Research shows slight shifts in the intestinal microbiome can have far-reaching effects

Scientists are slowly but surely unveiling the secret workings of the equine intestinal microbiome. Although it might sometimes seem more slow than sure, we must appreciate that studying the microbes that reside in the large intestine and cecum can be like shining a flashlight onto the surface of the Atlantic Ocean at 41.726931° N, 49.948253° W, hoping to study the Titanic.  

The Healthy Intestinal Microbiome

Originally, researchers believed the sole, or at least primary, role of the intestinal microbes was fiber fermentation. As we know, horses cannot digest the fiber that makes up the bulk of their diet with teeth, saliva, and gastric acids alone. Instead, they rely on the bacteria within the large intestine to ferment those feedstuffs and produce volatile fatty acids—short-chain fatty acids the horse then uses for energy.

Now, we know the intestinal microbiome comprises far more than just fiber-fermenting bacteria. In fact, the term microbiome refers to all the microbes and their genes that call the hindgut home, including bacteria, fungi, viruses, and protozoa.

“The gut microbiome is linked to so many parts of what we think of as host health,” says Grace Vaziri, a PhD candidate in ecology and evolutionary biology at the University of Connecticut, in Storrs. “In addition to its role in digestion, the microbiome influences immune system development and maturation. As well, gut microbes help maintain a constant gut-brain conversation going based on the chemical byproducts they produce during digestion.”

It is widely accepted that horses have a “normal” population of microbes in a healthy intestinal tract. This might vary somewhat from one horse to another but, overall, healthy horses generally have the same core microbiome.

According to a 2023 review article by Carolyn E. Arnold, DVM, PhD, Dipl. ACVS, published in Veterinary Clinics of North America: Equine Edition, healthy horses have about 17 to 20 different bacterial phyla in their fecal microbiome, which is reflective of the large colon (it’s difficult to impossible to sample the hindgut microbiome in the live horse), but three main phyla tend to be the most common in all equine microbiome studies: Firmicutes, Bacteroidetes, and ­Verrucomicrobia.

The Firmicutes, the most predominant phyla, play important roles in degrading complex plant materials. This phylum includes bacteria in the Lachnospiraceae and Ruminococcaceae families. These bacteria produce volatile fatty acids essential for equine health.

Bacteroidetes also ferment fiber to produce volatile fatty acids, while the Verrucomicrobia help maintain the gut barrier between cells lining the intestinal wall and the contents of the intestinal tract.

Research shows microbiome health is tenuous and relatively easily unbalanced, resulting in dysbiosis (alteration of the normal balance of the bacterial communities inhabiting the intestine).

“This dysbiosis results in changes in the bacterial communities and lack of important functions such as the production of short-chain fatty acids that are essential to maintain a healthy microbiome and the health of the intestinal cells,” says Diego Gomez, MV, MSc, MVSc, PhD, Dipl. ACVIM, an assistant professor in the Department of Clinical Studies at Ontario Veterinary College, in Guelph, Ontario, Canada.        

A Dysfunctional Hindgut

The intestinal microbiome can get blown out of kilter as easily as the wind blows west. In her review Arnold says any of the following can contribute to dysbiosis:

• Age, breed, and sex.
• Pregnancy status.
• Transport.
• Exercise intensity.
• Fasting.
• Season.
• Dietary variables (e.g., pasture, abrupt diet changes, feeding concentrates vs. forage).
• Medications, including antibiotics.

“Really, anything that stresses the horse can disrupt the intestinal microbiome,” says Gomez.

Recently, several research groups have delved into the depths of the equine ­intestinal microbiome, hoping to shed light on what affects its balance. Here are two such studies with comments from the authors explaining the significance of their findings.

The Microbiome in Horses With and Without Laminitis

Horses with colitis (inflammation of the colon, which results in diarrhea and other clinical signs) suffer dysbiosis. For example, study results show the abundance of Firmicutes decreases and bacteria such as Enterobacteriaceae (Escherichia, Shigella, Salmonella) increase in affected horses.

Gomez says myriad bacterial genera belong to the phylum Firmicutes. “Overall, the phylum Firmicutes decreases significantly during dysbiosis, especially genera from the families Clostridiaceae, Prevotellaceae, Ruminococcaceae, and Lachnospiraceae. During dysbiosis, however, some Firmicutes can increase, such as Lactobacillus,” he explains.

Horses that develop colitis are at risk of also developing the hoof disease laminitis and might not survive treatment. In turn, laminitis is associated with alterations of the gastrointestinal microbiota and absorption of bacteria and bacterial byproducts through the inflamed intestinal wall.

“Studying the microbiomes of horses with colitis will help to determine which bacteria are directly related to laminitis development,” says Gomez. “This will aid in developing targeted treatments to rapidly ameliorate or correct dysbiosis, which may include administering probiotics and/or prebiotics and performing fecal matter transplantations.”

In their 2022 study Gomez and colleagues collected and analyzed fecal samples from 36 healthy horses, 15 horses with colitis and laminitis, and 39 horses with colitis but no laminitis.

As expected, based on previous reports, horses with colitis suffered dysbiosis. The diarrheic horses included in this study had microbiomes enriched with (containing increased densities of) facultative anaerobes (Lactobacillus, Enterococcus) and decreased obligate anaerobes (Treponema, Ruminococcus, Prevotella). Horses with colitis that went on to develop laminitis had microbiomes enriched in Lactobacillus and Streptococcus and bacteria in the Enterobaceriaceae family. These results were in line with previous reports.

