Hostname: page-component-cd9895bd7-mkpzs Total loading time: 0 Render date: 2024-12-23T00:00:09.524Z Has data issue: false hasContentIssue false
  • English
  • Français

Supplementation of barley straw with Sesbania pachycarpa leaves in vitro: effects on fermentation variables and rumen microbial population structure quantified by ribosomal RNA-targeted probes

Published online by Cambridge University Press:  01 August 2008

S. Muetzel
S. Muetzel*
Affiliation:
Institute for Animal Production in the Tropics and Subtropics (480), University of Hohenheim, Fruwirthstr. 12, D-70593 Stuttgart, Germany
*
*Corresponding Author:Professor Dr K. Becker, fax +49 711 459 3702, email [email protected]
Rights & Permissions [Opens in a new window]

Abstract

Core share and HTML view are not available for this content. However, as you have access to this content, a full PDF is available via the ‘Save PDF’ action button.

Tropical livestock is often maintained on roughage-based diets deficient in N, and therefore requires supplementation with protein-rich substrates to achieve reasonable production levels. The optimum inclusion rate of a potential supplement is usually determined by in vivo feeding trials or by in vitro incubation of the diet components to estimate the feed value of the complete diet. The present work simulates a supplementation experiment in vitro, by incubating a pure roughage (barley straw), a pure supplement (Sesbania pachycarpa leaves) and mixtures of the two, with increasing inclusion levels of the supplement, in a short-term batch incubation system. Fermentation kinetics were followed by the release of fermentation endproducts (gas and short-chain fatty acids). Microbial biomass was estimated using ribosomal (r) RNA as internal marker for bacteria and eukaryotes separately. Cell-wall-degrading subpopulations were quantified by hybridisation with taxon-specific oligonucleotide probes targeting Chytridiomycetes, Fibrobacter spp., Ruminococcus albus and R. flavefaciens. Carboxymethylcellulase (CMCase) was assayed as an indicator for cell-wall-degrading activity. The addition of S. pachycarpa leaves stimulated fermentation in all cases. Gas production, and especially rRNA concentration, showed clear maxima at 40 % S. pachycarpa inclusion, rates that significantly exceeded the values interpolated from the incubations of the pure substrates. Short-chain fatty acid yield changed only slightly, but in the same way. The analysis of the microbial population structure showed that the positive effects were mainly mediated through enhanced growth of Ruminococcus spp. Increasing proportions of S. pachycarpa leaves in the diet led to a drastic decline in the total eukaryotic population. This points to a defaunation, which may also have added to the positive effects. The eukaryotic subpopulation of the rumen fungi were affected to a lesser degree. Although the cell-wall-degrading organisms showed positive responses to the supplementation, the CMCase activity was not affected significantly by the supplementation. The present work shows that it is possible to predict optimum inclusion levels for a new feed supplement in vitro and thus reduce in vivo experiments. It was also demonstrated that true supplementation effects occur particularly for the microbial biomass production, which is the primary source of amino acids for the ruminant animal. The analysis of microbial population structure in context with conventional metabolic measurements adds valuable information to interpret the observed effects on production-related variables.

Keywords

Microbial biomassGas testRibosomal RNA probes
Type
Research Article
Information
British Journal of Nutrition , Volume 89 , Issue 4 , April 2003 , pp. 445 - 453
Copyright
Copyright © The Nutrition Society 2003

