CAST Probiotics Issue Paper FINAL144

8/12/2019 CAST Probiotics Issue Paper FINAL144

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Number 36October 2007

Probiotics:Their Potential to Impact Human Health

A BSTRACTProbioticslive microorganisms

that when administered in adequateamounts confer a health benet onthe hosthave been studied for bothhuman and animal applications, andworldwide research on this topic hasaccelerated in recent years. This pa-per reviews the literature on probiot-ics, describes how probiotics work inhuman ecosystems, and outlines theimpact of probiotics on human healthand disease. The paper also addressessafety issues of probiotic use, sug-gests future developments in the eldof probiotics, and provides researchand policy recommendations. Productconsiderations and potential future de-velopments regarding probiotics alsoare discussed. The authors conclude

that controlled human studies haverevealed a diverse range of health ben-ets from consumption of probiotics,due largely to their impact on immunefunction or on microbes colonizingthe body. Additional, well-designedand properly controlled human andmechanistic studies with probioticswill advance the essential understand-ing of active principles, mechanisms

of action, and degree of effects thatcan be realized by specic consumergroups. Recommendations include es-tablishment of a standard of identityfor the term probiotic, adoption ofthird-party verication of label claims,use of probiotics selectively in clinicalconditions, and use of science-basedassessment of the benefits and risksof genetically engineered probioticmicrobes.

INTRODUCTIONProbiotics 1 are live microorgan-

isms that when administered in ad-equate amounts confer a health beneton the host (UNFAO/WHO 2001).Probiotics commonly are isolated fromhuman and animal intestinal tracts.Dead bacteria, products derived frombacteria, or end products of bacte-rial growth also may impart certainbenets, but these derivatives are notconsidered to be probiotics becausethey are not alive when administered.Native bacteria are not probiotics untilthe bacteria are isolated, puried, andproved to have a health benet whenadministered. Probiotics have been

This material is based upon work supported by the United States Department of Agriculture under Grant No. 2005-38902-02319, Grant No. 2006-38902-03539,and Grant No. 2007-31100-06019/ISU Project No. 413-40-02. Any opinions, ndings, conclusions, or recommendations expressed in this publication are those

of the author(s) and do not necessarily reect the view of the U.S. Department of Agriculture or Iowa State University.

Scanning electron micrographs (clockwise from upper left) of Lactobacillus paracasei , Lactobacillus plantarum , Strepto

coccus thermophilus , Bidobacterium infantis, Bidobacterium longum , and Bidobacterium breve . (Photos courtesyof Prof. Lorenzo Morelli, Instituto di Microbiologia, Piacenza, Italy.)

1 Italicized terms (except genus and speciesnames) are dened in the Glossary.


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CAST Issue Paper 36 Task Force Members

Authors

Reviewers

CAST Liaison

Mary Ellen Sanders (Chair) , Dairyand Food Culture Technologies,Centennial, ColoradoGlenn Gibson , Food MicrobialSciences Unit, School of Food Bio-sciences, The University of Reading,Reading, United KingdomHarsharnjit S. Gill , Primary Indus-tries Research Victoria, Departmentof Primary Industries, Werribee,Victoria, Australia

studied for both human and animal ap-plications. Key microbial species usedas human probiotics are listed in Table 1.

Worldwide, a diverse array of pro-biotic products is on the market. Yogurtis perhaps the most common probi-otic-carrying food, but the market hasexpanded beyond yogurt. Cheese, fer-mented and unfermented milks, juices,smoothies, cereal, nutrition bars, andinfant/toddler formula all are food ve-hicles for probiotic delivery. In addi-tion to being sold as foods, probioticsare sold as dietary supplements, medicalfoods, and drugs (although there are noprobiotics currently sold as drugs in the

United States). Often these productsare composed of concentrated, dried mi-crobes packaged into capsules, tablets,or sachets. This format is convenientfor the delivery of large numbers of mi-crobes that, if manufactured and storedproperly, can be quite stable even atroom temperature.

What motivates people to chooseone format over another has much todo with personal preference, productavailability in different geographic re-gions, or individual needs, althoughfewer types of probiotic foods are avail-able in the United States than in partsof Europe or Asia. Product formula-tion may impact greatly how a probi-otic survives during product storage andif it reaches the target site in the bodyphysiologically capable of exerting itsbenets. Furthermore, additive or evensynergistic activities of componentsin addition to probiotics in the prod-uct may enhance the products health-promoting properties. In the end, eachspecic product must be judged based

on its ability to deliver health benetsthrough a well-formulated probioticcontent.

Researchers have studied many pos-sible benets of probiotics (Table 2),and the pace of research in recent yearshas accelerated. More than four timesthe number of human clinical trials onprobiotics were published during the pe-riod from 2001 to 2005 than from 1996to 2000. To the uninitiated, the list ofbenets from probiotics seems too di-verse to be possible. But once it is un-derstood that probiotics can impact anycolonized regions of the body and thatbacteria have the potential to inuence

the body bothlocally

andsystemically

,the scope of benets can be appreciated.The authors of this Issue Paper have

reviewed the literature on probiotics en-compassing both U.S. and internation-al research reports. In this paper, theauthors describe the characteristics ofprobiotics, discuss what is known aboutthe microbes that colonize humans, andoutline the impacts of probiotics onhuman health and disease for specicconditions. The paper also addressesthe regulatory status of probiotic foodsin the United States, suggests futuredevelopments in the eld of probiotics,and provides research and policy rec-ommendations. A detailed discussion ofHow Probiotics Work can be found inAppendix 1.

C HARACTERISTICS OF P ROBIOTICS

Certain physiological characteristicsmay be important for probiotics target-ed toward particular applications. For

Francisco Guarner , DigestiveSystem Research Unit, UniversityHospital Vall dHebron, Barcelona,Spain

Stanley E. Gilliland, Food andMicrobiology Products Center,

Oklahoma State University,StillwaterTodd R. Klaenhammer , Depart-ment of Food Science, North Caro-lina State University, Raleigh

Gregor Reid , Canadian R&DCentre for Probiotics, LawsonHealth Research Institute, London,Ontario, CanadaGerald W. Tannock , Departmentof Microbiology and Immunology,University of Otago, Dunedin,New Zealand

Harold Swaisgood , ProfessorEmeritus, North Carolina StateUniversity, Raleigh

Table 1. Key genera and species of microbesstudied and used as probiotics

Genus Species

Lactobacillus acidophilus brevis delbrueckii a

fermentum gasseri johnsonii paracasei plantarum reuteri rhamnosus salivarius

Bidobacterium adolescentis animalis b

bidum

breve infantis longum

Streptococcus thermophilus salivarius

Enterococcus faecium

Escherichia coli

Bacillus coagulans c

clausii

Saccharomyces cerevisiae d

aL. delbrueckii subsp. bulgaricus is typicallyused as a starter culture for yogurt.

bA subspecies of B. animalis is B. animalis subsp. lactis . Several commercial probiotic strainsare members of this subspecies and commonlyare referred to as B. lactis , although the correctdesignation is B. animalis subsp. lactis .cSome manufacturers market a strain theycall Lactobacillus sporogenes. The microbemarketed under this name is likely a Bacilluscoagulans , not related to the true Lactobacillus genus (Sanders, Morelli, and Bush 2001).dA prominent probiotic strain marketed asSaccharomyces boulardii (not a validspecies name) is a strain of Saccharomycescerevisiae.


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wanted microbes from entering theblood stream, colonization resis-tance , and digestion are importantfunctions of colonizing microbes.

Regions of the Human BodyColonized by Microbes

Bacteria are prevalent in several re-

gions of the body, including the mouth,nose, pharynx, intestinal tract, vaginaltract, and skin (Willis et al. 1999) (SeeFigure 1). The stomach is not heav-ily colonized because of its low pH,and typically harbors up to 10 3 colony

forming units (CFU) per gram of con-tents, mainly consisting of lactobacilli,streptococci, and yeasts (Holzapfel et al.1998). In addition, Helicobacter pylori colonization of the stomach is endemicin certain geographical regions of theworld. The duodenum, or rst part ofthe small intestine, also has low micro-bial populations because of both thequick transit of contents through it andthe presence of pancreatic secretionsthat create a hostile environment for mi-

crobes. There is a progressive increasein both numbers and species of microbes,however, along the jejunum and ileum ,from approximately 10 4 to 10 6-7 CFU pergram of contents at the ileo-cecal region(Salminen et al. 1998). The colon is themost heavily populated area of the gas-trointestinal tract, with numbers typically

in the region of 1011

CFU per gram (wetweight) of contents. This environmentsupports greater bacterial growth with aslower transit time , ready availability ofnutrients, and favorable pH.

