Microbes in the gut can dramatically boost the effectiveness of a common anticancer drug, at least in mice. That’s the finding of Laurence Zitvogel of Gustave Roussy Cancer Campus in Vellejuif, France, and colleagues, who looked at the anticancer antibiotic cyclophosphamide in treated mice. In those mice lacking a protein that restricts the growth of two spices of gut bacteria (Enterococcus hirae and Barnesiella intestinihominis), the drug was almost twice as effective at reducing tumour size. The work suggests that gut bacteria could be used to optimise cancer therapies.

 

Recent research shows that gut bacteria affect cancer - some promote tumor growth while others hinder it. However, it has not been clear which species of gut microbes help and which do not. Now, a new study identifies two species of gut bacteria that boost the effect of a common chemotherapy drug by activating immune cells.

The researchers found two species of gut bacteria that boost the antitumor effect of chemotherapy by activating a type of immune cell that attacks cancer.

The researchers, including co-senior author Dr. Mathias Chamaillard, research director of the Center for Infection and Immunity of Lille (Inserm/CNRS/Lille University/Institut Pasteur de Lille) in France, report their findings in the journal Immunity.
The study concerns the complex interplay in the human body of three things in fighting cancer: chemotherapy, the immune system, and gut bacteria.
Chemotherapy is a type of cancer treatment that uses drugs. It works by stopping or slowing the growth of cancer cells, which grow and divide rapidly.
Chemotherapy can be used to cure cancer, reduce the chances it will return, or stop or slow its growth. It can also be used to shrink tumors that are causing pain and other problems.
The immune system also has mechanisms for fighting cancer. For instance, it has T cells that hunt down and kill cancer cells.
As technologies in microbiology and molecular biology advance, scientists are finding that the trillions of microbes that live in and on our bodies play an important role in health and disease.
In the intestines, for example, gut bacteria not only help to digest food, their byproducts (metabolites) also improve immune function and strengthen the lining of the gut to better defend against infection.

Two gut bacteria boost the effect of cyclophosphamide

In the new study, Dr. Chamaillard and colleagues show that two species of gut bacteria - Enterococcus hirae and Barnesiella intestinihominis - boost the effect of the commonly prescribed immunosuppressive chemotherapy drug cyclophosphamide by activating T cells.

Fast facts about chemotherapy

• Chemotherapy often accompanies other cancer treatments
• For example, it may be used to make a tumor smaller before surgery or radiation therapy
• The most common side effect is fatigue.

Learn more about chemotherapy

Moreover, they showed that the immune response boosted by these bacteria predicted longer progression-free survival in patients with advanced lung and ovarian cancers who were treated with chemo-immunotherapy.
Dr. Chamaillard says the finding shows it may be possible to improve the design of anticancer treatments, by "either optimizing the use of antibiotics, or implementing supplementation with so-called oncomicrobiotics or their bioactive metabolites, to promote the efficacy of anticancer drugs."
As a first step, the team used mouse models to investigate the effects of the two bacteria species separately with chemotherapy.

They found oral treatment with E. hirae activated antitumor T cell responses in the spleen, which, alongside toxic effects of the chemotherapy drug cisplatin, curbed tumor growth.
They also found similar results from oral treatment with B. intestinihominis - except in that case, the bacteria spurred T cells to infiltrate tumors in various mouse models.

Predicted progression-free survival

Moving on from mouse models, the team then analyzed blood T cell responses from 38 patients with advanced lung and ovarian cancer who were treated with chemo-immunotherapy.
The results showed memory T cell responses specific to E. hirae and B. intestinihominis predicted progression-free survival - that is, the amount of time after treatment that patients live with the disease without it getting worse.
The researchers are now planning further studies to find out which specific bacterial metabolites or immune-modulating molecules are responsible for boosting the effect of cyclophosphamide.

"Answering this question may provide a way to improve the survival of cyclophosphamide-treated cancer patients by supplementing them with bacterial-derived drugs instead of live microorganisms."

Dr. Mathias Chamaillard

Learn about the unusual way ultrasmall nanoparticles kill cancer cells.

