Skip to main content

Advertisement

SpringerLink
Log in

Metagenomic characterisation of ruminal bacterial diversity in buffaloes from birth to adulthood using 16S rRNA gene amplicon sequencing

  • Original Article
  • Published:
Functional & Integrative Genomics Aims and scope Submit manuscript

Abstract

Microbial colonisation in the forestomach of a ruminant is one of the most crucial factors in determining many of its physiological developments and digestive capabilities. The present study attempts to identify establishment pattern of microbes in relation to food, age and rumen development in the buffalo calves at every fortnight interval from birth to 6 months of age, followed by every month till animals became 1 year of age. Diversity study based on 16S rRNA gene sequencing identified rapidly changing bacterial population during initial 60 days of life, which got assemblage as rumen became physiologically mature with increasing age of animals. A lactate fermenting aerobic to facultative anaerobic genera found during initial 30 days of life were expeditiously replaced by strict anaerobic cellulolytic bacterial population with increasing age. The study confirms that initial colonisation mainly depends on the oral cavity and skin of the mother, followed by the surrounding environment and feed offered, which is reversed in order once animal gets older. Some of the well-described genera based on culture-dependent studies like Ruminococcus spp. were found to be in lesser proportion suggesting an additional role of other microbes or niche in cellulose degradation. We report the presence of Porphyromonas spp. and Mannheimia glucosidal for the first time in bovine infants.

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

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Data availability

All data generated or analysed during this study are included in this published article as supplementary information files. In addition, the datasets generated and analysed during the current study are available in the MG-RAST. The following link can be used by the reviewer to access data as a part of data share at MG-RAST. The data will be made publically available once paper will be accepted.

http://metagenomics.anl.gov/mgmain.html?mgpage=token&token=IvmrEitAji7z4n38Vt03ceovGO_ohm0mn6nWf9nf0a5BXn6QmO

Abbreviations

NGS:

next-generation sequencing

VFA:

volatile fatty acid

CP:

crude protein

EE:

ether extract

0D:

zero day

MG-RAST:

metagenome rapid annotation using subsystem technology

PAST:

paleontological statistics

PCoA:

principal coordinates analysis

PLS-DA:

partial least squares-discriminant analysis

TSV:

tab separated value

STAMP:

statistical analysis of metagenomic profiles

OUT:

operational taxonomic units

PC:

principal component

Fr:

French gauge

References

  • Arndt D, Xia J, Liu Y, Zhou Y, Guo AC, Cruz JA, Sinelnikov I, Budwill K, Nesbo CL, Wishart DS (2012) METAGENassist: a comprehensive web server for comparative metagenomics. Nucleic Acids Res 40:W88–W95

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Avgustin G, Wallace RJ, Flint HJ (1997) Phenotypic diversity among ruminal isolates of Prevotella ruminicola: proposal of Prevotella brevis sp. nov., Prevotella bryantii sp. nov., and Prevotella albensis sp. nov. and redefinition of Prevotella ruminicola. Int J Syst Bacteriol 47:284–288

    Article  CAS  PubMed  Google Scholar 

  • Bekele AZ, Koike S, Kobayashi Y (2010) Genetic diversity and diet specificity of ruminal Prevotella revealed by 16S rRNA gene-based analysis. FEMS Microbiol Lett 305:49–57

    Article  CAS  PubMed  Google Scholar 

  • Bergstrom A, Skov TH, Bahl MI, Roager HM, Christensen LB, Ejlerskov KT, Molgaard C, Michaelsen KF, Licht TR (2014) Establishment of intestinal microbiota during early life: a longitudinal, explorative study of a large cohort of Danish infants. Appl Environ Microbiol 80:2889–2900

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Connor EE, Baldwin RL, Li CJ, Li RW, Chung H (2013) Gene expression in bovine rumen epithelium during weaning identifies molecular regulators of rumen development and growth. Funct Integr Genomics 13:133–142

    Article  CAS  PubMed  Google Scholar 

  • Conroy ME, Shi HN, Walker WA (2009) The long-term health effects of neonatal microbial flora. Curr Opin Allergy Clin Immunol 9:197–201

    Article  PubMed  Google Scholar 

  • Cox MP, Peterson DA, Biggs PJ (2010) SolexaQA: at-a-glance quality assessment of Illumina second-generation sequencing data. BMC Bioinformatics 11:485

    Article  PubMed  PubMed Central  Google Scholar 

  • Curtis TP, Sloan WT (2004) Prokaryotic diversity and its limits: microbial community structure in nature and implications for microbial ecology. Curr Opin Microbiol 7:221–226

