LA LEVADURA ESTA CRECIENDO Nature 429, 224 - 225 (2004) doi:10.1038/nj6988-224a The yeast is rising Kendall Powell Kendall Powell is a freelance science writer based in Broomfield, Colorado. Makers of beer, wine and cheese need microbiologists to keep fermented products at their peak. Kendall Powell gets a taste of the career offerings. Every student who has ever crossed paths with Saccharomyces cerevisiae, the academic version of brewer's yeast, has pondered getting a job at the local brewery. Cody Reif is living the dream. As a microbiologist at the New Belgium Brewing Company in Fort Collins, Colorado, Reif and his colleagues start each day by tasting the latest batches of the company's eight brews. He says that the group's taste-test gives better statistics for determining if a batch is up to company standards. Yeah, sure, statistics. "Is working here as much fun as people think? Yes, it is," Reif says. The daily sampling illustrates a criterion for working with fermented food products not usually highlighted in a basic microbiology degree: critical sensory skills. It reflects the fact that the bottom line is not the science behind making fermented products, but rather the flavour and appeal of a product. Careers focusing on the microbiological processes that go into making beer, wine and dairy products demand solid backgrounds in microbiology and biochemistry, but they also incorporate special expertise in food science and sensory training. Research projects in this small but diverse field range from troubleshooting contaminants that might spoil a vat of wine to designing probiotic microbial cultures for 'functional foods' that aim to improve the immune system. An understanding of how fermenting microbes behave, and how their metabolites change the properties of foods and beverages, can be picked up in specialist degree programmes or through on-the-job experience. Reif, who has a microbiology degree, brags: "I can probably identify four or five different strains of yeast or bacteria just by smell." One place a microbiologist might seek work is in a microbrewery — a small regional enterprise. There, says Reif, it is necessary to know the ins and outs of the entire beer-making process because any microbial problems could bleed into the next phase. Also, scientists at smaller companies handle most analyses on site, from quality control and yeast propagation to monitoring for 'critical beer spoilers' — bugs that can crash the keg party. This holds true at smaller wineries, too, says Lars Bjorkman, an enologist at Flora Springs Winery in St Helena, California. His bachelor's degree in viticulture and enology from the University of California, Davis, combines classic microbiology, biochemistry and food science with sensory classes for the tailored enology half of the degree. These allow him to analyse his winery's varieties through each stage of production. Bjorkman says that a large part of his job is to ask each day, "What is growing in this wine?", and to find out if it is advantageous or a troublemaker. A Strange Brew Larger enterprises such as Miller Brewing Company in Milwaukee, Wisconsin, and E. & J. Gallo Winery in Modesto, California, have their own research and development departments that employ scientists at bachelor's, master's and PhD levels. Tom Pugh, director of enology research at Gallo, says that the company does both applied research, to optimize fermentation conditions, and basic research that strives to identify the microbial characteristics that affect a wine's feel, flavour and aroma. His research group looks at the way genetic differences between strains affect metabolism throughout the fermentation. Gallo also takes on microbiologists with degrees slanted towards wine-making as winemakers, he says, overseeing a wine's production from grape to bottle. Similarly, David Ryder, chief brewmaster at Miller, says that microbiologists may want to become brewmasters at individual breweries. There's also a small group of microbiologists doing R&D at the corporate headquarters in Milwaukee. These, says Ryder, track yeast physiology and use DNA microarrays to find out if the yeast strains are 'happy'. Happy yeast are healthy yeast that grow fast in a variety of environments and efficiently convert sugars to alcohol. Like Pugh, Ryder stresses that he looks for employees who are passionate about both the science and the product. Top Cheese Whizzes And what goes better after a fine wine than a nice hard cheese to cleanse the palate? Makers of cheese (and yoghurt) face an equally complex, though different, fermentation process. Microbiologists working on the dairy side of the fence also hunt for the microbes that bring the desired texture, flavour and aroma to fermented milk. Understanding how microbes will react under the different stresses of the cheese-making process is a key component not necessarily gleaned from working in an academic microbiology lab. This is where the practical side of a food-science degree comes in, says Kayla Polzin, a senior scientist and microbiologist at the Land O'Lakes dairy company's labs in Arden Hills, Minnesota. Polzin says that, in her job, a deep understanding of microbiology and metabolism is just as important as a broad understanding of the many microbes that can crop up in dairy cultures. She is one of two PhD-level microbiologists at Land O'Lakes, but many more dairy-food microbiologists are employed by the companies that provide the starter cultures. As well as developing cultures that are resistant to bacteriophages and viruses, or that work more efficiently, these scientists track down microbe characteristics responsible for specific flavours and textures sought by their customers. "Pizza companies can be very, very demanding as to what the cheese looks like on their pizza. It has to brown, stretch and melt just the right way," says Dennis Romero, a molecular microbiologist at the starter-culture company Rhodia in Madison, Wisconsin. He is one of about two dozen research scientists at Rhodia, but he notes that most of its competitors are based in Europe and are ten times as large. Romero stresses that researchers at starter-culture companies must do science that is practical at all times. The best way for microbiology students to gain this applied viewpoint of research, he says, is to work on an industry-funded collaboration. Starter-culture companies tend to 'outsource' the most basic research by setting up joint projects with an academic lab. Christian Hansen, a food-ingredient company based in Hoersholm, Denmark, employs about 80 scientists in its central research laboratories. Chief science officer Peter Olesen says that — because the company deals in colours, flavours and enzymes as well as cultures — it employs protein chemists, chemical engineers and scientists with medical and veterinary degrees (see Gut feeling for health). One of their recent basic microbiology research projects developed what Olesen calls a "revolution in lactic acid bacteria" — a strain engineered so that it can grow under aerobic conditions (with oxygen), resulting in cultures that ferment faster. All areas of food microbiology value knowledge of food processes and fermentations more than a narrow focus on molecular mechanisms of organisms. For that reason, most of the opportunities in this field do not require postdoctoral training. Industry-related experience is rated highly — whether an internship, working on a wine harvest or an industrial collaboration. Reif says that, all beer-drinking benefits aside, he gets job satisfaction from the critical thinking required in troubleshooting: "When problems come up it's an opportunity to use your head." Top Story from news@nature.com: http://news.nature.com//news/2004/040510/nj6988-224a.html

