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Volume 140, Issue 6

Virulence region of plasmid pNL2001 of Free

Abstract

The virulence region of the 55 kb plasmid pNL2001 was identified by Tn-insertion mutagenesis, DNA hybridization studies, and Western blot analysis of proteins encoded in the virulence region of the plasmid. DNA hybridization studies showed that the pNL2001 plasmid contained a 6·4 kb I-RI fragment homologous to the 6·4 kb I-RI plasmid virulence () region of the 50 kb plasmid (pKDSC50). One of the 247 Tn-insertion mutants of , designated strain TA19, showed a reduced mouse lethality, and the Tn-insertion of strain TA19 was located within this homologous 6·4 kb region, suggesting that the 6·4 kb I-RI fragment of pNL2001 contained the region. Two contiguous I-RI (2·3 kb) and RI-RI (4·1 kb) fragments derived from the 6·4 kb I-RI fragment were cloned into the expression vectors. By Western blot analysis using four Spv peptide antisera, each specific for individual proteins encoded in the and genes of pKDSC50, four proteins encoded in the 6·4 kb I-RI fragment of pNL2001 were identified. Protein SpvR with an apparent molecular mass of 32 kDa was produced from the 2·3 kb I-RI fragment, and proteins SpvA, SpvB and SpvC with apparent molecular masses of 32, 70 and 29 kDa, respectively, were produced from the 4·1 kb RI-RI fragment. From the 4·1 kb RI::Tn fragment of the TA19 plasmid, proteins SpvA and SpvB were expressed, but not SpvC. It was therefore suggested that the gene may contribute to the expression of virulence of . Furthermore, the nucleotide sequence of the 6·4 kb I-RI fragment encoding these four proteins was determined. Four open reading frames which encoded the four proteins with deduced molecular masses of 33906, 28200, 65349 and 27646 Da were detected. Deduced amino acid sequences of each protein showed a high degree of identity to corresponding sequences in the virulence region of and virulence plasmids. Therefore, we confirmed that the virulence plasmids of Salmonellae including share the highly conserved region responsible for virulence.

  • Received:
  • Accepted:
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  • Published Online:
Keyword(s): Salmonella enteritidis , spvABC genes and virulence plasmid pNL2001
© Society for General Microbiology 1994
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1994-06-01
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References

  1. Baird G. D., Manning E. J., Jones P. W. 1985; Evidence for related virulence sequences in plasmids of Salmonella dublinSalmonella typhimurium. J Gen Microbiol 131:1815–1823
     [Google Scholar]
  2. Beninger P. R., Chikami G., Tanabe K., Roudier C., Fierer J., Guiney D. G. 1988; Physicalgenetic mapping of the Salmonella dublin virulence plasmid pSDL2. J Clin Invest 81:1341–1347
     [Google Scholar]
  3. Bolivar F., Backman K. 1979; Plasmids of Escherichia coli as cloning vectors. Methods Enzymol 68:245–267
     [Google Scholar]
  4. Caldwell A.L., Gulig P. A. 1991; The Salmonella typhimurium virulence plasmid encodes a positive regulator of a plasmid- encoded virulence gene. J Bacteriol 173:7176–7185
     [Google Scholar]
  5. Chang A.C., Cohen S. N. 1978; Constructioncharacterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol 134:1141–1156
     [Google Scholar]
  6. Cohen S. N., Chang A. C. Y., Hsu L. 1972; Nonchromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA. Proc Natl Acad Sci USA 69:2110–2114
     [Google Scholar]
  7. Coynault C., Robbe-Saule V., Popoff M. Y., Norel F. 1992; Growth phaseSpy R regulation of transcription of Salmonella typhimurium spv ABC virulence genes. Microb Pathog 13:133–143
     [Google Scholar]
  8. Fang F. C., Krause M., Roudier C., Fierer J., Guiney D. G. 1991; Growth regulation of a Salmonella plasmid gene essential for virulence. J Bacteriol 173:6783–6789
     [Google Scholar]
  9. Fang F. C., Libby S. J., Buchmeier N. A., Loewen P. C., Switala J., Harwood J., Guiney D. G. 1992; The alternative a factor Kat F (Rpo S) regulates Salmonella virulence. Proc Natl Acad Sci USA 89:1978 1982
