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. 1996 May;178(9):2701–2708. doi: 10.1128/jb.178.9.2701-2708.1996

Transformation of Coxiella burnetii to ampicillin resistance.

M L Suhan 1, S Y Chen 1, H A Thompson 1
PMCID: PMC177998  PMID: 8626341

Abstract

A 5.8-kb chromosomal fragment isolated from Coxiella burnetii initiates plasmid replication in Escherichia coli and was characterized as an autonomous replication sequence, ars (M. Suhan, S.-Y. Chen, H.A. Thompson, T.A. Hoover, A. Hill, and J.C. Williams, J. Bacteriol. 176:5233-5243, 1994). In the present study, an ars replicon was used to transform C. burnetii to ampicillin resistance. Plasmid pSKO(+)1000 contained the C. burnetii ars sequence cloned into a ColE1-type replicon encoding beta-lactamase. pSKO(+)1000 was introduced into C. burnetii by electroporation. Ampicillin-resistant cells were selected, and survivors were examined for the transformed genotype by Southern hybridization. Transformants stably maintained the pSKO(+)1000 bla DNA sequence in the chromosome as a result of homologous recombination. The recombination event resulted in the duplication of the 5.8-kb ars sequence in the C. burnetii chromosome. The bla gene was also located in an episome. However, an ampicillin resistance plasmid lacking the C. burnetii ars sequence did not stably transform C. burnetii. A biological assay analyzing beta-lactamase activity of C. burnetii transformants during acid activation in vitro provided evidence for expression of the bla (beta-lactamase) gene.

