Skip to main content
SpringerLink
Log in

Quantitative qPCR Analysis of Ruminal Microorganisms in Beef Cattle Grazing in Pastures in the Rainy Season and Supplemented with Different Protein Levels

  • Published:
Current Microbiology Aims and scope Submit manuscript

Abstract

We tested the hypothesis that supplementation with three protein levels improves fermentation parameters without changing the rumen microbial population of grazing beef cattle in the rainy season. Four rumen-cannulated Nellore bulls (432 ± 21 kg of body weight) were used in a 4 × 4 Latin square design with four supplements and four experimental periods of 21 days each. The treatments were mineral supplement (ad libitum) and supplements with low, medium (MPS), and high protein supplement (HPS), supplying 106, 408, and 601 g/day of CP, respectively. The abundance of each target taxon was calculated as a fraction of the total 16S rRNA gene copies in the samples, using taxon-specific and domain bacteria primers. Supplemented animals showed lower (P < 0.05) proportions of Ruminococcus flavefaciens and greater (P < 0.05) proportions of Ruminococcus albus and Butyrivibrio fibrisolvens than animals that received only the mineral supplement. The HPS supplement resulted in higher (P < 0.05) proportions of Fibrobacter succinogenes, R. flavefaciens, and B. fibrisolvens and lower (P < 0.05) proportions of R. albus than the MPS supplement. Based on our results, high protein supplementation improves the ruminal conditions and facilitates the growth of cellulolytic bacteria in the rumen of bulls on pastures during the rainy season.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Allison MJ (1969) Biosynthesis of amino acids by ruminal microorganisms. J Anim Sci 29:797–807

    CAS  PubMed  Google Scholar 

  2. Association of Official Analytical Chemistry (AOAC). 1990. Official methods of analysis, 15th edn. AOAC International, Arlington

    Google Scholar 

  3. Attwood GT, Reilly K (1995) Identification of proteolytic rumen bacteria isolated from New Zealand cattle. J Appl Bacteriol 79:22–29

    CAS  PubMed  Google Scholar 

  4. Batista ED, Detmann E, Titgemeyer EC, ValadaresFilho SC, Valadares RFD, Prates LL, Rennó LN, Paulino MF (2016) Effects of varying ruminally undegradable protein supplementation on forage digestion, nitrogen metabolism, and urea kinetics in Nellore cattle fed low-quality tropical forage. J Anim Sci 94:201–216

    CAS  PubMed  Google Scholar 

  5. Barthram GT (1985) Experimental techniques: The HFRO sward stick. In: The hill farming research organization biennial report 1984/1985. Hill Farming Research Organization, Penicuik, pp 29–30

    Google Scholar 

  6. Bryant MP, Robinson IM (1962) Some nutritional characteristics of predominant culturable ruminal bacteria. J Bacteriol 84:605

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Canesin RC, Berchielli TT, Messana JD, Baldi F, Pires AV, Frighetto RTS, Reis RA (2014) Effects of supplementation frequency on the ruminal fermentation and enteric methane production of beef cattle grazing in tropical pastures. Revista Brasileira de Zootecnia 43:590–600

    Google Scholar 

  8. Chen J, Stevenson DM, Weimer PJ (2004) Albusin B, a bacteriocin from the ruminal bacterium Ruminococcus albus 7 that inhibits growth of Ruminococcus flavefaciens. Appl Environ Microbiol 70:3167–3170

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Cotta MA, Hespell RB (1986) Proteolytic activity of the ruminal bacterium Butyrivibrio fibrisolvens. Appl Environ Microbiol 52:51–58

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Detmann E, Paulino MF, Valadares Filho SC. 2010. Otimização do uso de recursos forrageiros basais. In: Proceeding of 3rd International Symposium on Beef Cattle Production. Universidade Federal de Viçosa, Viçosa, Brazil, pp 191–240

  11. Detmann E, Paulino MF, Valadares Filho SC, Huhtanen P (2014) Nutritional aspects applied to grazing cattle in the tropics: a review based on Brazilian results. SeminaCiênciasAgrárias 35:2829–2854

    CAS  Google Scholar 

  12. Detmann E, Valente EEL, Batista ED, Huhtanen P (2014) An evaluation of the performance and efficiency of nitrogen utilization in cattle fed tropical grasspastures with supplementation. Livest Sci 162:141–153

    Google Scholar 

  13. Detmann E, Paulino MF, Mantovani HC, Valadares Filho SC, Sampaio CB, Souza MA, Lazzarini I, Detmann KSC (2009) Parameterization of ruminal fibre degradation in low-quality tropical forage using Michaelis-Menten kinetics. Livest Sci 126:136–146

    Google Scholar 

  14. Detmann E, Paulino MF, Zervoudakis JT, Valadares Filho SC, Euclydes FF, Lana RP, Queiros DS (2001) Cromo e indicadores internos na estimação do consumo de novilhos mestiços, suplementados, a pasto. Revista Brasileira de Zootecnia 30:1600–1609

