The human microbiome has captured public interest over the last few decades. Naturally, scientists want to understand the microbiome within their own context of interest, leading to a menagerie of microbiome sub-fields focused on everything from inflammatory bowel disease to autism. Each has unique data on healthy vs. diseased states.

And yet, little is known about what exactly a 'healthy' microbiome is, and whether it can be generalized beyond broad strokes such as bacterial diversity or capacity for fiber fermentation. Moreover, microbiome research knows little standardization for specimen collection, processing and analysis thus far, making data sets even within the same field difficult to compare. Not to mention, many studies have been conducted within industrialized populations, often overrepresenting people of European descent, making it impossible to say with certainty that the data capture a true spectrum of health. These ideas are elegantly explored in 4 scientific presentations at World Microbe Forum 2021.

Global Industrialization and the Human Microbiome

The Global Microbiome Conservancy, founded by Mathilde Poyet, Ph.D., and Mathieu Groussin, Ph.D., of the Massachusetts Institute of Technology, aims to biobank a truly representative global sample of human microbiomes. In doing so, they hope to not only preserve biodiversity that is quickly diminishing in the industrialized world, but to protect microbiomes of indigenous groups whose lifestyles are under threat, and broaden microbiome research to diminish health inequities for underrepresented groups.

Poyet presented data about the microbial degradation of cholesterol in the gut. Normally, cholesterol is cycled in various forms through the liver, gut, bloodstream and back. However, cholesterol can be degraded microbially into another form called coprostanol, which is no longer reabsorbed and gets excreted with feces. In theory, microbial degradation in this manner could protect against high cholesterol levels in the bloodstream and their associated cardiovascular risk.

Poyet and her team found that the amount of coprostanol secreted in feces was inversely proportional to the level of industrialization of the subject. From hunter-gatherers to pastoralists to fully industrialized populations, the trend was consistent. The team then identified a candidate microbe, closely related to the non-human isolate Eubacterium coprostanoligenes, that expresses the IsmA cholesterol dehydrogenase and confirmed that it alone could convert cholesterol into coprostanol in vitro. From her vast population sampling, Poyet determined that this bacterial species was most commonly found among populations living in more ancestral ways, and less abundant in industrialized populations. Why the industrial lifestyle doesn't support this important organism's survival remains unknown. Poyet's findings underscore the importance of gathering representative global data, a mission echoed by the Global Microbiome Conservancy. A more conventional study that recruited people only from industrialized cultures or uniform populations might have missed this important story altogether.

Dietary Fiber and the Mucus Degradation Niche in the Gut

The mucus layer of the gut is an important glycoprotein scaffold constructed of interlocking mucin glycans, running along the surface of the intestine. This mucus layer exists in equilibrium, with constant growth and sloughing. It provides a niche for intestinal microbes while keeping them at a safe distance from the underlying intestinal epithelium. However, in times of stress, including acute lack of fiber in the diet, microbes can turn to the mucus layer for sustenance and eat away at this protective barrier. An acute lack of fiber can also lead to a loss of mucus-secreting goblet cells, which are responsible for creating the mucus layer, and a dramatic decrease in bacterial diversity within the colon, as not enough nutrients remain to feed the bacteria present.

David Berry, Ph.D., of the University of Vienna explored mucin degradation using stable isotope pulse-chase experiments in mice to understand which microbes are capable of consuming the mucus layer. Berry and his group measured labeled threonine as it was taken up by goblet cells of the gut, secreted as mucins and consumed by microbes. Combining fluorescence in-situ hybridization (FISH) and isotope imaging, they identified 2 primary species, Akkermansia muciniphila and Bacteroides acidifaciens, responsible for degrading most of the mucus, as well as a small collection of other microbes capable of doing so.

