Faecal Microbiota Of Horses Research Articles

Longitudinal effects of oral administration of antimicrobial drugs on fecal microbiota of horses.

Antimicrobial drug-associated diarrhea (AAD) is the most common adverse effect in horses receiving antimicrobials. Little information on how oral administration of antimicrobials alters intestinal microbiota in horses is available. Investigate changes of the fecal microbiota in response to oral administration of antimicrobials. Twenty healthy horses. Prospective, longitudinal study. Horses were randomly assigned to 4 groups comprising 4 horses each: group 1 (metronidazole); group 2 (erythromycin); group 3 (doxycycline); group 4 (sulfadiazine/trimethoprim, SMZ-TMP); and group 5 (control). Antimicrobials were administered for 5 days. Fecal samples were obtained before (day 0) and at 1, 2, 3, 4, 5, 6, and 30 days of the study period. Fecal microbiota was characterized by high throughput sequencing of the V4 region of the 16S rRNA. Horses remained healthy throughout the study. Richness and diversity in doxycycline, erythromycin, and metronidazole, but not SMZ-TMP groups, was significantly lower (P < .05) at multiple time points after administration of antimicrobials compared with samples from day 0. Main changes in the microbiota were observed during the time of antimicrobial administration (day 2-5; weighted and unweighted UniFrac PERMANOVA P < .05). Administration of erythromycin, doxycycline and, to a lesser extent, metronidazole produced a pronounced alteration in the microbiota compared with day 0 samples by decreasing the abundance of Treponema, Fibrobacter, and Lachnospiraceae and increasing Fusobacterium and Escherichia-Shigella. Oral administration of antimicrobials alters the intestinal microbiota of healthy horses resembling horses with dysbiosis, potentially resulting in intestinal inflammation and predisposition to diarrhea.

Effects of Starch Overload and Cecal Buffering on Fecal Microbiota of Horses.

Starch overload in horses causes gastrointestinal and metabolic disorders that are associated with microbiota changes. Therefore, we identified the fecal microbiota and hypothesized that intracecal injection of alkaline solution (buffer; Mg(OH)2 + Al(OH)3) could stabilize these microbiota and clinical changes in horses submitted to corn starch overload. Ten crossbred horses (females and geldings) were allocated to group I (water−saline and starch−buffer treatments) and group II (water−buffer and starch−saline treatments). Clinical signs, gross analysis of the feces, and fecal microbiota were evaluated through 72 h (T0; T8; T12; T24; T48; T72). Corn starch or water were administrated by nasogastric tube at T0, and the buffer injected into the cecum at T8 in starch−buffer and water−buffer treatments. Starch overload reduced the richness (p < 0.001) and diversity (p = 0.001) of the fecal microbiota. However, the starch−buffer treatment showed greater increase in amylolytic bacteria (Bifidobacterium 0.0% to 5.6%; Lactobacillus 0.1% to 7.4%; p < 0.05) and decrease in fibrolytic bacteria (Lachnospiraceae 10.2% to 5.0%; Ruminococcaceae 11.7% to 4.2%; p < 0.05) than starch−saline treatment. Additionally, animals that received starch−buffer treatment showed more signs of abdominal discomfort and lameness associated with dysbiosis (amylolytic r > 0.5; fribolytic r < 0.1; p < 0.05), showing that cecal infusion of buffer did not prevent, but intensified intestinal disturbances and the risk of laminitis.

Fecal microbiota of horses with colitis and its association with laminitis and survival during hospitalization.