     [Google Scholar]
  10. Gulig P.A. 1990; Virulence plasmids of Salmonella typhimurium and other salmonellae. Microb Pathog 8:3–11
     [Google Scholar]
  11. Gulig P.A., Chiodo V. A. 1990; GeneticDNA sequence analysis of the Salmonella typhimurium virulence plasmid gene encoding the 28 000-molecular-weight protein. Infect Immun 58:2651–2658
     [Google Scholar]
  12. Gulig P.A., Curtiss R. 1987; Plasmid-associated virulence of Salmonella typhimurium. Infect Immun 55:2891–2901
     [Google Scholar]
  13. Gulig P.A., Curtiss R. 1988; Cloningtransposon insertion mutagenesis of virulence genes of the 100-kilobase plasmid of Salmonella typhimurium. Infect Immun 56:3262–3271
     [Google Scholar]
  14. Gulig P.A., Doyle T. J. 1993; The Salmonella typhimurium virulence plasmid increases the growth rate of salmonellae in mice. Infect Immun 61:504–511
     [Google Scholar]
  15. Gulig P. A., Caldwell A. L., Chiodo V. A. 1992; Identification, genetic analysisDNA sequence of a 7'8-kb virulence region of the Salmonella typhimurium virulence plasmid. Mol Microbiol 6:1395–1411
     [Google Scholar]
  16. Gulig P. A., Danbara H., Guiney D. G., Lax A. J., Norel F., Rhen M. 1993; Molecular analysis of spv virulence genes of the salmonella virulence plasmids. Mol Microbiol 7:825–830
     [Google Scholar]
  17. Harayama S., Tsuda M., Iino T. 1980; High frequency mobilization of the chromosome of Escherichia coli by a mutant of plasmid RP4 temperature-sensitive for maintenance. MolGen Genet 180:47–56
     [Google Scholar]
  18. Heffernan E. J., Fierer J., Chikami G., Guiney D. 1987; Natural history of oral Salmonella dublin infection in BALB/c mice: effect of an 80-kilobase-pair plasmid on virulence. J Infect Dis 155:1254–1259
     [Google Scholar]
  19. Heffron F., McCarthy, B. J., Ohtsubo H., Ohtsubo E. 1979; DNA sequence analysis of the transposon TnJ: three genesthree sites involved in transposition of Tni. Cell 18:1153–1163
     [Google Scholar]
  20. Holmes D.S., Quigley M. 1981; The rapid boiling method for the preparation of bacterial plasmids. Anal Biochem 114:193–197
     [Google Scholar]
  21. Hovi M., Sukupolvi S., Edwards M. F., Rhen M. 1988; Plasmid-associated virulence of Salmonella enteritidis. Microb Pathog 4:385–391
     [Google Scholar]
  22. Jones C.S., Osborne D. J. 1991; Identification of contemporary plasmid virulence genes in ancestral isolates of Salmonella enteritidis and Salmonella typhimurium. FEMS Microbiol Lett 80:7–12
     [Google Scholar]
  23. Kado C.I., Liu S.-T. 1981; Rapid procedure for detectionisolation of largesmall plasmids. J Bacteriol 145:1365–1373
     [Google Scholar]
  24. Karber G. 1931; Beitrag zur kollektiven Behandlung phar- makologischer Reihenversuche. Arch Exp Pathol Pharmakol 162:480
     [Google Scholar]
  25. Kawahara K., Haraguchi Y., Tsuchimoto M., Terakado N., Danbara H. 1988; Evidence of correlation between 50-kilobase plasmid of Salmonella choleraesuis its virulence. Microb Pathog 4:155–163
     [Google Scholar]
  26. Kawahara K., Tsuchimoto M., Sudo K., Terakado N., Danbara H. 1990; Identificationmapping of mba regions of the Salmonella choleraesuis virulence plasmid pKDSC50 responsible for mouse bacteremia. Microb Pathog 8:13–21
     [Google Scholar]
  27. Korpela K., Ranki M., Sukupolvi S., Makela P. H., Rhen M. 1989; Occurrence of Salmonella typhimurium virulence plasmid- specific sequences in different serovars of Salmonella. FEMS Microbiol Lett 58:49–54
     [Google Scholar]
  28. Krause M., Roudier C., Fierer J., Harwood J., Guiney D. 1991; Molecular analysis of the virulence locus of the Salmonella dublin plasmid pSDL2. Mol Microbiol 5:307–316
     [Google Scholar]
  29. Krause M., Fang F. C., Guiney D. G. 1992; Regulation of plasmid virulence gene expression in Salmonella dublin involves an unusual operon structure. J Bacteriol 174:4482–4489
     [Google Scholar]
