1887

Abstract

Summary

Cytotoxic necrotising factors type 1 (CNF1) and type 2 (CNF2) are produced by many strains isolated from man and animals with intestinal or extra-intestinal colibacillosis. In most laboratories, CNF-producing strains are detected by a cell cytotoxicity assay and confirmed with a neutralisation assay or a mouse footpad assay. In this study, we sought to determine whether DNA probes could detect clinical isolates of producing CNF2 or CNF1, or both, without the need for cell cultures or animal assays. Two internal fragments of the gene encoding CNF2 were used as DNA probes: a 875-bp I-I DNA fragment and an adjacent 335-bp I-I fragment. A positive response with both DNA probes was associated with CNF2-producing strains, whereas a positive response with only the 335-bp probe was associated with CNF1-producing strains. Results of colony hybridisation experiments with 185 clinical isolates of demonstrated that these DNA probes detected CNF2-producing strains with a sensitivity and specificity of 100% and CNF1-producing strains with a sensitivity and specificity of 99 %. These two DNA probes should greatly facilitate epidemiological studies to assess the importance of CNF-producing strains as agents of diarrhoea and septicaemia.

Loading

Article metrics loading...

/content/journal/jmm/10.1099/00222615-40-6-428
1994-06-01
2024-04-18
Loading full text...

Full text loading...

/deliver/fulltext/jmm/40/6/medmicro-40-6-428.html?itemId=/content/journal/jmm/10.1099/00222615-40-6-428&mimeType=html&fmt=ahah