Written by Catharine Paddock PhD

 Gut flora (gut microbiota, or gastrointestinal microbiota) is the complex community of microorganisms that live in the digestive tracts of humans and other animals, including insects. The gut metagenome is the aggregate of all the genomes of gut microbiota.^[1] The gut is one niche that human microbiota inhabit.^[2]
In humans, the gut microbiota has the largest numbers of bacteria and the greatest number of species compared to other areas of the body.^[3] In humans the gut flora is established at one to two years after birth, and by that time the intestinal epithelium and the intestinal mucosal barrier that it secretes have co-developed in a way that is tolerant to, and even supportive of, the gut flora and that also provides a barrier to pathogenic organisms.^[4][5]
The relationship between some gut flora and humans is not merely commensal (a non-harmful coexistence), but rather a mutualistic relationship.^[2]:700 Some human gut microorganisms benefit the host by fermenting dietary fiber into short-chain fatty acids (SCFAs), such as acetic acid and butyric acid, which are then absorbed by the host.^[3][6] Intestinal bacteria also play a role in synthesizing vitamin B and vitamin K as well as metabolizing bile acids, sterols, and xenobiotics.^[2][6] The systemic importance of the SCFAs and other compounds they produce are like hormones and the gut flora itself appears to function like an endocrine organ,^[6] and dysregulation of the gut flora has been correlated with a host of inflammatory and autoimmune conditions.^[3][7]
The composition of human gut flora changes over time, when the diet changes, and as overall health changes.^[3][7] A systematic review from 2016 examined the preclinical and small human trials that have been conducted with certain commercially available strains of probiotic bacteria and identified those that had the most potential to be useful for certain central nervous system disorders.^[8]

Due to the high acidity of the stomach, most microorganisms cannot survive. The main bacterial inhabitants of the stomach include: Streptococcus, Staphylococcus, Lactobacillus, Peptostreptococcus, and types of yeast.^[2]:720 Helicobacter pylori is a Gram-negative spiral organism that establishes on gastric mucosa causing chronic gastritis and peptic ulcer disease and is a carcinogen for gastric cancer.^[2]:904

Intestinal flora

Bacteria commonly found in the human colon[24]
Bacterium	Incidence (%)
Bacteroides fragilis	100
Bacteroides melaninogenicus	100
Bacteroides oralis	100
Enterococcus faecalis	100
Escherichia coli	100
Enterobacter sp.	40–80
Klebsiella sp.	40–80
Bifidobacterium bifidum	30–70
Staphylococcus aureus	30–50
Lactobacillus	20–60
Clostridium perfringens	25–35
Proteus mirabilis	5–55
Clostridium tetani	1–35
Clostridium septicum	5–25
Pseudomonas aeruginosa	3–11
Salmonella enteritidis	3–7
Faecalibacterium prausnitzii	?common
Peptostreptococcus sp.	?common
Peptococcus sp.	?common

The small intestine contains a trace amount of microorganisms due to the proximity and influence of the stomach. Gram positive cocci and rod shaped bacteria are the predominant microorganisms found in the small intestine.^[2] However, in the distal portion of the small intestine alkaline conditions support gram-positive bacteria of the Enterobacteriaceae.^[2] The bacterial flora of the small intestine aid in a wide range of intestinal functions. The bacterial flora provide regulatory signals that enable the development and utility of the gut. Overgrowth of bacteria in the small intestine can lead to intestinal failure.^[25] In addition the large intestine contains the largest bacterial ecosystem in the human body.^[2] Factors that disrupt the microorganism population of the large intestine include antibiotics, stress, and parasites.^[2]
Bacteria make up most of the flora in the colon^[26] and 60% of the dry mass of feces.^[9] This fact makes feces an ideal source to test for gut flora for any tests and experiments by extracting the nucleic acid from fecal specimens, and bacterial 16S rRNA gene sequences are generated with bacterial primers. This form of testing is also often preferable to more invasive techniques, such as biopsies. Somewhere between 300^[9] and 1000 different species live in the gut,^[10] with most estimates at about 500.^[27][28] However, it is probable that 99% of the bacteria come from about 30 or 40 species, with Faecalibacterium prausnitzii being the most common species in healthy adults.^[11][29] Fungi and protozoa also make up a part of the gut flora, but little is known about their activities. The virome is mostly bacteriophages.^[30]
Research suggests that the relationship between gut flora and humans is not merely commensal (a non-harmful coexistence), but rather is a mutualistic, symbiotic relationship.^[10] Though people can (barely) survive with no gut flora,^[27] the microorganisms perform a host of useful functions, such as fermenting unused energy substrates, training the immune system via end products of metabolism like propionate and acetate, preventing growth of harmful species, regulating the development of the gut, producing vitamins for the host (such as biotin and vitamin K), and producing hormones to direct the host to store fats.^[2]:713ff Extensive modification and imbalances of the gut microbiota and its microbiome or gene collection are associated with obesity.^[31] However, in certain conditions, some species are thought to be capable of causing disease by causing infection or increasing cancer risk for the host.^[9][26]