    Google Scholar 

  15. Feng YL (2004) Ruminant animal nutrition. Science Press, Beijing

    Google Scholar 

  16. Fenner H (1965) Methods for determining total volatile bases in rumen fluid by steam distillation. J Dairy Sci 48:249–251

    CAS  PubMed  Google Scholar 

  17. Hall MB (2015) Comparisons of in vitro fermentation and high moisture forage processing methods for determination of neutral detergent fiber digestibility. Anim Feed Sci Technol 199:127–136

    CAS  Google Scholar 

  18. Hobson PN, Stewart CS (1997) The rumen microbial ecosystem, 2nd edn. Blackie Academic and Professionals, New York

    Google Scholar 

  19. Holleman DF, White RG (1989) Determination of digesta fill and passage rate from non-absorbed particulate phase markers using the single dose method. Can J Zool 67:488–494

    Google Scholar 

  20. Johnson AD (1978) Sample preparation and chemical analysis of vegetation. In: Manetje L (ed) Measurement of grassland vegetation and animal production. Commonwealth Agricultural Bureaux, Aberystwyth, pp 96–102

    Google Scholar 

  21. Johnson KA, Johnson DE (1995) Methane emissions from cattle. J Anim Sci 73:2483–2492

    CAS  PubMed  Google Scholar 

  22. Klieve AV, Hennessy D, Ouwerkerk D, Forster RJ, Mackie RI, Attwood GT (2003) Establishing populations of Megasphaera elsdenii YE34 and Butyrivibrio fibrisolvens YE44 in the rumen of cattle fed high grain diets. J Appl Microbiol 95:621–630

    CAS  PubMed  Google Scholar 

  23. Koike S, Kobayashi Y (2009) Fibrolytic rumen bacteria: their ecology and functions. Asian-Aust J Anim Sci 22:131–138

    CAS  Google Scholar 

  24. Koike S, Yoshitani S, Kobayashi Y, Tanaka YK (2003) Phylogenetic analysis of fiber-associated rumen bacterial community and PCR detection of uncultured bacteria. FEMS Microbiol Lett 229:23–30

    CAS  PubMed  Google Scholar 

  25. Lazzarini I, Detmann E, Sampaio CB, Paulino MF, Valadares Filho SC, Souza MA, Oliveira FA (2009) Intake and digestibility in cattle fed low-quality tropical forage and supplemented with nitrogenous compounds. Revista Brasileira de Zootecnia 38:2021–2030

    Google Scholar 

  26. Licitra G, Hernandez TM, Van Soest PJ (1996) Standardisation of procedures for nitrogen fractionation of ruminant feeds. Anim Feed Sci Technol 57:347–358

    Google Scholar 

  27. Martínez-Pérez MF, Calderón-Mendoza D, Islas A, Encinias AM, Loya-Olguín F, Soto-Navarro SA (2013) Effect of corn dry distiller grains plus solubles supplementation level on performance and digestion characteristics of steers grazing native range during forage growing season. J Anim Sci 91:1350–1361

    PubMed  Google Scholar 

  28. McAllister TA, Bae HD, Jones GA, Cheng KJ (1994) Microbial attachment and feed digestion in the rumen. J Anim Sci 72:3004–3018

    CAS  PubMed  Google Scholar 

  29. McAllister TA, Cheng KJ, Rode LM, Forsberg CW (1990) Digestion of barley, maize, and wheat by selected species of ruminal bacteria. Appl Environ Microbiol 56:3146–3153

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Mertens DR (2002) Gravimetric determination of amylase treated neutral detergent fibre in feeds with refluxing in beakers or crucibles: collaborative study. J AOAC Int 85:1212–1240

    Google Scholar 

  31. Michalet-Doreau B, Fernandez I, Peyron C, Millet L, Fonty G (2001) Fibrolytic activities and cellulolytic bacterial community structure in the solid and liquid phases of rumen contents. Reprod Nutr Dev 41:187–194

    CAS  PubMed  Google Scholar 

  32. Morgavi DP, Kelly WJ, Janssen PH, Attwood GT (2013) Rumen microbial (meta) genomics and its application to ruminant production. Animal 7:184–201

    CAS  PubMed  Google Scholar 

  33. Ohene-Adjei S, Chaves AV, McAllister TA, Benchaar C, Teather RM, Forster RJ (2008) Evidence of increased diversity of methanogenic archaea with plant extract supplementation. Microb Ecol 56:234–242

    CAS  PubMed  Google Scholar 

  34. Orskov ER (1982) Protein nutrition in ruminants, Academic Press, London

    Google Scholar 

  35. Paulino MF, Zervoudakis JT, Moraes EHBK (2002) Bovinocultura de Ciclo Curto em Pastagens. In: Simpósio de Produção de Gado de Corte, vol 3. Anais UFV, Viçosa-MG, pp 153–197

    Google Scholar 

  36. Paulino MF, Detmann E, Valadares Filho SC, Lana RP (2002) Soja grão e caroço de algodão em suplementos múltiplos para terminação de bovinos mestiços em pastejo. Revista Brasileira de Zootecnia 31:484–491