References

  1. Caprioli A, Falbo V, Roda LG, Ruggeri FM, Zona C. Partial purification and characterization of an Escherichia coli toxic factor that induces morphological cell alterations. Infect Immun 1983; 39:1300–1306
    [Google Scholar]
  2. De Rycke J, Gonzalez EA, Blanco J, Oswald E, Blanco M, Boivin R. Evidence of two types of cytotoxic necrotizing factor in human and animal clinical isolates of Escherichia coli. J Clin Microbiol 1990; 28:694–699
    [Google Scholar]
  3. De Rycke J, Guillot JF, Boivin R. Cytotoxins in non-enterotoxigenic strains of Escherichia coli isolated from faeces of diarrheic calves. Vet Microbiol 1987; 15:137–150
    [Google Scholar]
  4. Caprioli A, Donelli G, Falbo V. A cell division-active protein from E. coli. Biochem Biophys Res Commun 1984; 118:587–593
    [Google Scholar]
  5. De Rycke J, Phan-Thanh L, Bernard S. Immunochemical identification and biological characterization of cytotoxic necrotizing factor from Escherichia coli. J Clin Microbiol 1989; 27:983–988
    [Google Scholar]
  6. Oswald E, De Rycke J. A single protein of 110 kDa is associated with the multinucleating and necrotizing activity coded by the Vir plasmid of Escherichia coli . FEMS Microbiol Lett 1990; 68:278–284
    [Google Scholar]
  7. Falbo V, Famiglietti M, Caprioli A. Gene block encoding production of Cytotoxic Necrotizing Factor 1 and hemo-lysin in Escherichia coli isolates from extraintestinal infections. Infect Immun 1992; 60:2182–2187
    [Google Scholar]
  8. Oswald E, De Rycke J, Guillot JF, Boivin R. Cytotoxic effect of multinucleation in HeLa cell cultures associated with the presence of Vir plasmid in Escherichia coli strains. FEMS Microbiol Lett 1989; 58:95–100
    [Google Scholar]
  9. Lopez-Alvarez J, Gyles CL. Occurrence of the Vir plasmid among animal and human strains of invasive Escherichia coli . Am J Vet Res 1980; 41:169–11 A
    [Google Scholar]
  10. Lopez-Alvarez J, Gyles CL, Shipley PL, Falkow S. Genetic and molecular characteristics of the Vir plasmids of bovine septicemic Escherichia coli . J Bacteriol 1980; 141:758–769
    [Google Scholar]
  11. Smith HW. A search for transmissible pathogenic characters in invasive strains of Escherichia coli: the discovery of a plasmid-controlled toxin and a plasmid-controlled lethal character closely associated, or identical, with Colicine V. J Gen Microbiol 1974; 83:95–111
    [Google Scholar]
  12. Smith HW. Observations on Escherichia coli infection in calves. Rutter JM. Proceedings of the first seminar on “pathology” in the CEC programme of coordination of research in beef production Perinatal ill-health in calves 197547–59
    [Google Scholar]
  13. Wray C, Piercy DWT, Carroll PJ, Cooley WA. Experimental infection of neonatal pigs with CNF toxin-producing strains of Escherichia coli . Res Vet Sci 1993; 54:290–298
    [Google Scholar]
  14. Bisicchia R, Ciammarughi R, Caprioli A, Falbo V, Ruggeri FM. Toxin production and haemagglutination in strains of Escherichia coli from diarrhoea in Brescia, Italy. J Hyg 1985; 95:353–361
    [Google Scholar]
  15. Alonso MP, Blanco J, Blanco M, Gonzalez EA. Frequent production of toxins by Escherichia coli strains isolated from human urinary tract infections: relation with haemagglutination. FEMS Microbiol Lett 1987; 48:391–396
    [Google Scholar]
  16. Caprioli A, Falbo V, Ruggeri FM. Cytotoxic necrotizing factor production by hemolytic strains of Escherichia coli causing extraintestinal infections. J Clin Microbiol 1987; 25:146–149
    [Google Scholar]
  17. Blanco J, Alonso MP, Gonzales EA, Blanco M, Garabal JI. Virulence factors of bacteremic Escherichia coli with particular reference to production of cytotoxic necrotizing factor (CNF) by P-fimbriate strains. J Med Microbiol 1990; 31:175–183
    [Google Scholar]
  18. Cherifi A, Contrepois M, Picard B. Factors and markers of virulence in Escherichia coli from human septicemia. FEMS Microbiol Lett 1990; 70:279–284
    [Google Scholar]
  19. Pohl P, Mainil J, Devriese L. Escherichia coli productrices de la toxine cytotoxique necrosant de type 1 (CNF1) isolees a partir de processus pathologiques chez des chats et des chiens. Ann Med Vet 1992; 137:21–25
    [Google Scholar]
  20. Pohl P, Oswald E, Van Muylem K, Jacquemin E, Lintermans P, Mainil J. Escherichia coli producing CNF1 and CNF2 cytotoxins in animals with different disorders. Vet Res in press 1993; 24:311–315
    [Google Scholar]
  21. Prada J, Baljer G, De Rycke J. Characteristics of alpha-hemolitic strains of Escherichia coli isolated from dogs with gastroenteritis. Vet Microbiol 1991; 29:59–73
    [Google Scholar]
  22. Gonzalez EA, Blanco J. Production of cy to toxin VT in enteropathogenic and non-enteropathogenic Escherichia coli strains of porcine origin. FEMS Microbiol Lett 1985; 26:127–129
    [Google Scholar]
  23. Blanco J, Gonzalez EA, Blanco M, Regueiro B, Bernardez I. Production of toxins by Escherichia coli strains isolated from calves with diarrhoea in Galicia (north-western Spain). Vet Microbiol 1988; 18:297–311
    [Google Scholar]
  24. Pohl P, Daube G, Lintermans P, Kaeckenbeeck A, Mainil J, Oswald E. Facteurs de virulence et phenotypes de soixante et une souches d’Escherichia coli d’origine bovine, productrices de la toxine cytotoxique necrosante de type 1 (CNF1). Ann Rech Vet 1991; 23:83–91
    [Google Scholar]
  25. Oswald E, De Rycke J, Lintermans P. Virulence factors associated with the cytotoxic necrotizing factor type two in bovine diarrheic and septicemic strains of Escherichia coli . J Clin Microbiol 1991; 29:2522–2527
    [Google Scholar]
  26. De Rycke J, Oswald E, Boivin R. An in vivo assay for the detection of cytotoxic strains of Escherichia coli . Ann Rech Vet 1989; 20:39–46
    [Google Scholar]
  27. Blanco J, Blanco M, Gonzalez EA, Alonso MP, Garabal JI. Comparative evaluation of three tests for the detection of Escherichia coli cytotoxic necrotizing factors (CNF1 and CNF2) using filtrates of cultures treated with mitomycin C. FEMS Microbiol Lett 1990; 69:311–316
    [Google Scholar]
  28. Tabouret M, De Rycke J. Detection of cytotoxic necrotizing factor (CNF) in extracts of Escherichia coli strains by enzyme-linked immunosorbent assay. J Med Microbiol 1990; 32:73–81
    [Google Scholar]
  29. Oswald E, Tabouret M, Boivin R, De Rycke J. Detection of Escherichia coli strains producing cytotoxic necrotizing factor type two (CNF2) by enzyme-linked immunosorbent assay. Vet Microbiol (in press) 1994 in press
    [Google Scholar]
  30. Oswald E, Sugai M, Labigne A. Cytotoxic necrotizing factor type 2 produced by virulent Escherichica coli modifies the small GTP-binding proteins Rho involved in actin stress fibers assembly. Proc Natl Acad USA (in press) 1994; 91: in press
    [Google Scholar]
  31. Falbo V, Pace T, Picci L, Pizzi E, Caprioli A. Isolation and nucleotide sequence of the gene encoding cytotoxic necrot-izing factor 1 of Escherichia coli . Infect Immun 1993; 61:4909–4914
    [Google Scholar]
  32. Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory; 1989
    [Google Scholar]
  33. Mainil J, Bex G, Jacquemin E, Pohl P, Couturier M, Kaecken-beeck A. Prevalence of four enterotoxins (STaP,STaH, STb and LT) and four adhesin subunits (K99, K88, 987P, and F41) gene among Escherichia coli isolate from cattle. Am J Vet Res 1990; 51:187–190
    [Google Scholar]
  34. Kado CI, Liu S-T. Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol 1981; 145:1365–1373
    [Google Scholar]
  35. Broes A, Fairbrother JM, Mainil J, Harel J, Lariviere S. Phenotypic and genotypic characterization of entero-toxigenic Escherichia coli serotype O8:KX105 and 08 :K “2829” strains isolated from piglets with diarrhea. J Clin Microbiol 1988; 26:2402–2409
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jmm/10.1099/00222615-40-6-428
Loading
/content/journal/jmm/10.1099/00222615-40-6-428
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error