The association of microbiota with clinical outcomes and the taxa associated with colitis in horses remains generally unknown. Describe the fecal microbiota of horses with colitis and investigate the association of the fecal microbiota with the development of laminitis and survival. Thirty-six healthy and 55 colitis horses subdivided into laminitis (n=15) and non-laminitis (n=39, 1 horse with chronic laminitis was removed from this comparison) and survivors (n=27) and nonsurvivors (n=28). Unmatched case-control study. The Illumina MiSeq platform targeting the V4 region of the 16S ribosomal RNA gene was used to assess the microbiota. The community membership (Jaccard index) and structure (Yue and Clayton index) were different (analysis of molecular variance [AMOVA]; P < .001) between healthy and colitis horses. The linear discriminant analysis effect size (LEfSe; linear discriminant analysis [LDA] >3; P < .05) and random forest analyses found Enterobacteriaceae, Lactobacillus, Streptococcus, and Enterococcus enriched in colitis horses, whereas Treponema, Faecalibacterium, Ruminococcaceae, and Lachnospiraceae were enriched in healthy horses. The community membership and structure of colitis horses with or without laminitis was (AMOVA; P > .05). Enterobacteriaceae, Streptococcus, and Lactobacillus were enriched in horses with laminitis (LDA > 3; P < .05). The community membership (AMOVA; P=.008) of surviving and nonsurviving horses was different. Nonsurviving horses had an enrichment of Enterobacteriaceae, Pseudomonas, Streptococcus, and Enterococcus (LDA >3; P < .05). Differences in the microbiota of horses with colitis that survive or do not survive are minor and, similarly, the microbiota differences in horses with colitis that do or do not develop laminitis are minor.

EquineReview

Introduction: This month's Equine Review presents papers on a large prospective European study of the efficacy and safety of the glucocorticoid prodrug ciclesonide, changes in the faecal microbiota of horses hospitalised for colic and the comparison of chlorhexidine and alcohol-based antisepsis of the equine distal limb.

Resilience of Faecal Microbiota in Stabled Thoroughbred Horses Following Abrupt Dietary Transition between Freshly Cut Pasture and Three Forage-Based Diets.

Simple SummaryAbrupt changes in the dietary management of horses are a risk factor for gastrointestinal disturbances, potentially due to the negative effects observed on the population of gut microbiota. In this study, the faecal microbiota of horses was investigated to determine how quickly the bacterial communities respond and stabilise following sudden diet change. Six Thoroughbred mares were stabled for six weeks, consuming freshly cut pasture (on weeks 1, 3 and 5), before being abruptly fed three conserved forage-based diets (weeks 2, 4 and 6). The conserved forage-based diets were a chopped ensiled forage fed exclusively or with whole oats, and perennial ryegrass hay fed with whole oats. Faecal samples were collected at regular intervals from each horse following each diet change. High throughput 16S rRNA gene sequencing was used to evaluate the faecal microbiota. There were significant differences in the diversity of the microbiota (p < 0.001), with clustering of samples observed by diet group. A stable faecal microbiota profile was observed in the samples from 96 h after consumption of the treatment diets containing ensiled chopped forage. The present study confirmed that the diversity and community structure of the faecal bacteria in horses is diet-specific and resilient following dietary transition and emphasised the need for modern horse feeding management to incorporate large proportions of forage into diets.The management of competition horses in New Zealand often involves rotations of short periods of stall confinement and concentrate feeding, with periods of time at pasture. Under these systems, horses may undergo abrupt dietary changes, with the incorporation of grains or concentrate feeds to the diet to meet performance needs, or sudden changes in the type of forage fed in response to a lack of fresh or conserved forage. Abrupt changes in dietary management are a risk factor for gastrointestinal (GI) disturbances, potentially due to the negative effects observed on the population of GI microbiota. In the present study, the faecal microbiota of horses was investigated to determine how quickly the bacterial communities; (1) responded to dietary change, and (2) stabilised following abrupt dietary transition. Six Thoroughbred mares were stabled for six weeks, consuming freshly cut pasture (weeks 1, 3 and 5), before being abruptly transitioned to conserved forage-based diets, both offered ad libitum. Intestinal markers were administered to measure digesta transit time immediately before each diet change. The conserved forage-based diets were fed according to a 3 × 3 Latin square design (weeks 2, 4 and 6), and comprised a chopped ensiled forage fed exclusively (Diet FE) or with whole oats (Diet FE + O), and perennial ryegrass hay fed with whole oats (Diet H + O). Faecal samples were collected at regular intervals from each horse following the diet changes. High throughput 16S rRNA gene sequencing was used to evaluate the faecal microbiota. There were significant differences in alpha diversity across diets (p < 0.001), and a significant effect of diet on the beta diversity (ANOSIM, p = 0.001), with clustering of samples observed by diet group. There were differences in the bacterial phyla across diets (p < 0.003), with the highest relative abundances observed for Firmicutes (62–64%) in the two diets containing chopped ensiled forage, Bacteroidetes (32–38%) in the pasture diets, and Spirochaetes (17%) in the diet containing hay. Major changes in relative abundances of faecal bacteria appeared to correspond with the cumulative percentage of intestinal markers retrieved in the faeces as the increasing amounts of digesta from each new diet transited the animals. A stable faecal microbiota profile was observed in the samples from 96 h after abrupt transition to the treatment diets containing ensiled chopped forage. The present study confirmed that the diversity and community structure of the faecal bacteria in horses is diet-specific and resilient following dietary transition and emphasised the need to have modern horse feeding management that reflects the ecological niche, particularly by incorporating large proportions of forage into equine diets.