How viable are probiotic bacteria in yoghurts?

Foodinfo Online FSTA Reports -->8 October 2004

Probiotic bacteria such as Lactobacillus acidophilus and Bifidobacterium spp. have numerous health benefits, which have lead to their incorporation into dairy foods such as yoghurts. However, for probiotic bacteria to be therapeutically effective, a minimum viable count of 106 to 107 cfu/g of these bacteria is necessary.

Standards introduced by food authorities worldwide have encouraged manufacturers to guarantee adequate viability of these bacteria throughout product shelf life. However, several recent market surveys have indicated a steady decline in counts of L. acidophilus and Bifidobacterium spp. throughout the shelf life of probiotic yoghurts. Oxygen toxicity is considered a significant factor in influencing the viability of these probiotic bacteria in yoghurts.

The agitation and mixing steps during manufacture can incorporate high amounts of oxygen into the product. In addition, during storage, oxygen can diffuse into yoghurt through the high-impact polystyrene (HIPS) packaging. As a result, polystyrene-based gas barrier packaging and active packaging films have been developed. A study by Talwalkar et al.1 investigated the effects of packaging materials on the dissolved oxygen content and survival of the probiotic bacteria, L. acidophilus and Bifidobacterium spp., in yoghurt stored at 4--6°C for 42 days. The yoghurts were packaged in oxygen-permeable HIPS, oxygen-barrier material (Nupak) and Nupak with an oxygen-scavenging film (Zero2).

No significant decreases were observed in bacterial viability of any of the yoghurts tested, suggesting that oxygen alone is not a significant factor in causing poor viability of these bacteria in yoghurt.

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1 Talwalkar, A; Miller, CW; Kailasapathy, K; Nguyen, MH (2004).

Effect of packaging materials and dissolved oxygen on the survival of probiotic bacteria in yoghurt. International Journal of Food Science and Technology 39 (6) 605-611.

