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Evaluation of the bacterial diversity in the feces of cattle using 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP)
Open accessAbstract
BACKGROUND: The microbiota of an animal's intestinal tract plays important roles in the animal's overall health, productivity and well-being. There is still a scarcity of information on the microbial diversity in the gut of livestock species such as cattle. The primary reason for this lack of data relates to the expense of methods needed to generate such data. Here we have utilized a bacterial tag-encoded FLX 16s rDNA amplicon pyrosequencing (bTEFAP) approach that is able to perform diversity analyses of gastrointestinal populations. bTEFAP is relatively inexpensive in terms of both time and labor due to the implementation of a novel tag priming method and an efficient bioinformatics pipeline. We have evaluated the microbiome from the feces of 20 commercial, lactating dairy cows. RESULTS: Ubiquitous bacteria detected from the cattle feces included Clostridium, Bacteroides, Porpyhyromonas, Ruminococcus, Alistipes, Lachnospiraceae, Prevotella, Lachnospira, Enterococcus, Oscillospira, Cytophage, Anaerotruncus, and Acidaminococcus spp. Foodborne pathogenic bacteria were detected in several of the cattle, a total of 4 cows were found to be positive for Salmonella spp (tentative enterica) and 6 cows were positive for Campylobacter spp. (tentative lanienae). CONCLUSION: Using bTEFAP we have examined the microbiota in the feces of cattle. As these methods continue to mature we will better understand the ecology of the major populations of bacteria the lower intestinal tract. This in turn will allow for a better understanding of ways in which the intestinal microbiome contributes to animal health, productivity and wellbeing.
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“Prevotella, Bacteroides, Ruminococcus, Faecalibacterium, Roseburia and Clostridium are consistently identified in fecal samples from ruminants and are considered part of the core microbiota (Dowd et al., 2008; Durso et al., 2012).”
From Addressing global ruminant agricultural challenges through understanding the rumen microbiome: past, present, and future (2018) · cites this paper as Dowd et al., 2008
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“In the rumen of cattle, ubiquitous detected genera include Clostridium, Bacteroides, Succinivibrio, Prevotella and Ruminococcus (Dowd et al., 2008; Callaway et al., 2010).”
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Farnelid, H., Andersson, A. F., Bertilsson, S., Al‐Soud, W. A., Hansen, L. H., Sørensen, S. J., Steward, G. F., Hagström, Å., & Riemann, L. (2011) · PLoS ONE · doi:10.1371/journal.pone.0019223 · Cited by 220
“For 454 pyrosequencing of samples, adapters and samplespecific tags were added using custom primers (Table S1) in an additional PCR amplification of 10 cycles using the same PCR conditions [14].”
MethodGondalia, S. V., Palombo, E. A., Knowles, S. R., Cox, S. B., Meyer, D., & Austin, D. W. (2012) · Autism Research · doi:10.1002/aur.1253 · Cited by 208
Li, R., Khafipour, E., Krause, D. O., Entz, M. H., de Kievit, T. R., & Fernando, W. G. D. (2012) · PLoS ONE · doi:10.1371/journal.pone.0051897 · Cited by 207
“In brief, a mixture of Hot Start, HotStar high fidelity Taq polymerases, and Titanium reagents were used to perform a one-step PCR (35 cycles) with primers 28F (GAGTTTGATCMTGGCTCAG) and 519R (GTNTTACNGCGGCKGCTG), which covered the variable regions V1–V3 of the bacterial 16S rRNA genes [31].”
MethodBaleen whales host a unique gut microbiome with similarities to both carnivores and herbivores
Sanders, J. G., Beichman, A. C., Roman, J., Scott, J. J., Emerson, D., McCarthy, J. J., & Girguis, P. R. (2015) · Nature Communications · doi:10.1038/ncomms9285 · Cited by 208
“PCR amplification and 454 pyrosequencing of the V1–V3 regions were both performed at Research and Testing Laboratories (Lubbock, TX) using previously published protocols 35 .”
