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. 1987 Aug;169(8):3450–3457. doi: 10.1128/jb.169.8.3450-3457.1987

Transposition of Tn4551 in Bacteroides fragilis: identification and properties of a new transposon from Bacteroides spp.

C J Smith, H Spiegel
PMCID: PMC212416  PMID: 3038840

Abstract

Tn4551, a clindamycin resistance (Ccr) transposon from the R plasmid pBI136, was cloned onto an Escherichia coli-Bacteroides shuttle vector which could replicate normally in E. coli but was maintained unstably in Bacteroides fragilis. To aid in cloning and to ensure maintenance of Tn4551 in E. coli, a kanamycin resistance determinant (Kmr) was inserted in the transposon. The transposon-bearing shuttle vector pFD197 was transformed into B. fragilis 638, and putative insertions of Tn4551::Kmr were identified by screening for resistance to clindamycin and plasmid content. Southern hybridization analyses were used to verify integration of the transposon in the B. fragilis chromosome, and the frequency of insertion was estimated at 7.8 X 10(-5) events per generation. In 57% of the isolates tested a second integration event also occurred. This second insertion apparently involved just a single copy of the 1.2-kilobase repeat sequence which flanks the transposon. In addition, Tn4551::Kmr appeared to function as a transposon in E. coli. Evidence for this was obtained by the isolation of transposon insertions into the bacteriophage P1 genome. Finally, the transposon vector, pFD197, could be mobilized to other B. fragilis strains in which transposition was detected. Mobilization from the strain 638 background was via a conjugation like process, but occurred in the absence of known conjugative elements or other detectable plasmids. This result suggested the presence of a host-encoded transfer system in this B. fragilis strain.

