Review: Diet, Lifestyle, Gut Microbiota, and Human Health Relationship

This report, published in Nutrients in 2015, reviews the significant impact of diet and lifestyle on the human gut microbiota and its subsequent effects on health. The review explores the complex interactions within the gut, emphasizing the role of diet, particularly macronutrients, in shaping microbial composition and activity. It highlights the importance of both short- and long-term dietary changes, including infant nutrition, and discusses the influence of environmental factors and lifestyle choices like smoking and stress. The report also covers the use and potential benefits of prebiotics and probiotics, and identifies areas for future research, such as understanding the impact of lifestyle factors, the role of microbial products, and the long-term consequences of early-life nutrition on the gut microbiome. The review underscores the importance of a diverse and thriving gut microbiota in maintaining health and preventing disease.

Nutrients 2015, 7, 17-44; doi:10.3390/nu7010017
nutrients
ISSN 2072-6643
www.mdpi.com/journal/nutrients
Review
The Impact of Diet and Lifestyle on Gut Microbiota and
Human Health
Michael A. Conlon * and Anthony R. Bird
CSIRO Food and Nutrition Flagship, Kintore Ave, Adelaide, SA 5000, Australia;
E-Mail: [e-mail]
* Author to whom correspondence should be addressed; E-Mail: [e-mail];
Tel.: [phone]; Fax: [phone].
Received: 17 September 2014 / Accepted: 9 December 2014 / Published: 24 December 2014
Abstract: There is growing recognition of the role of diet and other environmental factors
in modulating the composition and metabolic activity of the human gut microbiota, which in
turn can impact health. This narrative review explores the relevant contemporary scientific
literature to provide a general perspective of this broad area. Molecular technologies have
greatly advanced our understanding of the complexity and diversity of the gut microbial
communities within and between individuals. Diet, particularly macronutrients, has a major
role in shaping the composition and activity of these complex populations. Despite the body
of knowledge that exists on the effects of carbohydrates there are still many unanswered
questions. The impacts of dietary fats and protein on the gut microbiota are less well defined.
Both short- and long-term dietary change can influence the microbial profiles, and infant
nutrition may have life-long consequences through microbial modulation of the immune
system. The impact of environmental factors, including aspects of lifestyle, on the microbiota
is particularly poorly understood but some of these factors are described. We also discuss
the use and potential benefits of prebiotics and probiotics to modify microbial populations.
A description of some areas that should be addressed in future research is also presented.
Keywords: diet; lifestyle; gut; microbiota; health
OPEN ACCESS

Nutrients 2015, 7 18
1. Introduction
There are approximately 10 times as many microorganisms within the gastro-intestinal (GI) tract of
humans (approximately 100 trillion) as there are somatic cells within the body. While most of the
microbes are bacteria, the gut can also harbor yeasts, single-cell eukaryotes, viruses and small parasitic
worms. The number, type and function of microbes vary along the length of the GI tract but the majority
is found within the large bowel where they contribute to the fermentation of undigested food components,
especially carbohydrates/fiber, and to fecal bulk. Some of the most commonly found or recognized genera
of gut bacteria in adults are Bifidobacterium, Lactobacillus, Bacteroides, Clostridium, Escherichia,
Streptococcus and Ruminococcus. Approximately 60% of the bacteria belong to the Bacteroidetes or
Firmicutes phyla [1]. Microbes which produce methane have been detected in about 50% of individuals
and are classified as Archaea and not bacteria [2]. Although individuals may have up to several hundred
species of microbes within their gut, recent findings from The Human Microbiome Project and
others [3,4] show that thousands of different microbes may inhabit the gut of human populations
collectively and confirm a high degree of variation in the composition of these populations between
individuals. Despite this variation in taxa the abundance of many of the microbial genes for basic or
house-keeping metabolic activities are quite similar between individuals [3]. There is growing evidence
that imbalances in gut microbial populations can be associated with disease, including inflammatory
bowel disease (IBD) [5], and could be contributing factors. Consequently, there is increased awareness
of the role of the microbiota in maintaining health and significant research and commercial investment
in this area. Gut microbes produce a large number of bioactive compounds that can influence health;
some like vitamins are beneficial, but some products are toxic. Host immune defenses along the intestine,
including a mucus barrier, help prevent potentially harmful bacteria from causing damage to tissues. The
maintenance of a diverse and thriving population of beneficial gut bacteria helps to keep harmful bacteria
at bay by competing for nutrients and sites of colonization. Dietary means, particularly the use of a range
of fibers, may be the best way of maintaining a healthy gut microbiota population. Strategies such as
ingestion of live beneficial bacteria (probiotics) may also assist in maintaining health. In this review, we
will expand upon these subjects relating to diet and lifestyle, the gut microbiota and health, and provide
some indication of opportunities and knowledge gaps in this area.
