Probiotics : An Innovative Globe

Probiotics are live microorganisms (in most cases, bacteria) that are similar to beneficial microorganisms found in the human gut.

They are also called “friendly bacteria” or “good bacteria”. Probiotics are available to consumers mainly in the form of dietary supplements and foods. They can be used as complementary and alternative medicine.

History of Probiotics:

Probiotics, which means "for life", have been used for centuries as natural components in health-promoting foods. The original observation of the positive role played by certain bacteria was first introduced by Russian scientist and Nobel laureate Eli Metchnikoff, who in the beginning of the 20^th century suggested that it would be possible to modify the gut flora and to replace harmful microbes by useful microbes¹.

Metchnikoff, at that time a professor at the Pasteur Institute in Paris, produced the notion that the ageing process results from the activity of putrefactive (proteolytic) microbes producing toxic substances in the large bowel. Proteolytic bacteria such as clostridia, which are part of the normal gut flora, produce toxic substances including phenols, indols and ammonia from the digestion of proteins. According to Metchnikoff these compounds were responsible for what he called “intestinal auto-intoxication”, which caused the physical changes associated with old age. It was at that time known that milk fermented with lactic-acid bacteria inhibits the growth of proteolytic bacteria because of the low pH produced by the fermentation of lactose. Metchnikoff had also observed that certain rural populations in Europe, for example in Bulgaria and the Russian Steppes who lived largely on milk fermented by lactic-acid bacteria were exceptionally long lived. Based on these facts, Metchnikoff proposed that consumption of fermented milk would “seed” the intestine with harmless lactic-acid bacteria and decrease the intestinal pH and that this would suppress the growth of proteolytic bacteria. Metchnikoff himself introduced in his diet sour milk fermented with the bacteria he called “Bulgarian Bacillus” and found his health benefited. Friends in Paris soon followed his example and physicians began prescribing the sour milk diet for their patients².

Henry Tissier, also from the Pasteur Institute, was the first to isolate a Bifidobacterium. He isolated the bacterium from a breast-fed infant and named it Bacillus bifidus communis³. This bacterium was later renamed Bifidobacterium bifidum. Tissier showed that bifidobacteria are predominant in the gut flora of breast-fed babies, and he recommended administration of bifidobacteria to infants suffering from diarrhea. The mechanism claimed was that bifidobacteria would displace the proteolytic bacteria that cause the disease.

German Professor Alfred Nissle, in 1917 isolated a strain of Escherichia coli from the feces of a First World War soldier who did not develop enterocolitis during a severe outbreak of shigellosis⁴. In those days, antibiotics were not yet discovered, and Nissle used the strain with considerable success in acute cases of infectious intestinal diseases (salmonellosis and shigellosis). Escherichia coli Nissle 1917 is still in use and is one of the few examples of a non-lab probiotic.

In 1920 Rettger demonstrated that Metchnikoff’s “Bulgarian Bacillus”, later called Lactobacillus bulgaricus, could not live in the human intestine⁵, and the fermented food phenomena petered out. Metchnikoff’s theory was disputable (at this stage), and people doubted his theory of longevity. After Metchnikoff’s death in 1916, the centre of activity moved to the US. It was reasoned that bacteria originating from the gut were more likely to produce the desired effect in the gut, and in 1935 certain strains of Lactobacillus acidophilus were found to be very active when implanted in the human digestive tract⁶. Trials were carried out using this organism, and encouraging results were obtained especially in the relief of chronic constipation.

The term “probiotics” was first introduced in 1953 by Kollath (see Hamilton-Miller et al 2003). Contrasting antibiotics, probiotics were defined as microbially derived factors that stimulate the growth of other microorganisms. In 1989, Roy Fuller suggested a definition of probiotics which has been widely used: “A live microbial feed supplement which beneficially affects the host animal by improving its intestinal microbial balance”⁷. Fuller’s definition emphasizes the requirement of viability for probiotics and introduces the aspect of a beneficial effect on the host.

In the 1960s the dairy industry began to promote fermented milk products containing Lactobacillus acidophilus. In subsequent decades other Lactobacillus species have been introduced including Lactobacillus rhamnosus, Lactobacillus casei, and Lactobacillus johnsonii, because they are intestinal species with beneficial properties⁸.

What Probiotics Are?^{9, 10}

Experts have debated how to define probiotics. One widely used definition, developed by the World Health Organization and the Food and Agriculture Organization of the United Nations, is that probiotics are “live microorganisms, which, when administered in adequate amounts, confer a health benefit on the host.” (Microorganisms are tiny living organisms—such as bacteria, viruses, and yeasts—that can be seen only under a microscope.)

Probiotics are not the same thing as prebiotics—non digestible food ingredients that selectively stimulate the growth and/or activity of beneficial microorganisms already in people's colons. When probiotics and prebiotics are mixed together, they form a synbiotic.