Age

It has been demonstrated that there are common patterns of microbiome composition evolution during life.^[32] In general, the diversity of microbiota composition of fecal samples is significantly higher in adults than in children, although interpersonal differences are higher in children than in adults.^[33] Much of the maturation of microbiota into an adult-like configuration happens during the three first years of life.^[33]
As the microbiome composition changes, so does the composition of bacterial proteins produced in the gut. In adult microbiomes, a high prevalence of enzymes involved in fermentation, methanogenesis and the metabolism of arginine, glutamate, aspartate and lysine have been found. In contrast, in infant microbiomes the dominant enzymes are involved in cysteine metabolism and fermentation pathways.^[33]

Diet

Studies and statistical analyses have identified the different bacterial genera in gut microbiota and their associations with nutrient intake. Gut microflora is mainly composed of three enterotypes: Prevotella, Bacteroides, and Ruminococcus. There is an association between the concentration of each microbial community and diet. For example, Prevotella is related to carbohydrates and simple sugars, while Bacteroides is associated with proteins, amino acids, and saturated fats. One enterotype will dominate depending on the diet. Altering the diet will result in a corresponding change in the numbers of species.^[22]
Malnourished human children have less mature and less diverse gut microbiota than healthy children, and changes in the microbiome associated with nutrient scarcity can in turn be a pathophysiological cause of malnutrition.^[34][35] Malnourished children also typically have more potentially pathogenic gut flora, and more yeast in their mouths and throats.^[36]

Geography

Gut microbiome composition depends on the geographic origin of populations. Variations in trade off of Prevotella, the representation of the urease gene, and the representation of genes encoding glutamate synthase/degradation or other enzymes involved in amino acids degradation or vitamin biosynthesis show significant differences between populations from USA, Malawi or Amerindian origin.^[33]
The US population has a high representation of enzymes encoding the degradation of glutamine and enzymes involved in vitamin and lipoic acid biosynthesis; whereas Malawi and Amerindian populations have a high representation of enzymes encoding glutamate synthase and they also have an overrepresentation of α-amylase in their microbiomes. As the US population has a diet richer in fats than Amerindian or Malawian populations which have a corn-rich diet, the diet is probably a main determinant of gut bacterial composition.^[33]
Further studies have indicated a large difference in the composition of microbiota between European and rural African children. The fecal bacteria of children from Florence were compared to that of children from the small rural village of Boulpon in Burkina Faso. The diet of a typical child living in this village is largely lacking in fats and animal proteins and rich in polysaccharides and plant proteins. The fecal bacteria of European children was dominated by Firmicutes and showed a marked reduction in biodiversity, while the fecal bacteria of the Boulpon children was dominated by Bacteroidetes. The increased biodiversity and different composition of gut flora in African populations may aid in the digestion of normally indigestible plant polysaccharides and also may result in a reduced incidence of non-infectious colonic diseases.^[37]
On a smaller scale, it has been shown that sharing numerous common environmental exposures in a family is a strong determinant of individual microbiome composition. This effect has no genetic influence and it is consistently observed in culturally different populations.^[33]

A new era in medical science has dawned with the realization of the critical role of the “forgotten organ,” the gut micro-biota, in health and disease. Central to this beneficial interaction between the microbiota and host is the manner in which bacteria and most likely other microorganisms contained within the gut communicate with the host’s immune system and participate in a variety of metabolic processes of mutual benefit to the host and the microbe. The advent of high-throughput methodologies and the elaboration of sophisticated analytic systems have facilitated the detailed description of the composition of the microbial constituents of the human gut, as never before, and are now enabling comparisons to be made between health and various disease states. Although the latter approach is still in its infancy, some important insights have already been gained about how the microbiota might influence a number of disease processes both within and distant from the gut. These discoveries also lay the groundwork for the development of therapeutic strategies that might modify the microbiota (eg, through the use of probiot-ics). Although this area holds much promise, more high-quality trials of probiotics, prebiotics, and other microbiota-modifying approaches in digestive disorders are needed, as well as laboratory investigations of their mechanisms of action.

Keywords: Gut flora, microbiota, probiotic, gut bacteria, microbial metabolism, mucosal immunology

Due largely to rapidly evolving advances in analytic techniques in microbiology, molecular biology, and bioinfor-matics, the true diversity of microorganisms that inhabit the gastrointestinal tract of humans (collectively referred to as the human gut microbiota) is being revealed and its contributions to homeostasis in health and to the pathogenesis of disease appreciated (Table 1). As a consequence, the study of gut ecology has emerged as one of the most active and exciting fields in biology and medicine. It is in this context that maneuvers to alter or modify the microbiota, either through dietary modifications or by the administration of antibiotics, probiotics, or prebiotics, must now be viewed.