Página WEB de referencia

AN: 2004-09-Pl1566
TI:
Effect of packaging materials and dissolved oxygen on the survival of probiotic bacteria in yoghurt.
DA:
7-May-2004
DT:
Journal Article
AU:
Talwalkar, A.; Miller, C. W.; Kailasapathy, K.; Nguyen, M. H.
PY:
2004
AD:
Correspondence (Reprint) address, K. Kailasapathy, Cent. for Advanced Food Res., UWS, Locked Bag 1797, NSW 1797, Australia.
SO:
International Journal of Food Science & Technology 39 (6) 605–611
RF:
22 ref.
LA:
English
SN:
0950-5423
AB:
Oxygen toxicity has been suggested to be a significant factor influencing the viability of probiotic bacteria, such as Lactobacillus acidophilus and Bifidobacterium spp., in yoghurts. Along with manufacturing steps (particularly agitation and mixing), which incorporate high amounts of oxygen into the yoghurt, the packaging materials used may also affect the level of oxygen diffusing into the product. This study investigated the effects of packaging materials having different oxygen permeabilities on the levels of dissolved oxygen and survival of the probiotic bacteria in yoghurt. Oxygen adapted and non-oxygen adapted strains of L. acidophilus and Bifidobacterium spp. were incorporated in yoghurts, which were packaged in oxygen permeable high-impact polystyrene (HIPS), oxygen-barrier material (NupakTM) and NupakTM with an oxygen scavenging film (Zero2TM). Results showed that, during storage, the level of dissolved oxygen increased steadily in HIPS packaged yoghurt, but remained low in yoghurts packaged in NupakTM and Zero2TM. In all yoghurts, no significant decreases were observed in the viability of either oxygen adapted or non-oxygen adapted cells of L. acidophilus and Bifidobacterium spp. It is suggested that, although the level of dissolved oxygen in yoghurt can be influenced by the type of packaging material, it may not affect the survival of probiotic bacteria in yoghurts.
SC:
Milk and dairy products
KW:
BIFIDOBACTERIUM; INHIBITION; LACTOBACILLUS; LACTOBACILLUS ACIDOPHILUS; MICROORGANISMS; OXYGEN; PACKAGING MATERIALS; PERMEABILITY; PROBIOTIC BACTERIA; YOGHURT

Yoghurt - part two

FoodInfo Online Features -->6 August 2004

In the second part of his review,

Ernest Mann concentrates on the scientific aspects of yoghurt production.

The second part of this review concentrates predominantly on more scientific aspects of yoghurt production. A recent study in Australia (30) looked at seasonal variations in physical characteristics of both set and stirred yoghurts. The effects of variations in solids-not-fat contents (10–14 per cent) on gel strengths, whey drainage and other properties were examined. It was found that standardisation of total solids content with dried skim milk was not sufficient to produce yoghurts with consistent physical characteristics throughout a season.

Another group of Australian scientists (31) studied the effects of fortification of milk with two per cent of either whey powder (WP), skim milk powder (SMP) or whey protein concentrate (WPC) on composition, pH, firmness, viscosity, syneresis and microstructure of yoghurt. The results indicated that WP and SMP supplementation reduced viscosity and firmness, while WPC supplementation increased these values.

Another report from the same laboratory (32) looks at the effects of adding proteolytic strains of Lactobacillus delbrueckii subspecies bulgaricus to commercial ABT starter cultures or a mixed starter on the texture of yoghurt and the effects on the survival of starter bacteria. Amongst other things, the results indicated that the viability of probiotic bacteria improved in yoghurt made with the mixed starter with the added Lactobacillus delbrueckii strain.

In the USA, researchers have reported (33) on the effects of stirred yoghurt thickness, flavour and colour on the sensory perceptions of 120 school children. A German study (34a) has reported on the enzymatic cross-linking of milk proteins on the functional properties of set-type yoghurt, involving transglutaminase treatment of milk prior to fermentation with a starter culture. The results indicated that, while the aroma and consistency of yoghurts from enzyme-treated milk were less 'yoghurt specific', they were more creamy than those produced from untreated milk. This suggests that a transglutaminase treatment may simulate fat in fermented products.

The effect of fermentation temperature between 37 and 46°C on the formation and rheology of yoghurt has been studied in the UK (34b) using reconstituted skim milk preheated and fermented with either a ropy or non-ropy starter culture. A study in Spain (35) investigated the effects of manothermosonication treatment (MTS) of milk on the rheological properties of yoghurt made from MTS milk. The results indicated that MTS treatment, which is the simultaneous application of heat and ultrasound under moderate pressure, could result in yoghurts with rheological properties superior to those found in control yoghurts made from untreated milk.

Studies in the USA (36) have compared yield stress, microstructure, shear values and water holding capacity (WHC) in full fat set yoghurt prepared from fortified milk. This had been subjected to either thermal processing (85°C for 35 minutes) or high hydrostatic pressure processing (193 or 676 MPa for 5 or 30 minutes). Amongst other things, the results indicated that yoghurts made from milk treated at 193 MPa and untreated milk (control) exhibited low yield stress, low WHC and large clusters of coalesced micelles. In a more general article (37) on the effects of different ingredients and manufacturing processes on the texture of set, stirred and probiotic yoghurts, the costs of various commercially available yoghurt texturisers are also considered.

