Pyrosequencing of 16S rRNA genes in fecal samples reveals high diversity of hindgut microflora in horses and potential links to chronic laminitis.

BMC Veterinary Research (Impact Factor: 1.74). 11/2012; 8(1):231. DOI: 10.1186/1746-6148-8-231
Source: PubMed

ABSTRACT BACKGROUND: The nutrition and health of horses is closely tied to their gastrointestinal microflora. Gut bacteria break down plant structural carbohydrates and produce volatile fatty acids, which are a major source of energy for horses. Bacterial communities are also essential for maintaining gut homeostasis and have been hypothesized to contribute to various diseases including laminitis. We performed pyrosequencing of 16S rRNA bacterial genes isolated from fecal material to characterize hindgut bacterial communities in healthy horses and those with chronic laminitis. RESULTS: Fecal samples were collected from 10 normal horses and 8 horses with chronic laminitis. Genomic DNA was extracted and the V4-V5 segment of the 16S rRNA gene was PCR amplified and sequenced on the 454 platform generating a mean of 2,425 reads per sample after quality trimming. The bacterial communities were dominated by Firmicutes (69.21% control, 56.72% laminitis) and Verrucomicrobia (18.13% control, 27.63% laminitis), followed by Bacteroidetes, Proteobacteria, and Spirochaetes. We observed more OTUs per individual in the laminitis group than the control group (419.6 and 355.2, respectively, P = 0.019) along with a difference in the abundance of two unassigned Clostridiales genera (P = 0.03 and P = 0.01). The most abundant bacteria were Streptococcus spp., Clostridium spp., and Treponema spp.; along with unassigned genera from Subdivision 5 of Verrucomicrobia, Ruminococcaceae, and Clostridiaceae, which together constituted ~ 80% of all OTUs. There was a high level of individual variation across all taxonomic ranks. CONCLUSIONS: Our exploration of the equine fecal microflora revealed higher bacterial diversity in horses with chronic laminitis and identification of two Clostridiales genera that differed in abundance from control horses. There was large individual variation in bacterial communities that was not explained in our study. The core hindgut microflora was dominated by Streptococcus spp., several cellulytic genera, and a large proportion of uncharacterized OTUs that warrant further investigation regarding their function. Our data provide a foundation for future investigations of hindgut bacterial factors that may influence the development and progression of chronic laminitis.

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    ABSTRACT: The effects of abrupt dietary transition on the faecal microbiota of forage-fed horses over a 3-week period were investigated. Yearling Thoroughbred fillies reared as a cohort were exclusively fed on either an ensiled conserved forage-grain diet ("Group A"; n = 6) or pasture ("Group B"; n = 6) for three weeks prior to the study. After the Day 0 faecal samples were collected, horses of Group A were abruptly transitioned to pasture. Both groups continued to graze similar pasture for three weeks, with faecal samples collected at 4-day intervals. DNA was isolated from the faeces and microbial 16S and 18S rRNA gene amplicons were generated and analysed by pyrosequencing. The faecal bacterial communities of both groups of horses were highly diverse (Simpson's index of diversity >0.8), with differences between the two groups on Day 0 (P<0.017 adjusted for multiple comparisons). There were differences between Groups A and B in the relative abundances of four genera, BF311 (family Bacteroidaceae; P = 0.003), CF231 (family Paraprevotellaceae; P = 0.004), and currently unclassified members within the order Clostridiales (P = 0.003) and within the family Lachnospiraceae (P = 0.006). The bacterial community of Group A horses became similar to Group B within four days of feeding on pasture, whereas the structure of the archaeal community remained constant pre- and post-dietary change. The community structure of the faecal microbiota (bacteria, archaea and ciliate protozoa) of pasture-fed horses was also identified. The initial differences observed appeared to be linked to recent dietary history, with the bacterial community of the forage-fed horses responding rapidly to abrupt dietary change.
    PLoS ONE 11/2014; 9(11):e112846. · 3.53 Impact Factor
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    ABSTRACT: Background The intestinal tract is a rich and complex environment and its microbiota has been shown to have an important role in health and disease in the host. Several factors can cause disruption of the normal intestinal microbiota, including antimicrobial therapy, which is an important cause of diarrhea in horses. This study aimed to characterize changes in the fecal bacterial populations of healthy horses associated with the administration of frequently used antimicrobial drugs.ResultsTwenty-four adult mares were assigned to receive procaine penicillin intramuscularly (IM), ceftiofur sodium IM, trimethoprim sulfadiazine (TMS) orally or to a control group. Treatment was given for 5 consecutive days and fecal samples were collected before drug administration (Day 1), at the end of treatment (Days 5), and on Days 14 and 30 of the trial. High throughput sequencing of the V4 region of the 16S rRNA gene was performed using an Illumina MiSeq sequencer. Significant changes of population structure and community membership were observed after the use of all drugs. TMS caused the most marked changes on fecal microbiota even at higher taxonomic levels including a significant decrease of richness and diversity. Those changes were mainly due to a drastic decrease of Verrucomicrobia, specifically the ¿5 genus incertae sedis¿. Changes in structure and membership caused by antimicrobial administration were specific for each drug and may be predictable. Twenty-five days after the end of treatment, bacterial profiles were more similar to pre-treatment patterns indicating a recovery from changes caused by antimicrobial administration, but differences were still evident, especially regarding community membership.Conclusions The use of systemic antimicrobials leads to changes in the intestinal microbiota, with different and specific responses to different antimicrobials. All antimicrobials tested here had some impact on the microbiota, but TMS significantly reduced bacterial species richness and diversity and had the greatest apparent impact on population structure, specifically targeting members of the Verrucomicrobia phylum.
    BMC Veterinary Research 02/2015; 11(1):19. · 1.74 Impact Factor
  • Equine Veterinary Journal 09/2014; 46(5). · 2.37 Impact Factor


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May 23, 2014