Table 1

Important Homeostatic Functions of the Gut Microbiota

Go to:

The Normal Gut Microbiota: An Essential Factor in Health

Basic Definitions and Development of the Microbiota

The term microbiota is to be preferred to the older term flora, as the latter fails to account for the many nonbacte-rial elements (eg, archea, viruses, and fungi) that are now known to be normal inhabitants of the gut. Given the relatively greater understanding that currently exists of the role of bacteria, in comparison with the other constituents of the microbiota in health and disease, gut bacteria will be the primary focus of this review. Within the human gastrointestinal microbiota exists a complex ecosystem of approximately 300 to 500 bacterial species, comprising nearly 2 million genes (the microbiome).¹ Indeed, the number of bacteria within the gut is approximately 10 times that of all of the cells in the human body, and the collective bacterial genome is vastly greater than the human genome.

At birth, the entire intestinal tract is sterile; the infant’s gut is first colonized by maternal and environmental bacteria during birth and continues to be populated through feeding and other contacts.² Factors known to influence colonization include gestational age, mode of delivery (vaginal birth vs assisted delivery), diet (breast milk vs formula), level of sanitation, and exposure to antibiotics.^3,4 The intestinal microbiota of newborns is characterized by low diversity and a relative dominance of the phyla Proteobacteria and Actinobacteria; thereafter, the microbiota becomes more diverse with the emergence of the dominance of Firmicutes and Bacteroidetes, which characterizes the adult microbiota.^5–7 By the end of the first year of life, the microbial profile is distinct for each infant; by the age of 2.5 years, the microbiota fully resembles the microbiota of an adult in terms of composition.^8,9 This period of maturation of the microbiota may be critical; there is accumulating evidence from a number of sources that disruption of the microbiota in early infancy may be a critical determinant of disease expression in later life. It follows that interventions directed at the microbiota later in life may, quite literally, be too late and potentially doomed to failure.

Following infancy, the composition of the intestinal microflora remains relatively constant until later life. Although it has been claimed that the composition of each individual’s flora is so distinctive that it could be used as an alternative to fingerprinting, more recently, 3 differ-ent enterotypes have been described in the adult human microbiome.¹⁰ These distinct enterotypes are dominated by Prevotella, Ruminococcus, and Bacteroides, respectively, and their appearance seems to be independent of sex, age, nationality, and body mass index. The microbiota is thought to remain stable until old age when changes are seen, possibly related to alterations in digestive physiology and diet.^11–13 Indeed, Claesson and colleagues were able to identify clear correlations in elderly individuals, not only between the composition of the gut microbiota and diet, but also in relation to health status.¹⁴

Regulation of the Microbiota

Because of the normal motility of the intestine (peristalsis and the migrating motor complex) and the antimicrobial effects of gastric acid, bile, and pancreatic and intestinal secretions, the stomach and proximal small intestine, although certainly not sterile, contain relatively small numbers of bacteria in healthy subjects.¹⁵ Interestingly, commensal organisms with probiotic properties have recently been isolated from the human stomach.¹⁶ The microbiology of the terminal ileum represents a transition zone between the jejunum, containing predominantly aerobic species, and the dense population of anaerobes found in the colon. Bacterial colony counts may be as high as 10⁹ colony-forming units (CFU)/mL in the terminal ileum immediately proximal to the ileocecal valve, with a predominance of gram-negative organisms and anaerobes. On crossing into the colon, the bacterial concentration and variety of the enteric flora change dramatically. Concentrations of 10¹² CFU/mL or greater may be found and are comprised mainly of anaerobes such as Bacteroides, Porphyromonas, Bifidobacterium, Lactobacillus, and Clos-tridium, with anaerobic bacteria outnumbering aerobic bacteria by a factor of 100 to 1000:1. The predominance of anaerobes in the colon reflects the fact that oxygen concentrations in the colon are very low; the flora has simply adapted to survive in this hostile environment.

