1887

Abstract

Ruminal bacteria and protozoa, and cell-free rumen fluid, were tested for the presence of enzymes involved in the degradation of the fungal cell wall. Protozoal homogenate obtained by ultrasonication showed chitinase (EC 3.2.1.14) and -acetyl-β-glucosaminidase (EC 3.2.1.52) activities when assayed with fluorogenic 4-methylumbelliferyl substrates. The chitinase activity was predominantly of the ‘exo’-type. Lysozyme (EC 3.2.1.17) and 1,3--glucanase (EC 3.2.1.39) activities were also present in this fraction. All these activities, except lysozyme activity, were recovered mainly in the supernatant fraction of the homogenate (approximately 85% of the total activity). Lysozyme showed the same amount of activity in the precipitate and supernatant fractions. Bacterial homogenates had -acetyl--glucosaminidase activity in both supernatant and precipitate fractions. The specific activity was one-third that of the protozoa. Bacteria able to grow in a medium with chitin as the sole carbon source were recognized and counted. Cell-free rumen fluid was unable to degrade any of the substrates tested.

Loading

Article metrics loading...

/content/journal/micro/10.1099/00221287-140-3-631
1994-03-01
2021-08-01
Loading full text...

Full text loading...

/deliver/fulltext/micro/140/3/mic-140-3-631.html?itemId=/content/journal/micro/10.1099/00221287-140-3-631&mimeType=html&fmt=ahah

