AnimalBiome lays out best practices for microbiome study design in animal research

The rapidly evolving field of research has sparked the interest of companies and researchers alike, with many hoping to to implement it. Despite the growing demand for clinical and dietary trials in companion animals to include microbiome investigations, best practices for doing so in veterinary medicine is lacking in scientific literature. 

In a new paper published in Frontiers in Veterinary Science​, AnimalBiome researchers aim to review best practices for conducting feeding trials or clinical trials that study the effects of an intervention on the microbiota. 

The paper addresses common mistakes that are made, such as cost-cutting decisions that reduce the value of the study and ultimately prevent brands from making particular product claims. Additionally, inexperienced researchers may miss ways to optimize the execution of a study that could have helped them obtain clear, unambiguous results, that in many cases would have been simple and inexpensive. 

“Through experience, we have learned that conducting microbiome analysis in pets has important aspects to consider in study design, which we haven’t seen discussed by others. We wanted to provide a clear, concise guide to help non-specialists get the most out of their investment of research dollars, and promote high-quality data in the published scientific literature,”​ explained Jessica Jarett, first author of the study and Computational Biologist at AnimalBiome. 

Complex interactions 

Jarett told NutraIngredients-USA​ that an animal’s gut microbiome is linked to its genes, immune system, the food it eats, lifestyle and treatment choices—and these interactions are complex and hard to predict.

“Individual breeds of dogs and cats have specific genetic differences and predispositions for certain diseases associated with the gut microbiome. For example, genetically susceptible Boxers get inflamed colons because their immune cells, called macrophages, allow the bacteria E. coli to adhere, invade and replicate within them. In general, pets and people that have intestinal inflammation have larger and more frequent changes within their microbiomes, again demonstrating the influence of the immune system,"​ explained Jarett.

“Cheap is expensive, and expensive is cheap”​

“‘Cheap is expensive, and expensive is cheap’ —an adage that couldn't be more applicable to investing in pet microbiome research,” ​the authors said. “ Limiting study costs poses a substantial risk of conducting an underpowered or fundamentally flawed study with inconclusive results. Conversely, a larger expenditure for a well-designed study of adequate size may be more expensive up front, but can pay dividends in credible, well-supported product claims and more successful products. Our strongest recommendations for best practices in companion animal studies with microbiome testing are to simplify the research question(s), perform microbiome pre-screening, include appropriate controls, consider a consistent pre-study diet for all animals, and allow 30 days for full microbiome response to diet change.”​

Jarett said that human medical research, including microbiome research, has much better funding mechanisms than do animal studies, so often animal science is leveraging human health advancements. 

“Only recently has the cost and ease of amplicon sequencing technology decreased enough to bring it within reach of smaller pet food companies,”​ she said.

Study design 

The guide covered experimental design, dietary considerations, recruitment, and microbiota sampling choices. The review also looked at a handful of basic and commonly used designs in intervention trials and microbiome studies, with general recommendations and caveats for each. 

“Beyond providing a brief overview of the two most common sequencing methods currently used to characterize microbial communities, we do not provide a comprehensive review of the diverse methods for sequencing or analyzing microbiome data, but rather refer readers to the many excellent reviews on this topic, including several written for non-bioinformaticians,”​ the report explained. 

Large, diverse group of healthy companion animals needed 

According to the authors, it’s easier to define an unhealthy microbiome than a healthy one. The report identifies a need for specific organisms that comprise a healthy microbiome, given their relative abundances can vary widely between individual animals. 

Jarett explained that a microbiome that is dominated by a single type of bacteria, or one that contains high abundances of bacteria associated with disease states, is easy to recognize as unhealthy. However, there are many possible ways a healthy microbiome can be assembled. For this reason, a large reference set of microbiomes from healthy animals is necessary for comparative research. 

Take-aways 

Simply put, Jarett suggests having a clear and simple focus for studies, which allow for a clear and unambiguous answer to the research question. She also advises researchers to perform microbiome analyses as a standard part of the pre-screening process. 

Another suggestion is to anticipate big influences on the microbiome and normalize them where possible. “For example, have all participants eating the same diet during the run-in period. Include appropriate control groups that match the treatment groups,” ​said Jarett. “Plan for an adequate duration of the study. A month is a good benchmark for dietary interventions.”​

What to know and how to apply it  

As evidence-based practices and microbiome testing in intervention trials become more common to support product claims, the authors advise veterinarians to stay current on microbiome studies. 

Supplement and probiotic manufacturers, as well as pet food companies that use probiotics as ingredients should be aware of how they work. Key understandings should include how probiotics work, if they colonize, how they impact microbiome test results and potential workarounds.

“We hope that this paper helps pet food companies, veterinarians and others understand the benefits of microbiome testing, and feel confident that they can implement best practices when they incorporate it into their research,”​ said Jarett.

Source: Frontiers in Veterinary Science​

09 April 2021 | https://doi.org/10.3389/fvets.2021.644836​

“Best Practices for Microbiome Study Design in Companion Animal Research”​

Authors: J. Jarett et al.