Summary

A great number of diverse microorganisms inhabit the human body and are collectively referred to as the human microbiome. Until recently, the role of the human microbiome in maintaining human health was not fully appreciated. Today, however, research is beginning to elucidate associations between perturbations in the human microbiome and human disease and the factors that might be responsible for the perturbations. Studies have indicated that the human microbiome could be affected by environmental chemicals or could modulate exposure to environmental chemicals. Given those findings, some fear that we might be missing or mischaracterizing health effects of exposure to environmental chemicals and have therefore argued that chemical–microbiome interactions should be considered in assessing human health risk associated with environmental-chemical exposure. Such considerations would add substantial complexity to an already complex analysis. Given the complexity and resource constraints, the US Environmental Protection Agency (EPA) and the National Institute of Environmental Health Sciences (NIEHS) asked the National Academies of Sciences, Engineering, and Medicine to develop a research strategy to improve our understanding of the interactions between environmental chemicals and the human microbiome and the implications of those interactions for human health risk. They also asked the National Academies to identify barriers to such research and opportunities for collaboration.¹ As a result of the request, the National Academies convened the Committee on Advancing Understanding of the Implications of Environmental-Chemical Interactions with the Human Microbiome, which prepared this report.

Here, the committee highlights key aspects of the human microbiome and its relation to health, describes potential interactions between environmental chemicals and the human microbiome, reviews the risk-assessment framework and reasons for incorporating chemical–microbiome interactions, and outlines its research strategy. The committee emphasizes that this report is not a comprehensive review of all microbiome research. The research strategy presented here focuses on addressing questions about the interactions of environmental chemicals with the human microbiome and the implications for human health risk. It is not a research strategy for directly investigating associations between the human microbiome and various diseases.

THE HUMAN MICROBIOME

The human microbiome is an all-encompassing term that refers to all microorganisms on or in the human body, their genes, and surrounding environmental conditions. Because of the vast diversity and sheer amount of microbial life that colonizes the human body, human beings are now regarded as ecosystems that are comprised of distinct ecologic niches or habitats, each housing a discrete collection of coevolved microorganisms that interact extensively with each other and with the human host. Coevolution has led to interdependence: the human microbiome contributes a vast array of essential functions to the human host and influences

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¹ The full statement of task is in Chapter 1 of this report.

a variety of physiologic, immunologic, and metabolic processes. Perturbations of the composition and function of niche-specific microbial communities have been implicated in an array of neurologic, gastrointestinal, metabolic, oncologic, hepatic, cardiovascular, psychologic, respiratory, and autoimmune disorders or diseases.

One key aspect of the human microbiome is the variation in its composition and function observed among populations, over the human life span, and between body sites. The variation between body sites is particularly noteworthy. Each body site is associated with the presence of a relatively conserved microbial community (a microbiome) that has adapted to the environmental conditions of the site. The site-specific differences in microbial composition yield differences in metabolic capacity and in the aggregate function of the human microbiome. Multiple factors also play roles in the variation observed among individual body sites. For example, age and diet play primary roles in the variation observed in the gut microbiome, and local ecologic conditions, particularly water and nutrient availability, drive the site-specific community states of the skin microbiome. Numerous physiologic and anatomic factors play roles in determining the composition and regional variation in the respiratory microbiome; research suggests that important factors include differences in oxygen tension, airway luminal temperature, mucociliary clearance mechanisms, and other innate defenses. All those factors and others—such as genetics, sex, socioeconomic status, disease state, geography, pregnancy status, diet, and environmental exposures—appear to play roles in shaping the composition and function of microbial communities.

As discussed throughout the present report, animal models provide valuable experimental platforms for studying microbiome structure and function, but it is important to note that the human microbiome differs from the microbiomes of other species in which microorganisms are present, in the relative abundance of dominant microorganisms, and in how the microbial community responds to a given perturbation. The degree to which microbiome composition differs between species (and between humans) depends partially on the taxonomic level at which microbiomes are characterized—whether at the strain, species, genus, family, order, class, or phylum level—and possibly on technical differences among study protocols, which can vary substantially. Although most studies have not compared functional attributes of the microbiomes, such comparison might indicate greater similarity than simply comparing microbial composition. However, given the differences between humans and animals, observations made in animal models, although informative and foundational, might not capture the full breadth of microbial interactions that occur in humans. The strengths and weaknesses of animal models for research into chemical–microbiome interactions are discussed further below.