Recently, Eckburg and colleagues(2005) characterized the microbiota offeces and the intestinal lining of the co-lon of three healthy humans and foundthat the majority of microbes presentwere not from species that have beencultured to date. These authors con-cluded that additional research still is

needed to understand fully the micro-ecology of the intestine.

In women, microbes (many of fe-cal origin) inhabit the vagina to aconcentration of approximately 10 7-8

CFU per milliliter (ml) of uid, with Lactobacillus species dominant inhealthy subjects and urinary and vagi-nal pathogens dominant in patients withinfection.

P ROBIOTIC I MPACT ON C OLONIZING M ICROBIOTA

The probiotic concept asserts thatadding the right live microbes to thiscomplex system can result in physiolog-ical benets. In some instances, theseeffects may result from alteration of thepopulation or activities of colonizingmicrobes. In other cases, effects maybe due to direct interaction of the probi-otic with host cells. The gut remains themost studied site of action for probiot-ics. Most human studies with probiot-ics have targeted specic health benets

associated with the gut microbiota, anda few studies have targeted the stomach,mouth, throat, or vagina.

Several laboratory scale modelshave been developed to simulate theintestine (Macfarlane, Macfarlane, andGibson 1998; Rumney and Rowland1992). These models are useful for es-timating probiotic impact on microbialpopulations or biochemical markers,probiotic survival during transit, andstrain-specic effects. But they cannotfully mimic host factors, and the effectsultimately must be measured in the tar-get host.

Analysis of fecal samples for thepresence of probiotics that have beenconsumed is used often as a proxy mea-sure of probiotic function. The rationaleis If they make it all the way throughthe body alive, they can do some good.Although easily obtained, fecal samplescannot be relied on as accurate indica-tors of the microecology upstream. Iffed in high enough numbers, most pro-

biotic strains can be recovered fromfeces. This observation is often termedcolonization , but commonly it is notdifferentiated from simple transient pas-sage. A more precise word to describethis situation is persistence, assessedby the number of organisms presentover time in feces following a feedingperiod. Persistence would be expectedto be a function of the time it takes theprobiotic to travel through the alimen-tary canal , as well as death and growthFigure 1. The human gastrointestinal tract.


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rates of the probiotic in the body.Discriminating the fed strain from

naturally occurring members of thesame genus or species in the gut can bea challenge. Culture methodologies fre-quently are not discriminatory enoughfor this type of differentiation unlessthe probiotic has a unique physiologi-

cal or biochemical trait that allows it tobe distinguished from background mi-crobes. It is possible, however, to applymodern deoxyribonucleic acid (DNA)-based methods to discriminate betweenfed and naturally occurring strains(McCartney 2002). But the signicanceof fecal recovery of probiotics must beinterpreted carefully because there is noknown clinical benet associated withhigher populations of lactobacilli or bi-dobacteria recovered in feces.

There also is debate about the im-

portance of any probiotic-inducedchanges observed in gut microbio-ta. The most consistently observedchange is the increase in the popula-tions of the genus of the fed probioticstrain. Probiotic-induced changes inthe populations of more dominant in-testinal groups (such as Bacteroides,Clostridium , or Enterobacteriaceae )vary from study to study and probioticto probiotic. Several factors complicateinterpretation of these ndings. Highintra-subject and day-to-day variabil-ity and a lack of understanding of thecomposition of the microbial commu-nity vertically and horizontally withinthe alimentary canal make it difcultto understand the signicance of suchndings. Perhaps more important thanthe microbe population changes are theresulting alterations in biochemistry andphysiology of the intestine.

P ROBIOTIC I MPACT ON H U -MAN H EALTH AND D ISEASE

Reviews on the impact of probiot-ics on human health and disease arenumerous and have emphasized dif-ferent components of the eld, such asuse of probiotics in medical practice(Montrose and Floch 2005; Picard etal. 2005), use in pediatric populations(Saavedra 2007), immunomodulation(Galdeano et al. 2007), and intestinaldiseases (OHara and Shanahan, 2007;Sheil et al. 2007). The following discus-

sion highlights specic areas of probi-otic intervention in human health anddisease.

Keeping Healthy PeopleHealthy

Early research evaluating probioticsin humans focused on relieving intes-tinal distress, frequently with subjectssuffering from an intestinal infection orantibiotic-associated complications. Asthis product concept developed further,the value of probiotics to prevent, ratherthan treat, disease was appreciated morefully. Toward this end, studies havebeen conducted in healthy populations,with end points such as decreasing theincidence of colds (de Vrese et al. 2005),winter infections (Turchet et al. 2003),or even absences from work (Tubelius,Stan, and Zachrisson 2005) or day care(Weizman, Asli, and Alsheikh 2005).These controlled human studies providesupport that certain probiotic strainsconsumed as part of a daily diet will in-crease the number of illness-free days.Infants were helped by Lactobacillusreuteri, which decreased crying time dueto colic (Savino et al. 2007).

Lactose MaldigestionLactose is a sugar found in milk,

composed of a glucose molecule linked

to a galactose molecule. Lactose canbe split into glucose and galactose bylactase, an enzyme produced by infants,children, and some adults. Most hu-mans, however, quit producing this en-zyme in childhood. If these people con-sume dairy products with lactose , theycan develop gastrointestinal symptomssuch as abdominal bloating, pain, atu-lence, and diarrhea. This situation isfound in 5 to 15% of adults in NorthernEuropean and American countries andin 50 to >90% of adults in African,Asian, and South American countries(de Vrese et al. 2001). These peopletend to eliminate milk and dairy prod-ucts from their diet, and consequently,their calcium intake may be compro-mised. The bacteria used as startercultures in yogurt ( Streptococcus ther-mophilus and Lactobacillus delbrueckiisubsp. bulgaricus ) also produce lactase,and when consumed with dairy prod-ucts can improve lactose digestion and

symptoms in these individuals (Kolarset al. 1984). A number of studies havedemonstrated better lactose digestion,as well as a decrease in gastrointestinalsymptoms, in people with this conditionwho consume yogurt with live cultures(deVrese et al. 2001).

Bowel TransitDaily consumption of one to threeservings of fermented milk contain-ing a probiotic strain, Bidobacteriumanimalis DN-173 010, decreased theamount of time it took food to travelfrom the mouth to the anus for peoplewho had longer-than-desired transittime (Marteau et al. 2002). The effectwas more pronounced in elderly sub-

jects and in women. A mixture of eightdifferent strains of lactobacilli, bido-bacteria, and S. thermophilus (productname, VSL#3) had no effect on gastro-intestinal transit time in irritable bowelsyndrome (IBS) subjects (Kim et al.2003). A recent controlled study showedthat L. rhamnosus Lcr35 improvedsymptoms of constipation in children(Bu et al. 2007).

Irritable Bowel SyndromeSymptoms of abdominal pain, bloat-

ing, and atulence commonly occurin patients with IBS. These symptoms

may result in part from fermentationstaking place in the colon that generategas. Certain gut bacteria process left-over food that reaches the colon with-out producing gas. Other species evenmay consume gas, particularly hydro-gen. But others produce gas, which iseliminated from the body through atu-lence. In a double-blind, clinical trialof 48 patients with bloating-predomi-nant IBS, the probiotic mixture VSL#3decreased atulence scores (Kim et al.2005). Likewise, two other placebo-controlled trials have shown relief ofabdominal bloating in patients with IBStreated with VSL#3 or Lactobacillus

plantarum 299V (Kim et al. 2003;Nobaek et al. 2000). In children, L.rhamnosus GG decreased perceived ab-dominal distension but not abdominalpain (Bausserman and Michail 2005).Finally, two large studies in adultsshowed that either Bidobacteriuminfantis 35624 or a strain mixture ( L.


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rhamnosus GG, L. rhamnosus LC705, Bidobacterium breve Bb99, andPropionibacterium freudenreichii subsp.shermanii JS) can be effective in allevi-ating symptoms of IBS (Kajander et al.2005; OMahony et al. 2005).

Some probiotics seem useful forcontrolling the symptoms related to

intestinal gas in this group of subjects.But the number of studies is small, andfurther focused research is needed.