2. Microbial Products that Impact Health—Beneficial and Harmful
Microbial mass is a significant contributor to fecal bulk, which in turn is an important determinant of
bowel health. Consumption of dietary fibers reduces the risk of colorectal cancer (CRC) [6] at least
partly as a consequence of dilution and elimination of toxins through fecal bulk, driven by increases in
fermentative bacteria and the presence of water-holding fibers [7–9]. Aspects of this will be discussed
in more detail later in the review.
Gut microbes are capable of producing a vast range of products, the generation of which can be
dependent on many factors, including nutrient availability and the luminal environment, particularly
pH [10]. A more in-depth review of gut microbial products can be found elsewhere [11]. Microbial
products can be taken up by GI tissues, potentially reach circulation and other tissues, and be excreted
in urine or breath. Fermentation of fiber and protein by large bowel bacteria results in some of the most

Nutrients 2015, 7 19
abundant and physiologically important products, namely short chain fatty acids (SCFA) which act as
key sources of energy for colorectal tissues and bacteria, and promote cellular mechanisms that maintain
tissue integrity [12–14]. SCFA can reach the circulation and impact immune function and inflammation
in tissues such as the lung [15]. However, some protein fermentation products such as ammonia, phenols
and hydrogen sulphide can also be toxic. There are many other products which deserve mention for their
influence on health. Bacteria such as Bifidobacterium can generate vitamins (e.g., K, B12, Biotin, Folate,
Thiamine) [11]. Synthesis of secondary bile acids, important components of lipid transport and turnover
in humans, is mediated via bacteria, including Lactobacillus, Bifidobacterium and Bacteroides [11].
Numerous lipids with biological activity are produced by bacteria, including lipopolysaccharide (LPS),
a component of the cell wall of gram negative bacteria that can cause tissue inflammation [16]. Also,
many enteropathogenic bacteria (e.g., some E. coli strains) can produce toxins or cause diahorrea under
the right conditions, but under normal circumstances other non-pathogenic commensal bacteria with
similar metabolic activities outcompete and eventually eliminate them [17]. Bacteria such as
Bifidobacterium can also help prevent pathogenic infection through production of acetate [18].
Many enzymes produced by microbes influence digestion and health. Indeed, much of the microbial
diversity in the human gut may be attributable to the spectrum of microbial enzymatic capacity needed
to degrade nutrients, particularly the many forms of complex polysaccharides that are consumed by
humans [19]. Some bacteria such as Bacteroides thetaiotamicron have the capacity to produce an array
of enzymes needed for carbohydrate breakdown [20], but in general numerous microbes appear to be
required in a step-wise breakdown and use of complex substrates. Bacterial phytases of the large intestine
degrade phytic acid present in grains, releasing minerals such as calcium, magnesium and phosphate that
are complexed with it [21], making these available to host tissues (e.g., bone). Enzymes which degrade
mucins help bacteria meet their energy needs and assist in the normal turnover of the mucus barrier
lining the gut.
Competition between bacteria for substrates has a significant influence on which products are
generated. Hydrogen is used by many bacteria and there is a hydrogen economy within the gut based
around production by some bacteria and its use by others, including methanogens and sulphate-reducing
bacteria (SRB) [22,23]. The use of hydrogen for production of methane by methanogenic Archaea may
limit acetate production by other microbes, thereby potentially limiting production of beneficial butyrate
and impacting health [2,23]. The role of methanogens in health is not yet clear. Breath methane correlates
with levels of constipation in irritable bowel syndrome (IBS) [24] but methanogens numbers are depleted
in IBD [2].
Production of gases such as methane, hydrogen, hydrogen sulphide and carbon dioxide is associated
with digestion and fermentation within the GI tract. While excess production may cause GI problems
such as bloating and pain, the gases may serve useful purposes. However, there is debate over whether
hydrogen sulphide is largely beneficial or detrimental [23].
There is a strong interaction between the host immune system and the microbiota, with both producing
compounds that influence the other. Some bacteria such as the key butyrate-producer Faecalibacterium
prausnitzii may produce anti-inflammatory compounds [25]. Microbes also produce substances that
allow communication between each other.

