Next Generation Sequencing to Define Prokaryotic and Fungal Diversity in the Bovine Rumen

Abstract

A combination of Sanger and 454 sequences of small subunit rRNA loci were used to interrogate microbial diversity in the bovine rumen of 12 cows consuming a forage diet. Observed bacterial species richness, based on the V1–V3 region of the 16S rRNA gene, was between 1,903 to 2,432 species-level operational taxonomic units (OTUs) when 5,520 reads were sampled per animal. Eighty percent of species-level OTUs were dominated by members of the order Clostridiales, Bacteroidales, Erysipelotrichales and unclassified TM7. Abundance of Prevotella species varied widely among the 12 animals. Archaeal species richness, also based on 16S rRNA, was between 8 and 13 OTUs, representing 5 genera. The majority of archaeal OTUs (84%) found in this study were previously observed in public databases with only two new OTUs discovered. Observed rumen fungal species richness, based on the 18S rRNA gene, was between 21 and 40 OTUs with 98.4–99.9% of OTUs represented by more than one read, using Good’s coverage. Examination of the fungal community identified numerous novel groups. Prevotella and Tannerella were overrepresented in the liquid fraction of the rumen while Butyrivibrio and Blautia were significantly overrepresented in the solid fraction of the rumen. No statistical difference was observed between the liquid and solid fractions in biodiversity of archaea and fungi. The survey of microbial communities and analysis of cross-domain correlations suggested there is a far greater extent of microbial diversity in the bovine rumen than previously appreciated, and that next generation sequencing technologies promise to reveal novel species, interactions and pathways that can be studied further in order to better understand how rumen microbial community structure and function affects ruminant feed efficiency, biofuel production, and environmental impact.

Intestinal fungi contribute to development of alcoholic liver disease

“…Sequence processing and analysis steps were performed by our open-access, mothur-based (49) workflow software, YAP (46,50). Mothur version 1.34 was used in this study.…”

“…To evaluate the human intestinal mycobiome, fungal-specific internal transcribed spacer (ITS) amplicon sequencing and analysis were conducted as previously described (4,(46)(47)(48). In brief, ITS sequence data were generated with Illumina MiSeq V2 kit using jci.org Volume 127 Number 7 July 2017…”

Genome sequencing of rumen bacteria and archaea and its application to methane mitigation strategies

“…Over the years .100, different bacterial species have been reported in the scientific literature. The development of cultivation-independent techniques led to a decline in isolation and cultivation studies, and results from 16S ribosomal RNA gene sequencing and metagenomic studies consistently emphasize that the majority of sequences are derived from organisms that are phylogenetically distinct from currently cultivated species Fouts et al, 2012). In recent years, there have been few attempts to systematically bring additional organisms into cultivation; however, recent studies in Japan (Koike et al, 2010;Nyonyo et al, 2013) and New Zealand (Kenters et al, 2011;S.J.…”

“…Although these sequenced bacteria include 21 species belonging to 14 different bacterial families, they only scratch the surface of the taxonomic and functional diversity present in the rumen. Estimates of the number of rumen microbial species based on 16S ribosomal RNA gene sequences vary from 300 to 1000 (Fouts et al, 2012). Consequently, genomic studies have yet to make a sizeable impact with respect to improving our understanding of rumen microbial ecology, and how specific rumen microbes interact with the animal, the feed or each other to influence animal production or the release of greenhouse gases.…”

Effects of Various Interference Intensities on the Soil Bacterial Communities Diversity in the Sanjiang Plain, Northeast China

“…The total number of bases determined was 1567,500,132 and the average read length was 404.68 bp, of which reads between 401 and 500 bp (the expected size for the VI-V3 region) accounted for 99.72% of the total number of reads. The rarefaction curve was constructed through randomly sampled optimized gene sequences, as it represents the relationship between the individuals and species by selecting a random subset (Fouts et al, 2012). A flat rarefaction curve indicates that sequences were sufficient, and further sequences would not produce other new OTUs.…”