  30. Kushner S.R. 1978; An improved method for transformation of Escherichia coli with Col El-derived plasmids. In Genetic Engineering pp. 17–23 Edited by Boyer H. B., Nicosia S. . Amsterdam: Elsevier;
     [Google Scholar]
  31. Laemmli U.K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
     [Google Scholar]
  32. Lax A. J., Pullinger G. D., Baird G. D., Williamson G. M. 1990; The virulence plasmid of Salmonella dublin: detailed restriction mapanalysis by transposon mutagenesis. J Gen Microbiol 136:1117–1123
     [Google Scholar]
  33. Lennox E.S. 1955; Transduction of linked genetic characters of the host by bacteriophage PL. Virology 1:190–206
     [Google Scholar]
  34. Matsui H., Kawahara K., Terakado N., Danbara H. 1990a; Nucleotide sequence of a gene encoding a 29 kDa polypeptide in mba region of the virulence plasmid, pKDSC50, of Salmonella choleraesuis. Nucleic Acids Res 18:1055
     [Google Scholar]
  35. Matsui H., Kawahara K., Terakado N., Danbara H. 1990b; Nucleotide sequences of genes encoding 32 kDa70 kDa polypeptides in mba region of the virulence plasmid, pKDSC50, of Salmonella choleraesuis. Nucleic Acids Res 18:2181–2182
     [Google Scholar]
  36. Matsui H., Abe A., Kawahara K., Terakado N., Danbara H. 1991; Positive regulator for the expression of Mba protein of the virulence plasmid, pKDSC50, of Salmonella choleraesuis. Microb Pathog 10:459–464
     [Google Scholar]
  37. Matsui H., Abe A., Suzuki S., Kijima M., Tamura Y., Nakamura M., Kawahara K., Danbara H. 1993; Molecular mechanism of the regulation of expression of plasmid-encoded mouse bacteremia (mba) genes in Salmonella serovar Choleraesuis. MolGen Genet 236:219–226
     [Google Scholar]
  38. Michiels T., Popoff M. Y., Durviaux S., Coynault C., Cornells G. 1987; A new method for the physicalgenetic mapping of large plasmids: application to the localisation of the virulence determinants on the 90 kb plasmid of Salmonella typhimurium. Microb Pathog 3:109–116
     [Google Scholar]
  39. Montenegro M. A., Morelli G., Helmuth R. 1991; Heteroduplex analysis of Salmonella virulence plasmidstheir prevalence in isolates of defined sources. Microb Pathog 11:391–397
     [Google Scholar]
  40. Nakamura M., Sato S., Ohya T., Suzuki S., Ikeda S. 1985; Possible relationship of a 36-Megadalton Salmonella enteritidis plasmid to virulence in mice. Infect Immun 47:831–833
     [Google Scholar]
  41. Norel F., Pisano M. R., Nicoli J., Popoff M. Y. 1989a; Nucleotide sequence of the plasmid-borne virulence gene mkf A encoding a 28 kDa polypeptide from Salmonella typhimurium. Res Microbiol 140:263–265
     [Google Scholar]
  42. Norel F., Pisano M. R., Nicoli J., Popoff M. Y. 1989b; Nucleotide sequence of the plasmid-borne virulence gene mkf B from Salmonella typhimurium. Res Microbiol 140:455–457
     [Google Scholar]
  43. Norel F., Pisano M. R., Nicoli J., Popoff M. Y. 1989c; A plasmid-borne virulence region (2'8 kb) from Salmonella typhimurium contains two open reading frames. Res Microbiol 140:627–630
     [Google Scholar]
  44. Norel F., Robbe-Saule V., Popoff M. Y., Coynault C. 1992; The putative sigma factor Kat F (Rpo S) is required for the transcription of the Salmonella typhimurium virulence gene spv B in Escherichia coli. FEMS Microbiol Lett 99:271–276
     [Google Scholar]
  45. Popoff M. Y., Miras I., Coynault C. L., Lasselin C., Pardon P. 1984; Molecular relationships between virulence plasmids of Salmonella serotypes typhimurium dublin large plasmids of other Salmonella serotypes. Ann Microbiol (Paris) 135A:389–398
     [Google Scholar]
  46. Poppe C., Curtiss R., Gulig P. A., Gyles C. L. 1989; Hybridization studies with a DNA probe derived from the virulence region of the 60 Mdal plasmid of Salmonella typhimurium. Can J Vet Res 53:378–384
     [Google Scholar]
  47. Pullinger G. D., Baird G. D., Williamson C. M., Lax A. J. 1989; Nucleotide sequence of a plasmid gene involved in the virulence of salmonellas. Nucleic Acids Res 17:7983
     [Google Scholar]
