Evaluation of the bacterial diversity in the feces of cattle using 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing (bTEFAP)

Abstract

BackgroundThe 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.ResultsUbiquitous 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).ConclusionUsing 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.

Inter-comparison of the potentially active prokaryotic communities in the halocline sediments of Mediterranean deep-sea hypersaline basins

“…Reverse transcription of the purified RNA samples was performed using the QuantiTect kit (Qiagen, USA). Tag--pyrosequencing of the 16S rRNA gene was performed using PCR amplification of the V4--V6 region of the 16S rRNA gene and the primer pair S--DBact--0341--b--S--17 (5'--CCTACGGGNGGCWGCAG--3') and S--D--Bact--0785--a--A--21 (5'--GACTACHVGGGTATCTAATCC--3') for Bacteria (Klindworth et al 2012), and archaea349F (5'--GYGCASCAGKCGMGAAW--3') and archaea806R (5'--GGACTACVSGGGTATCTAAT--3') for Archaea (Takai & Horikoshi 2000), as described in Dowd et al, (2008). A one--step, 30--cycle PCR reaction was performed using HotStarTaq Plus Master Mix Kit (Qiagen, USA).…”

Biotransformation of food spice curcumin by gut bacterium Bacillus megaterium DCMB-002 and its pharmacological implications

“…The identification of the isolated bacteria was performed using a molecular biological protocol by DNA amplification and sequencing of the 16S rDNA as described previously [18]. The resulting sequence data obtained from the bacterium strain has been deposited at GenBank (with accession number MF159101).…”

CD24 blunts oral squamous cancer development and dampens the functional expansion of myeloid-derived suppressor cells

“…58 Samples were sequenced utilizing Roche 454 FLX titanium instruments and reagents, following manufacturer's guidelines. The Q25 sequence data derived from the sequencing process was processed using a proprietary analysis pipeline (www.mrdnalab.com, MR DNA, Shallowater, TX).…”

“…Operational taxonomic units (OTUs) were defined after removal of singleton sequences, clustering at 3% divergence (97% similarity). 58,64 OTUs were then taxonomically classified using BLASTn against a curated Green Genes database 65 and compiled into each taxonomic level into both "counts" and "percentage" files. Counts files contain the actual number of sequences while the percent files contain the relative (proportion) percentage of sequences within each sample that map to the designated taxonomic classification.…”