References

Amann, RI, Krumholz, L & Stahl, DA (1990) Fluorescent oligonucleotide probing of whole cells for determinative, phylogenetic and environmental studies in microbiology. Journal of Bacteriology 172, 762–770.CrossRefGoogle ScholarPubMed
Association of Official Analytical Chemists International (1990) Official Methods of Analysis, 15th ed., Arlington, VA: AOAC.Google Scholar
Atasoglu, C, Valdes, C, Newbold, CJ & Wallace, RJ (1999) Influence of peptides and amino acids on fermentation rate and de novo synthesis of amino acids by mixed micro-organisms from the sheep rumen. British Journal of Nutrition 81, 307–314.CrossRefGoogle ScholarPubMed
Blümmel, M & Becker, K (1997) The degradability characteristics of fifty-four roughages and roughage neutral-detergent fibres as described by in vitro gas production and their relationship to voluntary feed intake. British Journal of Nutrition 77, 757–768.CrossRefGoogle ScholarPubMed
Blümmel, M, Makkar, HS & Becker, K (1997a). In vitro gas production – a technique revisited. Journal of Animal Physiology and Animal Nutrition 77, 24–34.CrossRefGoogle Scholar
Blümmel, M, Paul, C, Goodchild, A & Becker, K (1996) Grinding energy and in vitro gas technique for the assessment of Syrian barley straws – physical and microbial degradation and voluntary feed intake by sheep. Journal of Animal Physiology and Animal Nutrition 76, 132–140.CrossRefGoogle Scholar
Blümmel, M, Steingass, H & Becker, K (1997b) The relationship between in vitro gas production, in vitro microbial biomass yield and 15N incorporation and its implications for the prediction of voluntary feed intake of roughages. British Journal of Nutrition 77, 911–921.CrossRefGoogle ScholarPubMed
Bonsi, MLK, Osuji, PO, Tuah, AK & Umunna, NN (1995) Intake, digestibility, nitrogen balance and certain rumen characteristics of Ethiopian Menz sheep fed teff straw supplemented with cotton seed cake, dry Sesbania, dry Leucaena or fresh Leucaena. Agroforestry Systems 31, 243–256.CrossRefGoogle Scholar
Briesacher, SL, May, T, Grigsby, KN, Kerley, MS, Anthony, RV & Paterson, JA (1992) Use of DNA probes to monitor nutritional effects on ruminal prokaryotes and Fibrobacter succinogenes S85. Journal of Animal Science 70, 289–295.CrossRefGoogle ScholarPubMed
Broudiscou, L & Jouany, JP (1995) Reassessing the manipulation of protein synthesis by rumen microbes. Reproduction Nutrition Development 35, 517–535.CrossRefGoogle ScholarPubMed
Cone, JW & van Gelder, AH (1999) Influence of protein fermentation on gas production profiles. Animal Feed Science and Technology 7, 251–264.CrossRefGoogle Scholar
Dore, J, Brownlee, AG, Millet, L, Virlogeux, I, Saigne, M, Fonty, G & Gouet, P (1993) Ribosomal DNA-targeted hybridization probes for the detection, identification and quantitation of anaerobic rumen fungi. Proceedings of the Nutrition Society 52, 176.Google Scholar
Dryhurst, N & Wood, CD (1998) The effect of nitrogen source and concentration on in vitro gas production using rumen micro-organisms. Animal Feed Science and Technology 71, 131–143.CrossRefGoogle Scholar
Groleau, D & Forsberg, CW (1981) Cellulolytic activity of the rumen bacterium Bacteroides succinogenes. Canadian Journal of Microbiology 27, 517–530.CrossRefGoogle ScholarPubMed
Goehring, HK & Van Soest, PJ (1970) Forage Fiber Analyses Agricultural Handbook, no. 379. pp. 20–52. Washington, DC: USDA.Google Scholar
Groot, JCJ, Williams, BA, Oostdam, AJ, Boer, H & Tamminga, S (1998) The use of cumulative gas and volatile fatty acid production to predict in vitro fermentation kinetics of Italian ryegrass leaf cell walls and contents at various time intervals. British Journal of Nutrition 79, 519–525.CrossRefGoogle ScholarPubMed
Hespell, RB & Bryant, MP (1979) Efficiency of rumen microbial growth: Influence of some theoretical and experimental factors in YATP. Journal of Animal Science 49, 1640–1659.CrossRefGoogle ScholarPubMed
Hicks, RE, Amann, R & Stahl, DA (1992) Dual staining of natural bacterioplankton with 4′, 6-diamidino-2-phenylindole and fluorescent oligonucleotide probes targeting kingdom-level 16S rRNA sequences. Applied and Environmental Microbiology 58, 2158–2163.CrossRefGoogle ScholarPubMed
Holtershinken, M, Plitt, U, Tammen, FC, Hoffmann, P & Scholz, H (1997) Influence of mouldy grass on fermentation and thiamine metabolism in bovine rumen fluid (in vitro). Deutsche Tierarztliche Wochenschrift 104, 17–22.Google ScholarPubMed
Huhtanen, P, Vanhatalo, A & Varvikko, T (1998) Enzyme activities of rumen particles and feed samples incubated in situ with differing types of cloth. British Journal of Nutrition 79, 161–168.CrossRefGoogle ScholarPubMed
John, A (1984) Effects of feeding frequency and level of feed intake on chemical composition of rumen bacteria. Journal of Agricultural Science 102, 45–57.CrossRefGoogle Scholar
Khandaker, ZH, Steingass, H & Drochner, W (1998) Supplementation of wheat straw with Sesbania on voluntary intake, digestibility and ruminal fermentation in sheep. Small Ruminant Research 28, 23–29.CrossRefGoogle Scholar