    Article  PubMed  Google Scholar 

  • Dehority BA (1991) Effects of microbial synergism on fibre digestion in the rumen. Proc Nutr Soc 50:149–159

    Article  CAS  PubMed  Google Scholar 

  • Drackley JK (1999) ADSA Foundation Scholar Award. Biology of dairy cows during the transition period: the final frontier? J Dairy Sci 82:2259–2273

    Article  CAS  PubMed  Google Scholar 

  • Fanaro S, Chierici R, Guerrini P, Vigi V (2003) Intestinal microflora in early infancy: composition and development. Acta Paediatr 91:48–55

    CAS  Google Scholar 

  • Fonty G, Gouet P, Jouany JP, Senaud J (1983) Ecological factors determining establishment of cellulolytic bacteria and protozoa in the rumens of meroxenic lambs. J Gen Microbiol 129:213–223

    CAS  PubMed  Google Scholar 

  • Ghattargi VC, Nimonkar YS, Burse SA, Davray D, Kumbhare SV, Shetty SA, Gaikwad MA, Suryavanshi MV, Doijad SP, Utage B, Sharma OP, Shouche YS, Meti BS, Pawar SP (2018) Genomic and physiological analyses of an indigenous strain, Enterococcus faecium 17OM39. Funct Integr Genomics 18:385–399

    Article  CAS  PubMed  Google Scholar 

  • Hernandez-Sanabria E, Goonewardene LA, Wang Z, Durunna ON, Moore SS, Guan LL (2012) Impact of feed efficiency and diet on adaptive variations in the bacterial community in the rumen fluid of cattle. Appl Environ Microbiol 78:1203–1214

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hoover WH, Miller TK (1991) Rumen digestive physiology and microbial ecology. Vet Clin N Am Food Anim Pract 7:311–325

    Article  CAS  Google Scholar 

  • Hungate RE, Bryant MP, Mah RA (1964) The rumen bacteria and protozoa. Annu Rev Microbiol 18:131–166

    Article  CAS  PubMed  Google Scholar 

  • Jami E, Israel A, Kotser A, Mizrahi I (2013) Exploring the bovine rumen bacterial community from birth to adulthood. ISME J 7:1069–1079

    Article  PubMed  PubMed Central  Google Scholar 

  • Jami E, White BA, Mizrahi I (2014) Potential role of the bovine rumen microbiome in modulating milk composition and feed efficiency. PLoS One 9:e85423

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jost T, Lacroix C, Braegger CP, Chassard C (2012) New insights in gut microbiota establishment in healthy breast fed neonates. PLoS One 7:e44595

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Koenig JE, Spor A, Scalfone N, Fricker AD, Stombaugh J, Knight R, Angenent LT, Ley RE (2011) Succession of microbial consortia in the developing infant gut microbiome. Proc Natl Acad Sci U S A 108(Suppl 1):4578–4585

    Article  PubMed  Google Scholar 

  • Land M, Hauser L, Jun SR, Nookaew I, Leuze MR, Ahn TH, Karpinets T, Lund O, Kora G, Wassenaar T, Poudel S, Ussery DW (2015) Insights from 20 years of bacterial genome sequencing. Funct Integr Genomics 15:141–161

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lau JS, Omaleki L, Turni C, Barber SR, Browning GF, Francis MJ, Graham M, Korman TM (2015) Human wound infection with Mannheimia glucosida following lamb bite. J Clin Microbiol 53:3374–3376

    Article  PubMed  PubMed Central  Google Scholar 

  • Ley RE, Hamady M, Lozupone C, Turnbaugh PJ, Ramey RR, Bircher JS, Schlegel ML, Tucker TA, Schrenzel MD, Knight R, Gordon JI (2008a) Evolution of mammals and their gut microbes. Science 320:1647–1651

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ley RE, Lozupone CA, Hamady M, Knight R, Gordon JI (2008b) Worlds within worlds: evolution of the vertebrate gut microbiota nature reviews. Microbiology 6:776–788

    CAS  PubMed  Google Scholar 

  • Li RW, Connor EE, Li C, Baldwin Vi RL, Sparks ME (2012) Characterization of the rumen microbiota of pre-ruminant calves using metagenomic tools. Environ Microbiol 14:129–139

    Article  CAS  PubMed  Google Scholar 

  • Mackie RI (2002) Mutualistic fermentative digestion in the gastrointestinal tract: diversity and evolution. Integr Comp Biol 42:319–326

