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Antimicrobial Agents and Chemotherapy logoLink to Antimicrobial Agents and Chemotherapy
. 1997 Dec;41(12):2621–2628. doi: 10.1128/aac.41.12.2621

Cloning and sequence analysis of two copies of a 23S rRNA gene from Helicobacter pylori and association of clarithromycin resistance with 23S rRNA mutations.

D E Taylor 1, Z Ge 1, D Purych 1, T Lo 1, K Hiratsuka 1
PMCID: PMC164180  PMID: 9420030

Abstract

In this study, two identical copies of a 23S-5S gene cluster, which are separately situated within the Helicobacter pylori UA802 chromosome, were cloned and sequenced. Comparison of the DNA sequence of the H. pylori 23S rRNA gene with known sequences of other bacterial 23S rRNA genes indicated that the H. pylori UA802 23S rRNA genes are closely related to those of Campylobacter spp. and therefore belong in the proposed Proteobacteria subdivision. The 5'-terminal nucleotide T or A of the 23S rRNA is close to a Pribnow box which could be a -10 region of the transcription promoter for the 23S rRNA gene, suggesting that a posttranscriptional process is likely not involved in the maturation of the H. pylori 23S rRNA. Clinical isolates of H. pylori resistant to clarithromycin were examined by using natural transformation and pulsed-field gel electrophoresis. Cross-resistance to clarithromycin and erythromycin, which was transferred by natural transformation from the Cla(r) Ery(r) donor strain H. pylori E to the Cla(s) Ery(s) recipient strain H. pylori UA802, was associated with an single A-to-G transition mutation at position 2142 of both copies of the 23S rRNA in UA802 Cla(r) Ery(r) mutants. The transformation frequency for Cla(r) and Ery(r) was found to be approximately 2 x 10(-6) transformants per viable cell, and the MICs of both clarithromycin and erythromycin for the Cla(r) Ery(r) mutants were equal to those for the donor isolate. Our results confirmed the previous findings that mutations at positions 2142 and 2143 of the H. pylori 23S rRNA gene are responsible for clarithromycin resistance and suggest that acquisition of clarithromycin resistance in H. pylori could also result from horizontal transfer.