Krause, DO, Smith, WJM, Ryan, FME, Mackie, RI & McSweeney, CS (1999) Use of 16S-rRNA based techniques to investigate the ecological succession of microbial populations in the immature lamb rumen: Tracking of a specific strain of inoculated Ruminococcus and interactions with other microbial populations in vivo. Microbial Ecology 38, 365–376.CrossRefGoogle ScholarPubMed
Li, J & Heath, IB (1993) Chytridiomycetous gut fungi, oft over-looked contributors to herbivore digestion. Canadian Journal of Microbiology 39, 1003–1013.CrossRefGoogle Scholar
Malburg, LM & Fosberg, CW (1993) Fibrobacter succinogenes S85 possesses at least nine distinct glucanase genes. Canadian Journal of Microbiology 39, 882–891.CrossRefGoogle Scholar
May, T, Kerley, MS & Williams, JE (1993) Supplemental protein influences on carbohydrate degradation and bacterial 16S ribosomal ribonucleic acid. Journal of Dairy Science 76, 3479–3489.CrossRefGoogle ScholarPubMed
Menke, KH, Raab, L, Salewski, A, Steingass, H, Fritz, D & Schneider, W (1979) The estimation of the digestibility and metabolizable energy content of ruminant feedingstuffs from the gas production when they are incubated with rumen liquor in vitro. Journal of Agricultural Science 93, 217–222.CrossRefGoogle Scholar
Merry, RJ & McAllan, AB (1983) A comparison of the chemical composition of mixed bacteria harvested from the liquid and solid fractions of rumen digesta. British Journal of Nutrition 50, 701–709.CrossRefGoogle ScholarPubMed
Muetzel, S & Becker, K (2002) An optimised RNA extraction protocol for rumen samples. Reproduction Nutrition Development 42, Suppl. 1, S8.Google Scholar
Muetzel, S, Krishnamoorthy, U & Becker, K (2001) Effects of rumen fluid collection site on microbial population structure during in vitro fermentation of different substrates quantified by 16S rRNA hybridisation. Archives of Animal Nutrition 55, 103–120.Google ScholarPubMed
Newbold, CJ, Elhassan, SM, Wang, J, Ortega, ME & Wallace, RJ (1997) Influence of foliage from African multipurpose trees on activity of rumen protozoa and bacteria. British Journal of Nutrition 78, 237–249.CrossRefGoogle ScholarPubMed
Nierlich, DP (1978) Regulation of bacterial growth, RNA, and protein synthesis. Annual Reviews in Microbiology 32, 393–432.CrossRefGoogle ScholarPubMed
Odenyo, AA, Mackie, RI, Stahl, DA & White, BA (1994) The use of 16S rRNA-targeted oligonucleotide probes to study competition between ruminal fibrolytic bacteria: Pure-culture studies with cellulose and alkaline peroxide-treated wheat straw. Applied and Environmental Microbiology 60, 3697–3703.CrossRefGoogle ScholarPubMed
Odenyo, AA, Osuji, PO & Karanfil, O (1997) Effect of multipurpose tree (MPT) supplements on ruminal ciliate protozoa. Animal Feed Science and Technology 67, 169–180.CrossRefGoogle Scholar
Olsen, GJ, Lane, DJ, Giovannoni, J & Pace, NR (1986) Microbial ecology and evolution: a ribosomal RNA approach. Annual Review of Microbiology 40, 337–365.CrossRefGoogle ScholarPubMed
Pettipher, GL & Latham, MJ (1979) Characteristics of enzymes produced by Ruminococcus flavefaciens which degrade plant cell walls. Journal of General Microbiology 110, 21–27.CrossRefGoogle Scholar
Raskin, L, Amann, RI, Poulsen, LK, Rittmann, BE & Stahl, DA (1995) Use of ribosomal RNA based molecular probes for characterization of complex microbial communities in anaerobic biofilms. Water Science and Technology 31, 261–272.CrossRefGoogle Scholar
Robinson, PH, Fadel, JG & Ivan, M (1996) Critical evaluation of diaminopimelic acid and ribonucleic acid as markers to estimate rumen pools and duodenal flows of bacterial and protozoal nitrogen. Canadian Journal of Animal Science 76, 587–597.CrossRefGoogle Scholar
Sayler, GS & Layton, AC (1990) Environmental application of nucleic acid hybridization. Annual Review of Microbiology 44, 625–648.CrossRefGoogle ScholarPubMed
Stahl, OA, Flesher, B, Mansfield, HA & Montgomery, L (1988) Use of phylogenetically based hybridization probes for studies of ruminal microbial ecology. Applied and Environmental Microbiology 54, 1079–1084.CrossRefGoogle ScholarPubMed
Stewart, CS & Bryant, MP (1997) The rumen bacteria. In The Rumen Microbial Ecosystem, pp. 10–72 [Hobson, PN, editor]. London: Blackie Academic & Professional.CrossRefGoogle Scholar
Varga, GA & Kolver, ES (1997) Microbial and animal limitations to fiber digestion and utilization. Journal of Nutrition 127, 5-S.CrossRefGoogle ScholarPubMed
Volden, H, Harstad, OM & Mydland, LT (1999) Amino acid content and profile of protozoal and bacterial fractions isolated from ruminal contents of lactating dairy cows fed diets differing in nitrogen supplementation. Acta Agriculturae Scandinavica 49A, 245–250.CrossRefGoogle Scholar
Wolin, MJ (1960) A theoretical rumen fermentation balance. Journal of Dairy Science 40, 1452–1478.CrossRefGoogle Scholar
Wood, CO & Manyuchi, B (1997) Use of an in vitro gas production method to investigate interactions between Veld hay and Napier hay or groundnut hay supplements. Animal Feed Science and Technology 67, 265–278.CrossRefGoogle Scholar
Zheng, DD, Aim, EW, Stahl, OA & Aaskin, L (1996) Characterization of universal small-subunit rRNA hybridization probes for quantitative molecular microbial ecology studies. Applied and Environmental Microbiology 62, 4504–4513.CrossRefGoogle ScholarPubMed