The Equine Faecal Microbiota of Healthy Horses and Ponies in The Netherlands: Impact of Host and Environmental Factors.

Simple SummarySeveral studies have described the bacterial composition in the intestines of horses, and several factors of influence have been detected. Variation in the results between studies, however, is substantial. Therefore, the current study aimed to study the bacterial composition in the faeces of healthy horses and ponies kept under standard housing and management condition in The Netherlands. Seventy-nine horses and ponies originating from two farms were included. Several factors, such as location, age, the season of sampling, horse type (horses vs. ponies) and pasture access significantly affected the bacterial composition. The current study provides important baseline information on variation in the bacterial composition in healthy horses and ponies under standard housing and management conditions. The aforementioned factors identified in this study to affect the bacterial population of the gut should be considered in future studies regarding the bacterial population of the equine gut.Several studies have described the faecal microbiota of horses and the factors that influence its composition, but the variation in results is substantial. This study aimed to investigate the microbiota composition in healthy equids in The Netherlands under standard housing and management conditions and to evaluate the effect of age, gender, horse type, diet, pasture access, the season of sampling and location on it. Spontaneously produced faecal samples were collected from the stall floor of 79 healthy horses and ponies at two farms. The validity of this sampling technique was evaluated in a small pilot study including five ponies showing that the microbiota composition of faecal samples collected up to 6 h after spontaneous defaecation was similar to that of the samples collected rectally. After DNA extraction, Illumina Miseq 16S rRNA sequencing was performed to determine microbiota composition. The effect of host and environmental factors on microbiota composition were determined using several techniques (NMDS, PERMANOVA, DESeq2). Bacteroidetes was the largest phylum found in the faecal microbiota (50.1%), followed by Firmicutes (28.4%). Alpha-diversity and richness decreased significantly with increasing age. Location, age, season, horse type and pasture access had a significant effect on beta-diversity. The current study provides important baseline information on variation in faecal microbiota in healthy horses and ponies under standard housing and management conditions. These results indicate that faecal microbiota composition is affected by several horse-related and environment-related factors, and these factors should be considered in future studies of the equine faecal microbiota.

PSIV-14 Fecal Microbiome of Horses Grazing Integrated Warm- and Cool-Season Grass Rotational Pasture Systems