Lee, C., & Beauchemin, K. A. (2014) · Canadian Journal of Animal Science · doi:10.4141/cjas-2014-069 · Cited by 231
Studying Bacterial Multispecies Biofilms: Where to Start?
Røder, H. L., Sørensen, S. J., & Burmølle, M. (2016) · Trends in Microbiology · doi:10.1016/j.tim.2016.02.019 · Cited by 209
“community in the targeted environments [17,18].”
Pinloche, E., McEwan, N. R., Marden, J. P., Bayourthe, C., Auclair, E., & Newbold, C. J. (2013) · PLoS ONE · doi:10.1371/journal.pone.0067824 · Cited by 205
“A popular way to handle those large datasets is to cluster the sequences at a defined sequence identity threshold, select a representative sequence of each OTU and then identify the selected sequence using phylogeny [40,41,42].”
BackgroundJackson, C. R., Randolph, K. C., Osborn, S. L., & Tyler, H. L. (2013) · BMC Microbiology · doi:10.1186/1471-2180-13-274 · Cited by 206
“Bacterial tag-encoded FLX amplicon 454 pyrosequencing (bTEFAP) [45] was conducted on the 16S rRNA gene amplicons of each sample, through a dedicated sequencing facility (MR DNA, Shallowater, TX).”
MethodHu, H., Johani, K., Gosbell, I. B., Jacombs, A., Almatroudi, A., Whiteley, G. S., Deva, A. K., Jensen, S. O., & Vickery, K. (2015) · Journal of Hospital Infection · doi:10.1016/j.jhin.2015.05.016 · Cited by 198
“Pyrosequencing was performed using the Titanium platform (Roche, Basel, Switzerland) in a commercial facility (Molecular Research DNA Lab, Shallowater, TX, USA), as previously described.(23) Pyrosequencing data were analysed by QIIME software (Werner Lab, Cortland, NY, USA).”
MethodCulture Enriched Molecular Profiling of the Cystic Fibrosis Airway Microbiome
Sibley, C. D., Grinwis, M. E., Field, T. R., Eshaghurshan, C. S., Faria, M. M. P., Dowd, S. E., Parkins, M. D., Rabin, H. R., & Surette, M. G. (2011) · PLoS ONE · doi:10.1371/journal.pone.0022702 · Cited by 196
“NET and C# analysis pipeline the resulting BLASTn outputs were compiled and validated using sequence identity methods; data reduction analysis was performed as described previously [74–82].”
MethodMandaković, D., Rojas, C., Maldonado, J. E., Latorre, M., Travisany, D., Delage, E., Bihouée, A., Jean, G., Díaz, F. P., Fernández-Gómez, B., et al. (2018) · Scientific Reports · doi:10.1038/s41598-018-23931-0 · Cited by 184
Characterizing the microbiota across the gastrointestinal tract of a Brazilian Nelore steer
de Oliveira, M. N. V., Jewell, K. A., Freitas, F. S., Benjamin, L. A., Tótola, M. R., Borges, A. C., Moraes, C. A., & Suen, G. (2013) · Veterinary Microbiology · doi:10.1016/j.vetmic.2013.02.013 · Cited by 184
Gut microbiome composition is associated with temperament during early childhood
Christian, L. M., Galley, J. D., Hade, E. M., Schoppe-Sullivan, S., Kamp Dush, C., & Bailey, M. T. (2015) · Brain, Behavior, and Immunity · doi:10.1016/j.bbi.2014.10.018 · Cited by 191
“Bacterial tag-encoded FLX-Amplicon Pyrosequencing (bTEFAP) was performed as previously described (Dowd et al., 2008a,b).”
MethodMaternal Obesity Is Associated with Alterations in the Gut Microbiome in Toddlers
Galley, J. D., Bailey, M. T., Dush, C. M. K., Schoppe‐Sullivan, S. J., & Christian, L. M. (2014) · PLoS ONE · doi:10.1371/journal.pone.0113026 · Cited by 184
“bTEFAP Bacterial tag-encoded FLX-Amplicon Pyrosequencing (bTEFAP) was performed as previously described [21,22].”