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Selected References

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  1. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blaser M. J. Gastric Campylobacter-like organisms, gastritis, and peptic ulcer disease. Gastroenterology. 1987 Aug;93(2):371–383. doi: 10.1016/0016-5085(87)91028-6. [DOI] [PubMed] [Google Scholar]
  3. Bukanov N. O., Berg D. E. Ordered cosmid library and high-resolution physical-genetic map of Helicobacter pylori strain NCTC11638. Mol Microbiol. 1994 Feb;11(3):509–523. doi: 10.1111/j.1365-2958.1994.tb00332.x. [DOI] [PubMed] [Google Scholar]
  4. Chen X., Finch L. R. Novel arrangement of rRNA genes in Mycoplasma gallisepticum: separation of the 16S gene of one set from the 23S and 5S genes. J Bacteriol. 1989 May;171(5):2876–2878. doi: 10.1128/jb.171.5.2876-2878.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Debets-Ossenkopp Y. J., Sparrius M., Kusters J. G., Kolkman J. J., Vandenbroucke-Grauls C. M. Mechanism of clarithromycin resistance in clinical isolates of Helicobacter pylori. FEMS Microbiol Lett. 1996 Aug 15;142(1):37–42. doi: 10.1111/j.1574-6968.1996.tb08404.x. [DOI] [PubMed] [Google Scholar]
  6. Ezaki T., Takeuchi N., Liu S. L., Kai A., Yamamoto H., Yabuuchi E. Small-scale DNA preparation for rapid genetic identification of Campylobacter species without radioisotope. Microbiol Immunol. 1988;32(2):141–150. doi: 10.1111/j.1348-0421.1988.tb01373.x. [DOI] [PubMed] [Google Scholar]
  7. Fantry G. T., Zheng Q. X., Darwin P. E., Rosenstein A. H., James S. P. Mixed infection with cagA-positive and cagA-negative strains of Helicobacter pylori. Helicobacter. 1996 Jun;1(2):98–106. doi: 10.1111/j.1523-5378.1996.tb00018.x. [DOI] [PubMed] [Google Scholar]
  8. Fukunaga M., Mifuchi I. Unique organization of Leptospira interrogans rRNA genes. J Bacteriol. 1989 Nov;171(11):5763–5767. doi: 10.1128/jb.171.11.5763-5767.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ge Z., Hiratsuka K., Taylor D. E. Nucleotide sequence and mutational analysis indicate that two Helicobacter pylori genes encode a P-type ATPase and a cation-binding protein associated with copper transport. Mol Microbiol. 1995 Jan;15(1):97–106. doi: 10.1111/j.1365-2958.1995.tb02224.x. [DOI] [PubMed] [Google Scholar]
  10. Ge Z., Taylor D. E. Helicobacter pylori genes hpcopA and hpcopP constitute a cop operon involved in copper export. FEMS Microbiol Lett. 1996 Dec 1;145(2):181–188. doi: 10.1111/j.1574-6968.1996.tb08575.x. [DOI] [PubMed] [Google Scholar]
  11. Goddard A. F., Logan R. P. Antimicrobial resistance and Helicobacter pylori. J Antimicrob Chemother. 1996 Apr;37(4):639–643. doi: 10.1093/jac/37.4.639. [DOI] [PubMed] [Google Scholar]
  12. Green C. J., Stewart G. C., Hollis M. A., Vold B. S., Bott K. F. Nucleotide sequence of the Bacillus subtilis ribosomal RNA operon, rrnB. Gene. 1985;37(1-3):261–266. doi: 10.1016/0378-1119(85)90281-1. [DOI] [PubMed] [Google Scholar]
  13. Hartmann R. K., Erdmann V. A. Thermus thermophilus 16S rRNA is transcribed from an isolated transcription unit. J Bacteriol. 1989 Jun;171(6):2933–2941. doi: 10.1128/jb.171.6.2933-2941.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Jiang Q., Hiratsuka K., Taylor D. E. Variability of gene order in different Helicobacter pylori strains contributes to genome diversity. Mol Microbiol. 1996 May;20(4):833–842. doi: 10.1111/j.1365-2958.1996.tb02521.x. [DOI] [PubMed] [Google Scholar]
  15. Liesack W., Stackebrandt E. Evidence for unlinked rrn operons in the Planctomycete Pirellula marina. J Bacteriol. 1989 Sep;171(9):5025–5030. doi: 10.1128/jb.171.9.5025-5030.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Newnham E., Chang N., Taylor D. E. Expanded genomic map of Campylobacter jejuni UA580 and localization of 23S ribosomal rRNA genes by I-CeuI restriction endonuclease digestion. FEMS Microbiol Lett. 1996 Sep 1;142(2-3):223–229. doi: 10.1111/j.1574-6968.1996.tb08434.x. [DOI] [PubMed] [Google Scholar]
  17. Noller H. F. Structure of ribosomal RNA. Annu Rev Biochem. 1984;53:119–162. doi: 10.1146/annurev.bi.53.070184.001003. [DOI] [PubMed] [Google Scholar]