Abstract The objective of this study was to characterize shifts in the fecal microbiota of horses grazing different forage types within integrated cool- and warm-season grass (CSG; WSG) pasture systems and to explore relationships between forage nutrients and microbial composition. Eight mares grazed integrated rotational systems containing mixed CSG and one of two WSG: bermudagrass or crabgrass. Fecal samples were collected after 2–3 weeks grazing WSG, CSG, and following an orchardgrass hay diet (HAY). Forage nutrients were determined by near-infrared spectroscopy and analyzed by two-way ANOVA in SAS (v.9.4). Following DNA extraction, 16S rRNA gene sequence analysis was conducted in QIIME 2 (v.2020.2) with Kruskall-Wallis tests for alpha diversity, Spearman correlation with forage nutrients, and taxonomic assignment with Greengenes. A random forest classifier and regressor determined ability to predict forage type and nutrients based upon bacterial composition. Statistical significance was set at p ≤ 0.05. Forage water-soluble carbohydrates (WSC) were greatest in CSG and lowest in WSG; neutral detergent fiber (NDF) was greater in HAY than CSG or WSG. Species richness and evenness (Shannon Index) was greater in horses adapted to WSG vs. CSG or HAY and was correlated with WSC (rs = -0.49) and ethanol-soluble carbohydrates (ESCrs = -0.62). The random forest classifier resulted in model accuracy of 1.0 and identified amplicon sequence variants (ASV) most important in prediction of forage type. Sixteen ASV were from the order Clostridiales including Lachnospiraceae, Ruminococcacceae, and Veillonellaceae families and the genus Coprococcus. Other important taxa included Prevotella spp., Streptococcus luteciae, and Fibrobacter succinogenes. The regressor accurately predicted WSC (r2 = 0.95) and ESC (r2 = 0.84), but not NDF (r2 = 0.09). These results suggest that the equine hindgut microbiome is impacted by forage type and soluble carbohydrate content; however, further research is required to determine functional and physiological significance in grazing horses.

Impact of Dietary Cellobiose on the Fecal Microbiota of Horses

Cellobiose is a disaccharide with potential prebiotic effects, as demonstrated in different animal species, but not yet in horses. It was, therefore, the aim of the present study to evaluate the impact of dietary cellobiose on the fecal microbiota of horses. Eight healthy adult horses and two ponies were included in this study. The animals received a diet without or with 10 g and 20 g cellobiose per day for 14 days each. At the end of the feeding periods, fresh fecal samples were collected to measure bacterial metabolites and the microbial composition. For the microbiota analysis, 16S rRNA gene sequencing was used. Cellobiose was well accepted and tolerated by the animals. The lowest fecal concentrations of D-lactate, propionic acid, i-valeric acid, and total short-chain fatty acids were measured at the dose 10 g cellobiose per horse per day (quadratic effect: P < .05). A dose-dependent increase of the relative abundance of Firmicutes (P = .049), Coriobacteriales (P < .001), and Clostridium (P = .031) could be detected. In addition, a dose-dependent decrease of the relative abundance of Bacteroidetes (P = .035) was observed. In conclusion, the increase of Coriobacteriales and Clostridium indicates a bacterial fermentation of cellobiose in the equine intestine, as members of both groups exert saccharolytic activity. As clostridia have previously been assumed to be a key component of the intestinal microbiota in horses, the observed increase of Clostridium in the feces might indicate beneficial and potentially prebiotic effects of cellobiose in horses. However, this finding requires further investigation, particularly with regard to the Clostridium species that have been promoted by dietary cellobiose.

Changes in the faecal microbiota of horses and ponies during a two-year body weight gain programme.

Obesity is a major health concern in many domesticated equids animals since it is related to metabolic abnormalities such as insulin dysregulation, hyperlipidaemia or laminitis. Ponies especially are known as “easy keepers” and are often affected by obesity and its related metabolic disorders. Research in the last decade indicated that the intestinal microbiota may play an important role in the development of obesity, at least in humans. Therefore, the objective of our study was to characterize changes in the faecal microbiota during a two-year weight gain programme which compared ponies and warmblood horses. For this purpose, 10 Shetland ponies and ten warmblood horses were fed a ration which provided 200% of their maintenance energy requirement over two years. Feed intake, body weight, body condition and cresty neck score were recorded weekly. At three standardized time points faecal samples were collected to characterize the faecal microbiota and its fermentation products such as short chain fatty acids and lactate. Next generation sequencing was used for the analysis of the faecal microbiota. During body weight gain the richness of the faecal microbiota decreased in ponies. Besides changes in the phylum Firmicutes in ponies that were already described in human studies, we found a decrease of the phylum Fibrobacteres in horses and an increase of the phylum Actinobacteria. We were also able to show that the phylum Fibrobacteres is more common in the microbiota of horses than in the microbiota of ponies. Therefore, the fibrolytic phylum Fibrobacteres seems to be an interesting phylum in the equine microbiota that should receive more attention in future studies.