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Selected References

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  1. Akporiaye E. T., Rowatt J. D., Aragon A. A., Baca O. G. Lysosomal response of a murine macrophage-like cell line persistently infected with Coxiella burnetii. Infect Immun. 1983 Jun;40(3):1155–1162. doi: 10.1128/iai.40.3.1155-1162.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bachmann B. J. Linkage map of Escherichia coli K-12, edition 8. Microbiol Rev. 1990 Jun;54(2):130–197. doi: 10.1128/mr.54.2.130-197.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bramhill D., Kornberg A. A model for initiation at origins of DNA replication. Cell. 1988 Sep 23;54(7):915–918. doi: 10.1016/0092-8674(88)90102-x. [DOI] [PubMed] [Google Scholar]
  4. Burton P. R., Kordová N., Paretsky D. Electron microscopic studies of the rickettsia Coxiella burneti: entry, lysosomal response, and fate of rickettsial DNA in L-cells. Can J Microbiol. 1971 Feb;17(2):143–150. doi: 10.1139/m71-025. [DOI] [PubMed] [Google Scholar]
  5. Chen S. Y., Vodkin M., Thompson H. A., Williams J. C. Isolated Coxiella burnetii synthesizes DNA during acid activation in the absence of host cells. J Gen Microbiol. 1990 Jan;136(1):89–96. doi: 10.1099/00221287-136-1-89. [DOI] [PubMed] [Google Scholar]
  6. Dente L., Cesareni G., Cortese R. pEMBL: a new family of single stranded plasmids. Nucleic Acids Res. 1983 Mar 25;11(6):1645–1655. doi: 10.1093/nar/11.6.1645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Dunkin S. M., Wood D. O. Isolation and characterization of the Rickettsia prowazekii recA gene. J Bacteriol. 1994 Mar;176(6):1777–1781. doi: 10.1128/jb.176.6.1777-1781.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  9. GIMENEZ D. F. STAINING RICKETTSIAE IN YOLK-SAC CULTURES. Stain Technol. 1964 May;39:135–140. doi: 10.3109/10520296409061219. [DOI] [PubMed] [Google Scholar]
  10. Gille H., Egan J. B., Roth A., Messer W. The FIS protein binds and bends the origin of chromosomal DNA replication, oriC, of Escherichia coli. Nucleic Acids Res. 1991 Aug 11;19(15):4167–4172. doi: 10.1093/nar/19.15.4167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hackstadt T., Williams J. C. Biochemical stratagem for obligate parasitism of eukaryotic cells by Coxiella burnetii. Proc Natl Acad Sci U S A. 1981 May;78(5):3240–3244. doi: 10.1073/pnas.78.5.3240. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Harding N. E., Cleary J. M., Smith D. W., Michon J. J., Brusilow W. S., Zyskind J. W. Chromosomal replication origins (oriC) of Enterobacter aerogenes and Klebsiella pneumoniae are functional in Escherichia coli. J Bacteriol. 1982 Dec;152(3):983–993. doi: 10.1128/jb.152.3.983-993.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Holm C., Meeks-Wagner D. W., Fangman W. L., Botstein D. A rapid, efficient method for isolating DNA from yeast. Gene. 1986;42(2):169–173. doi: 10.1016/0378-1119(86)90293-3. [DOI] [PubMed] [Google Scholar]
  14. LENNOX E. S. Transduction of linked genetic characters of the host by bacteriophage P1. Virology. 1955 Jul;1(2):190–206. doi: 10.1016/0042-6822(55)90016-7. [DOI] [PubMed] [Google Scholar]
  15. Mandell G. L. Interaction of intraleukocytic bacteria and antibiotics. J Clin Invest. 1973 Jul;52(7):1673–1679. doi: 10.1172/JCI107348. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Masuda G., Tomioka S., Hasegawa M. Detection of beta-lactamase production by gram-negative bacteria. J Antibiot (Tokyo) 1976 Jun;29(6):662–664. doi: 10.7164/antibiotics.29.662. [DOI] [PubMed] [Google Scholar]
  17. Maurin M., Benoliel A. M., Bongrand P., Raoult D. Phagolysosomes of Coxiella burnetii-infected cell lines maintain an acidic pH during persistent infection. Infect Immun. 1992 Dec;60(12):5013–5016. doi: 10.1128/iai.60.12.5013-5016.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Miller J. F., Dower W. J., Tompkins L. S. High-voltage electroporation of bacteria: genetic transformation of Campylobacter jejuni with plasmid DNA. Proc Natl Acad Sci U S A. 1988 Feb;85(3):856–860. doi: 10.1073/pnas.85.3.856. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Norman E., Dellagostin O. A., McFadden J., Dale J. W. Gene replacement by homologous recombination in Mycobacterium bovis BCG. Mol Microbiol. 1995 May;16(4):755–760. doi: 10.1111/j.1365-2958.1995.tb02436.x. [DOI] [PubMed] [Google Scholar]
  20. Polaczek P. Bending of the origin of replication of E. coli by binding of IHF at a specific site. New Biol. 1990 Mar;2(3):265–271. [PubMed] [Google Scholar]
  21. Renard C., Vanderhaeghe H. J., Claes P. J., Zenebergh A., Tulkens P. M. Influence of conversion of penicillin G into a basic derivative on its accumulation and subcellular localization in cultured macrophages. Antimicrob Agents Chemother. 1987 Mar;31(3):410–416. doi: 10.1128/aac.31.3.410. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Reschke D. K., Frazier M. E., Mallavia L. P. Transformation of Rochalimaea quintana, a member of the family Rickettsiaceae. J Bacteriol. 1990 Sep;172(9):5130–5134. doi: 10.1128/jb.172.9.5130-5134.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. 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]
  24. Schowalter D. B., Sommer S. S. The generation of radiolabeled DNA and RNA probes with polymerase chain reaction. Anal Biochem. 1989 Feb 15;177(1):90–94. doi: 10.1016/0003-2697(89)90019-5. [DOI] [PubMed] [Google Scholar]
  25. Short J. M., Fernandez J. M., Sorge J. A., Huse W. D. Lambda ZAP: a bacteriophage lambda expression vector with in vivo excision properties. Nucleic Acids Res. 1988 Aug 11;16(15):7583–7600. doi: 10.1093/nar/16.15.7583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Smith D. W., Yee T. W., Baird C., Krishnapillai V. Pseudomonad replication origins: a paradigm for bacterial origins? Mol Microbiol. 1991 Nov;5(11):2581–2587. doi: 10.1111/j.1365-2958.1991.tb01966.x. [DOI] [PubMed] [Google Scholar]
  27. Suhan M., Chen S. Y., Thompson H. A., Hoover T. A., Hill A., Williams J. C. Cloning and characterization of an autonomous replication sequence from Coxiella burnetii. J Bacteriol. 1994 Sep;176(17):5233–5243. doi: 10.1128/jb.176.17.5233-5243.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Vodkin M. H., Williams J. C., Stephenson E. H. Genetic heterogeneity among isolates of Coxiella burnetii. J Gen Microbiol. 1986 Feb;132(2):455–463. doi: 10.1099/00221287-132-2-455. [DOI] [PubMed] [Google Scholar]
  29. Wiebe M. E., Burton P. R., Shankel D. M. Isolation and characterization of two cell types of Coxiella burneti phase I. J Bacteriol. 1972 Apr;110(1):368–377. doi: 10.1128/jb.110.1.368-377.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Williams J. C., Peacock M. G., McCaul T. F. Immunological and biological characterization of Coxiella burnetii, phases I and II, separated from host components. Infect Immun. 1981 May;32(2):840–851. doi: 10.1128/iai.32.2.840-851.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. 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]
  32. Yasuda S., Hirota Y. Cloning and mapping of the replication origin of Escherichia coli. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5458–5462. doi: 10.1073/pnas.74.12.5458. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Yee T. W., Smith D. W. Pseudomonas chromosomal replication origins: a bacterial class distinct from Escherichia coli-type origins. Proc Natl Acad Sci U S A. 1990 Feb;87(4):1278–1282. doi: 10.1073/pnas.87.4.1278. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zuerner R. L., Thompson H. A. Protein synthesis by intact Coxiella burnetii cells. J Bacteriol. 1983 Oct;156(1):186–191. doi: 10.1128/jb.156.1.186-191.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]

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