Nutrients 2015, 7 20
3. Lifestage and Lifetstyle Impacts on the Microbiota and the Influence of Nutrition
3.1. Lifestage
Microbes colonise the human gut during or shortly after birth. The fact that babies delivered naturally
have higher gut bacterial counts at 1 month of age than those delivered by caesarean section [26] suggests
gut colonization by microbes begins during, and is enhanced by, natural birth. The growth and
development of a robust gut microbiota is important for the development of the immune system [27] and
continues during breast-feeding, a stage which seems to be important for the long-term health of the
individual. Oligosaccharides present in breast milk promote the growth of Lactobacillus and
Bifidobacterium, which dominate the infant gut, and this can strengthen or promote development of the
immune system and may help prevent conditions such as eczema and asthma [28–30]. These bacteria
are undetectable in the stool of preterm infants in their first weeks of life [31]. A significant shift in the
populations of gut microbes occurs when infants switch to a more solid and varied diet, including a
decline in populations of Lactobacillus and Bifidobacterium to only a small percentage of the large bowel
microbiota [32]. A wide diversity of microorganisms is needed to utilize the many fibers and other
nutrients present in adult diets [19,33]. Functional maturation of the human microbiota, including the
capacity to produce vitamins, increases during the early years of life [34].
The complexities and variability of adult gut microbial populations have become increasingly evident
in recent years. The variability may relate to the influence of numerous factors, including diet and host
genetics. The composition and activity of gut bacteria can vary according to (and possibly a result of)
life events, including puberty, ovarian cycle, pregnancy and menopause [11]. The diets of children being
weaned may have particular influence on microbial diversity in later life. Another broad shift in gut
microbe populations occurs with age. The Bacteroidetes phylum bacteria tend to dominate numerically
during youth but numbers decline significantly by old age, whereas the reverse trend occurs for bacteria
of the Firmicutes phylum [11]. The consequences and reason for this change are not yet clear. However,
the gut microbiota profiles of the elderly may not be optimal. One study found a high prevalence of
potentially toxic Clostridium perfringens and lower numbers of Bifidobacterium and Lactobacillus in
those in long-term care [35]. The latter also have a reduced microbial diversity compared to the elderly
living in the community and this is related to increased frailty and changes in nutrition [36].
3.2. Lifestyle
The impact of non-dietary lifestyle factors on the gut microbiota has been largely ignored. Smoking
and lack of exercise can significantly impact the large bowel (and potentially the microbiota) as they are
risk factors for CRC [37]. Indeed, smoking has a significant influence on gut microbiota composition,
increasing Bacteroides-Prevotella in individuals with Crohn’s Disease (CD) and healthy individuals [38].
Smoking-induced changes in microbial populations could potentially contribute to increased risk of CD.
Air-borne toxic particles can reach the large bowel via mucociliary clearance from the lungs, and
increased environmental pollution associated with industrialization could contribute to concomitant
increases in IBD cases [39].
Another lifestyle factor, stress, has an impact on colonic motor activity via the gut-brain axis which
can alter gut microbiota profiles, including lower numbers of potentially beneficial Lactobacillus [40].

Nutrients 2015, 7 21
Stress may contribute to IBS, one of the most common functional bowel disorders, and the associated
changes in microbial populations via the central nervous system (CNS). The gut-brain axis is bi-directional,
involving both hormonal and neuronal pathways [41], and so changes in the gut microbiota may influence
brain activity, including mood [42]. Autism, a neurodevelopmental disorder, is associated with significant
shifts in gut microbiota populations [43–45].
Obesity is associated with excess energy intakes and sedentary lifestyles. Exercise (or rather a
lack of it) may be an important influence on any shifts in microbial populations that are associated
with obesity. This is highlighted by a recent study that showed an increase in the diversity of gut
microbial populations in professional athletes in response to exercise and the associated diet [46].
In humans and animal models with obesity, shifts in gut microbial populations occur, with increases in
the Firmicutes and decreases in the Bacteroidetes, which could potentially contribute to adiposity through
greater energy harvest [47–49]. However, other data suggests the shifts in microbial populations are
driven primarily by the high fat obesogenic diets [50,51]. Irrespective of the cause, there are associated
increases in gut bacteria linked with poor health outcomes (e.g., Staphylococcus, E. coli,
Enterobacteriaceae) [52,53]. Dietary saturated fats may increase numbers of pro-inflammatory gut
microbes by stimulating the formation of taurine-conjugated bile acids that promotes growth of these
pathogens [54].
Geography also has a strong bearing on the composition of gut microbial populations. The diversity
of fecal microbes in children from rural Africa is greater than that of children of developed communities
in the EU, as is the number of bacteria associated with breakdown of fiber [55], suggesting dietary
differences contributes significantly to the microbial differences. In another study, the type of fecal
bacteria and their functional genes differed between individuals in the USA and in rural areas of
Venezuela and Malawi [34].