Gastrointestinal InfectionsA number of clinical trials have

tested the efcacy of probiotics in theprevention of acute diarrhea, includingantibiotic-associated diarrhea. Both theshort- and long-term use of antibioticscan produce diarrhea, particularly whenmultiple drugs are used. Probioticsgiven along with antibiotic therapy havebeen shown to decrease the incidenceof antibiotic-associated diarrhea in chil-dren and in adults. Different strainshave been tested including L. rhamno-sus GG, the yeast Saccharomyces cere-visiae (boulardii) Lyo, and undenedstrains of Lactobacillus acidophilus and

L. delbrueckii subsp. bulgaricus . Meta-analysis of controlled trials concludedthat probiotics, particularly L. rham-nosus GG and S. cerevisiae (boulardii)Lyo, can be used to prevent antibiotic-associated diarrhea (McFarland 2006;Sazawal et al. 2006). An importantcomplication of antibiotic treatment canbe the establishment of Clostridium dif-

cile infection resulting in pseudomem-branous colitis. Hickson and colleagues(2007) demonstrated that a fermentedmilk containing Lactobacillus ca-sei DN-114 001, L. delbrueckii subsp.bulgaricus , and S. thermophilus candecrease the incidence of antibiotic-associated diarrhea and C. difficile -associated diarrhea, providing evidence

for the value of this treatment for thispotentially refractory condition.Probiotics are useful as treatment

of acute infectious diarrhea in children.Different strains, including L. reuteri SD2112, L. rhamnosus GG, L. casei DN-114 001, and S. cerevisiae (boular-dii) Lyo, tested in controlled clinical tri-als decreased the severity and durationof diarrhea. Meta-analysis concludesthat these probiotics are safe and ef-fective (Szajewska, Ruszczynski, and

Radzikowski 2006). Oral administra-tion of probiotics shortens the durationof acute diarrhea in children by approxi-mately 1 day.

Prophylactic use of probioticshas proved useful for the preventionof acute diarrhea in infants admittedto the hospital with chronic disease.

Supplementation of an infant formulawith B. animalis Bb12 and S. ther-mophilus TH4 signicantly decreasedthe incidence of diarrhea in hospitalizedinfants aged 5 to 24 months (Saavedraet al. 1994). A placebo-controlleddouble-blind study in infants aged 1 to36 months showed similar results for L.rhamnosus GG (Szajewska et al. 2001).In these two studies, control subjectswere more than four times more likelyto develop diarrhea than those treatedwith probiotics.

Several studies have investigatedprobiotics in the prevention of travel-lers diarrhea in adults, but methodolog-ical deciencies, such as low compli-ance with the treatment and problems inthe follow-up, limit the validity of theirconclusions (Marteau, Seksik, and Jian2002). This topic needs further study.

Probiotics have been tested as astrategy for eradication of H. pylori in-fection of the stomach. Some strains oflactic acid bacteria are known to inhibitthe growth of H. pylori in laboratoryexperiments. But results in human stud-ies with different probiotics are mixed.Eradication of H. pylori was attemptedby feeding yogurt containing probioticstrains selected for their ability to in-hibit H. pylori in laboratory studies. Thestrategy was not effective in patientsnot undergoing simultaneous antibiotictherapy (Wendakoon, Thomson, andOzimek 2002). In contrast, other clini-cal studies have tested the use of probi-otics as a supplement to antibiotic ther-apy for H. pylori eradication. In thesestudies, the use of probiotics decreasedthe side effects of antibiotics, improvedpatient compliance with taking the pre-scribed therapy, and increased the rate atwhich H. pylori was eradicated (Lionettiet al. 2006; Myllyluoma et al. 2005;Sheu et al. 2006; Sykora et al. 2005).

Prevention of Systemic InfectionsBacterial translocation is the pas-

sage of bacteria through the lining of

the intestine, which can lead to infectionof organs or the blood. This passagecan occur when patients have under-gone surgical procedures or are seri-ously ill with critical conditions, such assevere acute pancreatitis, advanced livercirrhosis, or multisystem organ failure.Probiotic organisms rarely translocate,

even through a disturbed epithelium(Daniel et al. 2006).In a study of patients with severe

acute pancreatitis, treatment with L. plantarum 299V signicantly decreasedthe incidence of infection (Olah et al.2002). In another study, liver transplantpatients received a synbiotic prepara-tion (including four probiotic strainsand four fermentable bers) or a pla-cebo consisting of the four bers only.Postoperative infection occurred in onlyone patient in the treatment group of 33

compared with 17 of 33 in the placebogroup (Rayes et al. 2005). The differ-ence was highly signicant. But anoth-er clinical study performed with patientswho submitted to elective abdominalsurgery found no effect of synbiotictreatment (four bacteria strains plus oli-gofructose ) on prevention of postopera-tive infections (Anderson et al. 2004).In that trial, synbiotic treatment aftersurgery was delayed until patients wereable to tolerate oral nutrition. In con-trast, the liver transplant studies intro-duced synbiotic therapy by naso-gastrictube immediately after surgery.

Necrotizing enterocolitis result-ing from immaturity and poor functionof the gut mucosal barrier is a severeclinical condition that may occur in lowbirth weight premature infants. Twocontrolled studies have demonstratedthat the use of probiotic mixtures inthese infants signicantly decreases theincidence and severity of necrotizingenterocolitis and prevents death (Bin-Nun et al. 2005; Lin et al. 2005). Datafrom these studies represent one of thefew examples of probiotics improv-ing survival rates. Currently, few otherstrategies have proved effective in de-creasing the incidence, morbidity, andmortality of necrotizing enterocolitis inpreterm infants.

Inammatory Bowel DiseasesUlcerative colitis , pouchitis , and

Crohns disease are chronic conditions


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of unknown cause characterized bypersistent inammation of the intes-tine. Evidence suggests that abnor-mal activation of the mucosal immunesystem against the gut microbiota is thekey event that triggers this abnormal in-ammatory response that in turn causesulcers in the gut that fail to heal, lead-

ing to chronic intestinal disease. Threestudies investigated the effectiveness ofan oral preparation of Escherichia coli Nissle 1917 compared with mesalazine,the standard treatment for maintenanceof remission in patients with ulcer-ative colitis (Kruis et al. 1997, 2004;Rembacken et al. 1999). These studiesconcluded that this strain was as effec-tive as mesalazine in maintaining remis-sion. A small pilot study suggested thata fermented milk containing B. breve strain Yakult, Bidobacterium bidum

strain Yakult, and L. acidophilus (un-dened strain) can be useful to induceremission in ulcerative colitis patientswith mild disease (Kato et al. 2004).

The VSL#3 probiotic mixtureproved highly effective for maintainingremission of chronic relapsing pouchi-tis (Gionchetti et al. 2000; Mimura etal. 2004). Treatment with VSL#3 alsois effective in the prevention of the on-set of pouchitis after surgery to formthe pouch (Gionchetti et al. 2003). InCrohns disease, however, clinical stud-ies with L. rhamnosus GG failed toshow efcacy in preventing postopera-tive recurrence of the disease (Pranteraet al. 2002) or as maintenance therapy(Bousvaros et al. 2005).

AllergyAtopic diseases are caused by ex-

aggerated or imbalanced immune re-sponses to environmental and harmlessantigens (allergens). The prevalence ofallergic diseases in western societies is

increasing at an alarming rate.The effectiveness of L. rhamnosus GG in the prevention of atopic der-matitis has been reported in random-ized, controlled trials (Kalliomaki et al.2001, 2003). In a subsequent study, thissame strain was combined with threeother probiotic strains, L. rhamnosus LC705, B. breve Bb99, and P. freuden-reichii subsp. shermanii JS, and a pre-biotic to determine the impact on the

cumulative incidence of allergic dis-eases (Kukkonen et al. 2007). This largestudy (925 subjects tracked through the2-year follow-up) showed no effect onincidence of all allergic diseases, butthe treatment did signicantly preventatopic eczema. Furthermore, severalwell-designed studies have provided

evidence that specic strains of probiot-ics can be somewhat effective in treat-ment of established atopic dermatitis(Rosenfeldt et al. 2003; Viljanen et al.2005; Weston et al. 2005). Effectivenessin the management of cows milk allergyin children is associated with the use ofprobiotics (Kirjavainen, Salminen, andIsolauri 2003). One study, however,failed to nd any effect for allergic in-fants with L. rhamnosus GG treatmentfor 3 months (Brouwer et al. 2006), em-phasizing the importance of conrma-

tory studies.