An Austrian study (38) on the influence of starter culture on the relationship between dry matter content and the physical properties of stirred yoghurt formed part of an EU project that aimed to develop a standard procedure for the laboratory-scale production of set-type and stirred yoghurt. The first of three reports from the USA (39) was concerned with the evaluation of the textural properties of two types of yoghurt (light or blended), using a trained sensory analysis panel and an instrumental compression/penetration test in combination with a novel data analysis method.

The second report (40) presented a comparison of the effects of aspartame and sucrose on the microstructure of yoghurt. The third study from the USA (41) reports the development of a method for manufacturing a milk protein powder rich in both casein and whey proteins, as well as being free of lactose. The powder was used successfully in the manufacture of non-fat yoghurts without the addition of gelling and stabilising agents. Russian workers (42) have developed a stabiliser for heat-treated yoghurt which is based on a modified maize starch and a texturising agent containing guar gum, xanthan gum and pectin.

The synthesis and utilisation of folate by 32 strains of yoghurt starter cultures and probiotic cultures was examined by Australian scientists (43). Brazilian workers (44) conducted a study of composition and sensory quality of whole stirred yoghurt made from milks with somatic cell counts ranging from below 400 000 to above 800 000 cells per ml. Cell counts higher than 400 000 per ml had a detrimental effect on the sensory quality of yoghurt. A report from Egypt (45) looked at the utilisation of laboratory produced xanthan gum in the manufacture of natural and soya yoghurt. It concluded that the highest sensory scores were obtained when xanthan gum additions of 0.01 and 0.005 per cent were employed with the two types of yoghurt respectively. Acetaldehyde is regarded as a major indicator of flavour in yoghurt.

In a recent Turkish study (46), the effects on acetaldehyde levels in yoghurts, of the use of viscous and non-viscous cultures plus amino acids, treatment with ß-galactosidase and the use of heat-shocked cultures were investigated. The results indicated that the highest acetaldehyde levels in yoghurt were obtained when non-viscous starter cultures were used alone.

The effects of processing and refrigerated storage on the distribution and stability of aflatoxin M1 in yoghurt, artificially contaminated with different levels of the toxin, has been investigated in Greece (47). Spanish scientists (48) have developed a new multiresidue method for determining trace amounts of organochlorine pesticides and polychlorinated biphenyls in yoghurt, the method proving quick, accurate and repeatable. In another Spanish study (49) the survival of E. coli 0157:H7 in yoghurt at different storage temperatures (4–22°C) was compared with that of a non-pathogenic strain of E. coli. The results showed that counts of the pathogenic strain during storage at 4 and 8°C were higher than those of the non-pathogenic strain. Turkish scientists (50) have studied the microbiological properties of labneh concentrated yoghurt stored under oil at room temperature and refrigerator temperature.

An Italian study looked into the occurrence of yeast contamination in yoghurt (51). Examination of 40 samples of fruit yoghurt with obvious signs of blowing, produced by two different firms, revealed the presence of three different species of yeast - Pichia fermentans, Torulospora delbrueckii and Claviospora lusitaniae. This is believed to be the first recorded time that these yeast species have been associated with yoghurt blowing. Several reports are associated with health-nutritional aspects of yoghurt. The first of these, from Argentina (52), gives the results of trials where mice were fed with different doses of yoghurt as part of a milk re-nutrition diet on recovery of the intestinal barrier and mucosal immune function. The results suggest that, although they were obtained with an animal model, the consumption of yoghurt by malnourished children may accelerate the restoration of gut function following intestinal malfunction.

In trials in the USA (53), the addition of three servings of yoghurt daily to the diet of 29 postmenopausal women with habitually low calcium intakes resulted in a significant reduction in the urinary excretion of N-telepeptide, a marker for bone resorption. The nutrients added with the yoghurt greatly improved overall diet quality as compared with a nutrient-poor snack diet. Somewhat inconclusive results were obtained in Italy (54) during a study of the effects of yoghurt feeding as a dietary supplement for 12 healthy, elderly people. However, significantly lower faecal clostridia counts were obtained in comparison with the controls, which suggests a beneficial effect of yoghurt feeding.