The fecal microbiota of healthy donor horses and geriatric recipients undergoing fecal microbial transplantation for the treatment of diarrhea.

Background and aimsFecal microbial transplantation (FMT), a treatment for certain gastrointestinal conditions associated with dysbiosis in people, is also empirically employed in horses with colitis. This study used microbiota high-throughput sequencing to compare the fecal microbial profile of healthy horses to that of geriatric microbial transplant recipients experiencing diarrhea and tested whether FMT restores microbiota diversity.MethodsTo evaluate the effect of environment and donor characteristics on the intestinal microbiota, fecal samples were collected per rectum from 15 healthy young-adult (2–12 years) and 15 geriatric (≥20 years) horses. Additionally, FMT was performed for 3 consecutive days in 5 geriatric horses with diarrhea using feces from the same healthy donor. Fecal samples were collected from both donor and recipient prior to each FMT and from recipients 24 hours following the last FMT. The profile of the fecal bacterial microbiota was compared using 16S amplicon sequencing.Results and conclusionsIn contrast to diet and farm location, age did not significantly affect the healthy equine fecal microbiota, indicating that both healthy geriatric and young-adult horses may serve as FMT donors. The fecal microbiota of horses with diarrhea was significantly more variable in terms of β-diversity than that of healthy horses. An inverse correlation between diarrhea score and relative abundance of Verrucomicrobia was identified in surviving FMT recipients. At study completion, the fecal microbiota of horses which responded to FMT had a higher α-diversity than prior to treatment and was phylogenetically more similar to that of the donor.

Acute changes in the colonic microbiota are associated with large intestinal forms of surgical colic

BackgroundHorses that undergo surgery for treatment of primary large colon disease have been reported to be at increased risk of developing recurrent colic episodes postoperatively. The reasons for this are currently unknown. The aim of the current study was to characterise the faecal microbiota of horses with colic signs associated with primary large colon lesions treated surgically and to compare the composition of their faecal microbiota to that of a control group of horses undergoing emergency orthopaedic treatment. Faecal samples were collected from horses in both groups on admission to hospital, during hospitalisation and following discharge from hospital for a total duration of 12 weeks. Additionally, colonic content samples were collected from surgical colic patients if pelvic flexure enterotomy was performed during laparotomy. A total of 12 samples were collected per horse. DNA was extracted from samples using a commercial kit. Amplicon mixtures were created by PCR amplification of the V1 – V2 regions of the bacterial 16S rRNA genes and submitted for sequencing using the Ion Torrent PGM next-generation sequencing system. Multivariate data analysis was used to characterise the faecal microbiota and to investigate differences between groups.ResultsReduced species richness was evident in the colonic samples of the colic group compared to concurrent sampling of the faeces. Alpha and beta diversity differed significantly between the faecal and colonic microbiota with 304 significantly differentially abundant OTUs identified. Only 46 OTUs varied significantly between the colic and control group. There were no significant differences in alpha and beta diversity of faecal microbiota between colic and control horses at admission. However, this lack of significant differences between groups should be interpreted with caution due to a small sample size.ConclusionsThe results of the current study suggest that faecal samples collected at hospital admission in colic cases may not accurately represent changes in upper gut microbiota in horses with colic due to large colon disease.

Selection of a candidate probiotic strain of Pediococcus pentosaceus from the faecal microbiota of horses by invitro testing and health claims in a mouse model of Salmonella infection.