Colon CancerSeveral experimental animal studies

clearly demonstrated a protective effectof prebiotics such as oligofructose, pro-biotics such as some Lactobacillus and

Bidobacterium strains, or the combi-nation of prebiotics and probiotics onthe establishment, growth, and metas-tasis of transplantable and chemicallyinduced tumors. Human intervention

trials to conrm these animal studiesare intrinsically difcult because of thenatural history of the disease (difcultyin selecting subjects at high risk andrequirement of long-term follow-up).A 4-year study of 398 subjects foundthat L. casei Shirota decreased the re-currence of atypical colonic polyps(Ishikawa et al. 2005). The EuropeanUnion (EU)-sponsored Synbiotics andCancer Prevention in Humans proj-ect tested a synbiotic (oligofructoseplus L. rhamnosus GG and B. anima-

lis subsp. lactis Bb12) in patients atrisk for colonic polyps. Among severalintermediate end points that were usedas biomarkers of colon cancer risk, thestudy found that the synbiotic decreaseduncontrolled growth of intestinal cells(Van Loo et al. 2005).

Vaginal InfectionVaginal infections are caused mostly

by fecal microbes ascending into the

vaginal tract and displacing the normallactobacilli microbiota. The poten-tial of using probiotic lactobacilli todecrease the risk of bacterial or yeastvaginal infections or to improve theclinical outcome during treatment forthese infections has captured the inter-est of researchers for decades (Reid and

Bocking 2003). Until recently, moststudies have been small and in needof conrmation. After preliminary as-sessment to document that L. rhamno-sus GR-1 and L. reuteri RC-14 admin-istered in milk could pass through theintestine, ascend to the vagina, andrestore a normal lactobacilli microbiotain women prone to infections (Reid etal. 2001), these strains were delivered inyogurt to African women with bacterialvaginosis and shown to improve thera-peutic outcome (Anukam et al. 2006

a, b). These studies have provided thebest evidence to date for successful pro-biotic intervention to improve vaginalhealth. Some other recent studies havenot shown positive results (Eriksson etal. 2005), highlighting the importanceof use of effective strains and deliverysystems.

S AFETY C ONSIDERATIONSAs probiotics increase in popularity,

it is prudent to consider whether there

are any safety concerns associated withthe resulting increased exposure to livemicrobes. It is tempting to presume thatthe question of safety is only about theprobiotic strain being used; in fact, howthe strain is used and who is consumingit are important considerations as well.For example, a safety assessment of astrain used in yogurt at 10 9 CFU/servingwould be different for the same strainadministered at 10 12 CFU/ enteric coat-ed capsule. A safety assessment for astrain administered intravaginally wouldagain be different. A safety assessment,therefore, must consider the nature ofthe specic microbe being consumed,how it is prepared, how it is adminis-tered, what dose is delivered, and thehealth status of the consumer.

Assessing the Safety ofProbiotics

The key components of assessingsafety include (1) identifying the probi-


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otic using the best genetic and physi-ological techniques to the strain level;(2) determining the antibiotic resistanceprole, including an evaluation of thelikelihood that any antibiotic resistancetraits would be transferred to membersof the colonic microbiota; (3) if appli-cable, establishing a history of safe use

based on the intended use of the speciesin question; and (4) conducting toxicityor pathogenicity assessments in validat-ed laboratory or animal models that arerelevant to the species being considered,as needed.

Most probiotic-containing foodproducts use lactic acid bacteria thathave a long history of safe use in foods.As long as the strain is devoid of anytransferable antibiotic resistance genes,members of the genera Lactococcus ,

Lactobacillus , and Bidobacterium are

considered safe; infections in humans bythese genera are extremely rare. Therehave been 180 cases of lactobacillemiaand 6 cases of bidobacteremia reportedduring the past 30 years (Borriello et al.2003). All cases of bidobacteremiaoccurred in subjects with abdominal in-fections or with obstetrical proceduresor infections (Gasser 1994). There havebeen 69 cases of infective endocarditisattributed to lactobacilli reported dur-ing the same period. In most cases ofendocarditis, dental surgery occurred inthe days or weeks preceding the disease.These infections resulted from nativesources of these genera and not fromconsumption of probiotic products.

Two cases of Lactobacillus infectionwere linked with probiotic consumption(Borriello et al. 2003). Increasing con-sumption of probiotic lactobacilli andbidobacteria has not led to an increasein such opportunistic infections in con-sumers (Salminen et al. 2002). Thus,the risk of infection by these genera isin the negligible range, taking into ac-count that exposure to them is universaland persistent, not only through probi-otic products but also as common colo-nizers of the human body (the digestivetract and oral and vaginal cavities). Thislack of pathogenicity extends across allage groups (including preterm infantsand pregnant women) (Lin et al. 2005;Saavedra et al. 2004).

Pharmaceutical or supplementforms of probiotics may be taken by

consumers with less-than-optimal healthstatus. These products also may en-compass a broader range of microbesthan typically used in foods, includingmembers of the genera Streptococcus ,

Enterococcus , and Escherichia . Somestrains or species of these genera arepotentially pathogenic. In these in-

stances, the accuracy of identicationto the strain level is a critical step in theassessment of safety. Although an un-common cause of infection in humans,a few cases of sepsis have been associ-ated with administration of the probioticS. cerevisiae (boulardii) (Munoz et al.2005), sometimes caused by inadvertentcontamination of intravenous lines.

Resistance to AntibioticsExpression of resistance to antibiot-

ics by candidate probiotic strains shouldbe considered from the perspective ofsafety. But intrinsic resistance to someantibiotics is inherent to the physiologyof certain probiotic strains because oftheir cell wall structure or other inher-ent physiological characteristics. Theexpression of resistance to an antibioticis considered a risk factor if strains aresuspected of harboring acquired, trans-ferable antibiotic resistance genes, assuggested by expression of resistanceto an antibiotic that exceeds the rangedetermined to be normal for the spe-cies. In many instances, resistance toantibiotics is not transmissible, and thespecies also are sensitive to many otherclinically used antibiotics. No particu-lar safety concern, therefore, is associ-ated with an intrinsic type of resistance.

Antibiotic resistance that is associat-ed with transmissible genetic elements,however, should be avoided in bacteriaintended for probiotic use because ofthe possibility of the resistance spread-ing to potentially harmful microbial

residents (e.g., transmission of entero-coccal resistance against the antibiotic,vancomycin). Vigilance regarding thedetection of possible rare instances ofinfection due to probiotics should bemaintained.

D-Lactate AcidosisAnother potential risk is the induc-

tion of D-lactate acidosis by D-lacticacid-producing probiotic bacteria. This

is a very rare clinical condition that hasnever been linked to probiotic consump-tion. In fact, the occurrence of D-lac-tate acidosis has been recognized almostexclusively in patients with signicantlydecreased small intestinal absorptivecapacity after intestinal bypass surgeryor with short bowel syndrome (Mack

2004). In these instances, ingestion ofcarbohydrates may be followed by amassive load of sugars into the co-lonic lumen followed by productionof D-lactate by commensal bacteria.Because humans metabolize D-lactateat a slower rate than the L-lactate iso-mer, increased absorption of D-lactatemay lead to acidosis. Most subjectswho have intestinal bypass operationsor short bowel syndrome, however, donot develop D-lactic acidosis becausethe population of D-lactic acid-produc-

ing bacteria in the colonic microbiotaof these subjects is not predominant.It is prudent, however, to recommendthat D-lactate-producing probiotics beavoided by subjects with short bowelsyndrome.

P RODUCT C ONSIDERATIONSProbiotic products are unique in

that keeping the microbes alive mustbe a consideration through the stages ofproduct concept, formulation, and the

sales/distribution process. The typicalissues surrounding product developmentalso apply: products should be tasty (ifin food form), convenient, and pricedcompetitively. But additional consid-erations must be addressed: optimiz-ing growth conditions for the probiotic,dening a product that can deliver theprobiotic successfully in a viable andfunctional form to the active site in thebody and through the end of shelf lifein the product, and determining the roleof the total product (including fermen-tation end products or other functionalingredients) on healthful properties.These considerations are not trivial, andunfortunately not all products marketedas probiotic suitably address them.

Several studies document examplesof foods and supplements that eitherdo not contain the amount of probioticstipulated on the label or do not use thecorrect scientic nomenclature to namethe microbes present. In addition, some


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products bear labels suggesting healtheffects that have not been documented.Some products (many yogurts fall intothis category) do not make any claims ofprobiotic potency or efcacy but simplylist the genus and species of additionallive bacteria. The implication is thatthese bacteria are good for you; infact, there may be little evidence that theproducts as formulated are efcacious.