  48. Rhen M., Virtanen M., Makela P. H. 1989; Localization by insertion mutagenesis of a virulence-associated region in the Salmonella typhimurium 96 kilobase pair virulence plasmid. Microb Pathog 6:153–158
     [Google Scholar]
  49. Riikonen P., Makela P. H., Saarilahti H., Sukupolvi S., Taira S., Rhen M. 1992; The virulence plasmid does not contribute to growth of Salmonella in cultured murine macrophages. Microb Pathog 13:281–291
     [Google Scholar]
  50. Roudier C., Krause M., Fierer J., Guiney D. G. 1990; Correlation between the presence of sequences homologous to the vir region of Salmonella dublin plasmid pSDL2the virulence of twenty-two Salmonella serotypes in mice. Infect Immun 58:1180–1185
     [Google Scholar]
  51. Roudier C., Fierer J., Guiney D. G. 1992; Characterization of translation termination mutations in the spv operon of the Salmonella virulence plasmid pSDL2. J Bacteriol 174:6418–6423
     [Google Scholar]
  52. Selander R. K., Smith N. H., Li J., Beltran P., Ferris K. E., Kopecko D. J., Rubin F. A. 1992; Molecular evolutionary genetics of the cattle-adapted serovar Salmonella dublin. J Bacteriol 174:3587–3592
     [Google Scholar]
  53. Short J. M., Fernandez J. M., Sorge J. A., Huse W. D. 1988; λZAP: a bacteriophage A expression vector with in vivo excision properties. Nucleic Acids Res 16:7583
     [Google Scholar]
  54. Suzuki S., Ohmae K., Nakamura M., Sato S., Koeda T., Ohishi K., Muramatsu M. 1989; Demonstration of the correlation of a 36-megada Iton Salmonella serovar enteritidis plasmid to virulence in mice by reintroduction of the plasmid. Jpn J Vet Sci 51:203–205
     [Google Scholar]
  55. Suzuki S., Ohishi K., Takahashi T., Tamura Y., Muramatsu M., Nakamura M., Sato S. 1992; The role of 36 megadalton plasmid of Salmonella Enteritidis for the pathogenesis in mice. J Vet Med Sci 54:845–850
     [Google Scholar]
  56. Taira S., Rhen M. 1989a; Identificationgenetic analysis of mka A - a gene of the Salmonella typhimurium virulence plasmid necessary for intracellular growth. Microb Pathog 7:165–173
     [Google Scholar]
  57. Taira S., Rhen M. 1989b; Molecular organization of genes constituting the virulence determinant on the Salmonella typhimurium 96 kilobase pair plasmid. FEBS Lett 257:274–278
     [Google Scholar]
  58. Taira S., Rhen M. 1990; Nucleotide sequence of mka D, a virulence-associated gene of Salmonella typhimurium containing variableconstant regions. Gene 93:147–150
     [Google Scholar]
  59. Taira S., Riikonen P., Saarilahti H., Sukupolvi S., Rhen M. 1991; The mka C virulence gene of the Salmonella serovar Typhimurium 96 kb plasmid encodes a transcriptional activator. MolGen Genet 228:381–384
     [Google Scholar]
  60. Takeshita S., Sato M., Toba M., Masahashi W., Hashimoto- Gotoh T. 1987; High-copy-numberlow-copy-number plasmid vectors for /rfc Za-complementationchloramphenicol- or kanamycin-resistance selection. Gene 61:63–74
     [Google Scholar]
  61. Viera J., Messing J. 1987; Production of single-strand plasmid DNA. Methods Enzymol 153:3–11
     [Google Scholar]
  62. Williamson G M, Baird G. D., Manning E. J. 1988a; A common virulence region on plasmids from eleven serotypes of Salmonella. J Gen Microbiol 134:975–982
     [Google Scholar]
  63. Williamson C. M., Pullinger G. D., Lax A. J. 1988b; Identification of an essential virulence region on Salmonella plasmids. Microb Pathog 5:469–473
     [Google Scholar]
  64. Woodward M. J., McLaren I., Wray C. 1989; Distribution of virulence plasmids within Salmonellae. J Gen Microbiol 135:503–511
     [Google Scholar]
  65. Yanisch-Perron G, Viera J., Messing J. 1985; Improved M3 cloning vectorshost strains: nucleotide sequences of the M13mpl8pUC19 vectors. Gene 33:103–119
     [Google Scholar]
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Virulence region of plasmid pNL2001 of Salmonella enteritidis
Microbiology 140, 1307 (1994); https://doi.org/10.1099/00221287-140-6-1307
/content/journal/micro/10.1099/00221287-140-6-1307
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