Other environmental factors may also influence health via gut microbes. Travel, particularly to
overseas destinations, increases the risk of contracting and spreading infectious diseases, including those
causing diarrhoea. Some infections may go undiagnosed but result in long-term GI problems, including
IBS [56]. Poor sanitary conditions in developing countries, and poor personal hygiene, can facilitate the
spread of infectious agents. Circadian disorganization, occurring because of travel, shift work or other
reasons, also impacts gut health and alters gut microbial populations [57].
4. Impacts of Macronutrients on the Gut Microbiota and Relevance to Health
4.1. Substrate Supply to the Colonic Microbiota
An adult colon contains approximately 500 g of contents, most of which is bacteria [58], and
about 100 g/day is voided as stool. A typical western type diet supplies the colonic microbiota with
about 50 g daily of potentially fermentable substrate, predominantly dietary fiber (DF). Non-starch
polysaccharides (NSP) are major components of DF and account for 20%–45% of the dry matter
supplied to the colon. Simple sugars and oligosaccharides each represent a further 10% whereas starch
(and starch hydrolysis products) supplies less than 8% of dry matter. Some sugar alcohols also escape
small intestine (SI) absorption and are minor dietary substrates for the colonic microbiota [59].
About 5–15 g of protein and 5–10 g of lipid passes into the proximal colon daily, largely of dietary

Nutrients 2015, 7 22
origin. Various other minor dietary constituents, including polyphenols, catechins, lignin, tannins and
micronutrients also nourish colonic microbes. About 90% of the approximately 1g/day of dietary
polyphenols escapes digestion and absorption in the SI [60,61] and can have significant influence on
microbial populations and activities [62–64].
4.2. Carbohydrates—Importance for Large Bowel Fermentation and Health
Carbohydrates are the principal carbon and energy source for colonic microbes. Collectively,
they have an immense capacity to hydrolyse a vast range of these nutrients, especially complex
polysaccharides [65].
DF is integral to a healthy diet and Australian adults consume ~27 g each day [66], which is greater
than in other high income countries, including the USA (<20 g/day). Epidemiological and experimental
studies show that DF is both preventative and therapeutic for many large bowel disorders and other
conditions or diseases, including cardiovascular diseases, type II diabetes and obesity [67–71].
One mechanism by which fiber promotes and maintains bowel health is through increasing digesta
mass. Incompletely fermented fiber (e.g., insoluble NSP such as cellulose), increases digesta mass
primarily though its physical presence and ability to adsorb water. An increase in digesta mass dilutes
toxins, reduces intracolonic pressure, shortens transit time and increases defecation frequency. Fibers
can also increase fecal mass to a lesser degree by stimulating fermentation, which leads to bacterial
proliferation and increased biomass [7].
Many of the health benefits ascribed to fiber are a consequence of their fermentation by the colonic
microbiota and the metabolites that are produced. Carbohydrates are fermented to organic acids that
provide energy for other bacteria, the bowel epithelium and peripheral tissues. SCFA are the major
endproducts of carbohydrate fermentation. These weak acids (pKa ~4.8) help lower the pH within the
colon thereby inhibiting the growth and activity of pathogenic bacteria. Other minor organic acids
produced include lactate, succinate and formate. Branched-chain SCFA (e.g., isobutyrate and isovalerate)
results from fermentation of branched chain amino acids [72].
There are spatial gradients in microorganisms along the length of the gut. Bacterial growth and
metabolic activity (fermentation) is greatest in the proximal colon where substrate availability is at a
maximum [13,73]. Accordingly, pH progressively increases as stool progresses from the proximal to
distal colon (from 5.8 to 7.0–7.5), largely because of the progressive depletion of carbohydrate substrates
and absorption of SCFA, and increasing efficiency of protein fermentation and production of alkaline
metabolites [72]. Total SCFA concentrations are highest in the proximal colon (~100 mM) and decline
progressively toward the distal colon. Acetate, propionate and butyrate are the major individual SCFA,
accounting for 90% of the total, with molar ratios approximating 65:20:15 [74].
Butyrate has attracted significant attention because it serves as the principal source of metabolic
energy for the colonocytes [75], is instrumental in maintaining mucosal integrity, modulates intestinal
inflammation and promotes genomic stability. The capacity of butyrate to regulate colonocyte
differentiation and apoptosis, promoting removal of dysfunctional cells, underscores its potential to
protect against colon cancer [76].
The SCFA also have roles beyond the gut and may improve risk of metabolic and immune system
diseases and disorders, such as osteoarthritis, obesity, type II diabetes and cardiovascular disease [13,76].