Table 3 lists considerations for pro-biotic product development and expandson a document developed by a work-ing group of the United Nations Foodand Agriculture Organization (UNFAO/ WHO 2002) that provides guidelinesfor probiotic products. This process in-volves choice of strain or combinationof strains (not all strains of probioticswould be expected to function equallywell in different roles), what amounts touse, what studies to conduct to docu-ment functionality, how to label theproduct, and how to communicate aboutthe product. All these issues are interre-lated and must start with an understand-ing of what health effect is envisionedfor a product.

R EGULATORY S TATUS OF P ROBIOTIC F OODS Labeling of Probiotics

There is neither a legally recognized

denition of, nor a standard of identityfor, the term probiotic in the UnitedStates or worldwide. Products contain-ing this label, therefore, currently arenot obligated to meet any standardsunique to probiotics. There is, how-ever, a growing understanding of thisterm among consumers and healthcareprofessionals. It is unfortunate thatproducts currently can be labeled asprobiotics but be neither well denednor substantiated with controlled humanstudies. Ideally, products labeled asprobiotics would conform to the guide-lines established by a working group ofthe FAO (UNFAO/WHO 2002). Therequirement in the United States is thatproducts be labeled in a truthful and notmisleading fashion; this requirementapplies to content as well as claims offunctionality.

Many commercial products likelydo not meet these criteria, as evidencedby several published surveys of com-mercial products. Most of these studiesdocument the degree of label non-compliance with numbers or types ofviable probiotic microbes recoveredfrom commercial products (Drisko etal. 2005; Temmerman et al. 2003a, b;Yeung et al. 2002). Some of these fail-ures are of more concern than others.Some instances of mislabeling the typesof microbes present can be attributed to

recent changes in scientic nomencla-ture; commercial product labels do notalways reect the most current, scien-tically recognized nomenclature. Forexample, the species of L. acidophi-lus was subdivided into six differentspecies ( L. acidophilus , L. gasseri, L.

johnsonii, L. crispatus, L. gallinarum ,and L. amylovoris ). Continued use ofthe L. acidophilus name instead of thenewer name, although it is incorrectand should be corrected, is not of greatsignicance. More important is that themicrobes bear a consistent strain desig-nation so that research can be trackedadequately for each strain even if thespecies name has been changed. Oneexample of mislabeling at the genus lev-el is with products claiming to containLactobacillus sporogenes. This namewas noted as a misclassication in 1939(Sanders, Morelli, and Bush 2001). Infact, microbes using this name are mostlikely Bacillus coagulans . Its continueduse commercially raises ethical, safety,and efcacy issues.

Studies also have documented fail-ure of products to meet label claimswith regard to numbers of viable mi-crobes present in the product, therebyconstituting illegally labeled products.Maintaining viability of microbes incommercial products through the end ofshelf life can be a challenge. Viability

Table 3. Considerations for probiotic product development

Considerations Comments

Probiotics should be described adequately and identied Biochemical, morphological, physiological, and DNA-based techniques canto the strain level. contribute to the description of commercial probiotic strains. Total genomic DNA

sequencing is becoming more common on fully characterized probiotic strains(Altermann et al. 2005).

Each probiotic strain should be able to be identied and This can be a challenge because culture microbiology methods often are notenumerated from the product. available to differentiate among different species of the same genus. DNA-based

approaches often can solve this problem.Probiotic strains and products should be supported by a dossier A dossier should be developed that is composed at least in part of peer-reviewedsubstantiating efcacy. publications documenting the ability of the probiotics to impact human health

positively and supporting any claims made.

Product formulation should be evidence-based. Decisions on product format (type of food or supplement), dose, and choice ofstrain(s) should be consistent with those used in clinical studies.

Viability of all probiotic strains in the product should be maintained Viability (generally assessed as CFU/g) should be maintained at the level shown toabove the target minimum level through the end of shelf life. deliver health effects. Although some decline in levels is common over time, initial

formulations should assure that the levels do not drop below what is indicatedon the label and what is known to be efcacious.

Product labeling should be done in a truthful and not Product labels and any supplementary communications should provide clear,misleading fashion accurate information on the types and levels of probiotics; any documented health

benets; and the amount of product that must be consumed for an effect.

Probiotic strain(s) must be safe for their intended use. (See main text)


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depends on many factors such as howthe microbe is grown and stabilized andhow it is handled (storage time, temper-ature, and exposure to moisture are im-portant factors) once it is in the product.Although manufacturers control overhow retailers and consumers handletheir products may be limited, storage

and handling recommendations shouldbe made; if those recommendations aremet, products should meet label claimsthrough the end of shelf life. But this isnot the approach taken by all manufac-turers and distributors of products cur-rently in the marketplace.

Some product manufacturers makeno claim to the quantities of microbesdelivered. This practice is common inmany food products, such as yogurt,that contain additional bacteria addedfor their health effects. For example,

the label may indicate that the productcontains L. acidophilus , but no com-munication is provided on quantities orif the quantities added were chosen tomatch a documented health effect. Insome instances, products might not in-dicate a quantity of probiotic on the la-bel, but scrutiny of the product websitemight give some indication of levelsdelivered per serving. Some supple-ment products are labeled stating thequantity of viable probiotic per dose attime of manufacture. This informationis not very helpful for the consumer.Some supplements indicate dose to con-sume as a function of a gram or capsuleamount but do not relate this to a viablecell count (CFU).

The most responsible approach tocommunicating the content of probioticin products is to include on the labelthe genus, species, and strain designa-tion for each probiotic in the productand the level of viable cells of eachprobiotic strain at the end of shelf life.Some manufacturers resist this ap-proach, claiming it confuses consumers.Another acceptable choice is to providethis information through advertising orwebsite communications. Additionally,the level needed to be consumed toachieve the documented health benetshould be disclosed.

Without any declaration of contents,however, consumers and healthcare pro-fessionals do not have adequate infor-mation to differentiate among products

on the market. Documented failures ofproducts to meet label claims with re-gard to numbers and types of viable mi-crobes present in the product and howmany must be consumed for a healthbenet suggest that there is a problemin the probiotic industry with regard toaccurate labeling. But not all such pub-

lished studies have used methods thatgive accurate measures of bacterial con-tent. Researchers unskilled in workingwith a particular species of probiotic orspecic product formats may concludeerroneously that products do not containindicated bacteria, whereas, in fact, theirmethods were inadequate.

A better approach for addressingthis situation would be for companiesto submit appropriate methods for as-saying the contents of their products toa reliable third party for independent

assessment. Making the results of suchanalyses available to consumers wouldassist in restoring condence in prod-ucts, a feeling that has eroded becauseof poor product formulation.

Safety StandardsAnother important regulatory issue

is safety. In the United States, thereare different safety standards for dif-ferent regulatory categories (foods, di-etary supplements, or drugs) (Table 4).For supplements marketed before theOctober 15, 1994 passage of the DietarySupplement Health and Education Act(DSHEA), it is the manufacturers re-sponsibility to ensure safety; no premar-ket approval is required. If a supple-ment product contains a new dietaryingredient or NDI (i.e., a dietary ingre-dient not sold in the United States in adietary supplement before October 15,1994 and not present in the food sup-ply), then the manufacturer must no-tify the Food and Drug Administration(FDA) at least 75 days before market-ing. If the FDA nds that the noticedoes not provide an adequate basis forbelieving that the ingredient is safe forits intended use, the Agency will warnthe notier that use of the ingredientin a dietary supplement may render itadulterated and liable to seizure.

The FDA has no authoritative listof dietary ingredients marketed be-fore October 15, 1994, so the burden of

proof rests with the manufacturer to de-termine if a new product is an NDI. Ifthe probiotic was not sold in the UnitedStates as a dietary supplement beforeOctober 1994 and was not present in thefood supply, then it is an NDI, and anydietary supplement containing it wouldbe considered adulterated unless an NDI

notication was led with the FDA.A dietary ingredient is dened bythe DSHEA as one or any combinationof the following substances: a vitamin;a mineral; an herb or other botanical;an amino acid; a dietary substance foruse by humans to supplement the dietby increasing the total dietary intake(e.g., enzymes or tissues from organsor glands); or a concentrate, metabo-lite, constituent, or extract (USFDACFSAN 2001). Although probioticsdo not fall intuitively into any of these

substance categories, the FDA has ap-proved six probiotic bacteria as new di-etary ingredients: L. casei, L. reuteri, L.

plantarum (combined with fructooligo-saccharide), L. bulgaricus, L. paracasei ,and B. infantis . Other probiotic bacteriaare sold in dietary supplements withouthaving been the subjects of NDI re-views and may be regarded by the FDAas being grandfathered by use beforeOctober 1994. Note that FDA approvalin this context is with regard to safety,not efcacy.