INTERACTIONS BETWEEEN ENVIRONMENTAL CHEMICALS AND THE HUMAN MICROBIOME

Scientific research is beginning to elucidate the various ways in which environmental chemicals might interact with the human microbiome. Studies suggest that exposure to environmental chemicals can alter the composition and potentially affect the function of the human microbiome. Other studies indicate that the human microbiome can modulate environmental-chemical exposure. For example, evidence of involvement of the gut microbiome in the metabolic transformation of environmental chemicals in broad chemical classes is compelling.

Many molecular mechanisms likely underlie microbiome interactions. However, research suggests that the human microbiome might modulate the exposure–response relationships of environmental chemicals by a few general mechanisms, as described below.

• Direct effect of a chemical on the human microbiome. Distinct microbial compositions can have specific effects on host biology. If exposure to an environmental chemical (or any other factor) causes a perturbation in the microbiome, that perturbation might have distinct effects on the host. It is also conceivable that changes induced by environmental-chemical exposures can result in an altered capacity of the microbiome to metabolize chemicals.

• Altered epithelial-barrier functions. Epithelial barriers form the interface between many host tissues and the external environment. Increasing evidence suggests that there are intimate bidirectional interactions between the microbiota and epithelial cells, wherein the composition and activity of the gut microbiota, for example, modulates the structure and function of the intestinal epithelium and vice versa. The ability to regulate epithelial permeability and integrity has important implications for the absorption, transport, and excretion of environmental chemicals.
• Direct chemical transformation. As noted, the gut microbiome has been shown to metabolize broad classes of environmental chemicals. Microbial metabolic transformations have been generally categorized into reduction and hydrolysis reactions and have been classified further into five major enzymatic families—azoreductases, nitroreductases, β-glucuronidases, sulfatases, and β-lyases.
• Transformation of host-generated metabolites. In some cases, detoxification and elimination of environmental chemicals by host liver enzymes might be reversed by microbial hydrolases in the gut. For example, deconjugation reactions by gut β-glucuronidases promote reabsorption of some drug metabolites, which potentially alters their pharmacokinetic profiles, toxicity, or efficacy. Because a wide array of environmental chemicals might be subject to elimination via β-glucuronidation, this mechanism might be more common than is now appreciated.
• Altered expression of host-tissue metabolic enzymes and pathways. Recent studies have demonstrated that the gut microbiota can regulate host genes involved in chemical metabolism, although more research is needed to understand the mechanisms by which the gut microbiome and its products interact with host nuclear receptors and whether similar processes can alter expression of other types of host-gene pathways that are involved in toxicity.

Although research has provided important clues regarding microbial transformation of environmental chemicals and vice versa, there are substantial gaps in the understanding of how chemical exposure changes activity or function of a microbiome and the breadth of potential pathways for metabolism of environmental chemicals represented in a given microbiome. Furthermore, it is important to note that each interaction can conceptually increase or decrease chemical exposure, and that the role of the interactions in modifying human susceptibility to toxicity at environmentally relevant exposures remains largely uncertain.

RISK ASSESSMENT: INCORPORATING CHEMICAL–MICROBIOME INTERACTIONS

Research indicates the important role that the human microbiome plays in human health and raises the question of whether some consideration needs to be incorporated into risk assessment. Risk assessment is a process that can be used to estimate the human health risk associated with exposure to an environmental chemical. Although risk assessment used in regulatory programs in the United States and globally has been reformed and advanced over the years, the core elements established in the 1980s—hazard identification, dose–response assessment, exposure assessment, and risk characterization—have remained the same (see Figure S-1). EPA has developed numerous guidelines for the conduct of risk assessment; the guidelines describe the optimal evaluation and use of data that often are inconsistent, and they indicate proper treatment of uncertainty in extrapolating results from animal or human studies of limited scope to policies designed to protect the general public.