Polish scientists (55) have reported on the effects of certain factors on the properties of set-type ewes' milk yoghurt. The factors studied included different starter cultures and fat contents and their effects on overall quality, rheological and sensory properties when fresh and following storage for up to 14 days at 5°C. The same team (56) has also studied the sensory quality and physico-chemical properties of ewes' milk yoghurt made from milk with fat contents ranging from zero to six per cent. Quality parameters varied with fat content and storage time, and optimum results for sensory quality were obtained for non-fat yoghurts.

A Greek study (57) looked at the effects of frozen storage of ewes' milk on the microbiological and physicochemical properties, as well as its yoghurt-making ability. The overall quality of yoghurt made from frozen stored ewes' milk was as high as that made from fresh milk. Another Greek report (58) described a method for the detection of bovine milk in ovine yoghurt by electrophoresis of para-k-casein.

Finally, it was concluded from a study carried out in Croatia (59), that goats' milk yoghurt fermented with probiotic bacteria and fortified with 1.5 per cent inulin shows good potential as a functional food.

References

(30) Cheng, L J et al (2002) Austral J Dairy Technol 57(3)187

(31) Bhullar, Y S et al (2002) Milchwissenschaft 57(6)328

(32) Shihata, A et al (2002) Internat Dairy J 12(9)765

(33) Brennan, E M et al (2002) J Food Sci 67(7)2785

(34a) Lorenzen, P C et al (2002) Internat J Dairy Technol 55(3)152

(34b) Haque, A et al (2001) Food Hydrocolloids 15(4/6)593

(35) Vercet, A et al (2002) J Agric & Food Chem 50(21)6165

(36) Harte, F et al (2002) J Food Sci 67(6)2245

(37) Meester, R 2002) Food Ingred & Anal Internat 24(1)12

(38) Jaros, D et al (2002) Milschwissenschaft 57(8)447

(39) Carson, K et al (2002) J Food Sci 67(3)1224

(40) Haque, Z Z et al (2002) Food Sci & Technol Res 8(1)21

(41) Mistry, V V (2002) Lait 82(4)515

(42) Dunchenko, N I et al (2002) Moloch Promyshl No 10 p27

(43) Crittenden, R G et al (2002) Internat J Food Microbiol 80(3)217

(44) Oliveira, C A F et al (2002) Austral J Dairy Technol 57(3)192

(45) El-Sayed, E M et al (2002) Europ Food Res & Technol 215(4)298

(46) Ozer, B et al (2002) Internat J Dairy Technol 55(4)166

(47) Govaris, A et al (2002) Food Additives and Contam 19(11)1043

(48) Yague, C et al (2002) J AOAC Internat 85(5)1181

(49) Bachrouri, M et al (2002) J Food Sci 67(5)1899

(50) Say, D et al (2002) Milchwissenschaft 57(9/10)528

(51) Vallone, L et al (2001) Industrie Alimentari 40(407)1001

(52) Gauffin-Cano, P et al (2002) J Dairy Res 69(2)303

(53) Heaney, R P et al (2002) J Amer Dietet Ass 102(11)1672

(54) Canzi, E et al (2002) Lait 82(6)713

(55) Bonczar, G et al (2002) Food Chem 79(1)85

(56) Bonczar, G et al (2002) Zywnosc 9(1)109

(57) Katsiari, M C et al (2002) Food Chem 77(4)413

(58) Kaminarides, S E et al (2002) Food Chem 78(1)53

(59) Bozanic, R et al (2002) Mljekarstvo 52(2)93

This review was originally published in the August 2003 edition of Dairy Industries International

Fuente WEB utilizado para el artículo

Bacteria banish fowl bugs

by Helen R. Pilchernews@nature.com
Publicado en: BioEd Online (http://www.BioEdOnline.org)

A probiotic diet makes chickens healthier and safer to eat.

Chickens could benefit from a daily dose of friendly bacteria, researchers say.

Probiotic bugs can destroy food-poisoning bacteria inside poultry, making the birds healthier and safer to eat.
The benefits to human health of probiotics are well known. The bacteria, found in yoghurt, are thought to out-compete other gut bacteria, including those that cause food poisoning.
Probiotics are thought to have similar benefits in animals, and are already included in some agricultural feeds. But such feeds contain a slew of bacteria, says Arjan Narbad from the Institute of Food Research, Norwich, so their effects are uncertain.