    Article  PubMed  Google Scholar 

  • Malmuthuge N, Griebel PJ, Guan le L (2015) The gut microbiome and its potential role in the development and function of newborn calf gastrointestinal tract. Front Vet Sci 2:36

    Article  PubMed  PubMed Central  Google Scholar 

  • Matsui H, Ogata K, Tajima K, Nakamura M, Nagamine T, Aminov RI, Benno Y (2000) Phenotypic characterization of polysaccharidases produced by four Prevotella type strains. Curr Microbiol 41:45–49

    Article  CAS  PubMed  Google Scholar 

  • McCann JC, Wickersham TA, Loor JJ (2014) High-throughput methods redefine the rumen microbiome and its relationship with nutrition and metabolism. Bioinf Biol Insights 8:109–125

    Article  CAS  Google Scholar 

  • Meyer F, Paarmann D, D'Souza M, Olson R, Glass EM, Kubal M, Paczian T, Rodriguez A, Stevens R, Wilke A, Wilkening J, Edwards RA (2008) The metagenomics RAST server - a public resource for the automatic phylogenetic and functional analysis of metagenomes. BMC Bioinformatics 9:386

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Naeem A, Drackley JK, Lanier JS, Everts RE, Rodriguez-Zas SL, Loor JJ (2014) Ruminal epithelium transcriptome dynamics in response to plane of nutrition and age in young Holstein calves. Funct Integr Genomics 14:261–273

    Article  CAS  PubMed  Google Scholar 

  • O'Neill BF, Deighton MH, O'Loughlin BM, Mulligan FJ, Boland TM, O'Donovan M, Lewis E (2011) Effects of a perennial ryegrass diet or total mixed ration diet offered to spring-calving Holstein-Friesian dairy cows on methane emissions, dry matter intake, and milk production. J Dairy Sci 94:1941–1951

    Article  CAS  PubMed  Google Scholar 

  • Parks DH, Tyson GW, Hugenholtz P, Beiko RG (2014) STAMP: statistical analysis of taxonomic and functional profiles. Bioinformatics 30:3123–3124

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Poulsen LL, Reinert TM, Sand RL, Bisgaard M, Christensen H, Olsen JE, Stuen S, Bojesen AM (2006) Occurrence of haemolytic Mannheimia spp. in apparently healthy sheep in Norway. Acta Vet Scand 48(19):19

    Article  PubMed  PubMed Central  Google Scholar 

  • Rey M, Enjalbert F, Combes S, Cauquil L, Bouchez O, Monteils V (2014) Establishment of ruminal bacterial community in dairy calves from birth to weaning is sequential. J Appl Microbiol 116:245–257

    Article  CAS  PubMed  Google Scholar 

  • Scholtens PA, Oozeer R, Martin R, Amor KB, Knol J (2012) The early settlers: intestinal microbiology in early life. Annu Rev Food Sci Technol 3:425–447

    Article  CAS  PubMed  Google Scholar 

  • 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

    Article  CAS  PubMed  Google Scholar 

  • Strobel HJ (1992) Vitamin B12-dependent propionate production by the ruminal bacterium Prevotella ruminicola 23. Appl Environ Microbiol 58:2331–2333

    CAS  PubMed  PubMed Central  Google Scholar 

  • Tissier H (1900) Recherches sur la flore intestinale des nourrissons:(état normal et pathologique)

  • van Nimwegen FA, Penders J, Stobberingh EE, Postma DS, Koppelman GH, Kerkhof M, Reijmerink NE, Dompeling E, van den Brandt PA, Ferreira I, Mommers M, Thijs C (2011) Mode and place of delivery, gastrointestinal microbiota, and their influence on asthma and atopy. J Allergy Clin Immunol 128(948–955):e941–e943

    Google Scholar 

  • Van Soest PJ (1994) Nutritional ecology of the ruminant. Cornell University Press

  • Zened A, Combes S, Cauquil L, Mariette J, Klopp C, Bouchez O, Troegeler-Meynadier A, Enjalbert F (2013) Microbial ecology of the rumen evaluated by 454 GS FLX pyrosequencing is affected by starch and oil supplementation of diets. FEMS Microbiol Ecol 83:504–514

    Article  CAS  PubMed  Google Scholar 

  • Zilber-Rosenberg I, Rosenberg E (2008) Role of microorganisms in the evolution of animals and plants: the hologenome theory of evolution. FEMS Microbiol Rev 32:723–735

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We are thankful to the Indian Council of Agricultural Research for providing financial support under Niche Area of Excellence Programme (Grant Letter F. No. 10 (2)/2011-EPD). We thank Dr. Ghanshyambhai Patel for providing excellent technical assistance in sample collection, maintaining animals and collecting nutritional data at Animal Nutrition Research Station, Anand Agricultural University, Anand, Gujarat, India.

