For personal use. Only reproduce with permission from The Lancet Publishing Group. REVIEW 512 THE LANCET ��� Vol 360 ��� February 8, 2003 ��� www.thelancet.com Many species of bacteria have evolved and adapted to live and grow in the human intestine. The intestinal habitat of an individual contains 300���500 different species of bacteria,1,2 and the number of microbial cells within the gut lumen is about 10 times larger than the number of eukaryotic cells in the human body.3 The stomach and small intestine contain only a few species of bacteria adhering to the epithelia and some other bacteria in transit. The scarcity of bacteria in the upper tract seems to be because of the composition of the luminal medium (acid, bile, pancreatic secretion), which kills most ingested microorganisms, and because of the phasic propulsive motor activity towards the ileal end, which impedes stable colonisation of bacteria in the lumen. By contrast, the large intestine contains a complex and dynamic microbial ecosystem with high densities of living bacteria, which achieve concentrations of up to 1011 or 1012 cells/g of luminal contents.1 These concentrations are similar to those found in colonies growing under optimum conditions over the surface of a laboratory plate.4 A large proportion of the faecal mass consists of bacteria (around 60% of faecal solids).5 Several hundred grams of bacteria living within the colonic lumen affect host homoeostasis. Some of these bacteria are potential pathogens and can be a source of infection and sepsis under some circumstances���for instance when the integrity of the bowel barrier is physically or functionally breached. However, the constant interaction between the host and its microbial guests can infer important health benefits to the human host.6 Recognition of these benefits is drawing particular attention to the functional implications of microflora in host physiology. Composition of the flora Colonisation of the gastrointestinal tract of newborn infants starts immediately after birth and occurs within a few days. Initially, the type of delivery (passage through Lancet 2003 361: 512���19 Digestive System Research Unit, Hospital General Vall d���Hebron, Barcelona, Spain (F Guarner MD, Prof J-R Malagelada MRCP) Correspondence to: Dr F Guarner, Digestive System Research Unit, Hospital General Vall d���Hebron, Autonomous University of Barcelona, 08035 Barcelona, Spain (e-mail: fguarnera@medynet.com) the birth canal versus caesarean section) and the type of diet (breast versus formula feeding) might affect the colonisation pattern.7���10 Other environmental factors also have a major role since differences exist between infants born in developed countries and those born in developing countries, and between infants from different hospital wards.11���13 Pioneer bacteria can modulate expression of genes in host epithelial cells,14 thus creating a favourable habitat for themselves, and can prevent growth of other bacteria introduced later in the ecosystem. The initial colonisation is therefore very relevant to the final composition of the permanent flora in adults.15 Conventional bacteriological analysis of faecal flora requires meticulous techniques for cultivation of bacteria on various growth media and an array of methods for taxonomic identification of the isolates. Results of such studies1 have shown that anaerobic bacteria outnumber aerobic bacteria by a factor of 100���1000. The genera bacteroides, bifidobacterium, eubacterium, clostridium, peptococcus, peptostreptococcus, and ruminococcus are predominant in human beings,1,6 whereas aerobes (facultative anaerobes) such as escherichia, enterobacter, enterococcus, klebsiella, lactobacillus, proteus, etc are among the subdominant genera. Every individual has several hundreds of species belonging to these genera, with a particular combination of predominant species that is distinct from that found in other individuals.1,16 The species vary greatly between individuals.16 The comp- osition of the individual���s flora can fluctuate under some circumstances, for instance acute diarrhoeal illnesses, antibiotic treatment, or to lesser extent induced by dietary interventions, but individuals��� flora composition pattern usually remain constant.1,16 Several bacteria that can be seen by direct microscopic examination of diluted faecal specimens cannot be grown in culture media. Unicellular organisms need biodiversity for growth. Thus, 40���80% of the total microscopic counts Gut flora in health and disease Francisco Guarner, Juan-R Malagelada Review The human gut is the natural habitat for a large and dynamic bacterial community, but a substantial part of these bacterial populations are still to be described. However, the relevance and effect of resident bacteria on a host���s physiology and pathology has been well documented. Major functions of the gut microflora include metabolic activities that result in salvage of energy and absorbable nutrients, important trophic effects on intestinal epithelia and on immune structure and function, and protection of the colonised host against invasion by alien microbes. Gut flora might also be an essential factor in certain pathological disorders, including multisystem organ failure, colon cancer, and inflammatory bowel diseases. Nevertheless, bacteria are also useful in promotion of human health. Probiotics and prebiotics are known to have a role in prevention or treatment of some diseases. Search strategy and selection criteria In writing this review, we relied on original articles and reviews that were published in scientific journals and are searchable in database libraries (OVID, PubMed, Medline Plus Databases), and on our current readings on the topic. Due to space limitations, the number of studies quoted has been restricted. We chose articles for citation on the basis of the relevance of its contents without any bias toward author or journal.