    Google Scholar 

  37. Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29:2002–2007

    Google Scholar 

  38. Powell JE, Martinson VG, Urban-Mead K, Moran NA (2014) Routes of acquisition of the gut microbiota of the honey bee Apismellifera. Appl Environ Microbiol 80:7378–7387

    PubMed  PubMed Central  Google Scholar 

  39. Russell JB (2002) Rúmen microbiology and its role in ruminant nutrition. James B Russell, New York, p 139

    Google Scholar 

  40. Russell JP (1984) Factors influencing competitions and compositions of rúmen bacterial flora. In: Proceedings of the Symposium on Herbivore Nutrition in the Sub-Tropics and Tropics. The Science Press, Graighall, South Africa, p 313

  41. Sawanon S, Kobayashi Y (2006) Synergistic fibrolysis in the rumen by cellulolytic Ruminococcus flavefaciens and noncellulolytic Selenomonas ruminantium: evidence in defined cultures. Anim Sci J 77:208–214

    CAS  Google Scholar 

  42. Scheiffinger CC, Wolin MJ (1973) Propionate formation from cellulose and soluble sugars by combined cultures of Bacteroides succinogenes and Selenomonas ruminantium. Appl Microbiol 26:789–795

    Google Scholar 

  43. Sharp R, Ziemer CJ, Stern MD, Stahl DA (1998) Taxon-specific association between protozoal and methanogen populations in the rúmen and a model rúmen system. FEMS Microbiol Ecol 26:71–78

    CAS  Google Scholar 

  44. Shi Y, Weimer PJ (1996) Utilization of individual cellodextrins by three predominant ruminal cellulolytic bacteria. Appl Environ Microbiol 62:1084–1088

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Stevenson DM, Weimer PJ (2007) Dominance of Prevotella and low abundance of classical ruminal bacterial species in the bovine rumen revealed by relative quantification real-time PCR. Appl Microbiol Biotechnol 75:165–174

    CAS  PubMed  Google Scholar 

  46. Tajima K, Aminov RI, Nagamine T, Matsui H, Nakamura M, Benno Y (2001) Diet-dependent shifts in the bacterial population of the rumen revealed with real-time PCR. Appl Environ Microbiol 67:2766–2774

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Valadares Filho SC, Marcondes MI, Chizzotti ML, Paulino PVR (2010) Nutrient requirements of zebu beef cattle BR-CORTE, 2nd edn. UFV-Departamento de Zootecnia, Viçosa

    Google Scholar 

  48. Valente TNP, Detmann E, Valadares Filho SC, Cunha M, Queiroz AC, Sampaio CB (2011) In situ estimation of indigestible compounds contents in cattle feed and feces using bags made from different textiles. Revista Brasileira de Zootecnia 40:666–675

    Google Scholar 

  49. Van Soest PJ (1994) Nutritional ecology of the ruminant. Cornell University Press, Ithaca

    Google Scholar 

  50. Yu Z, Michel F Jr, Hansen G, Wittum T, Morrison M (2005) Development and application of real-time PCR assays for quantification of genes encoding tetracycline resistance. Appl Environ Microbiol 71(11):6926–6933

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Ziemer CJ, Sharp R, Stern MD, Cotta MA, Whitehead TR, Stahl DA (2000) Comparison of microbial populations in model and natural rúmens using 16S ribosomal RNA-targeted probes. Environ Microbiol 2:632–643

    CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Faculdade de Agronomia e Zootecnia, Universidade Federal de Mato Grosso, Cuiaba, Mato Grosso, Brazil

    Renata Pereira da Silva-Marques, Luciano da Silva Cabral & Maria Isabel Leite da Silva

  2. Faculdade de Medicina Veterinária, Universidade Federal de Mato Grosso, Cuiaba, Mato Grosso, Brazil

    Joanis Tilemahos Zervoudakis, Luciano Nakazato, Luciana Keiko Hatamoto-Zervoudakis, Núbia Bezerra do Nascimento Matos & Letícia Camara Pitchenin

Authors
  1. Renata Pereira da Silva-Marques
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Joanis Tilemahos Zervoudakis
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Luciano Nakazato
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Luciano da Silva Cabral
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Luciana Keiko Hatamoto-Zervoudakis
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Maria Isabel Leite da Silva
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Núbia Bezerra do Nascimento Matos
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Letícia Camara Pitchenin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Renata Pereira da Silva-Marques.

Ethics declarations

Conflict of interest

None of the authors have any conflicts of interest to declare.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

da Silva-Marques, R.P., Zervoudakis, J.T., Nakazato, L. et al. Quantitative qPCR Analysis of Ruminal Microorganisms in Beef Cattle Grazing in Pastures in the Rainy Season and Supplemented with Different Protein Levels. Curr Microbiol 75, 1025–1032 (2018). https://doi.org/10.1007/s00284-018-1484-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00284-018-1484-2

Search

Navigation