NE2202: The Equine Microbiome

(Multistate Research Project)

Status: Active

NE2202: The Equine Microbiome

Duration: 10/01/2022 to 09/30/2027

Administrative Advisor(s):


NIFA Reps:


Statement of Issues and Justification

The stakeholders who will be served by the Equine Microbiome Multistate Research Project include: microbiome researchers, undergraduate and graduate students, horse owners/managers, veterinarians, equine industries (e.g., feed producers, nutritionists, behaviorists, competitors) equine enthusiasts (e.g., youth groups and spectators), and underserved communities (e.g., rural horse owners, undergraduate and graduate students with limited access to bioinformatics, biostatistics, or data resources).

The impact of the gut microbiome has been a popular press topic for human and animal care. Despite the popularity and availability of products purporting to positively impact health through modulating the gut microbiome, research to understand the details of the functional impact of the microbiome on horse health and disease is still in its infancy. A search of PubMed using the terms “equine” and “microbiome” yielded only 320 results. Publication in this area has increased exponentially since 2007.

This project will address the following needs to bridge the gaps between equine microbiome researchers, equine industries, students, and the public:

  • Advance the knowledgebase of equine microbiome science with robust, statistically significant sample numbers and diversity.
  • Communicate advances in microbiome science among researchers, students, and the public.
  • Refine and test protocols and standardized methods for researchers to enable collaboration, data sharing, and greater utilization of samples and data.
  • Provide translational connections between the equine gut microbiome and health parameters, nutrition, management, and performance that are based on research specific to horses.

 

 

The importance of this work is to establish a coordinated and collaborative network of equine microbiome researchers and to provide resources for students and the public that reflect the state of the science.

  The consequences of not establishing the Equine Microbiome Multistate Group are:

  • Confusion regarding the state of equine microbiome science among stakeholders leading to the spread of misleading information, poor purchasing and management decisions, uncertainty regarding efficacy of products and research strategies.
  • The continued lack of coordination among researchers, which is slowing the progress and efficiency of the science due to smaller sample sizes (horsekeeping is expensive relative to other species), inconsistent sampling and analysis protocols, inconsistent training of students, inability to share data.
  • Lack of understanding of the unique aspects of the equine microbiome and continued application of therapeutic and nutritional strategies from research on other species such as ruminants and humans that may be ineffective or have adverse effects on the unique equine gastro-intestinal tract.

 

The Equine Microbiome Multistate Group will leverage the depth of technical expertise, range of laboratory, sequencing, and animal facilities, diversity of horse herds, extensive archival sampling efforts, training capacities, and stakeholder support of each member institution. As a nation-wide effort, this group will maximize impact of data/samples for multiple studies, reach numbers needed for statistical power, mitigate geographic bias, and coordinate host and microbial connections.

 

  As a Multistate effort, this project will be able to:

  • Investigate the role of the microbiome in equine health by describing the structural and functional properties of the “normal” equine microbiome.
  • Test hypotheses of microbiome structure and function related to management, life stages, and disease states.
  • Build and test statistically rigorous models of host-microbiome interactions.
  • Establish a network for training students, outreach to the public, veterinary and equine industry education in microbiome science.
  • Create print and web-based resources for the equine community (public, education, veterinary and equine industry) to increase understanding of microbiome science and its applications to improving horse health and management.

 

  The likely impacts of successful completion of this work are:

  • For Researchers,
    1. The opportunity to work together to develop grant proposals and experimental collaborations to explore research questions focused on understanding the structure and function of the equine microbiome in health and disease. Through this multistate we will share expertise, horse samples, and technical assistance for basic and translational experimental approaches.
    2. The establishment of a shared database of metadata, sequence data and samples will increase the statistical power and efficiency of hypothesis testing and enable a baseline for the “normal” equine gut microbiome.
  • For Students,
    1. The shared database will provide opportunities to formulate and test hypotheses and increase research participation, especially for students from institutions with limited access to research facilities.
    2. Training in bioinformatics, biostatistics, microbial ecology, and host physiology will enable students to apply knowledge and techniques to real-world questions in the equine industry.
    3. The opportunity to participate in conferences and workshops around equine microbiome science will enable students to widen their network, practice presentation skills, and gain confidence in their futures as scientists.
  • For Veterinarians and Nutritionists,
    1. Equine microbiome insights will inform clinical practice, specifically understanding the impact of the microbiome on horse health and welfare and advice given to horse owners regarding management, feeding, and modulating the microbiome via pre/probiotics or antibiotics.
    2. Observations made by clinical veterinarians will provide essential questions and insights for microbiome researchers.
    3. Conference and workshop participation will enable veterinarians to share insights, clinical strategies, questions, and case studies.
    4. Creation of evidence-based web and print resources to translate equine microbiome science for better health outcomes.
    5. Equine nutritionists and feed manufacturers will gain insight to how the diet, microbiome, and equids interact with the potential to guide feeding recommendations and feed formulation for improved outcomes (health, welfare, and performance).
  • For Horse Owners and Enthusiasts,
    1. Greater understanding of the impacts of diet, management, and other factors on microbiome health, and ultimately the welfare of horses.
    2. Deeper knowledge of specific feed ingredients, pre/pro/postbiotics will enable horse owners to make informed decisions in purchasing horse care products, supplements, and feeds.
    3. Conference and workshop participation will help to dispel myths about the impacts of feeding and management practices on the microbiome, and promote equine welfare through communication of research based findings.
    4. Creation of evidence-based web and print resources to translate equine microbiome science for better management decisions.