Narbad and colleagues tested a single probiotic dose in the lab. The good bacterium Lactobacillus johnsonii ousted the harmful Clostridium perfringens from chicks' guts, they report in Letters in Applied Microbiology 1 .

"We have used a single strain and shown that it can be targeted to eliminate a specific pathogen," says Narbad.
Clostridium can flare up chickens, making them sickly and thin. It's also one of the top five bacterial causes of food poisoning in humans. In the United Kingdom, the bug poisons about 200 people through undercooked chicken each year.

Health food
"We also have preliminary data suggesting that Lactobacillus may be effective against Campylobacter," Narbad says. This is a much nastier bug, causing about 63,000 UK cases of food poisoning each year. Lactobacillus also has a weak effect against the sometimes-deadly gut bacterium E. coli.

Probiotics could have other benefits too. They may also increase chicken growth rate, for example, says microbiologist Anne McCartney from the University of Reading.
The probiotic bacteria are easy to give to animals, as they can be put in animal feed or drinking water, says Narbad.
They should also help to reduce the use of antibiotics in animals. Farmers are being encouraged to cut their use of antibiotics to reduce the chance of bacteria evolving resistance.
The team is now seeking other probiotic species to help combat different pathogens. They plan to test the treatments on farms to see if results are as good as they are in the lab.

1 La Ragione, R.M., La Narbad, M.J., Gasson, M.J. & Woodward, M.J.. Letters in Applied Microbiology, 28, 197 - 205, (2004).

Microbiology: Gut reaction

Nature 427, 284 - 286 (22 January 2004); doi:10.1038/427284a

Consumers are stocking up on live yoghurts and fermented drinks that claim to improve health. But is there any science behind the marketing of these 'probiotic' products? Alison Abbott investigates.

Glenn Gibson's wife prefers him to tell dinner-party guests that he works as a painter and decorator. That's understandable, because if he talks about his real job as a researcher of gut bacteria at theUniversity of Reading, UK, the conversation all too easily turns to the source of his research material — human excrement.

For better or worse, faeces provide the best window into the microbial life of the human gut, a subject that is attracting more funding nowthan ever before. Partly in reaction to commercial claims that the bacteria in some yoghurts and other 'probiotic' products can boost health, the European Union (EU) has invested more than 15 million (US$19 million) since 1995 to research this poorly explored frontier.

As a result, a growing number of microbiologists are taking an interest in the ecology of the human gut. They are adapting tools previously developed for the study of microbes in oceans and soil to answer a range of questions. What lives in our gut? Do some natural gut microbes predispose us to diseases such as colon cancer? And can we change the make-up of our intestinal residents to improve our health?
Gibson has even built a collection of artificial guts to study our internal microbial ecology under controlled laboratory conditions (see 'Roboguts').

Hidden world
The average human intestine contains about 1.2 kilograms of bacteria plus a smattering of yeasts. So far, few of these microbes have been characterized or even identified. But this dearth of information hasn't kept companies from promoting the health value of probiotics, which contain living bacteria, and prebiotics — nutrients designed to boost populations of beneficial bacteria already living in the gut.

Probiotic dietary supplements are available in just about any form imaginable, from tubes of liquid to capsules. Some yoghurts and fermented milk drinks also promote their living contents. A typical online shop claims that its probiotic products can "strengthen the immune system, reverse the negative effects on the digestive tract of infections, antibiotics, alcohol ... treat symptoms of irritable bowel disease" and more.

Worldwide, the pro- and prebiotics market is now worth about US$6 billion.
But so far, the science behind these commercial boasts is rather limited. "There are a lot of bogus claims and vested interests," says Michael Blaut, head of gastrointestinal microbiology at the German Institute of Human Nutrition in Potsdam, and one of the researchers who helped to convince the EU to fund probiotics research.

Although some clinical trials of probiotics have suggested a benefit, Gibson adds, few of these have been sufficiently rigorous. And even when probiotics seem to work, he says, we know too little about the normal gut ecosystem to understand why.

Soon after it was established, the EU-funded network, which includes scientists from 16 countries, discovered that the gut ecosystem is much more diverse than previously thought. Microbiologists knew that their traditional techniques of isolating and cultivating individual microorganisms were not pulling out all of the species that we live with. Many gut bacteria are notoriously difficult to grow in culture — largely because they depend on the presence of other bacterial species. But few scientists had anticipated just how diverse the ecosystem would turn out to be.