For personal use. Only reproduce with permission from The Lancet Publishing Group. with high production of short-chain fatty acids, an acidic pH (5���6), and rapid bacterial growth.26,32,33 By contrast, the substrate in the left or distal colon is less available, the pH is close to neutral, putrefactive processes become quantitatively more important, and bacterial populations are close to static (figure 1). Colonic microoganisms also play a part in vitamin synthesis34,35 and in absorption of calcium, magnesium, and iron.25,36,37 Absorption of ions in the caecum is improved by carbohydrate fermentation and production of short-chain fatty acids, especially acetate, propionate, and butyrate. All of these fatty acids have important functions in host physiology. Butyrate is almost completely consumed by the colonic epithelium, and it is a major source of energy for colonocytes.26 Acetate and propionate are found in portal blood and are eventually metabolised by the liver (propionate) or peripheral tissues, particularly muscle (acetate).26,30 Acetate and propionate might also have a role as modulators of glucose metabolism: absorption of these short-chain fatty acids would result in lower glycaemic responses to oral glucose or standard meal���a response consistent with an ameliorated sensitivity to insulin.38,39 In fact, foods with high proportion of non-digestible carbohydrates all have a low glycaemic index.40,41 However, results of one study42 showed no effect of colonic fermentation of carbohydrates on insulin resistance. Trophic functions Epithelial cell growth and differentiation���Possibly, the most important role of short-chain fatty acids on colonic physiology is their trophic effect on the intestinal epithelium. The rate of production of crypt cells is reduced in the colon of rats bred in germ-free environments, and their crypts contain fewer cells than do those of rats colonised by conventional flora, suggesting that intraluminal bacteria affect cell proliferation in the colon.43 Differentiation of epithelial cells is greatly affected by interaction with resident microorganisms.14,44 All three major short-chain fatty acids stimulate epithelial cell proliferation and differentiation in the large and small bowel in vivo.45 However, butyrate inhibits cell proliferation and stimulates cell differentiation in epithelial cell lines of neoplastic origin in vitro.46 Moreover, butyrate promotes reversion of cells from neoplastic to non-neoplastic phenotypes.47 A role for REVIEW THE LANCET ��� Vol 360 ��� February 8, 2003 ��� www.thelancet.com 513 are not recoverable by culture,17,18 although estimates vary between individuals and between studies. Molecular biological procedures can now also be used to investigate the microbial ecology in the colon without use of cultures.19 Results of an analysis18 of bacterial genes in human faeces showed that many DNA sequences correspond to previously undescribed microorganisms, and some data20 suggest that every individual has unique strains of bacteria. Quantitative analysis21 of faecal bacteria shows important differences between individuals and over time within the same individual that are not always detectable by conventional culture techniques.22 Molecular procedures have shown that aerobes, including Escherichia coli, enterococci, and lactobacilli, achieve very high densities and metabolic activity in the human caecum, since 50% of total bacteria ribosomal RNA in caecal contents correspond to these species.23 By contrast, these species account for only 7% of bacteria ribosomal RNA in faecal samples.23 Such species could have an important role in caecal fermentations. Main functions of microflora Use of animals bred under germ-free conditions has provided important information about the effect of the microbial community of the gut on host physiology and pathology.24 Evidence obtained through such studies25 suggests that microflora have important and specific metabolic, trophic, and protective functions (panel). Metabolic functions A major metabolic function of colonic microflora is the fermentation of non-digestible dietary residue and endogenous mucus produced by the epithelia.25 Gene diversity in the microbial community provides various enzymes and biochemical pathways that are distinct from the host���s own constitutive resources. Overall outcomes of this complex metabolic activity are recovery of metabolic energy and absorbable substrates for the host, and supply of energy and nutritive products for bacterial growth and proliferation. Fermentation of carbohydrates is a major source of energy in the colon. Non-digestible carbohydrates include large polysaccharides (resistant starches, cellulose, hemicellulose, pectins, and gums), some oligosaccharides that escape digestion, and unabsorbed sugars and alcohols.26,27 The metabolic endpoint is generation of short-chain fatty acids. Anaerobic metabolism of peptides and proteins (putrefaction) by the microflora also produces short-chain fatty acids but, at the same time, it generates a series of potentially toxic substances including ammonia, amines, phenols, thiols, and indols.28,29 Available proteins include elastin and collagen from dietary sources, pancreatic enzymes, sloughed epithelial cells and lysed bacteria.6 Substrate availability in the human adult colon is about 20���60 g carbohydrates and 5���20 g protein per day.30,31 In the caecum and right colon, fermentation is very intense Main functions of gut flora Metabolic Fermentation of non-digestible dietary residue and endogenous mucus: salvage of energy as short-chain fatty acids, production of vitamin K, absorption of ions Trophic Control of epithelial cell proliferation and differentiation development and homoeostasis of the immune system Protective Protection against pathogens (the barrier effect) Distal colon Low substrate availability Proteolysis Neutral pH Slow bacterial growth Proximal colon High concentration of substrates Saccharolysis Acid pH (5���6) Rapid bacterial growth Figure 1: Fermentation in the colon