Related, Current and Previous Work

  The equine gut microbiome plays a crucial role for the horse in accessing nutrients and energy. As hindgut fermenting animals, horses have limited endogenous enzyme capacity in the foregut to digest and absorb nutrients, instead relying on communities of microbes in the caecum and colon to break down complex plant material and ferment simple sugars and amino acids to short chain fatty acids that can be used by the horse for energy. Unlike ruminants that are known for digestive efficiency (50-60% DM digestibility of forage), the horse is less successful at extracting nutrients (39-48% DM digestibility of forage)[1]. To maintain energy levels for their large body size, a high rate of intake is required. Since approximately 70% of the equine energy budget is supplied by microbial fermentation products [2,3], this activity is a crucial component of the horses’ energy budget.

  Studies to characterize the structure of the equine gut microbiome show that the equine gut microbiome is dominated by Firmicutes and Bacteriodetes, with lesser abundance of other groups such as Spirochaeta, Proteobacteria, Fibrobacter, and Archaea [4–7]. Relative abundances of members of equine gut microbiome are sensitive to diet [4–7], stress [8–10], immune status [11], and age [12,13], and prebiotics and probiotics formulated and marketed to modulate the equine gut microbiome show variable results [14–17].

  Most published equine microbiome studies are based on relatively few samples. Efforts to combine datasets to increase statistical power are constrained by inconsistent sampling and/or sequencing methods, confounding factors (diet or environment), and differences in duration or other conditions. Efforts such as the Human Microbiome Project [18,19], the American Gut Project [18–20], and the development of the Dog Dysbiosis Index [21] demonstrate the power of a coordinated, collaborative approach to microbiome research.

  As equine microbiome science moves from compositional surveys to functional studies seeking to understand the details of the conversations between the microbiome and the horse host, the formation of this Multistate Research group will enable us to expand our sampling and experimental design possibilities through leveraging the diversity of horse herds, environmental conditions and dietary strategies available to our members. At the same time, we bring a depth and breadth of expertise to bear in microbial ecology, genomics, genetics, bioinformatics, clinical veterinary practice, equine management, nutrition, and biostatistics. The coordination of sampling protocols and the establishment of a shared database of samples and metadata will provide resources and lasting benefits to the equine science community.   

Objectives

  1. Identify the components and functionalities of the “normal” equine gut microbiome.
  2. Identify how specific factors: diet, age, metabolic diseases, inflammation, stress and others impact the structure and function of the equine microbiome, and examine the role of the equine microbiome in management, maintaining horse health, and disease prevention.
  3. Explore how the genetic factors of horses are related to and interact with the gut microbiome.
  4. Disseminate results and applications of microbiome science for equine welfare and management through stakeholder outreach and education, including youth groups, horse owners, nutritionists, and veterinarians.
  5. 5. Create print and web-based resources for the equine community (public, education, veterinary and equine industry) to increase understanding of microbiome science and its applications to improving horse health and management.

Methods

Objective 1: Identify the components and functionalities of the “normal” equine gut microbiome through comparative surveys, in vitro and in vivo approaches.

Core community structure will be explored through sequence-based 16S and shotgun metagenomic surveys. Culture-based approaches will be used to investigate functional aspects of equine microbiome communities including in vitro challenge trials and in vivo measurements of nutrient acquisition, digestibility, and metabolite production.

To leverage the maximum number of horses and regions, we will establish methods for equine microbiome sampling, reporting metadata, and data analysis to enable robust comparisons between studies, efficiency in sample utilization and synthesis of datasets. Datasets will be provided to the research community through an equine microbiome database to facilitate sharing of data from a range of samples. Sampling and reporting methods will be with established metadata standards and ontologies tailored to the unique aspects of equine management and use.