At any given level of the gut, the composition of the flora also demonstrates variation along its diameter, with certain bacteria tending to be adherent to the mucosal surface, while others predominate in the lumen. It stands to reason that bacterial species residing at the mucosal surface or within the mucus layer are those most likely to participate in interactions with the host immune system, whereas those that populate the lumen may be more relevant to metabolic interactions with food or the products of digestion. It is now evident that different bacterial populations may inhabit these distinct domains. Their relative contributions to health and disease have been explored to a limited extent, though, because of the relative inaccessibility of the juxtamucosal populations in the colon and, especially, in the small intestine. However, most studies of the human gut microbiota have been based on analyses of fecal samples, therefore representing a major limitation. Indeed, a number of studies have already shown differ-ences between luminal (fecal) and juxtamucosal populations in disorders such as inflammatory bowel disease (IBD) and irritable bowel syndrome (IBS).^17,18

In humans, the composition of the flora is influenced not only by age but also by diet and socioeconomic conditions. In a recent study of elderly individuals, the interaction of diet and age was demonstrated, firstly, by a close relationship between diet and microbiota composition in the subjects and, secondly, by interactions between diet, the microbiota, and health status.¹⁴ It must also be remembered that nondigestible or undigested components of the diet may contribute substantially to bacterial metabolism; for example, much of the increase in stool volume resulting from the ingestion of dietary fiber is based on an augmentation of bacterial mass. The subtleties of interaction between other components of diet and the microbiota are now being explored and will, undoubtedly, yield important information. For example, data indicating a potential role of certain products of bacterial metabolism in colon carcinogenesis have already provided strong hints of the relevance of diet-microbiota interactions to disease. Antibiotics, whether prescribed or in the food chain as a result of their administration to animals, have the potential to profoundly impact the microbiota.¹⁹ In the past, it was thought that these effects were relatively transient, with complete recovery of the microbiota occurring very soon after the course of antibiotic therapy was complete. However, while recent studies have confirmed that recovery is fairly rapid for many species, some species and strains show more sustained effects.²⁰

Host-Microbiota Interactions

Gut-commensal microbiota interactions play a fundamental role in promoting homeostatic functions such as immunomodulation, upregulation of cytoprotec-tive genes, prevention and regulation of apoptosis, and maintenance of barrier function.²¹ The critical role of the microbiota on the development of gut function is amply demonstrated by the fate of the germ-free animal.^22,23 Not only are virtually all components of the gut-associated and systemic immune systems affected in these animals, but the development of the epithelium, vasculature, neu-romuscular apparatus, and gut endocrine system also is impaired. The subtleties of the interactions between the microbiota and the host are exemplified by studies that demonstrate the ability of a polysaccharide elaborated by the bacterium Bacteroides fragilis to correct T-cell deficien-cies and Th1/Th2 imbalances and direct the development of lymphoid organs in the germ-free animal.²⁴ Intestinal dendritic cells appear to play a central role in these critical immunologic interactions.^24,25

How does the gut immune system differentiate between friend and foe when it comes to the bacteria it encounters?²⁶ At the epithelial level, for example, a number of factors may allow the epithelium to tolerate commensal (and thus probiotic) organisms. These include the masking or modification of microbial-associated molecular patterns that are usually recognized by pattern recognition receptors, such as Toll-like receptors,²⁷ and the inhibition of the NFκB inflammatory pathway.²⁸ Responses to commensals and pathogens also may be distinctly different within the mucosal and systemic immune systems. For example, commensals such as Bifidobacterium infantis and Faecalibacterium prausnitzii have been shown to differentially induce regulatory T cells and result in the production of the anti-inflammatory cytokine interleukin (IL)-10.²⁹ Other commensals may promote the development of T-helper cells, including T_H17 cells, and result in a controlled inflammatory response that is protective against pathogens in part, at least, through the production of IL-17.³⁰ The induction of a low-grade inflammatory response (physiologic inflammation) by commensals could be seen to prime the host’s immune system to deal more aggressively with the arrival of a pathogen.³¹

Through these and other mechanisms, the microbiota can be seen to play a critical role in protecting the host from colonization by pathogenic species.³² Some intestinal bacteria produce a variety of substances, ranging from relatively nonspecifc fatty acids and peroxides to highly specific bacteriocins,^33,34 which can inhibit or kill other potentially pathogenic bacteria,³⁵ while certain strains produce proteases capable of denaturing bacterial toxins.³⁶

The Microbiota and Metabolism

Although the immunologic interactions between the microbiota and the host have been studied in great detail for some time, it has been only recently that the true extent of the metabolic potential of the microbiota has begun to be grasped. Some of these metabolic functions were well known, such as the ability of bacterial disac-charidases to salvage unabsorbed dietary sugars, such as lactose, and alcohols and convert them into short-chain fatty acids (SCFAs) that are then used as an energy source by the colonic mucosa. SCFAs promote the growth of intestinal epithelial cells and control their proliferation and differentiation. It has also been known for some time that enteric bacteria can produce nutrients and vitamins, such as folate and vitamin K, deconjugate bile salts,³⁷ and metabolize some medications (such as sul-fasalazine) within the intestinal lumen, thereby releasing their active moieties. However, it is only recently that the full metabolic potential of the microbiome has come to be recognized and the potential contributions of the microbiota to the metabolic status of the host in health and in relation to obesity and related disorders have been appreciated. The application of genomics, metabolomics, and transcriptomics can now reveal, in immense detail, the metabolic potential of a given organism.^38–41