MethodShepherd, M. L., Swecker, W. S., Jensen, R. V., & Ponder, M. A. (2011) · FEMS Microbiology Letters · doi:10.1111/j.1574-6968.2011.02434.x · Cited by 187
“93%) (Dowd et al., 2008).”
BackgroundResultInfluentialLópez‐Velasco, G., Welbaum, G. E., Boyer, R. R., Mane, S. P., & Ponder, M. A. (2011) · Journal of Applied Microbiology · doi:10.1111/j.1365-2672.2011.04969.x · Cited by 176
“2007), intestinal tract (Andersson et al. 2008; Dowd et al. 2008), soils (Roesch et al.”
BackgroundLladó, S., Žifčáková, L., Větrovský, T., Eichlerová, I., & Baldrián, P. (2015) · Biology and Fertility of Soils · doi:10.1007/s00374-015-1072-6 · Cited by 176
“The V4–V6 hypervariable regions of the 16S rRNA gene of each bacterial isolate were amplified using the eubacterial primers eub530F/eub1100aR, modified from Dowd et al. (2008).”
MethodAssessing the Impact of Copper and Zinc Oxide Nanoparticles on Soil: A Field Study
Collins, D., Luxton, T. P., Kumar, N., Shah, S., Walker, V. K., & Shah, V. (2012) · PLoS ONE · doi:10.1371/journal.pone.0042663 · Cited by 174
Wolcott, R. D., Gontcharova, V., Sun, Y., & Dowd, S. E. (2009) · BMC Microbiology · doi:10.1186/1471-2180-9-226 · Cited by 174
“The current study was undertaken to better characterize the bacterial ecology of VLU using modern next-generation approaches [8-13].”
BackgroundMethodGontcharova, V., Youn, E., Wolcott, R. D., Hollister, E. B., Gentry, T. J., & Dowd, S. E. (2010) · The Open Microbiology Journal · doi:10.2174/1874285801004010047 · Cited by 176
“Bacterial tag-encoded FLX Amplicon Pyrosequencing (bTEFAP) is a newly described method for evaluating microbial diversity within complex environments [3-5], based upon the 16S rRNA gene or any other gene.”
MethodSteelman, S. M., Chowdhary, B. P., Dowd, S. E., Suchodolski, J. S., & Janečka, J. E. (2012) · BMC Veterinary Research · doi:10.1186/1746-6148-8-231 · Cited by 173
“Sequences were depleted of chimeras and assignments to putative species (>97% similarity) were done with BlastN [58] against a manually curated database compiled from NCBI by the Research and Testing Laboratory (Lubbock, TX) [38].”
BackgroundMethodInfluentialInvestigation of bacterial diversity in the feces of cattle fed different diets1
Kim, M.-H., Kim, J., Kuehn, L. A., Bono, J. L., Berry, E. D., Kalchayanand, N., Freetly, H. C., Benson, A. K., & Wells, J. E. (2013) · Journal of Animal Science · doi:10.2527/jas.2013-6841 · Cited by 176
Animal-to-animal variation in fecal microbial diversity among beef cattle
Durso, L. M., Harhay, G. P., Smith, T. P. L., Bono, J. L., DeSantis, T. Z., Harhay, D. M., Andersen, G. L., Keen, J. E., Laegreid, W. W., & Clawson, M. L. (2010) · Applied and Environmental Microbiology · doi:10.1128/aem.00207-10 · Cited by 168
“5% of the total beef sequences but 19% of the dairy pyrosequences (1).”