  18. Nomura A., Stemmermann G. N., Chyou P. H., Kato I., Perez-Perez G. I., Blaser M. J. Helicobacter pylori infection and gastric carcinoma among Japanese Americans in Hawaii. N Engl J Med. 1991 Oct 17;325(16):1132–1136. doi: 10.1056/NEJM199110173251604. [DOI] [PubMed] [Google Scholar]
  19. Ogasawara N., Nakai S., Yoshikawa H. Systematic sequencing of the 180 kilobase region of the Bacillus subtilis chromosome containing the replication origin. DNA Res. 1994;1(1):1–14. doi: 10.1093/dnares/1.1.1. [DOI] [PubMed] [Google Scholar]
  20. Parsonnet J., Friedman G. D., Vandersteen D. P., Chang Y., Vogelman J. H., Orentreich N., Sibley R. K. Helicobacter pylori infection and the risk of gastric carcinoma. N Engl J Med. 1991 Oct 17;325(16):1127–1131. doi: 10.1056/NEJM199110173251603. [DOI] [PubMed] [Google Scholar]
  21. Ree H. K., Zimmermann R. A. Organization and expression of the 16S, 23S and 5S ribosomal RNA genes from the archaebacterium Thermoplasma acidophilum. Nucleic Acids Res. 1990 Aug 11;18(15):4471–4478. doi: 10.1093/nar/18.15.4471. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schwartz J. J., Gazumyan A., Schwartz I. rRNA gene organization in the Lyme disease spirochete, Borrelia burgdorferi. J Bacteriol. 1992 Jun;174(11):3757–3765. doi: 10.1128/jb.174.11.3757-3765.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Stone G. G., Shortridge D., Flamm R. K., Versalovic J., Beyer J., Idler K., Zulawinski L., Tanaka S. K. Identification of a 23S rRNA gene mutation in clarithromycin-resistant Helicobacter pylori. Helicobacter. 1996 Dec;1(4):227–228. doi: 10.1111/j.1523-5378.1996.tb00043.x. [DOI] [PubMed] [Google Scholar]
  24. Stone G. G., Shortridge D., Versalovic J., Beyer J., Flamm R. K., Graham D. Y., Ghoneim A. T., Tanaka S. K. A PCR-oligonucleotide ligation assay to determine the prevalence of 23S rRNA gene mutations in clarithromycin-resistant Helicobacter pylori. Antimicrob Agents Chemother. 1997 Mar;41(3):712–714. doi: 10.1128/aac.41.3.712. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Taschke C., Klinkert M. Q., Wolters J., Herrmann R. Organization of the ribosomal RNA genes in Mycoplasma hyopneumoniae: the 5S rRNA gene is separated from the 16S and 23S rRNA genes. Mol Gen Genet. 1986 Dec;205(3):428–433. doi: 10.1007/BF00338078. [DOI] [PubMed] [Google Scholar]
  26. 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]
  27. Taylor D. E., Eaton M., Yan W., Chang N. Genome maps of Campylobacter jejuni and Campylobacter coli. J Bacteriol. 1992 Apr;174(7):2332–2337. doi: 10.1128/jb.174.7.2332-2337.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Trust T. J., Logan S. M., Gustafson C. E., Romaniuk P. J., Kim N. W., Chan V. L., Ragan M. A., Guerry P., Gutell R. R. Phylogenetic and molecular characterization of a 23S rRNA gene positions the genus Campylobacter in the epsilon subdivision of the Proteobacteria and shows that the presence of transcribed spacers is common in Campylobacter spp. J Bacteriol. 1994 Aug;176(15):4597–4609. doi: 10.1128/jb.176.15.4597-4609.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Versalovic J., Shortridge D., Kibler K., Griffy M. V., Beyer J., Flamm R. K., Tanaka S. K., Graham D. Y., Go M. F. Mutations in 23S rRNA are associated with clarithromycin resistance in Helicobacter pylori. Antimicrob Agents Chemother. 1996 Feb;40(2):477–480. doi: 10.1128/aac.40.2.477. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wang Y., Roos K. P., Taylor D. E. Transformation of Helicobacter pylori by chromosomal metronidazole resistance and by a plasmid with a selectable chloramphenicol resistance marker. J Gen Microbiol. 1993 Oct;139(10):2485–2493. doi: 10.1099/00221287-139-10-2485. [DOI] [PubMed] [Google Scholar]
  31. de Boer H. A., Gilbert S. F., Nomura M. DNA sequences of promoter regions for rRNA operons rrnE and rrnA in E. coli. Cell. 1979 May;17(1):201–209. doi: 10.1016/0092-8674(79)90308-8. [DOI] [PubMed] [Google Scholar]
  32. van der Hulst R. W., Keller J. J., Rauws E. A., Tytgat G. N. Treatment of Helicobacter pylori infection: a review of the world literature. Helicobacter. 1996 Mar;1(1):6–19. doi: 10.1111/j.1523-5378.1996.tb00003.x. [DOI] [PubMed] [Google Scholar]

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