It is now also known that certain commensal organisms also produce other chemicals, including neurotrans-mitters and neuromodulators, which can modify other gut functions, such as motility or sensation.^42–44 Most recently and perhaps most surprisingly, it has been proposed that the microbiota can influence the development⁴⁵ and func-tion⁴⁶ of the central nervous system, thereby leading to the concept of the microbiota-gut-brain axis.^47–49

Go to:

The Gut Microbiota in Disease

Just as we are only now beginning to understand the key role of the flora in health, it has only been in very recent years that the true extent of the consequences of disturbances in the flora, or in the interaction between the flora and the host, has been recognized. Some of these consequences are relatively obvious. For example, when many components of the normal flora are eliminated or suppressed by a course of broad-spectrum antibiotics, the stage is set for other organisms that may be pathogenic to step in and cause disease.^1,2,32 The classic example of this is antibiotic-associated diarrhea and its deadliest manifestation, Clostridium difficile colitis. Similar perturbations in the flora are thought to be involved in a devastating form of intestinal inflammation that may occur in newborns and especially premature infants: necrotizing enteroco-litis. In other situations, bacteria may simply be where they should not be. If motility of the bowel is impaired and/or acid secretion from the stomach is drastically reduced, an environment conducive to the proliferation of organisms in the small intestine that are normally con-fined to the colon results; the consequence is the syndrome of small bowel bacterial overgrowth. In other situations, the immunologic interaction between the flora and the host is disturbed, and the host may, for example, begin to recognize the constituents of the normal flora not as friend but as foe and may mount an inappropriate inflammatory response, which, some believe, may ultimately lead to conditions such as IBD.^1,2,32 In other situations, damage to the intestinal epithelium renders the gut wall leaky and permits bacteria (in whole or in part) from the gut to gain access to the submucosal compartments or even to the systemic circulation, with the associated potential to cause catastrophic sepsis. This mechanism is thought to account for many of the infections that occur in critically ill patients in the intensive care unit, for example.

Most recently, qualitative changes in the microbiota have been invoked in the pathogenesis of a global epidemic: obesity.⁴¹ It has been postulated that a shift in the composition of the flora toward a population dominated by bacteria that are more avid extractors of absorbable nutrients—which are then available for assimilation by the host—could play a major role in obesity.⁴¹ Such studies rely on the application of modern technologies (genomics, metagenomics, and metabolomics) to the study of the colonic flora and have the potential to expose the true diversity and metabolic profile of the microbiota and the real extent of changes in disease. Rather than provide an exhausting survey of all the disease states that might be influenced by the microbiota, a brief overview of current information on the role of the microbiota in a few common diseases/disorders will be provided below.

Inflammatory Bowel Disease

There is a considerable body of evidence to support the hypothesis that the endogenous intestinal microflora plays a crucial role in the pathogenesis of IBD and its variants and related disorders.^50,51 Some of this evidence is time-honored, such as the predilection of IBD for areas of high bacterial numbers and the role of contact with the fecal stream in sustaining inflammation. Other evidence is more recent and includes studies described above that illustrate the key roles of the microbiota in host immune responses and the generation of inflammatory responses. This evidence is supplemented by experimental observations on the ability of strategies that modify the microbiota (eg, the administration of probiotics) to modulate the inflamma-tory response in experimental models of IBD.^52–58 Studies of the gut microbiota in IBD have revealed quantitative and qualitative changes,⁵⁹ including the intriguing finding in some studies⁶⁰ that a bacterium with anti-inflammatory properties, F prausnitzii, is less abundant in patients with IBD than in healthy individuals. The importance of microbiota-host interactions in IBD is further supported by the many studies of IBD genetics that have identified a host of changes in genes that code for molecules involved in bacterial recognition, host-bacteria engagement, and the resultant inflammatory cascade.⁶¹ On a more clinical level, the role of the microbiota is supported by the efficacy, albeit variable, of antibiotics in IBD⁶² and the suggestion, not always supported by high-quality clinical trials, that a number of probiotic organisms, including nonpathogenic Escherichia coli, Saccharomyces boulardii, and a Bifidobacte-rium, have efficacy in maintaining remission and in treating mild to moderate flare-ups in ulcerative colitis.^63–70 There are some preliminary data to suggest that fecal transplan-tation,⁷¹ a strategy used with considerable success in the treatment of resistant and recurrent C difficile infection,⁷² may be effective in ulcerative colitis.^73,74