Generally Recognized asSafe

For conventional foods, it also isthe manufacturers responsibility toensure safety; no premarket approvalis required. All ingredients used infoods must be considered generallyrecognized as safe (GRAS), be a PriorSanctioned Substance (sanctionedin a letter written by the FDA or theU.S. Department of Agriculture before1958), or be a food additive approvedby the FDA and used in compliancewith an FDA food additive regula-tion. The GRAS substances are foodsubstances judged by qualied subjectexperts as safe under the intended con-ditions of use. This determination maybe based on data existing in the pub-lished scientic literature or througha long history of safe use. Althougha limited list of GRAS substances is


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published by the FDA, this list is notcomplete. Companies are allowed toconduct a self-determination of GRASstatus. The company may opt to notifythe FDA of its ndings, or it may keepthe ndings as an in-house documentsurrendered to the FDA only on theirrequest. If a food substance is judgedto be GRAS, either through the self-af-rmation of GRAS status approach orthrough the FDA notication process,no premarket approval is required.

Only a few probiotic microbes areincluded on any ofcial GRAS list. Acommon misconception is that probi-otics are GRAS. It should be kept inmind that the term GRAS applies to afood substance for a specied use. Theterm does not apply to drug uses andit does not apply to non-oral routes ofadministration. Manufacturers cannotassume that all probiotics are GRAS,even if they are composed of speciesof Lactobacillus or Bidobacterium .

A responsible company should reectseriously on all intended uses for allprobiotic strains and strain blends itseeks to market and provide a consid-ered rationale for GRAS status self-de-termination. Prior history of safe use ofthe species, negligible associations withinfection or toxicity, and the absence oftransferable antibiotic resistance genesfrom the specic strains being consid-ered may comprise the bulk of the ra-tionale. This endeavor becomes morecomplicated when dealing with otherprobiotic genera such as Enterococcus,Streptococcus, Escherichia , or Bacillus .

One difference in safety consider-ations for conventional foods comparedwith dietary supplements is that conven-tional foods are marketed to the gen-eral population. It is allowable to labeldietary supplements for use in particularsubpopulations (e.g., adults, or chil-dren over the age of 3 years), but thispractice is not allowed on conventional

foods. Also, foods may be consumedfor an entire lifetime, so cumulative ef-fects need to be considered. Further,dose amounts may be recommended fordietary supplements, but food ingredi-ents must be safe even for people whomay consume far more than averageamounts.

The European Food SafetyAuthority (EFSA) recently proposed thequalied presumption of safety (QPS)approach as an operating tool for safetyassessment of microorganisms in foodand feed (EFSASC 2004). This systemis based on the taxonomic identica-tion of the candidate microorganismsand the body of knowledge concerningthe particular species of microorgan-ism, including history of use, scienticliterature and databases, clinical aspects,ecology, and industrial applications.The result of this effort will be a list ofQPS microbes that will be consideredsafe for use in foods.

Table 4. Differences among regulatory categories available for marketing probiotic products in the United States

Regulatory Categories

Actions Food Supplement (must be ingested) Drug

Premarket approval by the FDA Not required for GRAS Not required for microorganisms used before Requiredmicroorganisms October 1994

Disease claim (describes the effect Not allowed Not allowed Allowed if approvedof a drug on the diagnosis, treatment, by the FDAmitigation, cure, or prevention of disease)

Health claim (describes the effect of Allowed if approved Allowed if approved by the FDA Not used, althougha dietary substance on the reduction by the FDA (may be (may be unqualied or qualied) can use strongerof risk of disease by the currently unqualied or qualied) prevention claimshealthy population)

Structure function claim (describes the Allowed if truthful and not Allowed if truthful and not misleading; commonly Not usedeffect of a dietary substance on the misleading; effect must used; label must say this statement has notstructure or function of the body) derive from the nutritive been reviewed by the FDA; must notify the

value of the food; no FDA of intent to use this claim within 60 daysrequirement for label of marketing the productdisclaimer or FDAnotication

Safety standards a Reasonable certainty No signicant or unreasonable risk of illness or The FDA assessesof no harm under the injury; target consumer group can be stipulated safety and mustintended conditions on the label determine that benetof use; GRAS status outweighs riskcan be self-determinedor submitted throughGRAS noticationprocess; must be safefor general populationand all subgroups

Product examples Yogurt Capsules No probiotic productscurrently are regulatedas drugs for human usein the United States

aThese are not zero-risk standards.


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Regulating Claims of EfcacyThe U.S. regulatory status of claims

that can be made for conventional foodsand dietary supplements is summarizedin an online document from the U.S.Food and Drug AdministrationCenterfor Food Safety and Applied Nutrition(USFDACFSAN 2003). The respon-sibility for ensuring the validity of theseclaims rests with the manufacturer, theFDA, orwith regard to advertisingthe Federal Trade Commission (FTC);the validity is determined on a case-by-case basis. Although probiotics mostcommonly are considered as compo-nents of functional foods or dietary sup-plements, they also may be suitable fortreatment or mitigation of disease and,as such, could be marketed as drugs. Nosuch products, however, have enteredthe U.S. market for human use. Twoimportant differences among these cat-egories are how statements about healthor disease can be made on the label andwhat can be said in advertising of thesedifferent product categories (Sanderset al. 2005). The FTC has long used areasonable consumer standard for de-termining whether messages are truthfuland not misleading. The FDA adoptedthe reasonable consumer standard in2002, superseding the previous standardof the ignorant, the unthinking, and

the credulous. The FDA has providedguidance on what constitutes substan-tiation for claims of efcacy on dietarysupplements (USFDACFSAN 2004).

Since the enactment of the DSHEAin 1994, a commonly expressed senti-ment is that dietary supplements areunregulated. This point is illustratedby comments made in a television newsreport aired May 8, 2006 on probiot-ics in which a University of CaliforniaBerkeley professor was quoted as say-ing, Right now, its the Wild West.

There is no quality control. There is nostandardization. There is no proof of ef-cacy, and there is no proof of safety(Mulvihill 2006).

Although it is true that dietary sup-plements are not regulated in the samemanner as drugswith preapprovalfor efcacy and safety and mandatoryanalyses of every batch produced toassure potency and puritythere areregulations issued and enforced by the

FDA and the FTC to prevent the use ofuntested or unsafe ingredients, to requirethe use of good manufacturing prac-tices, and to compel truth in labelingand marketing. Unfortunately, enforce-ment rigor on the part of these agenciesis limited. An examination of warningletters reveals that the focus of enforce-

ment efforts is on the mislabeling ofsupplements as drugs through use ofunapproved drug claims and removal ofunsafe products; no warning letters wereissued to a food or supplement manufac-turer for claims of efcacy that were un-substantiated. This circumstance reectsthe FDAs priorities: rst, safety, includ-ing hazards presented by leading con-sumers to forego needed pharmacologi-cal intervention through false claims;second, lth; anda distant thirdeco-nomic deception.

The lack of FDA enforcement, how-ever, does not diminish the responsi-bility of companies to substantiate anyclaims of efcacy. Even structure/func-tion claims such as improves microbialbalance or enhances natural immunefunction must be based on evidencederived from human studies on the strainor strains and levels used. Strain- anddose-specicity of probiotic functionmust be presumed unless demonstrat-ed otherwise, and simply providing asource of live cultures is not sufcient tosupport such claims.

F UTURE D EVELOPMENTSThe eld of probiotics is growing

rapidly with concomitant developmentsin the research, commercial, and medi-cal sectors. The complete genomic se-quences are known for several impor-tant probiotic bacteria, and functionalgenomics ndings will be instrumentalin identifying many features responsiblefor probiotic functionality. Accordingto Klaenhammer and colleagues (2005),This information is providing an im-portant platform for understanding coremechanisms that control and regulatebacterial growth, survival, signaling,andunderlying probiotic activitieswithin complex microbial and host eco-systems. Genetic approaches also willenable design of genetically modiedprobiotic strains with specic thera-peutic capabilities such as delivery of

anti-inammatory cytokines, vaccineepitopes, or antipathogenic molecules.Well-designed and properly controlledhuman and mechanistic studies willadvance the essential understanding ofactive principles, mechanisms of action,and degree of effects that can be real-ized by specic consumer groups.