Animal toxicology studies have traditionally provided the data for hazard identification and dose–response assessment, but epidemiology (human) studies have provided the primary evidence on a few chemicals, such as arsenic and formaldehyde. In vitro assays and computational approaches are also being developed in light of scientific and technologic advances in biology and related fields and substantial increases in computational power. The hope is that the new approaches can predict toxicity on the basis of an understanding of the biologic processes that lead to adverse effects. Exposure science has also undergone remarkable advances in the last few decades; technologies for

developing rapid and comprehensive exposure profiles, from the use of remote and personal sensors to identification and sampling of key biomarkers, are contributing copious new data for risk assessment. Regardless of the approaches used to provide data for various risk-assessment elements, none has explicitly considered or incorporated the human microbiome. Therefore, risk assessments might mischaracterize the nature of a hazard associated with an exposure or overestimate or underestimate the risk associated with the exposure, particularly when the results from studies in animals or in a specific population are used to characterize risk to another species or population that has a microbiome different from that of the studied population.

Studies on chemical–microbiome interactions and their consequences suggest that further research could substantially advance understanding of human health risk posed by exposure to environmental chemicals. Specifically, research might explain differences between animal toxicology studies and human responses, provide greater confidence in extrapolating findings of animal studies to humans, and identify unrecognized health consequences of environmental exposures. Furthermore, differences in responses to chemical exposure reported in epidemiology studies conducted on different populations might be explained by the population variation in microbiome composition and function. Given the recent research on the human microbiome, it is reasonable to hypothesize that its adequate consideration in risk assessment could improve the understanding of health risks posed by exposures to environmental chemicals.

RESEARCH STRATEGY

Development of a research strategy to understand the interactions between environmental chemicals and the human microbiome and the implications of those interactions for human health risk is a complex task. One reason is that our understanding of how perturbations of the human microbiome might cause or contribute to the development of various diseases is in its infancy, so the task of understanding how environmental chemicals fit into the picture is even more difficult than it might otherwise be. Initially, the committee envisioned a research strategy that was similar to a flowchart or decision tree in which the results of one or more experiments would lead naturally to a next set of experiments. However, such a straightforward approach is not feasible given the state of the science. Thus, the committee determined that the research strategy should focus broadly on the three general topics: the effects of environmental chemicals on

the human microbiome, the role of the human microbiome in modulating environmental-chemical exposure, and the importance of variation in the human microbiome in modulating chemical–microbiome interactions. The discussion below provides the primary goals of the research, identifies some possible barriers, and highlights the need for collaboration. A more detailed discussion of experimental approaches and barriers related to each topic can be found in Chapter 6 of the committee’s report with criteria for selecting chemicals for experimental approaches. It is important to note that the committee is not recommending that all the research described in this report be undertaken at once. Discoveries made in trying to understand the relationships between microbiome perturbations and disease will influence the course of the committee’s proposed research strategy, and various agencies and organizations will have different priorities and interests in pursuing various research topics described here. The committee hopes that the near-term research will help to elucidate whether the microbiome is an important contributor to human health risks associated with exposure to environmental chemicals and the need for and direction of research in this area.

The Effects of Environmental Chemicals on the Human Microbiome

The question for this research to answer is whether environmental-chemical exposures or doses that are in the range of known or anticipated human exposures can induce microbiome alterations that modulate adverse health effects. As noted, recent evidence indicates that exposures to some environmental chemicals can alter the microbiome, but there is little evidence that the alterations have adverse effects on health status. To address the question posed, the research program should focus on defining toxicity end points for the microbiome, on identifying environmental chemicals that can perturb the microbiome structurally and functionally, and on using animal and epidemiology studies to demonstrate that microbiome perturbations by environmental chemicals cause or modulate a change in health. Although individual microbial physiology can be detailed robustly, no end points for microbiome toxicity have been established. Thus, defining quantifiable end points that reflect toxicity to the microbiome are of paramount importance, and comprehensive approaches will be needed to capture all aspects of microbiome responses to a given toxicant. Establishing toxicity end points for the microbiome will enable the development of high-throughput bioreactors that can screen environmental chemicals in a uniform manner for their ability to perturb microbiomes. Once chemicals that perturb microbiomes have been identified, they can be investigated in animal models and in epidemiology studies.