To begin to quantify the diversity, gut researchers borrowed a method from soil and ocean microbiologists that relies on comparisons of the gene for a portion of the ribosome — the cellular machine that manufactures new proteins — known as 16S. The ribosome is so fundamental to the workings of the cell that it has changed little during evolution. That makes it easy to extract the 16S genes from all microorganisms in a single faecal sample using the DNA-amplifying polymerase chain reaction. By looking for subtle differences between the sequences of these genes, microbiologists can gauge the number of different species present in the sample.

One surprise was that no two people have quite the same complement of bacteria. In unpublished work, molecular biologist Joël Doré of the INRA, the French agricultural research agency in Jouy-en-Josas, near Paris, has so far analysed the faeces of more than a dozen healthy adults and found the contents of each to be quite different. Although thousands of microbes can live in the gut, each person has only about 100 different species. "There is remarkably little overlap in the gut bacterial species between individuals," he says.

This is partly because of the haphazard way in which the bacteria arrive. Our guts start off in the womb completely sterile, but they are rapidly colonized with vaginal and faecal bacteria during birth.
Microorganisms from food and other environmental sources contribute to the mix during the first months of life. By the age of two at the latest, the average human gut hosts its full complement of microbial species, mixed and matched from a dozen or so dominant groups of bacteria and a longer list of rarer bacteria and yeasts.

From this point on, little changes until old age — a person's microbial complement seems to remain stable throughout adulthood, Doré says.
But he has found that the faeces of people over 60 contain a much larger number of different bacteria than younger people. He suspects that the weakening barrier to new species may help explain why the elderly are more susceptible to gut infections and certain forms of cancer.

Colonic closed shop
The basis of the microbial stability that persists throughout most of our lives is still poorly understood, but is probably related to nutrient supply. By the time an infant is two years old, resident bacteria have monopolized every source of nutrients in the gut. They have also become interdependent, supplying nutrients to each other — one cell's waste is another's food. With all the nutrients accounted for, newcomers may find it hard to gain a toehold.

The stability of this ecosystem benefits not only the gut microbes, but also the human host. It prevents pathogenic bacteria, such as the various species of Salmonella that cause food poisoning, from taking up long-term residence. But it also means that probiotics cannot permanently change the make-up of the gut — they must be taken daily to have any effect.

What are the possible benefits? Individual species of bacteria are informally classified on a sliding scale of 'goodness' and 'badness'.
Collectively, gut bacteria aid digestion by breaking down tough fibres, enzymes and other proteins. In addition, 'good' bacteria, such as species of Lactobacillus, Bifidobacterium and Eubacterium, are involved in fermentation reactions that produce organic acids that can be absorbed into the body and used as an energy source. 'Bad' bacteria, such as some members of the genus Clostridium, generate as by-products compounds including nitrosamines and cresols, which are possible carcinogens.

Commercial probiotic strains are, of course, 'good' bacteria. The probiotic milk products, yoghurts and capsules on the market generally contain Lactobacillus and Bifidobacterium. Most studies of their efficacy have been poorly controlled and have produced contradictory results, says Gibson. But a handful of well-designed clinical trials indicates that some such bacteria may help ameliorate diarrhoea1-3 and some types
of inflammatory bowel disease4, 5.
Less well documented are the claims for beneficial stimulation of the immune system. The gut, with its massive blood supply, is the immune system's primary contact with the outside world, and gut bacteria seem to play a role in teaching the immune system to differentiate between dangerous invaders and non-hostile challenges.
Although some studies suggest that probiotics can affect features of the immune system, few have shown that these changes are beneficial to health.
A notable exception is a long-term study supported by the Finnish Academy of Sciences, in which pregnant women from families prone to allergies ate Lactobacillus rhamnosus daily. After delivery, the bacteria were given daily to the babies for the first six months of their lives.

The treated infants were much less prone to allergic reactions such as eczema than controls who did not get the bacteria6, 7. Erika Isolauri, an immunologist at th University of Turku who led the study, is now trying to determine how the treatment works. She suspects that the probiotics shift the balance between pro- and anti-inflammatory factors in the developing gut.

Friend or foe?
Other studies to assess the health benefits of probiotics are under way. With funding from the EU, for example, Doré is setting out to test a combination of Bifidobacterium animalis and a type of prebiotic sugar known as FOS on the gut ecosystems of young and old people in France, Germany, Sweden and Italy. "We are testing faecal samples to see whether the level of Bifidobacterium really does rise with this treatment, as would be expected," he says. His team will also assess how the levels of toxic and potentially carcinogenic compounds in the gut rise and fall with treatment by exposing cell cultures to extracts from the subjects' faeces. The researchers hope to determine whether suppressing 'bad' bacteria with pre- and probiotics might protect against colon cancer.