 

Objective 2: Identify how specific factors: diet, age, metabolic diseases, inflammation, stress, environment and others impact the structure and function of the equine microbiome and examine the role of the equine microbiome in management, maintaining horse health, and disease prevention.

Associations between microbiome differences and horse health parameters will have greater statistical power as the database grows. Sequence and culture-based experiments based on hypotheses generated from the survey data will seek to understand the roles of the microbiome in maintaining equine health. This effort will focus on three areas: a) Normal life stage factors such as age, breed, gender and others  b) Management factors such as diet, workload, stress, environment and others, c) Disease factors such as parasitism, metabolic diseases, inflammation, and others.

 

Objective 3: Study how the genetic factors of horses are related to and interact with the gut microbiome.

To explore the horse side of the host-gut microbiome conversation, genotype and phenotype analysis of equine samples including feces, blood, hair, saliva, gastric fluid, gastric endoscopy results, vaginal swabs, semen and others will be correlated with sequence and culture-based data to develop and test models of host-gut microbiome interactions.

 

Objective 4: Disseminate results and applications of microbiome science for equine welfare and management through stakeholder outreach and education, including youth groups, horse owners, nutritionists and veterinarians.

We will develop and share workshop series/other educational events for stakeholders in the area of equine microbiome science. Additionally, we will organize a yearly conference segmented to target multiple stakeholder audiences to develop collaborations, share data analysis strategies, disseminate and interpret findings.

Objective 5:  Create print and web-based resources for the equine community (public, education, veterinary and equine industry) to increase understanding of microbiome science and its applications to improving horse health and management.

As understandings of the impacts of the equine microbiome on horse health are shown by the data (generated by this project or elsewhere), we will create and disseminate print and web based materials to enable each stakeholder group to translate findings into more informed decision making for care and management.

Measurement of Progress and Results

Outputs

  • 1. Written protocols (SOPs) for Equine Microbiome studies: sampling, metadata collection and analysis
  • 2. Database of samples/ metadata that is searchable and shared with the equine microbiome research community.
  • 3. Organization of workshops for stakeholders and a biyearly Equine Microbiome Conference for the scientific, general and clinical community.
  • 4. To support grant writing and experimental design efforts, compilation of a list of available horses, environments, and facilities to provide a diversity of samples as well as available technical resources and facilities to provide analytical depth and strength for cutting edge research plans.
  • 5. Collaborative grant proposals and manuscripts identifying the structure and functionality of the core microbiome for horses.
  • 6. Collaborative grant proposals and manuscripts reporting the role of the microbiome in equine health, management, and disease prevention.
  • 7. Collaborative grant proposals and manuscripts reporting the interactions between the equine gut microbiome and host genetics, behavior, and immune function.
  • 3. Informational materials (print and web-based) for non-academic stakeholders to translate microbiome science into better care and management decisions.

Outcomes or Projected Impacts

  • 1. Increased communication, collaboration, networking among scientists working on Equine Microbiome topics.
  • 2. Increased rate of progress in equine microbiome science from survey-based studies to a more functional understanding
  • 3. Greater depth and breadth of communication of equine microbiome topics to the equine science community at large.
  • 4. Greater level of funding for equine microbiome grant proposals.
  • 5. Increased integration of microbiome insights into equine nutrition and veterinary publications and products.
  • 6. Increased integration of knowledge of the equine microbiome and the host.

Milestones

(2022):Establishment of protocols (SOPs) for Equine Microbiome Studies.

(2022):Compilation of lists of horses/facilities and technical/analytical capabilities.

(2022):Identification of questions and hypotheses for collaborative study and submission of collaborative grant proposals.

(2022):Identify stakeholder questions and confusions around the equine gut microbiome.

(2023):Organization and establishment of the Equine Microbiome Database.

(2023):Initial addition of data/samples and metadata to the database.

(2023):Planning and executing the first Equine Microbiome Conference

(2023):Studies and grant proposals to identify the role of the microbiome in equine health, management, nutrition, and disease prevention.

(2023):Studies and grant proposals to identify host/microbiome factors related to behavior, immune health, performance.

(2023):Development and dissemination of educational materials about the equine microbiome in health, management, and nutrition.

(2024):Evaluation of depth and breadth of Equine Microbiome Database.

(2024):Expansion of the Equine Microbiome Database in sample types, horses and numbers as identified.

(2024):Continue studies to identify the role of the microbiome in equine health, management, nutrition, and disease prevention.

(2024):Continue studies to identify host/microbiome factors related to behavior, immune health, performance.

(2024):Development and dissemination of educational materials about the equine microbiome in health, management, and nutrition.