A more convincing clinical illustration of the impact of modulation of the microbiota is provided by the example of pouchitis, an IBD variant that occurs in the neorectum in patients with ulcerative colitis who have undergone a total colectomy and ileo-anal pouch procedure. Here, VSL#3 (Sigma Tau Pharmaceuticals), a probiotic cocktail containing 8 different strains of lactic acid bacteria, has proven to be effective in the primary prevention and maintenance of remission of patients with pouchitis. In one study, remission was maintained in 85% of patients on VSL#3 compared with 6% of patients receiving placebo.⁷⁵

Irritable Bowel Syndrome

A variety of strands of evidence suggest a role for the gut microbiota in IBS⁷⁶ (Table 2). First and foremost among these is the clinical observation that IBS can develop in individuals de novo following exposure to enteric infections and infestations (ie, postinfectious IBS).⁷⁷ More contentious has been the suggestion that patients with IBS may harbor small intestinal bacterial overgrowth (SIBO).⁷⁸ More indirect evidence of a role for the micro-biota can be gleaned from some of the metabolic functions of the components of the microbiota. Thus, given the effects of bile salts on colonic secretion, changes in bile salt deconjugation could lead to changes in stool volume and consistency. Similarly, changes in bacterial fermentation could result in alterations in gas volume and/or composition. Further evidence comes from the clinical impact of therapeutic interventions, such as antibiotics, prebiotics, or probiotics, which can alter or modify the microbiota. Thus, the poorly absorbed antibiotic rifaximin (Xifaxan, Salix) has been shown to alleviate symptoms in diarrhea-predominant IBS,⁷⁹ and some probiotics (B infantis 35624 [Align][Procter & Gamble] in particular⁸⁰) have been shown to exert substantial clinical responses. The latter is of interest, given its demonstrated ability to modulate the systemic immune response in humans.^25,81 Also gaining currency is the suggestion that the colonic microbiota may demonstrate qualitative and/or quantitative changes in IBS.⁸²

Table 2

Evidence for a Role for the Gut Flora in Irritable Bowel Syndrome

Modern molecular microbiologic methods are now being applied to this complex issue and have, indeed, confirmed that patients with IBS, regardless of subtype, do exhibit a fecal flora that is clearly different from that of control subjects.⁸³ Studies by my colleagues and I have demonstrated, firstly, a reduced microbial diversity in IBS⁸⁴ and, secondly, using high-throughput pyrosequencing, the existence of different IBS subgroups based on a detailed examination of the microbiota.⁸⁵ At the phylum level, 1 of these subgroups resembled control subjects, whereas another demonstrated a shift in the relative proportion of the 2 major phyla, Firmicutes and Bacteroidetes, as well as significant changes at species and strain levels.^82,84 The primacy of these microbial shifts and their potential to disturb mucosal or myoneural function in the gut wall, impact the brain-gut axis, or induce local or systemic immune responses remains to be defined (Figure 1). Most intriguing has been the suggestion, from animal studies, that the gut microbiota can influence brain function and morphology.⁴⁹

Figure 1

A schema to summarize the possible role of the microbiota in irritable bowel syndrome. An altered microbiota in concert with a leaky epithelial barrier allows bacteria and/or bacterial products access to the submucosal compartment where mast cells and ...

Several experimental observations provide a sci-entifc basis for the use of therapies that might modify the microbiota in IBS.^85–87 Thus, oral administration of B infantis 35624 has been shown to attenuate interferon γ, tumor necrosis factor α (TNF-α), and IL-6 responses following mitogen stimulation, increase plasma levels of tryptophan and kynurenic acid, and, most strikingly, reduce concentrations of 5-hydroxyindoleacetic acid and dihydroxyphenylacetic acid concentrations in the frontal cortex and amygdala, respectively.⁸⁸

These observations were taken one step further by the same research group by demonstrating normalization of immune responses, reversal of behavioral deficits, and restoration of basal norepinephrine concentrations in the brainstem in an animal model of depression (the maternally separated rat).⁸⁹ While these latter observations could address some of the proposed pathophysiologic mechanisms associated with symptom development in IBS, namely, immune activation and disturbances in the brain-gut axis, other studies suggest that the same strain can also modify peripheral mechanisms linked with IBS, such as visceral hypersensitivity.⁹⁰