AimsThe aim of this study was to verify the suitable use of candidate ‘probiotics’ selected by in vitro tests and the importance of in vivo assays to nominate micro‐organisms as probiotics and alternative prophylactic treatments for Salmonella Typhimurium infection.Methods and ResultsThirty‐three lactic acid bacteria (LAB) isolated from foal's faeces were assessed based on the main desirable functional in vitro criteria. Based on these results, Pediococcus pentosaceus strain 40 was chosen to evaluate its putative probiotic features in a mouse model of Salmonella infection. Daily intragastric doses of Ped. pentosaceus 40 for 10 days before and 10 days after Salmonella challenge (106 CFU of Salm. Typhimurium per mouse) led to a significant aggravation in mouse health by increasing weight loss, worsening clinical symptoms and anticipating the time and the number of deaths by Salmonella. Pediococcus pentosaceus modulated cell‐mediated immune responses by up‐regulation of the gene expression of the proinflammatory cytokines IFN‐γ and TNF‐α in the small intestine.ConclusionThe usual criteria were used for in vitro screening of a large number of LAB for desirable probiotic functional properties. However, the best candidate probiotic strain identified, Ped. pentosaceus #40, aggravated the experimental disease in mice.Significance and Impact of the StudyThese findings emphasize the need for prophylactic or therapeutic effectiveness to be demonstrated in in vivo models to make precise health claims.

Analysis of the large intestinal and faecal microbiota of horses with grass sickness using denaturing gradient gel electrophoresis

within the insect, the infectious virus can remain in its mouthparts for several hours after a bite. Several studies have shown that EIAV infection has occurred among horses in North and South America, France, Germany, Italy and Romania. So far no evidence of the presence of EIAV in horses was reported in Serbia. In order to determine EIA prevalence among the 5000 horses of the Vojvodina region of Serbia, 316 horse sera have been collected during 2013 and 2014. Serological analysis was performed using Agar Gel Immuno Diffusion test (AGID) as described by the World Organization of Animal Health (OIE) chapter 2.5.6 and by Enzyme Linked Immuno Sorbent Assay (ELISA). Those sera have been tested with 3 different commercial AGID test (Idexx, ID-Vet and VMRD) and two different commercial ELISA tests (Synbiotics and VMRD). With the three AGID kit 311 (98,4%) among the 316 tested sera were found negative and only five (1,6%) sera were found positive for EIA. With ELISA kits results were slightly different and were dependent of the kit producer. Indeed, Synbiotics ELISA gave the same results as AGID tests but VMRD ELISA found 295 negative samples, 5 positive samples and 16 samples were doubtfull. Our study shows for the first time that EIA is present in Serbia and more specifically in the Vojvodina region. Euthanasia and viral characterization of field isolate collected in Serbian horses will be investigated.

Analysis of the large intestinal and faecal microbiota of horses with grass sickness using denaturing gradient gel electrophoresis

Analysis of the large intestinal and faecal microbiota of horses with grass sickness using denaturing gradient gel electrophoresis

Fecal microbiota of horses in the clinical setting: Potential effects of penicillin and general anesthesia

Antimicrobial treatment is associated with the spread of antimicrobial resistance and disturbances in the ecological balance of intestinal microbiota. In horses, the main adverse effect of antimicrobial treatment is colitis. We used culture and 16S rRNA gene based molecular methods to monitor the prevalence of antimicrobial resistance and changes in predominant fecal populations during penicillin treatment and general anesthesia of horses in the clinical setting. After 5 days of parenteral administration of penicillin, fecal Escherichia coli were resistant to multiple unrelated antimicrobial agents when compared to the pre-exposure situation. Denaturing gradient gel electrophoresis (DGGE) profiles indicated that horses have an extremely diverse fecal microbiota, with marked differences between individual horses. Most of the variation in DGGE profiles could be attributed to horse-specific factors, and penicillin, general anesthesia or both could not explain the remaining variation. Within-animal variation remained less than between-animal variation despite treatment. However, real-time PCR quantification (qPCR) indicated subclinical changes in selected bacterial groups of the penicillin treated horses.