In the commercial realm, the suc-cess of probiotics in Europe and Asialikely will be realized in the UnitedStates. The number and types of prod-ucts will increase in the food, supple-ment, and pharmaceutical categories.As competition intensies in the mar-ketplace, companies providing respon-sibly formulated and promoted productswill prevail. Lastly, as this eld ad-vances, look for new types of probioticstrains with benets not yet exploredthat may surpass the value of those cur-

rently in commercial use.

R ECOMMENDATIONSEstablish a standard of identityfor the term probiotic basedon the FAO denition (UNFAO/ WHO 2001). Adoption of a stan-dard of identity for use of the termwould assure that the word retainssome meaning in the marketplaceand that only well-dened prod-ucts with documented efcacy

could use this term on labels.Regulate probiotics based ontheir intended use, but expandregulatory conceptualizationof health benet claims. In theUnited States, probiotics havebeen used as food ingredients,medical foods, and supplementsand are in development as drugs.Different regulatory requirementsexist for each of these categoriesand are imposed from the researchand development stages throughto marketing of a product. TheFDA should be encouraged toexplore how it might expand itsapproach to appropriate targets forhealth benet claims, and relatedresearch to substantiate thoseclaims, for foods. Foods (includ-ing those containing probiotics)often are useful to provide supportto improve tolerance or otherwisecomplement drug therapy. Probi-

1.

2.


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otic foods also may be of benetto help at-risk individuals avoiddisease, including acute condi-tions such as bacterial vaginosis,necrotizing enterocolitis, and an-tibiotic-associated diarrhea. Stud-ies that validate such effects, andcommunication of these benets

for foods , should be facilitated,not suppressed, by regulatoryagencies.Adopt the use of third partyverication of label claims. Thispractice would go far towardincreasing consumer condencein probiotic products and woulddecrease the oversight needed bygovernmental agencies. Verica-tion could address both contentand efcacy claims, resultingin the ability of consumers andhealthcare professionals to betterdifferentiate among commercialproducts.Use probiotics selectively in clin-ical conditions. It is common toeld questions from some peoplewho are desperately hoping thatprobiotics will solve a certaincritical health concern. The useof probiotics for new medicalapplications is not recommendedin clinical conditions in which

there is no evidence of benetthat is based on positive results inwell-designed and well-performedclinical trials. The use of probi-otics in a new clinical setting orcondition should be accepted onlyafter consultation and approval bythe ethics committee of a clinicalresearch institution.Consider multiple factors whenevaluating probiotics. Whenconsidering the body of evidencesupporting probiotics and theirimpact on human health, it shouldbe kept in mind that human stud-ies are necessary for conrmingbenets. Research evaluatingprobiotics should follow standardbest clinical practice guidelines.But there are additional factorsto consider such as the contri-bution of the delivery vehicle,including bioactive ingredientsintrinsic to the carrier or result-

3.

4.

5.

ing from growth or metabolismof the probiotic during productpreparation; the need to specifyand verify the dose of eachcomponent of the probiotic beingtested; and the need to identify thegenus, species, and strain of eachstrain tested. Editors of journals

publishing papers on probioticsshould be aware of the importanceof these disclosures.Focus research on the importantrole of human native microbiotain health. The role that native,colonizing microbes play (such asimproving vaccine efciency, im-proving outcomes of drug therapy,improving tolerance to drugtherapy, or improving resistanceto infectionsespecially ones ofglobal magnitude) is an importantpublic health concern. Inuenc-ing these microbial populationsor activities through the use ofprobiotics, and the causal relation-ship between these changes andvalidated health biomarkers orclinical effects, is a recommendedline of research.Use a science-based assessmentof the benets and risks ofgenetically engineered probi-otic microbes. Although not

addressed in this review, the useof genetically engineered probi-otic microbes to treat or preventdisease shows potential. Althoughsome people advocate an outrightban on such use, a more appropri-ate approach is a science-basedassessment of safety and environ-mental exposure risk comparedwith the benets that recombinantorganisms might provide.Provide better information toconsumers. Consumers are ask-ing basic questions about probiot-ics such as: What are they? Howmany do I need to consume?What kinds are best? How oftendo I need to consume them? Arethey safe? What effects can Iexpect? Unbiased, straightfor-ward answers to these types ofquestions need to be developedand disseminated to the generalpublic.

6.

7.

8.

A PPENDIX 1.

H OW P ROBIOTICS W ORKThe benecial effects of probiot-

ics likely result from several complex,interacting mechanisms that will differfor different strains and sites of ac-tion. These mechanisms may include

competition for binding sites to theintestinal wall, competition for essen-tial nutrients, production of antimicro-bial substances, stimulation of mucin production, stabilization of the intesti-nal barrier, improvement of gut transit,metabolism of nutrients to volatile fattyacids, and immunomodulation (immunestimulation and immunoregulation).Some of these mechanisms have beendemonstrated only through laboratoryexperiments or animal models and arenot substantiated in humans.

Contrary to popular belief, probi-otics generally have not been shownto have a large or consistent impact onpopulations of intestinal microbes, andany effect they do exert is generallytransient. This is likely because thenative microbiota are a well-adapted,naturally stable association of microbesthat effectively resist change (see re-view issued by the French Food SafetyAgency [AFSSA 2005]). Probiotics,however, have been shown to impactthe biochemistry of the intestinal envi-ronment, and administration of probi-otics during periods of disruption ofnormal microbial balance (such as dur-ing antibiotic therapy) seems to provideopportunity for probiotics to inuencehealth.

The recognition during the pastdecade of the limitations of scientistsknowledge of the composition of theintestinal populations of microbes hasresulted in a shift of focus from probi-otic-induced changes in populations of

microbes to more readily measurableend points. Perhaps more importantthan documenting an impact on popula-tions of intestinal microbes is the abil-ity to demonstrate that probiotics canpromote the recovery of disturbed mi-crobial populations (Engelbrektson etal. 2006), improve the character of theintestinal environment, or impact vali-dated indicators of human health.


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proof that pathogen inhibition resultsdirectly from bacteriocin productionby the probiotic. Pathogens including

E. coli, Salmonella, Campylobacter,Shigella, Vibrio, Clostridium, Candidaalbicans , human immunodeciency vi-rus, and other viruses have been inhib-ited in laboratory tests or animal stud-

ies by bacteriocin-producing probioticstrains (OSullivan and Kullen 1998).But proof that the bacteriocinand notsome other cell functionmediates ob-served antipathogenic effects in vivostill is needed for most examples ofbacteriocins.

Stimulation of Mucin ProductionA gel-like mucus layer composed

of complex proteins bound with sugarmolecules (mucins) covers the intestinal

surfaces. The mucus layer shields thegut surface from direct contact with thecontents passing through the intestineand acts as a barrier against invasionby pathogenic organisms and toxins(Sanderson and Walker 1994; Yolkenet al. 1994). Mucus can protect againstenteric pathogens by serving as a physi-cal barrier, by housing antibodies thatcan bind potentially harmful antigens,and by releasing mucins into the in-testinal tract thereby removing boundpathogens from the intestinal cells (Daiet al. 2000).

It is well documented that somemembers of the natural microbiotaare able to degrade intestinal mucin,whereas others are able to stimulate mu-cus secretion (Kohler, McCormick, andWalker 2003). Mack and colleagues(1999) showed that certain probioticsare able to inuence the regulation ofmucin production. Growth of probiot-ics ( L. rhamnosus GG and L. plantarum 299V) with colon cells in laboratorystudies resulted in stimulation of mucinproduction. Furthermore, these strainswere effective at inhibiting enterohem-orrhagic E. coli attachment to mucus-producing intestinal cells, but not tonon-mucus-secreting cells, thus sug-gesting a protective role of mucin. Theability of probiotic strains to impact mu-cus production is an elegant example ofcommunication of the bacteria with thehost and may provide a mechanism by

which probiotics can decrease the likeli-hood of infection by pathogens.

Stabilization of Gut MucosalBarrier

The native microbes colonizing hu-man bodies play an important role inpreserving the integrity and function ofthe barrier between the contents of theintestine and the inside of bodies. Thisintegrity can be compromised by patho-genic bacteria, toxins, inammation, orstress, leading to intestinal permeabilityand unwanted transfer of bacterial (in-cluding resident microbiota) and dietaryantigens across the gut wall. This trans-fer leads to activation of immune re-sponses that can result in inammatoryand autoimmune disorders. Increasedgut permeability is a characteristicfeature of food allergies and immuno-inammatory gut diseases.