“Experimental studies inducing laminitis with high amounts of dietary carbohydrates also had increased Lactobacillus and Streptococcus,” says Gomez.

He says the enrichment of these bacteria in laminitic horses could be related to the microbiota’s effect on the immune system and changes in the intestinal pH leading to an acidic colonic environment. His team hypothesized that these microorganisms induce critical damage to the intestinal tract lining, allowing pathogenic (disease-­causing) bacteria to invade and multiply in the gut, causing more intestinal wall damage, and releasing bacteria and toxins into the horse’s bloodstream. This can cause inflammation of and damage to the tissue connecting the hoof and the underlying coffin bone and consequent laminitis.

“Together, these data suggest that the hindgut microbiota could play a role in the development of laminitis and may affect the severity of disease in horses with colitis and that treatment approaches to ameliorate dysbiosis in horses with colitis could have a positive impact in the prevention of laminitis,” reports Gomez.

Microbiome Shifts With Social Disruptions

Recognizing the role of stress in the stability of the intestinal microbiome, Vaziri et al. (2023) examined the fecal microbiomes of free-living feral mares as they changed social groups.

“Feral horses organize themselves in bands that are normally long-lasting,” explains the project’s principal investigator, Cassandra Nuñez, PhD, of the Department of Biological Sciences at The University of Memphis, in Tennessee. “Stability of these groups is important to the overall well-being of all group members. Decreased stability has been associated with decreased body condition score and reproductive success, increased parasite load, and increased offspring mortality.

“These changes, in addition to the fact that mares moving into a new band are often subject to aggressive behaviors such as chasing, kicking, and biting, could act as stressors, potentially affecting the intestinal microbiome,” she adds. “Stress has been linked to changes in the microbiome in several species including laboratory mice, rhesus monkeys, bullfrogs, coral, and domestic horses.”

Nuñez says horses changing bands could also experience new environments, forage types, water access, and exposure to the microbial communities of their new herdmates, all while establishing rank.

To evaluate whether social instability (i.e., changing bands) affected mares’ intestinal microbiomes, her team studied feral mares on Shackleford Banks, North Carolina. They collected fecal samples from mares with known identities, tracking which band they were part of or changing to. They gathered 16 samples from mares that changed groups and 36 samples from mares that did not.

The researchers found the composition of the microbiomes from mares that did change bands—and, therefore, experienced social instability—differed from mares that did not change bands.

“One of the analytical methods we used to compare the microbiomes between mares that did and didn’t change bands is called permutational multivariate analysis of variance (PERMANOVA),” says Vaziri. “This analysis can tell us if the centers of two clusters of points (one cluster representing all the microbiomes of horses that did change groups, and the other cluster representing the microbiomes of mares that didn’t change groups) are located in significantly different locations. While this analysis doesn’t tell us about the nature of the differences, it did show us that the centers of those two clusters were significantly different from each other.

“We also used a few methods to specifically characterize the differences between communities,” she continues. “First, we found that the overall bacterial community was simply more unstable (or less consistent) in mares that changed bands compared to mares that didn’t. In other words, the microbiomes of two mares who changed groups had less in common with each other than the microbiomes of two mares that didn’t change groups.”

Vaziri et al. also assessed whether the presence and abundance of any bacteria were strongly associated with mares that changed groups. They found that several were, including bacteria in the genus ­Blautia.

“Blautia were consistently less present in feces from mares that did change groups than in feces from mares that didn’t,” she says. “In general, Blautia seem to be important to host diet and metabolic processes. While our study didn’t track mare diet or nutritional intake in association with group-changing behavior, our observation that Blautia are less common in mares that change groups is intriguing and suggests that future work could consider diet as a possible mechanism for how group-­changing behavior may precipitate changes to the gut microbiota.”

In addition, the researchers reported no differences between the alpha diversity of mares that did and did not change bands.

“This means that the microbial communities of mares that did and did not change bands were similarly diverse: The number of species making up their microbial communities and the abundance and phylogenetic diversity of those species were not associated with group-changing behavior by mares,” explains Nuñez. “Thus, social perturbations, such as changing groups, may not precipitate major shifts to the microbiome. Instead, more subtle changes in the structure of the microbial community structure may occur.”

Vaziri adds, “The subtle changes we saw offer interesting hints about what other challenges mares might face when they change groups.”

“Mares that change groups more frequently may be less able to ‘develop’ stable microbial communities in their guts because they keep changing groups and therefore are subject to the new environments, forage type, water access, and exposure to the microbial communities of their new bandmates,” says Nuñez.

In conclusion, she says, “The observed changes in the mares’ microbiomes could prove critical to horses—animals that are heavily dependent on their microbiota for nutrient absorption. Still, whether the changes in the microbiomes observed in mares changing bands are beneficial, detrimental, or neutral remains unclear. But these results do demonstrate a link between animal behavior and physiology.” 

Take-Home Message

The sheer size of the equine hindgut and the relative inability to collect samples from it currently hampers our capacity to study the intestinal microbiome. Further, the available tests are limited to identifying ­bacteria (not the other types of microbes) and only at the family or, maybe, genus level but not typically at the species level. 

This means we can’t truly see what’s in there and we have limited tools to more importantly ask, “What are they doing in there?” “Nonetheless, the recent microbiota studies have demonstrated the importance of the gut microbial communities for host health and provide insights into the potential benefits of administering probiotics, prebiotics, and fecal matter transplantation to prevent and treat life-threatening diseases, including colitis and laminitis,” Gomez says.