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Selected References

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

  1. Bachmann B. J. Pedigrees of some mutant strains of Escherichia coli K-12. Bacteriol Rev. 1972 Dec;36(4):525–557. doi: 10.1128/br.36.4.525-557.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. Callihan D. R., Young F. E., Clark V. L. Identification of three homology classes of small, cryptic plasmids in intestinal Bacteroides species. Plasmid. 1983 Jan;9(1):17–30. doi: 10.1016/0147-619x(83)90028-8. [DOI] [PubMed] [Google Scholar]
  4. Currier T. C., Nester E. W. Isolation of covalently closed circular DNA of high molecular weight from bacteria. Anal Biochem. 1976 Dec;76(2):431–441. doi: 10.1016/0003-2697(76)90338-9. [DOI] [PubMed] [Google Scholar]
  5. Finegold S. M., Attebery H. R., Sutter V. L. Effect of diet on human fecal flora: comparison of Japanese and American diets. Am J Clin Nutr. 1974 Dec;27(12):1456–1469. doi: 10.1093/ajcn/27.12.1456. [DOI] [PubMed] [Google Scholar]
  6. Guerry P., LeBlanc D. J., Falkow S. General method for the isolation of plasmid deoxyribonucleic acid. J Bacteriol. 1973 Nov;116(2):1064–1066. doi: 10.1128/jb.116.2.1064-1066.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Guiney D. G., Jr, Hasegawa P., Davis C. E. Expression in Escherichia coli of cryptic tetracycline resistance genes from bacteroides R plasmids. Plasmid. 1984 May;11(3):248–252. doi: 10.1016/0147-619x(84)90031-3. [DOI] [PubMed] [Google Scholar]
  8. Guiney D. G., Jr, Hasegawa P., Davis C. E. Homology between clindamycin resistance plasmids in Bacteroides. Plasmid. 1984 May;11(3):268–271. doi: 10.1016/0147-619x(84)90035-0. [DOI] [PubMed] [Google Scholar]
  9. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  10. Kleckner N. Transposable elements in prokaryotes. Annu Rev Genet. 1981;15:341–404. doi: 10.1146/annurev.ge.15.120181.002013. [DOI] [PubMed] [Google Scholar]
  11. LeBlanc D. J., Inamine J. M., Lee L. N. Broad geographical distribution of homologous erythromycin, kanamycin, and streptomycin resistance determinants among group D streptococci of human and animal origin. Antimicrob Agents Chemother. 1986 Apr;29(4):549–555. doi: 10.1128/aac.29.4.549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Malamy M. H., Tally F. P. Mechanisms of drug-resistance transfer in Bacteroides fragilis. J Antimicrob Chemother. 1981 Dec;8 (Suppl 500):59–75. doi: 10.1093/jac/8.suppl_d.59. [DOI] [PubMed] [Google Scholar]
  13. Matthews B. G., Guiney D. G. Characterization and mapping of regions encoding clindamycin resistance, tetracycline resistance, and a replication function on the Bacteroides R plasmid pCP1. J Bacteriol. 1986 Aug;167(2):517–521. doi: 10.1128/jb.167.2.517-521.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Mays T. D., Smith C. J., Welch R. A., Delfini C., Macrina F. L. Novel antibiotic resistance transfer in Bacteroides. Antimicrob Agents Chemother. 1982 Jan;21(1):110–118. doi: 10.1128/aac.21.1.110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Moore W. E., Holdeman L. V. Human fecal flora: the normal flora of 20 Japanese-Hawaiians. Appl Microbiol. 1974 May;27(5):961–979. doi: 10.1128/am.27.5.961-979.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Privitera G., Dublanchet A., Sebald M. Transfer of multiple antibiotic resistance between subspecies of Bacteroides fragilis. J Infect Dis. 1979 Jan;139(1):97–101. doi: 10.1093/infdis/139.1.97. [DOI] [PubMed] [Google Scholar]
  17. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  18. Robillard N. J., Tally F. P., Malamy M. H. Tn4400, a compound transposon isolated from Bacteroides fragilis, functions in Escherichia coli. J Bacteriol. 1985 Dec;164(3):1248–1255. doi: 10.1128/jb.164.3.1248-1255.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rosner J. L. Formation, induction, and curing of bacteriophage P1 lysogens. Virology. 1972 Jun;48(3):679–689. doi: 10.1016/0042-6822(72)90152-3. [DOI] [PubMed] [Google Scholar]
  20. Shimell M. J., Smith C. J., Tally F. P., Macrina F. L., Malamy M. H. Hybridization studies reveal homologies between pBF4 and pBFTM10, Two clindamycin-erythromycin resistance transfer plasmids of Bacteroides fragilis. J Bacteriol. 1982 Nov;152(2):950–953. doi: 10.1128/jb.152.2.950-953.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Shoemaker N. B., Getty C., Gardner J. F., Salyers A. A. Tn4351 transposes in Bacteroides spp. and mediates the integration of plasmid R751 into the Bacteroides chromosome. J Bacteriol. 1986 Mar;165(3):929–936. doi: 10.1128/jb.165.3.929-936.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Shoemaker N. B., Getty C., Guthrie E. P., Salyers A. A. Regions in Bacteroides plasmids pBFTM10 and pB8-51 that allow Escherichia coli-Bacteroides shuttle vectors to be mobilized by IncP plasmids and by a conjugative Bacteroides tetracycline resistance element. J Bacteriol. 1986 Jun;166(3):959–965. doi: 10.1128/jb.166.3.959-965.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Shoemaker N. B., Guthrie E. P., Salyers A. A., Gardner J. F. Evidence that the clindamycin-erythromycin resistance gene of Bacteroides plasmid pBF4 is on a transposable element. J Bacteriol. 1985 May;162(2):626–632. doi: 10.1128/jb.162.2.626-632.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Smith C. J. Characterization of Bacteroides ovatus plasmid pBI136 and structure of its clindamycin resistance region. J Bacteriol. 1985 Mar;161(3):1069–1073. doi: 10.1128/jb.161.3.1069-1073.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Smith C. J. Development and use of cloning systems for Bacteroides fragilis: cloning of a plasmid-encoded clindamycin resistance determinant. J Bacteriol. 1985 Oct;164(1):294–301. doi: 10.1128/jb.164.1.294-301.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Smith C. J., Gonda M. A. Comparison of the transposon-like structures encoding clindamycin resistance in Bacteroides R-plasmids. Plasmid. 1985 May;13(3):182–192. doi: 10.1016/0147-619x(85)90041-1. [DOI] [PubMed] [Google Scholar]
  27. Smith C. J., Macrina F. L. Large transmissible clindamycin resistance plasmid in Bacteroides ovatus. J Bacteriol. 1984 May;158(2):739–741. doi: 10.1128/jb.158.2.739-741.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Smith C. J. Polyethylene glycol-facilitated transformation of Bacteroides fragilis with plasmid DNA. J Bacteriol. 1985 Oct;164(1):466–469. doi: 10.1128/jb.164.1.466-469.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Smith C. J., Welch R. A., Macrina F. L. Two independent conjugal transfer systems operating in Bacteroides fragilis V479-1. J Bacteriol. 1982 Jul;151(1):281–287. doi: 10.1128/jb.151.1.281-287.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  31. Tally F. P., Snydman D. R., Shimell M. J., Malamy M. H. Characterization of pBFTM10, a clindamycin-erythromycin resistance transfer factor from Bacteroides fragilis. J Bacteriol. 1982 Aug;151(2):686–691. doi: 10.1128/jb.151.2.686-691.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Thayer R. E. An improved method for detecting foreign DNA in plasmids of Escherichia coli. Anal Biochem. 1979 Sep 15;98(1):60–63. doi: 10.1016/0003-2697(79)90705-x. [DOI] [PubMed] [Google Scholar]
  33. Trieu-Cuot P., Courvalin P. Nucleotide sequence of the Streptococcus faecalis plasmid gene encoding the 3'5"-aminoglycoside phosphotransferase type III. Gene. 1983 Sep;23(3):331–341. doi: 10.1016/0378-1119(83)90022-7. [DOI] [PubMed] [Google Scholar]
  34. Welch R. A., Macrina F. L. Physical characterization of Bacteroides fragilis R plasmid pBF4. J Bacteriol. 1981 Feb;145(2):867–872. doi: 10.1128/jb.145.2.867-872.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Willetts N., Wilkins B. Processing of plasmid DNA during bacterial conjugation. Microbiol Rev. 1984 Mar;48(1):24–41. doi: 10.1128/mr.48.1.24-41.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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