1887

Abstract

Monoclonal antibodies were produced to spore surface antigens. One particular hybridoma cell line, 43H6, produced a monoclonal antibody that reacted exclusively with cluster group 21 in an enzyme-linked immunosorbent assay (ELISA). Antibody 43H6 was found to be of subclass lgG1, kappa light chain. Western blot (immunoblot) analysis revealed that 43H6 recognized a major outer spore polypeptide of about 37000 Da. The epitope was stably maintained in spores over at least seven sporulation cycles on laboratory medium and for at least 14 weeks in sterile soil systems. The species group specificity of antibody 43H6 was exploited in the development of an immunocapture technique for the isolation of streptomycetes from soil. Magnetic beads coated with antibody 43H6 were mixed with soil samples seeded with spores. Spore-bead complexes were recovered using magnets. Treatment of beads with blocking agents and the inclusion of detergents in the recovery system lessened non-specific binding of spores to beads and improved recovery. In buffer solutions decreasing the spore concentration increased the recovery values for a fixed bead concentration. At a spore concentration of 5 × 10 ml the recovery was 4.3% whilst at 5 × 10 ml it was 76% for a fixed bead concentration of 0.6 mg ml. Using a bead concentration of 2 mg per 10 g soil, approximately 30% of the target spore population of 10 c.f.u. was recovered from sterile soil and 4% from non-sterile soil. This method offers a rapid means of selectively recovering and concentrating spores from soil samples.

Loading

Article metrics loading...

/content/journal/micro/10.1099/13500872-140-8-2067
1994-08-01
2021-05-07
Loading full text...

Full text loading...

/deliver/fulltext/micro/140/8/mic-140-8-2067.html?itemId=/content/journal/micro/10.1099/13500872-140-8-2067&mimeType=html&fmt=ahah