References

  1. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 1976; 72:248–254
    [Google Scholar]
  2. Caldwell D. R., Bryant M. P. Medium without rumen fluid for nonselective enumeration and isolation of rumen bacteria. Appl Microbiol 1966; 14:794–801
    [Google Scholar]
  3. Clesceri L. S., Greenberg A. E., Trussell R. R. 1989; Multipletube fermentation technique for members of the coliform group. In Standard Methods for the Examination of Water and Wastewater, 17th edn. pp. 9-66–9-80 Washington, DC: American Public Health Association, American Water Works Association, and Water Pollution Control Federation;
    [Google Scholar]
  4. Delmotte F. M., Privât J.-P. D. J., Monsigny M. L. P. Interactions glycanne-protéine. Synthèse des 4-méthylombelliféryl- (2-acétamido-2-désoxy-β-d-glucopyranoside), -di-N-acétyl-β-chito-bioside et-tri-N-acétyl-β-chitotrioside. Interaction de ces osides avec le lysozyme. Carbohydr Res 1975; 40:353–364
    [Google Scholar]
  5. Dygert S., Li L. H., Florida D., Thoma J. A. De¬termination of reducing sugar with improved precision. Anal Biochem 1965; 13:367–374
    [Google Scholar]
  6. Eadie J. M. The development of rumen microbial populations in lambs and calves under various conditions of man-agement. J Gen Microbiol 1962; 29:563–578
    [Google Scholar]
  7. Farkas V. Fungal cell walls: their structure, biosynthesis and biotechnological aspects. Acta Biotechnol 1990; 10:225–238
    [Google Scholar]
  8. Ghuysen J.-M., , Tipper D. J., Strominger J. L. Enzymes that degrade bacterial cell walls. Methods Enzymol 1966; 8:685–699
    [Google Scholar]
  9. Gooday G. W. Chitinases. In Enzymes in Biomass Conversion (1990a) Edited by Leatham G., Himmel M. Washington, DC: American Chemical Society; pp 478–485
    [Google Scholar]
  10. Gooday G. W. The ecology of chitin degradation. Adv Microb Ecol (1990b); 11:387–430
    [Google Scholar]
  11. Hood M. A. Comparison of four methods for measuring chitinase activity and the application of the 4-MUF assay in aquatic environments. J Microbiol Methods 1991; 13:151–160
    [Google Scholar]
  12. Hungate R. E. A roll tube method for cultivation of strict anaerobes. Methods Microbiol 1969; 3B:117–132
    [Google Scholar]
  13. Ling J. R. Digestion of bacterial cell walls in the rumen. In The Rumen Ecosystem. The Microbial Metabolism and Its Regulation 1990 Edited by Hoshino S., Onodera R., Minato H., Itabashi H. Tokyo: Japan Scientific Societies Press; /italic> pp 83–90
    [Google Scholar]
  14. Majeau N., Trudel J., Asselin A. Diversity of cucumber chitinase isoforms and characterization of one seed basic chitinase with lysozyme activity. Plant Sci 1990; 68:9–16
    [Google Scholar]
  15. McCreath K. J., Gooday G. W. A rapid and sensitive microassay for determination of chitinolytic activity. J Microbiol Methods 1992; 14:229–237
    [Google Scholar]
  16. Morgavi D. P., Onodera R., Nagasawa T. In vitro metabolism of chitin and protein from ruminal fungi by ruminal protozoa. Anim Sci Technol 1993; 64:584–592
    [Google Scholar]
  17. Nagasawa T., Tamaru K., Onodera R. A new intracellular proteolytic activity in mixed rumen ciliate protozoa. Asian J Anim Sci 1989; 2:446–448
    [Google Scholar]
  18. Newbold C. J., Hillman K. The effect of ciliate protozoa on the turnover of bacterial and fungal protein in the rumen of sheep. Eett Appl Microbiol 1990; 11:100–102
    [Google Scholar]
  19. O’Brien M., Colwell R. R. A rapid test for chitinase activity that uses 4-methylumbelliferyl-N-acetyl-β-d-glucosamin- ide. Appl Environ Microbiol 1987; 53:1718–1720
    [Google Scholar]
  20. Ogimoto K., Imai S. 1981 Atlas of Rumen Microbiology Tokyo: Japan Scientific Societies Press;
    [Google Scholar]
  21. Onodera R., 8t Henderson C. Growth factors of bacterial origin for the culture of the rumen oligotrich protozoon, Entodinium caudatum . J Appl Bacteriol 1980; 48:125–134
    [Google Scholar]
  22. Onodera R., Yamaguchi H., Eguchi C., Kandatsu M. Limits of survival of the mingled rumen bacteria in the washed cell suspension of rumen ciliate protozoa. Agric Biol Chem 1977; 41:2465–2466
    [Google Scholar]
  23. Orpin C. G. The occurrence of chitin in the cell walls of the rumen organisms Neocallimastix frontalis, Piromonas communis and Sphaeromonas communis . J Gen Microbiol 1977; 99:215–218
    [Google Scholar]
  24. Orpin C. G. The role of ciliate protozoa and fungi in the rumen digestion of plant cell walls. Anim Feed Sci Technol 1984; 10:121–143
    [Google Scholar]
  25. Patton R. S., Chandler P. T. In vivo digestibility evaluation of chitinous materials. J Dairy Sci 1975; 58:397–401
    [Google Scholar]
  26. Robbins P. W., Albright C., Benfield B. Cloning and expression of a Streptomyces plicatus chitinase (chitinase-63) in Escherichia coli . J Biol Chem 1988; 263:443–447
    [Google Scholar]
  27. Romulo B., Bird S. H., Leng R. A. Combined effects of defaunation and protein supplementation on intake, digestibility, N retention and fungi counts in sheep fed straw based diets. In The Roles of Protozoa and Fungi in Ruminant Digestion 1989 Edited by Nolan J. V., Leng R. A., Demeyer D. I. Armidale, NSW: Penambul Books; pp 285–288
    [Google Scholar]
  28. Shimahara K., Takiguchi Y. Preparation of crustacean chitin. Methods Ensymol 1988; 161:417–423
    [Google Scholar]
  29. Snedecor G. W., Cochran W. G. 1980 Statistical Methods, 7th edn.. Ames, Iowa: Iowa State University Press;
    [Google Scholar]
  30. Somogyi M. Notes on sugar determination. J Biol Chem 1952; 195:19–23
    [Google Scholar]
  31. Spiro R. G. Analysis of sugars found in glycoproteins. Methods Ensymol 1966; 8:3–26
    [Google Scholar]
  32. Theodorou M. K., Gill M., King-Spooner C., Beever D. E. Enumeration of anaerobic chytridiomycetes as thallus-forming units: novel method for quantification of fibrolytic fungal populations from the digestive tract ecosystem. Appl Environ Microbiol 1990; 54:1073–1078
    [Google Scholar]
  33. Ushida K., Tanaka H., Kojima Y. A simple in situ method for estimating fungal population size in the rumen. Eett Appl Microbiol 1989; 9:109–111
    [Google Scholar]
  34. Williams A. G., Strachan N. H. The distribution of polysaccharide-degrading enzymes in the bovine rumen digesta ecosystem. Curr Microbiol 1984; 10:215–220
    [Google Scholar]
  35. Williams A. G., Withers S. E., Coleman G. S. Glycoside hydrolases of rumen bacteria and protozoa. Curr Microbiol 1984; 10:287–294
    [Google Scholar]
  36. Williams A. G., Ellis A. B., Coleman G. S. Subcellular distribution of polysaccharide depolymerase and glycoside hydrolase enzymes in rumen ciliate protozoa. Curr Microbiol 1986; 13:139–147
    [Google Scholar]
  37. Williams A. G., Withers S. E., Strachan N.H. Postprandial variations in the activity of polysaccharide-degrading enzymes in microbial populations from the digesta solids and liquor fractions of rumen contents. J Appl Bacteriol 1989; 66:15–26
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/00221287-140-3-631
Loading
/content/journal/micro/10.1099/00221287-140-3-631
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