Epidemiology studies constitute a considerable undertaking, so it is important to note that existing epidemiology and population studies could be leveraged for this research. For example, one could identify a human population in which a chemical exposure of interest has been tracked and collect new samples appropriate for microbiome analyses, one could generate new microbiome-relevant data from stored samples from such a cohort, or one could add measurements of environmental-chemical exposures to a human population that is being followed for other purposes, including microbiome measurements. Simple measures of microbiome structure might be sufficient to identify cases in which a perturbation occurs in tandem with or after chemical exposure and manifestation of adverse health outcomes; the microbiome changes would then need to be investigated in more detail to characterize their functional or clinical consequences, if any. In such cases, it will also be crucial to separate health effects mediated by microbial activity from those induced directly by chemical exposures of the host.

The Role of the Human Microbiome in Modulating Environmental-Chemical Exposure

The question for this research to answer is, What is the role of the human microbiome in modulating absorption, distribution, metabolism (activation or inactivation), and elimination (ADME) of environmental chemicals? The research pro-

gram would focus on generating pharmacokinetic–pharmacodynamic data from animal and in vitro experiments. The animal experiments would assess the effects of the microbiome on ADME processes in vivo and the magnitude of the effects. The in vitro experiments would be used to define functional traits for a microbial community that transforms an environmental chemical, to identify microorganisms and microbial interactions implicated in chemical transformations, to identify microorganism-modified metabolites, and to obtain microorganism-specific chemical transformation rates. The data generated from the experiments could be used to develop a microbiome component for physiologically based pharmacokinetic or pharmacodynamic models that would permit better assessment of human responses to chemical exposures.

Another aspect of the research program would be identification of specific microorganisms and their enzymes that mediate chemical transformation processes by using new chemical probes and chemical screening technologies. Ultimately, linking the specific microorganisms, genes, and enzymes to particular chemical transformation processes is essential if substantive progress is to be made in addressing individual susceptibility and interspecies extrapolation at a mechanistic level and in understanding the degree of functional redundancy that exists within a microbiome.

The Importance of Microbiome Variation

Two aspects of microbiome variation need to be investigated. The first is the microbiome variation in the human population; the question is whether knowledge of population variation in the human microbiome improves understanding of individual health risks and susceptibility to effects of environmental chemicals. The research goals are to understand the importance of human microbiome variation at any given life stage or among specific populations and ultimately to ensure that studies consider such variation adequately and appropriately when assessing the human health risks posed by exposure to environmental chemicals. Variation will be best understood by conducting comparative studies that assess functional similarities and differences of the factors known or hypothesized to affect microbiome diversity. The studies should emphasize populations that represent key windows of potential vulnerability—such as pregnant women, infants, adolescents, and geriatric populations—and resilience, such as healthy adults. As discussed above, existing epidemiology and population studies could be leveraged for this research to obtain results in the near term.

The second aspect of variation that needs to be explored is that between species. One question is whether the differences are so great that effects are being missed or mischaracterized by using animal models to predict human health risk associated with environmental-chemical exposure. Another question is whether the interspecies uncertainty factors that are used to extrapolate effects in animals to humans account adequately for the microbiome variation. The research program would focus on comparative studies that ultimately could reveal the functional capacity encoded by the human microbiome so that animal species and study designs that are most appropriate for extrapolating to humans could be identified. Specifically, near-term research could focus on identifying functional pathways that are uniquely encoded by microbiomes of select model organisms and humans, on understanding differences and similarities between model-organism and human-host responses to environmental-chemical exposures, and on assessing the redundancy in the microbiomes of various model organisms and humans.

Barriers to Research

To accomplish the research described in the committee’s report, tools will need to be developed, and barriers will need to be overcome. Some barriers are specific to the research described, and others are broadly applicable. A few overarching barriers are highlighted below (further details are provided in Chapter 6 of this report).