Nutrients 2015, 7 23
More than 90% of the total SCFA produced in the colon is absorbed by the epithelium, through
mechanisms that are not fully elucidated. SCFA-stimulated sodium-coupled transport in the apical
membrane of colonocytes is especially important as it mediates (co)absorption of water and helps
recover electrolytes as well as energy [77]. The SCFA can bind to G-protein coupled receptors in
colorectal tissues, particularly GPR 41 and 43, which may influence immune function and tumour
suppression, but these pathways are still relatively poorly characterized [76].
Most of the absorbed acetate reaches the liver via the portal vein, whereas propionate, and butyrate
to an even larger extent, is metabolized extensively by colonocytes. Acetate and propionate are used by
the liver for oxidation, and for lipogenesis and gluconeogenesis, respectively. Hepatic metabolic
clearance of SCFA is very high and so concentrations in the systemic bloodstream are about 100-fold
lower than those in colonic digesta and feces (~50 μM versus 100 mM, respectively) [13].
4.3. Protein
Dietary proteins are an important part of a balanced diet. Humans are unable to synthesize numerous
amino acids and must obtain them from proteins in food to maintain health. Some protein-rich foods such
as meat, eggs and nuts are also good sources of vitamins or nutrients such as iron. There is good evidence
that a diet containing moderate to high amounts of protein can also contribute to weight loss in
overweight individuals, particularly if combined with exercise [78], thereby minimising the health risks
associated with obesity. Dietary proteins also have a significant impact on gut health. Depending on the
type of protein and the other nutrients present in the food this can be beneficial or harmful. Some
epidemiological studies, particularly large studies (up to 500,000 people), indicate a slight but significant
association between CRC risk and the consumption of high levels of red and processed meats [79–82].
Not all epidemiological studies show such an association and the inconsistent findings may relate to the
many factors which may contribute to CRC [83,84].
The potential for protein to harm colorectal tissues is explicable using current knowledge. An increase
in protein intake usually results in more of the macronutrient, and hence fermentable substrate, reaching
the colon. Although protein digestibility has an important influence on how much reaches the colon,
most common dietary protein sources are highly susceptible to hydrolysis by SI enzymes. Dietary protein
serves as the major source of nitrogen for colonic microbial growth and is essential to their assimilation
of carbohydrates and the production of beneficial products such as SCFA. Hence, a combination of
protein and carbohydrates in the large bowel can contribute to bowel health. However, unlike
carbohydrates, fermentation of protein sources by the microbiota produces a much greater diversity of
gases and metabolites, and increasing the nitrogenous substrate for the microbiota can also increase
putrefactive fermentation products [85]. As digesta passes down the bowel its carbohydrate content
dwindles and protein fermentation becomes progressively more important. Putrefactive fermentation has
been implicated in the development and progression of many common bowel diseases given their greater
prevalence in the distal colon [86], including CRC and IBD. Many of these protein fermentation
endproducts, which include ammonia, hydrogen sulphide, amines, phenols, thiols and indoles, have been
shown to be cytotoxins, genotoxins and carcinogens [87], in in vitro and animal models [88]. Generally,
fecal levels of protein fermentation products, such as sulphide, are positively associated with dietary
protein consumption in humans and there is evidence from rat studies that higher dietary protein intake

Nutrients 2015, 7 24
(including higher red meat intake) is associated with greater DNA damage in colonic mucosa when
dietary levels of fermentable carbohydrate are low [88–91]. Recently completed studies suggest that this
relationship holds true for humans [92–94]. However, higher protein intake does not always result in
higher fecal levels of protein fermentation products [95] nor does it necessarily increase the genotoxicity
of fecal water in humans [96].
Although ammonia is a well-known toxin [97] it is used as an N source by the microbiota and
most is excreted via stool or absorbed in the gut and eliminated in urine. Diets promoting microbial
protein synthesis (and concomitant increased utilisation of ammonia), effectively reroute systemic
N excretion from the kidneys to the fecal stream, which has benefits for renal health [98]. Other
components derived from dietary protein sources such as red meat may also influence the gut microbiota
and health. Microbial metabolism of L-carnitine, abundant in red meat, may generate products such as
trimethylamine-N-oxide that could increase risk of atherosclerosis [99].
4.4. Fat
Dietary fat also influences the composition and metabolic activity of the gut microbiota and some
evidence for this has been described earlier in relation to obesity.
High fat diets induce increased circulating levels of bacteria-derived LPS in humans, possibly as a
consequence of increased intestinal permeability [100]. LPS is an immune system modulator and potent
inflammatory agent linked to the development of common metabolic diseases.