Addressing another gut abnormality identified in IBS, Zeng and colleagues partially reversed changes in small intestinal permeability with a probiotic cocktail.⁹¹ Another organism, Lactobacillus acidophilus, has been shown to produce visceral analgesic effects through the induction of μ-opioid and cannabinoid receptors,⁹² and Lactobacillus paracasei has been shown to attenuate gut muscle hypercontractility in an animal model of post-infectious IBS.⁹³ Again, this effect was strain-dependent and appeared to be mediated, in part, through a modulation of the immunologic response to the initial infection and, in part, through the direct effects of the organism, or a metabolite thereof, on gut muscle. In other experiments, this same organism was capable of attenuating antibiotic-induced visceral hypersensitivity in mice.⁹⁴

Lactobacillus reuteri also has been shown to inhibit visceral pain induced by colorectal distension in the rat.⁹⁵ Of clinical relevance, this same probiotic organism has been shown to readily colonize and induce an immune response in the small intestine in humans.⁹⁶ Interestingly, in view of the relevance of techoic acid biosynthesis in the immunologic responses to certain lactobacilli, it has been shown by Duncker and colleagues that a Lactobacil-lus mutant (leading to D-alanine depletion of lipotechoic acid) also significantly inhibited visceral pain perception in healthy noninflamed rats.⁹⁷

Functional and morphologic changes in the enteric neuromuscular apparatus develop in mice infected with Trichinella spiralis long after the worms have been expelled and the related inflammatory response has subsided, thus providing an animal model of postinfectious IBS.^98,99 L paracasei, but not other strains, has been shown to attenuate gut muscle hypercontractility, reduce immune activation,⁹³ and normalize the metabolic profile of mice in this model.¹⁰⁰

These experimental observations are now supported by clinical studies with probiotics in IBS in humans. Results in IBS continue to be variable with a number of organisms, such as Lactobacillus GG, Lactobacillus plantarum, L acidophilus, Lactobacillus casei, the probiotic cocktail VSL#3, and Bifidobacterium animalis,^101,102 alleviating individual IBS symptoms (eg, bloating, flatulence, and constipation) and only a few products affecting pain and global symptoms.^80,103–105 Other products have shown no benefit.^106,107

Obesity, Metabolic Syndrome, Nonalcoholic Fatty Liver Disease, and Nonalcoholic Steatohepatitis

Several mechanisms involving the microbiota in the patho-genesis of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) have been identified. In particular, a role for the microbiota in relation to diet in the pathogenesis of obesity per se has been extensively investigated.^108,109 Pertinent findings include the ability of gram-negative anaerobes, such as Bacteriodes thetaiotami-cron, to cleave most glycosidic linkages and degrade plant polysaccharides, thereby supplying the host with 10% to 15% of its calorific requirement.^108–111 The microbiota of obese individuals, as well as the cecal microbiota of ob/ob mice, is more efficient at the extraction of energy from the diet and in the production of SCFAs.^110,112 Furthermore, the microbiota has been shown to stimulate hepatic triglyceride production through suppression of the lipoprotein lipase (LPL) inhibitor, fasting-induced adipose factor (also known as angiopoietin-like 4), thereby leading to continued expression of LPL, a key regulator of fatty acid release from triglycerides in the liver.¹¹³ The gut microbiota also can modulate systemic lipid metabolism through modification of bile acid metabolic patterns, also impacting directly on the emulsification and absorption properties of bile acids and, thus, indirectly on the storage of fatty acids in the liver. The microbiota also has been implicated in the development of insulin resistance,¹¹³ a fundamental abnormality in metabolic syndrome, by affecting energy balance, glucose metabolism, and the low-grade inflammatory state that has been associated with obesity and related metabolic disorders. Its role in choline metabolism,^114–116 as well as inactivation of pro-inflammatory cytokines (eg, TNF-α), appears relevant to the development of NAFLD and progression to NASH. Most recently, studies in experimental models have shown that defective/deficient inflammasome sensing and related dysbiosis result in an abnormal accumulation of bacterial products in the portal circulation and promote progression of NAFLD/NASH.¹¹⁷

A more fundamental role for SIBO has been proposed in NAFLD by promoting both steatosis and inflammation^118,119 (Figure 2). The potential of microbes of enteric origin to induce a progressive and even fatal steatohepatitis had been recognized several years ago in relation to the liver injury that complicated jejuno-ileal bypass operations for morbid obesity; indeed, that procedure has provided a valuable experimental model for exploring the impact of the microbiota in liver disease.

Figure 2

The gut flora (microbiota) and the liver. Small intestinal bacterial overgrowth (SIBO), present in a variety of liver diseases, and/or an altered composition of the colonic microbiota lead to an enhanced release of proinflammatory cytokines. Increased ...