References

  1. Altenbucher J., Cullum J. DNA amplification and an unstable arginine gene in Streptomyces lividans 66. Mol A Gen Genet 1984; 195:134–138
    [Google Scholar]
  2. Betzler M., Dyson P., Schrempf H. Relationship of an unstable tfrjG gene to a 5'7-kilobasc amplifiable DNA sequence in Streptomyces Iividans 66. J batterioI 1987; 169:4804–4810
    [Google Scholar]
  3. Clark M., Waldmann H. Production of murine monoclonal antibodies. In Methods in Haematology 1985 Edited by Beverly P., Livingston C. London: Academic Press; pp 1–19
    [Google Scholar]
  4. Cresswell N., Herron P.R., Saunders V.A., Wellington E.M.H. The fate of introduced strcptomycctes, plasmid and phage populations in a dynamic soil system. J Gen Microbiol 1992; 138:659–666
    [Google Scholar]
  5. Cross T., Spooner D.F. The serological identification of strcptomvcctcs bv agar diffusion techniques. J Gen Microbiol 1963; 33:275–282
    [Google Scholar]
  6. Dyson P., Schrempf H. Genetic instability and DNA amplification in Streptomyces Iividans 66. J bacteriol 1987; 169:4796–4803
    [Google Scholar]
  7. Gates J.E., Pham K.D. An indirect fluorescent antibody staining technique for determining population levels of Thiobacillus ferrooxidans in acid mine drainage waters. Microb Eco 1979; 15:121–127
    [Google Scholar]
  8. Guijarro J.A., Suarez J.E., Salas J.A., Hardisson C. Pattern of protein degradation during germination of Streptomyces antibiotici spores. Can J Microbiol 1983; 29:637–643
    [Google Scholar]
  9. Guijarro J., Santamaria R., Schauer A., Losick R. Promoter determining the timing and spatial localization of transcription of a cloned Streptomyces coelicolor gene encoding a spore-associated polypeptide. J bacteriol 1988; 170:1895–1901
    [Google Scholar]
  10. Harlow E., Lane D. Aatibodies-A Laboratory Manual 1988 Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  11. Hopwood D.A., Bibb M.J., Chater K.F., Kieser T., Bruton C.J., Kieser H.M., Lydiate O.J., Smith C.D., Ward J.M., Scbompf H. Gettiu Mmipuiitiai tfSlrtptnmyiti: A Ldiorn/ery Maraal 198S Norwich: John Innes Foundation;
    [Google Scholar]
  12. Laemmli U.K. Cleavage of structural proteins during the assembly of rite head of bacteriophage T4. Nuture 1970; 227:680–685
    [Google Scholar]
  13. Levdsseur S., Husson M.-O., Leitz R., Merlin F., Laurent F., Peladan F., Drocourt J.L., Ledcc H., Van Hoegaerden M. Rapid detection of members of the family Enterobacteriaceae by a monoclonal antibody. Appi Emirem Mitrtbhl 1992; 58:1524–1529
    [Google Scholar]
  14. Lund A., Hellemann A.L., Vartdal F. Rapid isolation of K88+ Escbericbia coli by using immunomagnciic particles. J Che Micrehui 1988; 26:2572–2575
    [Google Scholar]
  15. Mason J., Bums R.G. Production of a monoclonal antibody specific for a Flavobacterium species isolated from soil. FEMS Microbiol Lett 1990; 73:290–308
    [Google Scholar]
  16. Mayfield C.I., Williams S.T., Ruddlck S.M., Hatfield H.L. Studies on the ecology of actinomyccies in soil IV Obictvations on the form and growth of streptomycetes in soil. Soil BiolBiochem 1972; 4:79–91
    [Google Scholar]
  17. McGroaty E.J., Rivera M. Growth-dependent alteration in production of serotype-specific and common antigen lipopolysaccharides in Pseudomonad aeruginosa PAOI. Infect Immun 1990; 57:1030–1037
    [Google Scholar]
  18. Morgan J.A.W., Winstanley C., Pickup R.W., Jones J.G., Saunders J.R. Direct phenotypic and genotypic derecrioo of a recombinant pseudomonad population released into lake water. Appl Environ Microbiol 1989; 55:2537–2544
    [Google Scholar]
  19. Morgan J.A.W., Winstanley C., Pickup R.W., Saunders J.R. Rapid immunocapture of Pseudomonas cells from lake water by using bacterial flagella. Appl Environ Microbiol 1991; 57:503–509
    [Google Scholar]
  20. Nelson O., Bathgate A.J., Poxton I.R. Monoclonal antibodies as ptobcs for detecting tipopolysacchandc expression on Eifhaiehid ah from different growth conditions. J Gen Microbiol 1991; 137:2741–2751
    [Google Scholar]
  21. Ngo T.T., Lenhoff H.M. Enzyme-mediated Immunoassay 1985 New York: Plenum Press;
    [Google Scholar]
  22. Ramos-Gonzilez M.L., Ruiz-Cabello F., Brettar I., Garrido F., Ramos J.L. Tracking genetically engineered bacteria: monoclonal antibodies against surface determinants of soil bic-tcrium Pntveetooi paSida 2440. J iUW 1992; 174:2978–2985
    [Google Scholar]
  23. Ridell M., Williams S.T. Serotaxonomic analysis of some Streptomyces and related organisms. J Gen Microbiol 1983; 129:2857–2861
    [Google Scholar]
  24. Ridell M., Wallerstrom G., Williams S.T. Immunodiffusion analyses of phonetically defined strains of Strepfonryeet, StreptoverticiUum and Nocardiopsis. Syst AppI Microbiol 1986; 8:24–27
    [Google Scholar]
  25. Smith A.W., Wilton J., Clark S.A., Alpar O., Melling J., Brown M.R.W. Production and characterization of monoclonal antibodies to outer membrane proteins of Pseudomonas aeruginosa grown in iron depleted media. J Gen Microbiol 1991; 137:227–236
    [Google Scholar]
  26. Strayer R.F., Tiedje J.M. Application of the fluorescent antibody technique to the study of methanogenic bacterium in lake sediments. Appl Environ Microbiol 1978; 35:192–197
    [Google Scholar]
  27. Tjissen P. Laboratory Techniques in biochemistry and Molecular Biology, Practice and Theory of Ensjmt Immunoassays 1985 New York: Elsevier; 15
    [Google Scholar]
  28. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from SDS and acid/urea polyacrylamide gels to nitrocellulose sheets, procedure and some applications. Proc Natl Acad Sci USA 1979; 76:4350–4354
    [Google Scholar]
  29. Ward J.M., Janssen G.R., Keiser T., Bibb M.J., Bibb M.J. Construction and characterisation of a series of multi-copy promotcr-probc vectors for Streptomyces using the aminoglycoside phosphotransferase gene from Tn5 as indicator. Mol & Gen Genet 1986; 203:468–478
    [Google Scholar]
  30. Wellington E.M.H., Cresswell N., Saunders V.A. Growth and survival of strcptomycctc inoculants and extent of plasmid transfer in sterile and nonsterile soil. Appl Environ Microbiol 1990; 56:1413–1419
    [Google Scholar]
  31. Williams S.T., Goodfellow M., Alderson G., Wellington E.M.H., Sneath P.H.A., Sackin M.J. Numerical classification of Streptomyces and related genera. J Gen Microbiol 1983; 129:1743–1813
    [Google Scholar]
  32. Wipat A., Wellington E.M.H., Saunders V.A. Streptomyces marker plasmids for monitoring survival and spread of streptomycetes in soil. Appl Environ Microbiol 1991; 57:3322–3330
    [Google Scholar]
  33. Wright S.F., Morton J.B., Sworobuk J.E. Identification of a vcsicular-arbuscular mycorrhizal fungus by using monoclonal antibodies in an enzyme linked immunosorbent assay. Appl Environ Microbiol 1987; 53:2222–2225
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/13500872-140-8-2067
Loading
/content/journal/micro/10.1099/13500872-140-8-2067
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