BackgroundMethodResultInfluentialHunter, R. C., Klepac‐Ceraj, V., Lorenzi, M. M., Grotzinger, H., Martin, T. R., & Newman, D. K. (2012) · American Journal of Respiratory Cell and Molecular Biology · doi:10.1165/rcmb.2012-0088oc · Cited by 169
RetractedLiu, Z., & Liu, J. (2013) · MicrobiologyOpen · doi:10.1002/mbo3.89 · Cited by 166
“Bacterial diversity was studied via the bacterial tagencoded FLX amplicon pyrosequencing (bTEFAP) using eubacterial primers 28F 5′TTTGATCNTGGCTCAG-3′ and 519R 5′-GTNTTACNGCGGCKGCTG-3′ to amplify the 500 bp region of 16S rRNA gene (Dowd et al. 2008; Smith et al. 2010).”
MethodOshkin, I. Y., Beck, D. A. C., Lamb, A. E., Tchesnokova, V. L., Benuska, G., McTaggart, T. L., Kalyuzhnaya, M., Dedysh, S. N., Lidstrom, M. E., & Chistoserdova, L. (2014) · The ISME Journal · doi:10.1038/ismej.2014.203 · Cited by 166
“16S rRNA gene amplicon pyrosequencing was carried out as previously described (Dowd et al., 2008).”
MethodZarkasi, K. Z., Abell, G. C. J., Taylor, R. S., Neuman, C., Hatje, E., Tamplin, M. L., Katouli, M., & Bowman, J. P. (2014) · Journal of Applied Microbiology · doi:10.1111/jam.12514 · Cited by 168
“1993) was carried out by Research and Testing Laboratories (Lubbock, Texas) using a Roche 454 FLX instruments with titanium reagents as previously detailed by Dowd et al., (2008). Approximately 3000 raw reads were obtained per sample.”
MethodZarkasi, K. Z., Taylor, R. S., Abell, G. C. J., Tamplin, M. L., Glencross, B. D., & Bowman, J. P. (2016) · Microbial Ecology · doi:10.1007/s00248-015-0728-y · Cited by 165
Board-invited review: Rumen microbiology: Leading the way in microbial ecology1,2
Krause, D. O., Nagaraja, T. G., Wright, A.-D. G., & Callaway, T. R. (2013) · Journal of Animal Science · doi:10.2527/jas.2012-5567 · Cited by 162
Community fingerprinting in a sequencing world
van Dorst, J. M., Bissett, A. B., Palmer, A. S., Brown, M. V., Snape, I., Stark, J. S., Raymond, B., McKinlay, J. P., Ji, M., Winsley, T. J., et al. (2014) · FEMS Microbiology Ecology · doi:10.1111/1574-6941.12308 · Cited by 172
“The DNA was used as a template for amplification of the 16S rDNA gene with the universal primers, 28F and 519R (Dowd et al., 2008).”
MethodA Comparison of Bacterial Composition in Diabetic Ulcers and Contralateral Intact Skin
Gontcharova, V., Youn, E., Sun, Y., Wolcott, R. D., & Dowd, S. E. (2010) · The Open Microbiology Journal · doi:10.2174/1874285801004010008 · Cited by 156
Self-citationModulation of the faecal microbiome of healthy adult dogs by inclusion of potato fibre in the diet
Panasevich, M. R., Kerr, K. R., Dilger, R. N., Fahey, G. C., Jr., Guérin‐Deremaux, L., Lynch, G. L., Wils, D., Suchodolski, J. S., Steer, J. M., Dowd, S. E., et al. (2014) · British Journal Of Nutrition · doi:10.1017/s0007114514003274 · Cited by 166
“The physiological relevance of these results was further investigated by conducting post hoc Pearson’s correlation analysis with faecal SCFA data reported previously((14)) and qPCR data.”
MethodBacterial Succession in the Broiler Gastrointestinal Tract
Ranjitkar, S., Lawley, B., Tannock, G. W., & Engberg, R. M. (2016) · Applied and Environmental Microbiology · doi:10.1128/aem.02549-15 · Cited by 157
“from DNA using a two-step protocol as described previously (30).”
MethodHumblot, C., & Guyot, J. (2009) · Applied and Environmental Microbiology · doi:10.1128/aem.00451-09 · Cited by 155
“also showed good representation of all samples of cattle feces with 1,732 to 3,224 sequences (4).”