(2025):Planning and executing the second Equine Microbiome Conference

(2025):Continue identification of underrepresented sample groups and expansion of Equine Microbiome Database.

(2025):Summarize outcomes and determine directions for new and ongoing research questions.

(2025):Complete and write materials summarizing collaborative studies.

(2025):Make practical recommendations about management decision-making and maintaining horse health through microbiome interventions

(2025):Development and dissemination of educational materials about the equine microbiome in health, management, and nutrition.

Projected Participation

View Appendix E: Participation

Outreach Plan

  1. Identification of questions and misinformation about the equine microbiome by all stakeholder groups.
  2. Creation of education materials (webinars, fact sheets, speaker series, infographics) accessible to the general public regarding equine management in relation to the microbiome. These would be freely available to the public and accessible to under-served and underrepresented communities.
  3. Opportunities for stakeholder submission of samples and contribution of metadata to the Equine Microbiome Database.
  4. The Equine Microbiome protocols and database will be made available to researchers and the public through an accessible web portal.
  5. Biyearly conference will have segments targeted to the more general equine owning/managing public.
  6. Workshops for youth groups (e.g. FFA, 4-H, and Pony Club) and undergraduate students on microbiome science.
  7. Manuscripts of experimental results submitted to peer review journals.

Organization/Governance

Organization/Governance

This regional project NE-18869 was initiated by the University of Delaware and the University of Connecticut. The following are the current members of the project:

 

Amy Biddle, University of Delaware

Jenifer Nadeau, University of Connecticut

Sarah Reed, University of Connecticut

Aaron Ericsson, University of Missouri

Robert Coleman, University of Kentucky

Stephen Coleman, Colorado State University

Carissa Wickens, University of Florida

Samantha Brooks, University of Florida

Carolyn Hammer, North Dakota State University

Sara Mastellar, The Ohio State University’s Agricultural Technical Institute

Sarah Springer-White Texas A&M University

 

 

The recommended Standard Governance for multistate research activities including the election of a Chair, a Chair-elect, and a Secretary. All officers will be elected for at least two-year terms to provide continuity. Administrative guidance will be provided by an assigned Administrative Advisor and a CSREES Representative. As of this date, Amy Biddle, University of Delaware, has been serving as chair and will continue so until the initiation of the next project. Subcommittee/research teams will be chosen from the objective areas: 1. Evaluate existing methods for equine microbiome sampling, reporting metadata, and data analysis to enable robust comparisons between studies, efficiency in sample utilization and synthesis of datasets. 2. Establish an equine microbiome database to facilitate sharing of data from a range of samples. 3. Compile a list of available expertise, horses, and facility resources at participants’ locations to leverage institutional strengths to address microbiome questions using cutting edge techniques. 4. Compile a list of available technical and analytical resources: Core facilities and expertise for sequencing, diagnostic labs, specialized analytical tools to leverage institutional strengths to address microbiome questions using cutting edge techniques. 5. Identify the components of the “normal” equine gut microbiome. 6. Disseminate results and applications of microbiome science for equine welfare and management through stakeholder outreach and education, including youth groups, horse owners, nutritionists and veterinarians. 7. Examine the role of the equine microbiome in management, maintaining horse health and disease prevention. 8. Study how the genetic, immunological, and behavioral factors of horses are related to and interact with the gut microbiome.

Literature Cited

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  4. Diether N, Willing B. Microbial Fermentation of Dietary Protein: An Important Factor in Diet–Microbe–Host Interaction. Microorganisms. 2019;7: 19. doi:10.3390/microorganisms7010019
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  7. Dougal K, Harris PA, Girdwood SE, Creevey CJ, Curtis GC, Barfoot CF, et al. Changes in the Total Fecal Bacterial Population in Individual Horses Maintained on a Restricted Diet Over 6 Weeks. Frontiers in Microbiology. 2017;8. doi:10.3389/fmicb.2017.01502
  8. Alexander S, Irvine CHG. Stress in the Racing Horse: Coping vs Not Coping. JES. 1998;9: 77–81. doi:10.1294/jes.9.77
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  15. Tanabe S, Suzuki T, Wasano Y, Nakajima F, Kawasaki H, Tsuda T, et al. Anti-inflammatory and Intestinal Barrier–protective Activities of Commensal Lactobacilli and Bifidobacteria in Thoroughbreds: Role of Probiotics in Diarrhea Prevention in Neonatal Thoroughbreds. Journal of Equine Science. 2014;25: 37–43. doi:10.1294/jes.25.37
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Attachments

Land Grant Participating States/Institutions

CO, CT, DE, FL, ND

Non Land Grant Participating States/Institutions

Ohio State University
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