Using a range of techniques, including the use of deuterium (heavy water) to measure microbe metabolic activity, he and his group identified numerous other bacteria capable of degrading sugars found in mucin glycans. They determined that mucin degraders hail from across the phylogenetic tree, but seem to be especially enriched among the Bacteroidetes phylum, specifically, the Muribacilaceae family. In the absence of their preferred energy sources (such as those derived from fiber), these species attack proteins within the mucus layer. By identifying the sugar degradation abilities of different microbial families, Berry's group also identified microbes that could directly compete with Clostridioides difficile (C. diff) by filling its preferred niche for sialic acid metabolism.

Berry's work illuminates the utility of identifying the metabolic niches of commensal organisms and the potential to develop useful bacteriotherapies to combat opportunistic or antibiotic-induced pathogens. It also highlights how lifestyle factors such as diet can lead to pathogenic shifts in the microbiota by impacting their metabolic function in the absence of a preferred nutrient source. Notably, an acute lack of fiber is more prevalent among industrialized populations, which are more likely to consume low-fiber, processed foods.

Common Signatures of Health and Disease in the Gut Microbiome

In human microbiome research, 'dysbiosis' refers to microbial communities that are out of balance but has become something of a catch-all term for the microbiome in a state of unhealth. What does dysbiosis actually mean in different contexts? Saad Khan of the Albert Einstein College of Medicine noted that microbiome studies are plagued by inconsistencies. Further still, meta analyses can help identify microbes associated with a particular disease process, but the same microbe may be affiliated with multiple conditions, making clinical predictions imprecise. Without consistent and specific disease biomarkers, collecting clinically useful data about the microbiome is challenging.

To better understand what a 'healthy' microbiome means, Khan started by asking, What is health? Rather than just seeking disease markers for a particular condition, he trained a graph convolutional neural network (a type of artificial intelligence) to seek markers for 17 different conditions, including that of healthy controls across studies. While developing disease-specific markers for multiple conditions, Khan recognized that he had the most data for the 'healthy' group, providing powerful associations for what a microbiome looks like in the presumed absence of disease. Khan and his colleagues found a core set of microbial taxa that consistently associate with health across 21 datasets that they evaluated.

Where so many studies ask, "What is disease?," flipping the question provided a rich dataset with useful clinical markers for the microbiome. Several trends emerged that spoke to what 'dysbiosis' could mean in a more universal sense. For example, a higher abundance of oral microbes in the gut was a consistent, non-specific indicator of disease. Conversely, Khan found that the more abundant an organism was within the gut, the more likely it was to be a consistent marker of health. Moving forward, he hopes to identify core microbial genes and pathways associated with health, as well as understand the underpinnings of what drives dysbiosis.

Co-Diversification of Gut Microbes and Their Human Hosts

For a microbiome to be healthy, is the locality of its host important? Some have previously published evidence that specific microbial organisms such as Helicobacter pylori reflect human migration patterns, but is the overall microbial community 'healthiest' in the geographic context of their host? These are the questions of Ruth Ley, Ph.D., and her postdoc, Taichi Suzuki, Ph.D.

By seeking patterns of co-phylogeny between humans and specific microbe strains, Ley and her team investigated whether there is sufficient evidence to suggest that the human microbiome has evolved alongside its human hosts and their environment. Ley generated gut metagenomes from mothers and their children in 3 global regions: Europe (Germany and the United Kingdom), Gabon and Vietnam. By matching the host genetic phylogeny to the phylogenies of different strains of bacteria, Ley identified some species that showed clear signs of co-diversification and others that appeared more varied. The top 7 taxa demonstrated strong evidence for co-phylogeny with their human hosts. Applying these findings to public metagenomic datasets, Ley recapitulated a co-diversification phenotype. The data provide an intriguing story of human microbial heritage, and raise questions about vertical transmissibility, as well as the importance of the microbiome in adapting to the local environment.

Altogether, the session "What Makes a Microbiome 'Healthy'?" at World Microbe Forum provided a rich context to challenge some of the assumptions within the microbiome field for how to frame health and disease. Researchers highlighted a deep need for more diverse microbiome sampling, thinking about local context and being critical of hazy or nebulous definitions within the field. Moreover, the studies provided unique and creative tools for studying microbes, some of which show promise as potential microbial therapies of the future.