Recent studies have shown that in-take of specic probiotics is effectivein preventing and repairing damage tothe lining of the intestine. Some ani-mal studies have demonstrated reducedgut permeability associated with cowsmilk feeding (Isolauri et al. 1993) andinammatory bowel disease (Madsenet al. 2001). Probiotics may mediatethese effects in several ways: by inhib-iting damage to intestinal cell junc-

tions (Luyer et al. 2005; Montalto et al.2004); improving cell growth and sur-vival (Otte and Podolsky 2004); induc-ing mucin secretion (Mack et al. 2003);promoting tissue repair (Yamaguchi,Yan, and Polk 2003); decreasing bac-terial adhesion (Sherman et al. 2005);and secreting repair factors and nu-trients (e.g., short-chain fatty acids,polyamines, nitric acid, and stimulatingproduction of secretory immunoglobu-lin A [IgA]) (Viljanen et al. 2005). Theability of probiotics to produce factors

that directly strengthen intestinal barrierintegrity and protect against pathogenicbacteria also has been reported (Madsenet al. 2001).

G LOSSARYAcquired immunity . Immunity result-

ing from previous exposure to aninfectious agent or antigen (e.g.,vaccine).

Alimentary canal . The system, or or-gans, from the mouth to the anus thattakes in food, digests it to extractenergy and nutrients, and expels theremaining waste.

Arginine . A naturally occurring aminoacid.

Bacteriocin . A high molecular weight

protein or protein complex producedby bacteria that kills bacteria closelyrelated to the producer bacteria.

Colonization . The state of microbes be-coming established and growing ina particular environmental niche. Inthe intestinal tract, this state may beachieved through direct attachmentof the microbe to intestinal cells (ad-herence), or may be due to associa-tion with the mucus layer lining theintestinal wall.

Colonization resistance . Limiting ac-tion of the normal ora on coloni-zation of the bowel by potentiallypathogenic microorganisms.

Colony forming units (CFU) . Themeasure of the count of bacteriawhen assayed on solid growth me-dia, or an agar plate. The bacterialsuspension is diluted serially so thata small amount spread on the surfaceof an agar plate leaves between 30and 300 bacteria. The plate is in-cubated until each bacterial cell orchain of cells forms a colony visiblewith the naked eye. After guring inthe dilution factor, this assay resultsin a count (CFU) per ml or g of testproduct.

Crohns disease . A chronic, episodic,inammatory condition of the gas-trointestinal tract (mouth to anus)characterized by inammation andlesions.

Culture methodologies . Methods toenumerate microorganisms based

on the process of dilution to extinc-tion and subsequent plating on agarplates containing nutrients necessaryfor the growth and incubation underthe appropriate conditions of the par-ticular microbe being assayed.

Cytokines . Regulatory proteins andpeptides released by many differentcell types for the purpose of signal-ing other cells. Cytokines are partic-ularly important for the initiation and


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regulation of both innate and adap-tive immune responses. Cytokinesalso are involved in several devel-opmental processes during embryo-genesis.

Dairy products with lactose . Lactoseis a carbohydrate (composed of oneglucose molecule bound to one ga-

lactose molecule) unique to mam-malian milk. Its biological role is toprovide a carbohydrate source fornursing mammals. It is present atabout 5% by weight in cows milk.Lactose is naturally present in manydairy foods such as milk, yogurt, icecream, buttermilk, cottage cheese,and other soft cheeses. Aged cheesesusually do not contain signicantlactose because much of the lac-tose comes off with the whey duringmanufacture and the remaining lac-tose is fermented by cultures duringthe aging process. Lactose in yogurtis only partially fermented so thatyogurt has approximately the samelactose content as milk.

Dendritic cells . Immune cells that pro-cess and present antigens on theirsurface to other cells of the immunesystem. They are present in smallnumbers in tissues that are in contactwith the external environment, in-cluding skin, the inner lining of the

nose, lungs, stomach, and intestines.They also can be found at an imma-ture state in the blood. Once acti-vated, dendritic cells migrate to thelymphoid tissues where they interactwith T cells and B cells to initiateand shape the immune response.

Double-blind, clinical trial . Double-blind describes a clinical trial inwhich neither the subject nor theinvestigator (researcher) knows whattreatment/supplement a subject isreceiving.

Enteric coated . Enteric coating is abarrier applied to oral supplements/ drugs that controls the location in thedigestive system where the coatedsubstance is released. This processcan protect probiotics from exposureto harmful gastric acid and pancre-atic secretions and improve survivalthrough intestinal transit.

Genus . The taxonomic level of divisionbetween the Family and the Species.

Gram-positive , Gram-negative .Named by the Danish bacteriolo-gist who developed the test, theGram reaction is a color-bind-ing assay (purple Gram positive;pink Gram negative) that revealsfundamental cell surface character-istics of bacteria. All lactic cultures

(e.g., Lactobacillus, Bidobacterium,Streptococcus ) and most probioticgenera are Gram-positive.

Ileum . The most distal region of thesmall intestine.

Innate (nonspecic) immune system. The rst line of defense against in-fections, innate immunity does notdemonstrate immunological memo-ry. The innate immune system givesrise to the acute inammatory re-sponse and has limited specicity formicrobes. Phagocytes dominate thisresponse.

Irritable bowel syndrome . An intesti-nal disorder characterized most com-monly by cramping, abdominal pain,bloating, constipation, and diarrheathat cannot be attributed to any otherknown disease.

Jejunum . The middle section of thesmall intestine extending from theduodenum to the ileum. The jejunumis the longest section of the smallintestine.

Locally . In this context, locally refersto effects that occur at the site of ex-posure to probiotics. For example,the production of organic acids in thecolon by probiotic bacteria can in-hibit pathogens in the colon.

Luminal contents . The contents of thelumen, or the interior of the intesti-nal tract.

Meta-analysis . The process of usingstatistical methods to combine theresults of different studies on the

same topic to determine the strengthof overall conclusions that can bemade on the effect of the treatment.

Microbiota . Term that refers to thecollective body of microorganismsinhabiting an ecosystem. Sometimecalled microora, this latter termis not taxonomically correct becausebacteria are not plants or ora.

Mucin. A family of large, heavily gly-cosylated proteins, membrane-bound

or secreted on mucosal surfaces.Mucosal immune system . The im-

mune system associated with mu-cous membranes of the respiratory,gastrointestinal, and urogenitaltracts.

Oligofructose . Also known as fructool-igosaccharide, the prebiotic, oligo-

fructose, is a short chain of fructosemolecules that reaches the colon andserves as a substrate for growth ofcolonic microbes.

Pancreatic secretions . Secretions fromthe pancreas (pancreatic juice) con-taining digestive enzymes, includ-ing trypsinogen, chymotrypsinogen,elastase, carboxypeptidase, pancre-atic lipase, and amylase. The alkalinenature of pancreatic juice neutralizesgastric acid, improving digestive en-zyme effectiveness.

Peyers patch cell . Peyers patches,named after the 17th-century Swissanatomist Hans Conrad Peyer, areorganized lymphoid tissue (second-ary lymphoid organs) found in thewall of the small intestine.

Pouchitis . Inammation of a surgical-ly constructed pouch after removalof the large bowel, often in patientswith ulcerative colitis.

Prebiotics . Nondigestible substancesthat, when consumed, provide a ben-ecial physiological effect on thehost by selectively stimulating thefavorable growth or activity of a lim-ited number of indigenous bacteria.

Probiotics . Live microorganisms thatconfer a health benet on the hostwhen administered in adequateamounts.

Species . A taxonomic unit that for bac-teria refers to the degree of related-ness among individuals assigned tothe species, and the degree of dif-

ference among individuals of otherspecies. In general, members of thesame bacterial species are 95% simi-lar to each other based on their chro-mosomal DNA.

Strain . A designation for a specic indi-vidual member of a species. Strainsare genetically homogeneous. Manydifferent strains can be part of thesame species.

Synbiotic . A product containing both a


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probiotic and a prebiotic. Evidencefor synergy or specicity of the probi-otic for the prebiotic is not essential.

Systemically . In this context, systemi-cally refers to effects that occurdistant from the site of exposure toprobiotics (extra-intestinally). Theseeffects can be mediated by activated,

circulating immune cells.Transit time . The time it takes for

food and resulting digesta to movethrough the alimentary canal.

Ulcerative colitis . A form of inam-matory bowel disease that affects thelarge intestine or colon and is char-acterized by inamation and ulcersin the colon.

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