The influence of dietary fat on the gut microbiota may be indirectly mediated by bile acids. Hepatic
production and release of bile acids from the gall bladder into the SI, and the amount that escapes
enterohepatic recycling and enters the colon, is increased with fat intake. Secondary bile acids, produced
by 7 α-dehydroxylation of primary bile acids by colonic microbiota, are potentially carcinogenic and have
been implicated in the aetiology of CRC and other GI diseases [101,102]. Further research is required
on the interactions between dietary fat, the type and amount of bile acids that reach the large bowel, and
the population structure and function of the microbiota in that viscus.
5. Effects of Polyphenols on the Microbiota
Dietary polyphenols, sourced from many foods including grapes, grains, tea, cocoa and berries,
generally promote health and are linked to prevention of diseases such as cancer and cardiovascular
disease [103]. Although many dietary polyphenols may have biological impacts through anti-oxidant
effects or anti-inflammatory pathways [103], polyphenols which reach the colon can be metabolized by
the resident microbiota and result in bioactive products, but our understanding of the microbial
bioconversion processes is limited [104–106]. Metabolic profiling of polyphenolic products in excreta
and blood using tools such as NMR is enabling greater insights into effects of dietary polyphenols
in humans [107] but linking the metabolic changes to health outcomes remains a challenge [108].
Individual differences in microbiota populations may result in different capacities for polyphenol
bioconversion [109] with potential consequences for health. In this context, it is noteworthy that the gut
microbiota population profiles of individuals with IBD are significantly different from healthy
individuals, and also that the polyphenolic metabolite profiles are also different between the two
groups [110].

Nutrients 2015, 7 25
6. Western-Style Diets
The Western lifestyle, including diet, is associated with high incidences of chronic diseases, such as
cardiovascular disease, CRC and type II diabetes which individually and collectively carry a hefty
socioeconomic burden [111]. Most Western populations over-consume highly refined, omnivorous diets
of poor nutritional quality. Those diets are energy dense, high in animal protein, total and saturated fats,
and simple sugars but low in fruits, vegetables and other plant-based foods. Consequently, they are
typically low in DF, NSP in general and RS in particular. For Western civilisations, refined cereal
products (e.g., white bread) are the main DF source. Overfeeding (and sedentary behaviour) is also a
hallmark of these populations.
Much of what is known about the diversity and complexity of human gut microbiota comes
from molecular analysis of fecal samples obtained mainly from small cohorts of Caucasian adults
habitually consuming Western style diets. Considerably less is understood about how other dietary
patterns (e.g., vegetarian, Mediterranean) might influence the community structure and metabolic
activity of microbiota.
7. Diet and Dietary Change
In humans, the microbial gene set is 150 times larger than the gene complement of the host [112].
However, only about 50 species belonging to just five or six genera and two phyla account for 99% of
biomass. Of the genera Bacteroides, Bifidobacterium and Eubacterium are numerically the most
important and may account for more than 60% of culturable bacteria present in human stool. Clostridium,
Enterobacteriaceae and Streptococcus are also important but less numerous. Nearly all (~90%) of the
bacteria in the human gut can be mapped to just two phyla, Bacterioidetes and Firmicutes. The relative
proportions of the two dominant phyla vary, and can be influenced by a range of factors, but most people
have similar proportions of each [113].
Long-term, habitual diet (i.e., dietary pattern) and shorter term dietary variation influences gut
microbiota composition. The population structure is responsive to acute dietary change (daily variation),
as evidenced by rapid and substantial increases in populations at the genus and species level. However,
dietary change does not necessarily result in a permanent (paradigm) compositional shift, at least at
phylum level, although evidence for this assertion is limited [114].
8. Dietary Patterns, Macronutrients and Microbiota Taxonomic Composition
8.1. Observational Studies
Cross-sectional studies have shown some evidence that Western-style diets are associated with gut
microbial populations that are typified by a Bacteroides enterotype whereas traditional diets rich in plant
polysaccharides are associated with a Prevotella enterotype [114]. The Prevotella enterotype was only
weakly associated with components that typify Western diets but strongly linked to carbohydrates and
simple sugars. The fecal microbiota of children in the USA is dominated by Bacteroides [34,115].
