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Current Microbiology

, Volume 75, Issue 8, pp 1025–1032 | Cite as

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

  • Renata Pereira da Silva-Marques
  • Joanis Tilemahos Zervoudakis
  • Luciano Nakazato
  • Luciano da Silva Cabral
  • Luciana Keiko Hatamoto-Zervoudakis
  • Maria Isabel Leite da Silva
  • Núbia Bezerra do Nascimento Matos
  • Letícia Camara Pitchenin
Article
  • 192 Downloads

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.

Notes

Compliance with Ethical Standards

Conflict of interest

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

References

  1. 1.
    Allison MJ (1969) Biosynthesis of amino acids by ruminal microorganisms. J Anim Sci 29:797–807CrossRefPubMedGoogle Scholar
  2. 2.
    Association of Official Analytical Chemistry (AOAC). 1990. Official methods of analysis, 15th edn. AOAC International, ArlingtonGoogle Scholar
  3. 3.
    Attwood GT, Reilly K (1995) Identification of proteolytic rumen bacteria isolated from New Zealand cattle. J Appl Bacteriol 79:22–29CrossRefPubMedGoogle Scholar
  4. 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 94:201–216CrossRefPubMedGoogle Scholar
  5. 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 29–30Google Scholar
  6. 6.
    Bryant MP, Robinson IM (1962) Some nutritional characteristics of predominant culturable ruminal bacteria. J Bacteriol 84:605PubMedPubMedCentralGoogle Scholar
  7. 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 43:590–600CrossRefGoogle Scholar
  8. 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 70:3167–3170CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Cotta MA, Hespell RB (1986) Proteolytic activity of the ruminal bacterium Butyrivibrio fibrisolvens. Appl Environ Microbiol 52:51–58PubMedPubMedCentralGoogle Scholar
  10. 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 191–240Google Scholar
  11. 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 35:2829–2854Google Scholar
  12. 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 162:141–153CrossRefGoogle Scholar
  13. 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 126:136–146CrossRefGoogle Scholar
  14. 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 30:1600–1609CrossRefGoogle Scholar
  15. 15.
    Feng YL (2004) Ruminant animal nutrition. Science Press, BeijingGoogle Scholar
  16. 16.
    Fenner H (1965) Methods for determining total volatile bases in rumen fluid by steam distillation. J Dairy Sci 48:249–251CrossRefPubMedGoogle Scholar
  17. 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 199:127–136CrossRefGoogle Scholar
  18. 18.
    Hobson PN, Stewart CS (1997) The rumen microbial ecosystem, 2nd edn. Blackie Academic and Professionals, New YorkCrossRefGoogle Scholar
  19. 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 67:488–494CrossRefGoogle Scholar
  20. 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 96–102Google Scholar
  21. 21.
    Johnson KA, Johnson DE (1995) Methane emissions from cattle. J Anim Sci 73:2483–2492CrossRefPubMedGoogle Scholar
  22. 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 95:621–630CrossRefPubMedGoogle Scholar
  23. 23.
    Koike S, Kobayashi Y (2009) Fibrolytic rumen bacteria: their ecology and functions. Asian-Aust J Anim Sci 22:131–138CrossRefGoogle Scholar
  24. 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 229:23–30CrossRefPubMedGoogle Scholar
  25. 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 38:2021–2030CrossRefGoogle Scholar
  26. 26.
    Licitra G, Hernandez TM, Van Soest PJ (1996) Standardisation of procedures for nitrogen fractionation of ruminant feeds. Anim Feed Sci Technol 57:347–358CrossRefGoogle Scholar
  27. 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 91:1350–1361CrossRefPubMedGoogle Scholar
  28. 28.
    McAllister TA, Bae HD, Jones GA, Cheng KJ (1994) Microbial attachment and feed digestion in the rumen. J Anim Sci 72:3004–3018CrossRefPubMedGoogle Scholar
  29. 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 56:3146–3153PubMedPubMedCentralGoogle Scholar
  30. 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 85:1212–1240Google Scholar
  31. 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 41:187–194CrossRefPubMedGoogle Scholar
  32. 32.
    Morgavi DP, Kelly WJ, Janssen PH, Attwood GT (2013) Rumen microbial (meta) genomics and its application to ruminant production. Animal 7:184–201CrossRefPubMedGoogle Scholar
  33. 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 56:234–242CrossRefPubMedGoogle Scholar
  34. 34.
    Orskov ER (1982) Protein nutrition in ruminants, Academic Press, LondonGoogle Scholar
  35. 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 153–197Google Scholar
  36. 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 31:484–491CrossRefGoogle Scholar
  37. 37.
    Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29:2002–2007CrossRefGoogle Scholar
  38. 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 80:7378–7387CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Russell JB (2002) Rúmen microbiology and its role in ruminant nutrition. James B Russell, New York, p 139Google Scholar
  40. 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 313Google Scholar
  41. 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 77:208–214CrossRefGoogle Scholar
  42. 42.
    Scheiffinger CC, Wolin MJ (1973) Propionate formation from cellulose and soluble sugars by combined cultures of Bacteroides succinogenes and Selenomonas ruminantium. Appl Microbiol 26:789–795Google Scholar
  43. 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 26:71–78CrossRefGoogle Scholar
  44. 44.
    Shi Y, Weimer PJ (1996) Utilization of individual cellodextrins by three predominant ruminal cellulolytic bacteria. Appl Environ Microbiol 62:1084–1088PubMedPubMedCentralGoogle Scholar
  45. 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 75:165–174CrossRefPubMedGoogle Scholar
  46. 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 67:2766–2774CrossRefPubMedPubMedCentralGoogle Scholar
  47. 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, ViçosaGoogle Scholar
  48. 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 40:666–675CrossRefGoogle Scholar
  49. 49.
    Van Soest PJ (1994) Nutritional ecology of the ruminant. Cornell University Press, IthacaGoogle Scholar
  50. 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):6926–6933CrossRefPubMedPubMedCentralGoogle Scholar
  51. 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 2:632–643CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Renata Pereira da Silva-Marques
    • 1
  • Joanis Tilemahos Zervoudakis
    • 2
  • Luciano Nakazato
    • 2
  • Luciano da Silva Cabral
    • 1
  • Luciana Keiko Hatamoto-Zervoudakis
    • 2
  • Maria Isabel Leite da Silva
    • 1
  • Núbia Bezerra do Nascimento Matos
    • 2
  • Letícia Camara Pitchenin
    • 2
  1. 1.Faculdade de Agronomia e ZootecniaUniversidade Federal de Mato GrossoCuiabaBrazil
  2. 2.Faculdade de Medicina VeterináriaUniversidade Federal de Mato GrossoCuiabaBrazil

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