi: 10.1128/jb.174.21.6800-6806.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Veldhuyzen van Zanten S. J., Tytgat K. M., Hollingsworth J., Jalali S., Rshid F. A., Bowen B. M., Goldie J., Goodacre R. L., Riddell R. H., Hunt R. H. 14C-urea breath test for the detection of Helicobacter pylori. Am J Gastroenterol. 1990 Apr;85(4):399–403. [PubMed] [Google Scholar]
  35. Weiss J., Mecca J., da Silva E., Gassner D. Comparison of PCR and other diagnostic techniques for detection of Helicobacter pylori infection in dyspeptic patients. J Clin Microbiol. 1994 Jul;32(7):1663–1668. doi: 10.1128/jcm.32.7.1663-1668.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Westblom T. U., Phadnis S., Yang P., Czinn S. J. Diagnosis of Helicobacter pylori infection by means of a polymerase chain reaction assay for gastric juice aspirates. Clin Infect Dis. 1993 Mar;16(3):367–371. doi: 10.1093/clind/16.3.367. [DOI] [PubMed] [Google Scholar]
  37. Xia H. X., Keane C. T., O'Morain C. A. Pre-formed urease activity of Helicobacter pylori as determined by a viable cell count technique--clinical implications. J Med Microbiol. 1994 Jun;40(6):435–439. doi: 10.1099/00222615-40-6-435. [DOI] [PubMed] [Google Scholar]
  38. von Wulffen H., Heesemann J., Bützow G. H., Löning T., Laufs R. Detection of Campylobacter pyloridis in patients with antrum gastritis and peptic ulcers by culture, complement fixation test, and immunoblot. J Clin Microbiol. 1986 Nov;24(5):716–720. doi: 10.1128/jcm.24.5.716-720.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]

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