Probiotics are available in foods and dietary supplements (for example, capsules, tablets, and powders) and in some other forms as well. Examples of foods containing probiotics are yogurt, fermented and unfermented milk and some juices and soy beverages. In probiotic foods and supplements, the bacteria may have been present originally or added during preparation. Most probiotics are bacteria similar to those naturally found in people's guts, especially in those of breastfed infants (who have natural protection against many diseases). Most often, the bacteria come from two groups, Lactobacillus or Bifidobacterium. Within each group, there are different species (for example, Lactobacillus acidophilus and Bifidobacterium bifidus), and within each species, different strains (or varieties). A few common probiotics, such as Saccharomyces boulardii, are yeasts, which are different from bacteria.

Scanning electron micrograph (SEM) of Bifidobacterium bifidum.

Color enhanced scanning electron micrograph (SEM) Saccharomyces boulardi (large cells) found of the bacteria Lactobacillus acidophilus. A spirochete along with bacteria in fermented fruit juice.
Bacterium can also be seen at center.

Table 1 Successful Probiotic Bacteria and their Reported Effects²⁰

What the Science Says:

Scientific understanding of probiotics and their potential for preventing and treating health conditions is at an early stage, but moving ahead. In November 2005, a conference that was cofunded by the National Center for Complementary and Alternative Medicine (NCCAM) and convened by the American Society for Microbiology explored this topic. According to the conference report, some uses of probiotics for which there is some encouraging evidence from the study of specific probiotic formulations are as follows:

• To treat diarrhea (this is the strongest area of evidence, especially for diarrhea from rotavirus)
• To prevent and treat infections of the urinary tract or female genital tract
• To treat irritable bowel syndrome
• To reduce recurrence of bladder cancer
• To shorten how long an intestinal infection lasts that is caused by a bacterium called Clostridium difficile
• To prevent and treat pouchitis (a condition that can follow surgery to remove the colon)
• To prevent and manage atopic dermatitis (eczema) in children.

Some other areas of interest to researchers on probiotics are:

• What is going on at the molecular level with the bacteria themselves and how they may interact with the body (such as the gut and its bacteria) to prevent and treat diseases. Advances in technology and medicine are making it possible to study these areas much better than in the past.
• Issues of quality for example, what happens when probiotic bacteria are treated or are added to foods—is their ability to survive, grow, and have a therapeutic effect altered?
• The best ways to administer probiotics for therapeutic purposes, as well as the best doses and schedules.
• Probiotics’ potential to help with the problem of antibiotic-resistant bacteria in the gut.
• Whether they can prevent unfriendly bacteria from getting through the skin or mucous membranes and traveling through the body (e.g., which can happen with burns, shock, trauma, or suppressed immunity).

Microorganisms Used As Probiotics:

The most frequently used probiotics found in dairy-based food products are Lactobacilli and bifidobacteria. Since these belong to the indigenous human microflora, they have a long history of safe use and there are evidences to support their positive roles, some of these have been designated as GRAS (generally regarded as safe) by the FDA (Food and Drug Administration) due to their long history of use in food fermentations¹⁴. Lactobacillus, first identified by Pasteur, was the first genus of bacteria suspected to have health benefits, rather than to be agents of disease. Metchnikoff had introduced his intoxication theory, which stated that the main cause of ageing are ‘toxicants’ formed by intestinal putrefaction and fermentation, and proposed a diet containing milk fermented by lactic acid bacteria. Thus Lactobacilli suddenly attracted attention of the scientific community as well as common man. Lactobacillus acidophilus, found in yogurt, was believed to be responsible for the longevity of some eastern Europeans who traditionally consumed it a s a staple food.

In the 1920’s and 1930’s, Lactobacillus acidophilus was used in the form of acidophilus milk to treat constipation and diarrhea in the USA. Although Lactobacillus acidophilus is the most popular species among the Lactobacilli group of probiotic bacteria, other Lactobacillus sp. Have been found to be beneficial as well e.g. L. rhamnosus, L. plantarum, L. casei. Among Bifidobacterium species, the important ones are B. bifidum, B. breve and B. lactis. There are three major guardians that live and work inside the human intestinal tract. Lactobacillus acidophilus guards the small intestine, Bifidobacteria protect the large intestine, whereas Lactobacillus bulgaricus is a traveling transient bacterium that aids the other two as it passes through the body. The interest in the use of strains other than the traditional lactic acid bacteria (Lactobacilli and Bifidobacteria) is increasing. For Example, the yeast Saccharomyces boulardii, nonpathogenic Escherichia coli, streptococcus thermophilus and complex mixture of intestinal microbes have been studied for their use as probiotics¹¹.

Beneficial Effects of Probiotics:

Probiotics are purported to impart numerous nutritional and therapeutic benefits to the host^{11, 13}. In some instance, the probiotic microbes function for the benefit of the host, such as antimicrobial mechanisms. While in other cases, the probiotic microbes may trigger the indigenous microbes or the host physiology to induce the action. One probiotic preparation maybe, but need not be, multi-functional. The beneficial attributes of probiotics, presented in Table 2, can be broadly grouped as: antimicrobial, biochemical, physiological and immunological.