BackgroundLópez‐Mondéjar, R., Voříšková, J., Větrovský, T., & Baldrián, P. (2015) · Soil Biology and Biochemistry · doi:10.1016/j.soilbio.2015.04.008 · Cited by 153
Lekberg, Y., Schnoor, T. K., Kjøller, R., Gibbons, S. M., Hansen, L. H., Al‐Soud, W. A., Sørensen, S. J., & Rosendahl, S. (2011) · Journal of Ecology · doi:10.1111/j.1365-2745.2011.01894.x · Cited by 160
“Additionally, three different 10-base tags were included between linker A and the glo454 primer (Dowd et al. 2008), allowing all 48 samples to be distributed among sixteen 454-sectors with three samples in each sector.”
BackgroundThe alligator gut microbiome and implications for archosaur symbioses
Keenan, S. W., Engel, A. S., & Elsey, R. M. (2013) · Scientific Reports · doi:10.1038/srep02877 · Cited by 153
Zhou, Z., Meng, H., Liu, Y., Gu, J., & Li, M. (2017) · Frontiers in Microbiology · doi:10.3389/fmicb.2017.02148 · Cited by 151
“…community with sufficient sequence coverage and equal sampling scale, which enables robust and comprehensive assessment on reflecting microbial distribution pattern (Dowd et al., 2008; Schloss et al., 2009; Caporaso et al., 2010, 2012; Angiuoli et al., 2011; Quast et al., 2013; Cole et al., 2014).”
BackgroundTabor, A. E., Ali, A., Rehman, G., Garcia, G. R., Zangirolamo, A. F., Malardo, T., & Jonsson, N. N. (2017) · Frontiers in Cellular and Infection Microbiology · doi:10.3389/fcimb.2017.00506 · Cited by 158
“The differences in the composition of their microbiota (Dowd et al., 2008; Durso et al., 2010, 2012; de Oliveira et al., 2013; Mao et al., 2015), as well as the distinct volatile organic compounds (VOC) produced by B. t. taurus and B. t. indicus cattle may corroborate the contrasting tick…”
Background
Metadata
- Authors
- Dowd, S. E., Callaway, T. R., Wolcott, R. D., Sun, Y., McKeehan, T., Hagevoort, R. G., & Edrington, T. S.
- Affiliations
- Agricultural Research Service (US)
- Corresponding author
- Dowd, Scot E
- Venue
- BMC Microbiology
- Published
- 2008
- Volume
- 8
- Issue
- 1
- Pages
- 125
- ISSN
- 1471-2180 (ISSN-L)
- PMC
- PMC2515157
- OpenAlex
- W2105807803
- Cited by
- 1,121
- References
- 80
- Citations by year
- 2026: 16 · 2025: 23 · 2024: 32 · 2023: 32 · 2022: 45 · 2021: 65 · 2020: 52 · 2019: 60 · 2018: 68 · 2017: 105 · 2016: 116 · 2015: 121 · 2014: 129 · 2013: 99 · 2012: 75
- FWCI
- 11.37
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- cc-by
- Locations
- BMC Microbiology (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion)
- Funders
- U.S. Department of Agriculture
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Gut microbiota and health
- Life Sciences › Agricultural and Biological Sciences › Animal Science and Zoology › Ruminant Nutrition and Digestive Physiology
- Life Sciences › Agricultural and Biological Sciences › Animal Science and Zoology › Animal Nutrition and Physiology
- Keywords
- fecal microbiota, bacterial diversity, dairy cattle, Clostridium, Bacteroides, Ruminococcus, Lachnospiraceae, foodborne pathogens, Salmonella, Campylobacter, gut microbiota
- MeSH terms
- Animals, Bacteria, Cattle, DNA, Bacterial, DNA, Ribosomal, Feces, Female, RNA, Ribosomal, 16S, Polymerase Chain Reaction, Sequence Analysis, DNA, Gastrointestinal Tract, Biodiversity
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