• Resources. Many experiments that the committee describes are likely to require substantial investments of time and resources, are exploratory and thus unlikely to be supported through

• traditional funding mechanisms, and require multidisciplinary expertise not found within a single laboratory.

• In vitro model systems. Despite advances, in vitro model systems that faithfully model, for example, the gut environment have not yet been developed. Current in vitro model systems are unable to incorporate microbial communities that represent naturally occurring microbiomes fully, and researchers do not yet understand how various factors change microbiome gene expression and metabolism and which factors need to be recapitulated in an in vitro system. Furthermore, in vitro systems are not yet able to capture fully all the functional diversity of a microbiome and its interactions with its host.
• Standardization. Lack of standardization in experimental approaches results in an inability to reproduce findings related to chemical–microbiome interactions. Investigators need to control and disclose variables relevant to microbiome assessments, including animal-care procedures and conditions, choices in laboratory reagents, and methods for processing samples and measuring outcomes.
• Microbial reference communities. There is no consensus regarding reference strains or microbial communities. Past initiatives have provided data on the composition of microbial communities from healthy adults, but additional microbial reference communities and standardized microbial populations that faithfully recapitulate the variation present in the human microbiome are needed; their development and use will allow comparison of study results among institutions and increase reproducibility of results.
• Reference information. The vastness and complexity of the microbiome has resulted in genomic databases that contain scores of unannotated genes about which scientists know almost nothing. Similarly, much in metabolomics databases remains to be annotated and identified, including chemical structure, metabolite source (human vs microbe), and metabolic pathway. Genomic, transcriptomic, and metabolic databases and libraries will need to expand their coverage of relevant strains, genes, enzymes, metabolite identities and function, and associated characteristics of microbiome sources to enable understanding of microbiome dynamics. Large-scale data generation and data-integration efforts will be required to develop computational models that can predict chemical–microbiome interactions and their consequences.

Collaboration

In the United States, several agencies play roles in assessing health risks associated with exposures to environmental pollutants. Similarly, microbiome-related research is being conducted by several agencies and sectors. Progress in fields related to risk assessment and in microbiome research has occurred largely independently, and the segregation of such research programs poses a major barrier to advancing knowledge on interactions between environmental chemicals and the human microbiome and the implications of the interactions for human health risk. Funding mechanisms that promote interdisciplinary research and specifically encourage collaboration are vital for implementing the research strategy detailed in the committee’s report.

To support such efforts effectively, agencies and research entities that conduct microbiome and human-health research are encouraged to develop collaborations with their counterparts in risk-assessment fields and vice versa. For example, collaborations between the National Institutes of Health and EPA or state agencies that have a long history of assessing the health risks posed by environmental-chemical exposures are encouraged. That type of interdisciplinary collaboration should be sought out, encouraged, and supported to make the best use of available knowledge and resources in each agency or organization. Likewise, initiatives similar to the Center for Children’s Health, the Environment, the Microbiome and Metabolomics at Emory University, jointly funded by EPA and NIEHS, should be considered as vehicles for stimulating and fostering the types of interdisciplinary research needed. The participation of experts in diverse research disciplines during the entire research cycle—planning and designing studies, conducting the experiments, and analyz-

ing the data—is likely to result in studies that are well suited to address the research recommended by the committee. Such interdisciplinary initiatives could also serve as an ideal training environment for the next generation of researchers whose expertise spans several fields.

CONCLUDING REMARKS

Implementation of the committee’s proposed research strategy should substantially advance understanding of whether and to what extent the human microbiome affects the nature and magnitude of adverse health effects caused by exposures to environmental chemicals. In the relatively near term (2–4 years), results of the proposed research should allow judgments to be made about whether explicit consideration of microbiome interactions in the study of environmental-chemical toxicity yields information that is not available from traditional studies (ones that do not explicitly consider microbiomes). Within a similar time frame, it should also be possible to determine whether new information is gained by studying the effects of chemicals on the human microbiome, the role of the human microbiome in modulating chemical exposures, or both. The research should lead to the type of information needed to assess the importance of the human microbiome as a contributor to human health risks associated with exposures to environmental chemicals and thus permit informed decisions about the need for and nature of continuing research in this field.