Funding

All the funding for the current research work was provided by Indian Council of Agricultural Research under Niche Area of Excellence programme wide NO. ICAR letter No. 10(2)/2011-EPD dated 15-07-2014 under research project entitled “Metagenomic analysis of ruminal microbes”.

Author information

Author notes

  1. Prakash G. Koringa and Jalpa R. Thakkar contributed equally to this work.

Authors and Affiliations

  1. Department of Animal Biotechnology, College of Veterinary Science and Animal Husbandry, Anand Agricultural University, Anand, Gujarat, 388001, India

    Prakash G. Koringa, Jalpa R. Thakkar, Ramesh J. Pandit, Ankit T. Hinsu, Mithil J. Parekh, Ravi K. Shah, Subhash J. Jakhesara & Chaitanya G. Joshi

Authors
  1. Prakash G. Koringa
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jalpa R. Thakkar
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ramesh J. Pandit
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ankit T. Hinsu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mithil J. Parekh
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ravi K. Shah
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Subhash J. Jakhesara
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Chaitanya G. Joshi
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

PGK conceptualised the project, participated in sample collection and drafted the manuscript; JRT performed DNA extraction, amplicon generation and data analysis; RJP assisted in data analysis; ATH assisted technically during experimental study; MJP performed sequencing and generated data on MiSeq; RKS maintained animals and did sample collection; SJJ drafted actual research project and improved manuscript; CGJ conceptualised research idea and provided all facilities to carryout research. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Subhash J. Jakhesara.

Ethics declarations

Ethics approval and consent to participate

All animal ethics guidelines were followed and complied as per permission from Ethical Committee norms and letter No. IAEC 525-2015.

Conflict of interest

None of the authors has any financial or personal relationships that could inappropriately influence or bias the content of the paper. The authors declare no conflict of interest.

Electronic supplementary material

Supplementary Table 1

Details PCR primers used for detection of rumen bacterial species in this study by amplicon generation followed by NGS. (XLSX 8 kb)

Supplementary Table 2

Details of numbers of reads and bases generated for all 19 samples. (XLSX 10 kb)

Supplementary Table 3

Relative abundance of each bacterial taxa at phylum level (> 1% level of abundance) as well as ratio of Bacteriodetes:Firmicutes (XLSX 19 kb)

Supplementary Table 4

Relative abundance of each bacterial taxa at genus level (> 1% level of abundance) among seven groups consist of a total 19 samples collected at different age. (XLSX 72 kb)

Supplementary Table 5

Details of individually shared genera across different age groups. (XLSX 18 kb)

Supplementary Table 6

Relative abundance of each bacterial taxa at species level (> 1% level of abundance) among seven groups consist of a total 19 samples collected at different age. (XLSX 274 kb)

Supplementary Figure 1

Rarefaction analysis for bacterial operational taxonomic units (OTUs) in each sampling. Individual rarefaction curves for each rumen sample taken to evaluate the depth of sequencing for each sample. Each age group is distinguished by different colour of trend lines. (DOCX 1583 kb)

Supplementary Figure 2

This figure shows the relative abundance of each bacterial taxa at phylum level. X-axis, samples from different age; Y-axis, the relative abundance of bacterial taxa at each phylum level (%). (DOCX 1197 kb)

Supplementary Figure 3

This figures shows the relative abundance of each bacterial taxa at genus level. X-axis, seven age groups consist of a total 19 sampling at different age: Y-axis, the relative abundance of bacterial taxa at each genus level (%). (DOCX 737 kb)

Supplementary Figure 4

PCoA analysis of genus level bacterial composition of ruminal samples taken at different age. PC1, PC2 and PC3 together explains 82% of total variation at genus level composition. It is clearly defined to make seven different groups showing different bacterial composition from all 19 samples collected at different age. (DOCX 420 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Koringa, P.G., Thakkar, J.R., Pandit, R.J. et al. Metagenomic characterisation of ruminal bacterial diversity in buffaloes from birth to adulthood using 16S rRNA gene amplicon sequencing. Funct Integr Genomics 19, 237–247 (2019). https://doi.org/10.1007/s10142-018-0640-x

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10142-018-0640-x

Keywords

Search

Navigation