Similarly, Italian children have high levels of Enterobacteriaceae (mainly Shigella, Escherichia and
Salmonella). In contrast, the stool of children in rural Africa and South America consuming traditional

Nutrients 2015, 7 26
plant-based diets was enriched in Bacteroidetes, in particular the Prevotella enterotype and species
associated with fiber utilization (e.g., Xylanibacter) [55]. Prevotella and (Xylanibacter) are known to
use cellulose and xylans as substrates [55,116]. Diets of North American and Italian urban children are
much richer in animal protein and saturated fats whereas the diets for the other two populations are
plant-based and have higher levels of fiber. The Bacteroidetes:Firmicutes ratio was lower for children
in the Western countries.
As stated earlier, there is a paucity of data on the association between vegetarian dietary patterns and
the gut microbiota, especially using molecular methods. A study that used PCR-denaturing gradient gel
electrophoresis (DGGE) for microbial population fingerprinting found no significant differences in the
fecal microbiota of vegetarians and omnivores, although the abundance of Clostridium cluster IV in the
latter tended to be greater [117]. In a cohort of female college students from rural India, the fecal
microbiota of those whose dietary pattern was omnivorous had a greater relative abundance of
Clostridium cluster XIVa bacteria, specifically Roseburia-E. rectale (butyrate-producing bacteria),
compared to the lacto-vegetarians [118]. There were no differences in the relative proportions of other
major bacterial groups targeted. A gene encoding for a pivotal enzyme (butyryl-CoA CoA-transferase)
involved in butyrate synthesis was also upregulated in the omnivores. The study demonstrates
differences in the composition and functional capacity of the microbiota of individuals with two
markedly diverse dietary patterns.
The taxonomic diversity of the fecal microbiota of individuals on habitual Western diets appears
to be less than for those consuming plant-based diets. Also, individuals who are obese or have
type II diabetes, inflammatory diseases (osteoarthritis) and other major health problems (prevalent in
Western societies) have a sub-optimal fecal microbiota profile. Specifically, it is less diverse than
that of healthy controls [119,120] and there are also major compositional differences at the phylum
level. Obesity is associated with an increased fecal Bacteroidetes:Firmicutes ratio relative to lean
subjects [121]. Whether a microbiota with lower compositional diversity is less resilient to environmental
challenges and is less “healthier” for the host is not yet known [122].
The fecal hydrogenotrophic microbiota of native Africans, whose diet is low in animal products,
compared to that of African and European Americans consuming a typical Western diet was more
diverse and contained different populations of hydrogenotrophic Archaea and methanogenic Archaea as
well as SRB populations [123]. The differences in bacterial community structures of native African
populations were reflective of the diets of the hosts. Those on Western diets, characterized by higher
intakes of dietary animal proteins (as meat, milk and eggs), may deliver greater amounts of sulphur
compounds to the colonic microbiota [124], thus favouring sulfidogenic hydrogen disposal whereas in
native Africans methane is the major hydrogen sink. Native African populations have lower intake of
animal products and higher breath methane concentrations than westernized populations [123,125].
8.2. Dietary Interventions
Replacing a habitual Western diet with one high in fiber elicited rapid (within 24 h) and marked
alterations in fecal microbiota composition, although the changes were insufficient to produce a broad
switch from Bacteroides to Prevotella enterotype [114].

Nutrients 2015, 7 27
In an inpatient study [126], altering dietary energy load in lean and obese adults induced rapid changes
in the proportional abundance of Bacteroidetes and Firmicutes. The former decreased whereas the latter
increased with increasing energy intake. Further studies are required to determine if the changes in
microbiota composition were the result of the increase in dietary fat or another macronutrient. High fat
diets are also associated with substantial compositional changes in the colonic microbiota at the phylum
and genus levels, including reductions in both Gram positive (e.g., Bifidobacterium spp.) and Gram
negative bacteria (e.g., Bacteroides) [123].
Animal models are also proving useful in understanding factors that impact the gut microbiota,
particularly in regards to high fat diets and obesity. A study using a murine (RELMβ) knockout
model showed that dietary fat-induced changes to gut microbiome composition were independent of
obesity [127]. In conventional mice, increased dietary fat intake resulted in fewer numbers of
Bacteroidetes and increases in Firmicutes and Proteobacteria. A high fat diet also reduced cecal
Bifidobacterium numbers and increased circulating LPS concentrations [128,129] and has also been
shown to reduce the abundance of Clostridium cluster XIVa, including Roseburia spp. [130].
Diet-induced changes in mucosal integrity have been shown to promote metabolic endotoxemia and
trigger systemic low grade inflammatory responses in a range of tissues [100,128,129].