Francisco Guarner, a gastroenterologist at the Vall d'Hebron Hospital in Barcelona, Spain, is helping to organize an EU-backed clinical study involving 360 patients chronically suffering from one of two types of inflammatory bowel disease — ulcerative colitis or Crohn's disease — at centres in Ireland, Spain, Finland and France. The patients, all in remission, receive either Lactobacillus salivarius or Bifidobacterium infantis, two species that reduce gut inflammation in lab animals. The researchers then test the patients' saliva for marker molecules associated with inflammation. "Animal studies show that inflammatory disorders of the bowel may be helped by making the gut microbes less aggressive," says Guarner. He hopes that the probiotics will extend the patients' remission so they can reduce their reliance on immunosuppressive drugs, which have severe side effects.
Outside the EU network, Gibson is running trials at six British centres using Lactobacillus plantarum together with a second type of prebiotic sugar called GOS in various kinds of inflammatory bowel disease.

Gibson's hypothesis is that the yeast Candida causes the symptoms, and he hopes that the probiotics will outcompete its growth. In a separate study, he is testing his hypothesis that certain 'bad' bacteria contribute to ulcerative colitis by generating toxic sulphur compounds such hydrogen sulphide, which smells of rotten eggs. He is giving patients FOS and GOS to stimulate the growth of competing 'good' bacteria to see whether this eases the symptoms.

Microbes on trial
In the next few years, these and other studies will help to determine how beneficial probiotics actually are. In the meantime, other salient questions are being addressed. Can we, for example, assume that probiotics are safe? One potential problem is that many probiotic strains have genes that allow them to resist antibiotics, which they might pass on to pathogenic bacteria. To address these concerns, Herman Goossens of the University of Antwerp in Belgium has acquired more than 200 commercial probiotic strains — the world's largest collection. He is systematically analysing them for their potential to transfer antibiotic resistance genes, and is also testing for any direct toxic effects that they may have.

Some experts believe that another important step will be to read the genomic sequence of every species of microbe that can colonize the human gut. They argue that a complete genomic databank would make it much easier to select species for specific probiotic effects.

To that end, the Defense Advanced Research Projects Agency, a research arm of the US military, is sponsoring a project to read all the genomes in the gut ecosystem with the same 'shotgun' method used for the privately funded effort to sequence the human genome. This approach avoids the need to separate out individual organisms.

Instead, fragments of all the genomes are read off together. Computer algorithms then reassemble the fragments on the basis of overlapping sequences into complete genomes. "It's possible to conceive of doing this because the cost of sequencing has come right down," says Claire Fraser, director of The Institute for Genomic Research in Rockville, Maryland, where the work will be done.

Even if probiotics and prebiotics prove to have only modest health benefits, some scientists are considering the possibility of souping them up with genetic engineering. Many proponents of probiotics reject this idea, saying that it would be too hard to convince the public to eat live, genetically modified bacteria. But among the traits that would be useful to engineer are the ability to survive the acid environment of the stomach, a bit of 'stickiness' to help bacteria adhere to the gut lining, and so take residence for longer, and the ability to produce organic acids. Bacteria might even be engineered to deliver drugs, vitamins or vaccines8, 9.

There is already evidence that some bacteria can serve as efficient delivery vehicles. For example, Lothar Steidler of Ghent University in Belgium and his colleagues have shown that Lactococcus lactis genetically modified to secrete the anti -inflammatory molecule interleukin-10 can reduce colitis in mice10. A version for humans has also been developed that includes safety features to prevent the escape of the inserted gene into the environment11. A small clinical trial of this microbe is planned in Amsterdam, marking the first use of a genetically engineered bacterium as a therapeutic agent.

Unfortunately for the scientists studying probiotics, the only way forward is to delve into more human waste. Doré says he recently felt a pang of regret over that fact on a trip to visit some oceanographer friends in Marseille. "I looked out into the Mediterranean and thought:
'I'm obviously working on the wrong ecosystem'," he sighs. But the scientific challenges presented by gut bacteria are interesting enough to keep him going, he says. "It makes up for the unpleasantness."

ALISON ABBOTT
Alison Abbott is Nature's senior European correspondent.
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