9. Microbes and Mucosal Health
A layer of mucus, produced by goblet cells, lines the epithelium of the GI tract and acts as a barrier
to microbial invasion of tissues and can contribute to intestinal homeostasis [131,132]. The basic
component of mucus is mucin. Some bacterial products (SCFA) stimulate the production of mucus in
response to dietary components such as NSP [133]. Over-utilization of the mucus by bacteria or reduced
production can lead to thinning of the barrier under certain dietary conditions [88]. In the colon, “mucin-
depleted foci” may develop as one of the features associated with tumorigenesis in rodents and humans in
response to carcinogens [134]. However, the role of mucin depletion in oncogenesis is not clear as a
recent study in rats showed that inflammation associated with mucin-depleted foci was not due to
infiltration of bacteria, whereas colonic tumors did appear to be colonized by bacteria [135]. Many
bacteria can adhere to and degrade the outer layer of colonic mucus but the inner layer is generally bacteria
free [136]. Although break-down of mucus by bacteria is a normal part of mucus barrier turnover, an
overabundance of mucus-degrading bacteria, such as Akkermansia muciniphila in the adherent mucus
layer of individuals with IBD [137,138], could contribute to tissue inflammation by weakening
the barrier.
Tight junctions between cells also helps prevent translocation of bacteria and molecules (including
toxins) across gut epithelial tissues. A loss of this integrity (a so-called “leaky gut”) may have serious
consequence for health. In the first few years of life, interactions between the gut microbiota and the
mucosal barrier appear important and perturbations in the relationship that lead to excessive gut
permeability and immune changes may result in susceptibility to a range of diseases in later life [139].
A significant proportion of the activities of the immune system occur within the gut. Gut-associated
lymphatics contribute substantially to this defense but other cells lining the gut also produce a range of
molecules which can neutralise pathogenic microbes. Dendritic cells sample the gut luminal environment
for harmful bacteria and can induce a suite of responses including the activation of macrophages,

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B cells and T cells within mucosal tissues and the release of broad specificity ant-microbial agents such
as Immunoglobulin A and α-defensins into the luminal environment [140].
A loss of gut barrier function may contribute to numerous diseases. An example is Parkinsons disease
(PD), a multi-system disease in which there is dysfunction of the GI tract, including changes in the
enteric nervous system which appear before obvious degeneration of the CNS [141,142]. Individuals
with PD have increased intestinal permeability, greater intestinal infiltration of E. coli and greater
endotoxin (LPS) exposure, and these changes correlate with the enteric neuronal damage [143], leading
to suggestions that a pathogen may be responsible for PD [144] and a breakdown in mucosal barrier
function may play a central role. An impaired gut barrier may also contribute to symptoms or complications
of autism, kidney disease, type 2 diabetes, cardiovascular disease, metabolic syndrome, obesity, and
liver diseases [45,100,145–149].
10. Inter-Individual Variation in Gut Microbiota and Responses to Diet
Each individual has a distinct combination of gut microbial species. This has become increasingly
evident from molecular analyses of recent decades, including The Human Microbiome Project. One
metagenomic analysis also suggested that the gut microbiota of each human is typified by one of
three enterotypes, with each enterotype characterised by distinct dominant groups of microbes [150],
namely Bacteroides, Prevotella and Ruminococcus. However, subsequent studies, including those of
The Human Microbiome Project, have been unable to provide clear support for the concept as initially
proposed [4,114]. More recent findings and analysis of the evidence roughly support typing with
Prevotella or Bacteroides dominance of the microbiota but the numerous factors, especially dietary, that
impact gut microbial populations means there is considerable variation in numbers of these genera,
making it difficult to classify populations as a particular “type” [113].
Inter-individual differences in populations of the gut microbiota may lead to different capacities to
utilize dietary components and to different levels of disease risk. For example, some individuals have
consistently low stool levels of the microbial fermentation product butyrate, levels which generally
remain lower relative to others despite concentrations increasing in response to a diet high in RS [151].
Butyrate production is important for the maintenance of colorectal tissue integrity and may protect
against colorectal diseases [13,76]. Individual differences in numbers and functions of bacteria such as
Ruminococcus bromii, important for the generation of SCFA in response to RS in humans [152,153],
could potentially influence colorectal health.
11. Use of Probiotics and Prebiotics as Nutritional Strategies to Improve Health
Probiosis and prebiosis are diet-based processes/strategies for promoting the health of the host
through improving the composition of the colonic microbiota. Although both prebiotics and probiotics
have been shown to increase numbers of selected bacteria at the species and genus level, typically
Bifidobacterium and Lactobacillus, changes in the overall composition of the gut microbiota are often
relatively small, and generally persist only for as long as the period of the intervention. Also, definitive
proof that the identified compositional alterations are directly responsible for an improvement in host
health generally remains elusive. While the concepts have practical relevance they are simplistic given