Table 2 The Beneficial Attributes of Probiotics¹⁰

• Enhancement of resistance against pathogens.
• Stimulation of immune system.
• Improve lactose absorption in lactose intolerant.
• Prevention and treatment of diarrhea (traveller’s, pediatric, antibiotic-associated and diarrhea secondary to Clostridiumdifficile).
• Prevention or reduction of constipation.
• Reduction of irritable bowel syndrome.
• Regression of tumors and reduction in carcinogen and mutagen production.
• Reduction in recurrence rate of female urinary tract and vaginal infections.
• Production of nutraceuticals and vitamins.
• Reduction in serum cholesterol, blood pressure, management of diabetes and prevention of osteoporosis.
• Probiotics are considered good pharmaceutical delivery systems. For example, the use to Lactobacillus sp. as live vectors in oral vaccination.

Antimicrobial Mechanisms:

These refer to the action of the probiotic preparation on another microbe or group of microbes. This may be helpful directly for enhanced resistance against intestinal pathogens and in the prevention of diarrhea. The interactions may include competitive colonization (the probiotic strain can successfully out compete the pathogen for either nutrients or the site of colonization) as well as adhesion and growth inhibition. As the first step in infection by most gastrointestinal pathogens is attachment to the intestinal mucosa, the inhibition of adhesion would be beneficial to the host. During growth of the probiotic, various compounds may be produced which are inhibitory to the pathogen’s growth. These include organic acids such as lactic and acetic acids, bacteriocins and reuterin. The organic acids not only lower the pH, thereby affecting the growth of the pathogen, but they can also be toxic to the microbes. Reuterin, produced by Lactobacillus reuteri, is inhibitory to a broad range of cells. Lactobacilli are known to produce many types of bacteriocins like acidophilin, acidolin, lactocidin, bulgarican, lactolin, lactobacillin and lactobrevin. They can either have a broad range of activity or may specifically inhibit the growth of a very limited range of closely related microbes. Probiotic microbe s also affect urinogenital health in women, since urinary and genital tract infections are often associated with colonic bacteria. The consumption of probiotics has been linked to a reduced recurrence of Candida infections and bacterial vaginities.

Table 3 Microbial probiotics and their safety status¹⁹

Biochemical Effects:

These include;

(i)the reduction of faecal enzymes that can convert pro-carcinogens to caricinogens in the digestive tract,

(ii)decrease of lactose intolerance,

(iii)reduction of serum cholesterol,

(iv)relieve constipation

(v) Improve the quantity, availability and digestibility of some dietary nutrients.

The ingestion of Lactobacilli is known to result in the reduction of faecal enzymes such as b-glucuronidase, azoreductase and nitroreductase in humans, which are capable of converting pro-carcinogens to carcinogens in the digestive tract. Thus, they lower the chances for tumor development. Lactose intolerance is seen in persons lacking the enzyme lactase (b-galactosidase). The symptoms include abdominal pain and osmotic diarrhea after eating foods high in lactose, since the lactose is not degraded and absorbed in the upper regions of the small intestine and thus used by the indigenous microbiota, resulting in production of gases and organic acids responsible for the characteristic symptoms of lactoseintolerance. Probiotic microbes containing lactase (S. thermophilus, L. bulgaricus, etc.) can degrade lactose before it reaches the indigenous microbes in the lower part of the small intestine.

Probiotic microbes can also be useful in the treatment of hypercholesterolemia. There are reports that probiotics like Lactobacilli can assimilate cholesterol and deconjugate bile acids, leading to a reduction in the serum cholesterol levels. There is also some preliminary evidence on antihypertensive effects of probiotics, and thus documenting their role in blood pressure control. Fermented milk containing Saccharomyces cerevisiae and Lactobacillus helveticus has been found to be effective. Thus, regular consumption of probiotics may provide a modest prophylactic effect against heart disease. There is an evidence to support the role of lactic acid bacteria in improving intestinal mobility and treating constipation, possibly through a reduction in gut pH. Microbial fermentations tend to increase the content of various nutrients in the food. Termed as bio-enrichement, it involves enrichment of food value by supplementation with proteins, essential amino acids, minerals and vitamins. For example, lactic acid bacteria increase folic acid in yogurt, bifidus milk and kefir, niacin and riboflavin levels in yogurt, vitamin B₁₂ in Cottage cheese, and vitamin B₆ in Cheddar cheese. Probiotic microbes may release various enzymes into the intestinal lumen, thus aiding the digestion process.

Physiological Mechanisms:

These refer to the influence of probiotic microbes on the host response, including stimulation of immune system and reduction of risk of colon cancer. Probiotics can enhance both specific and nonspecific immune responses, without eliciting a harmful inflammatory response. Their effects are mediated through activation of macrophages by increasing levels of cytokines, natural killer cell activity and/or levels of immunoglobulin. The cell components of Lactobacillus are known to stimulate the immune response, which tends to protect the host from infection as well as from conditions involving the immune response, such as irritable bowel syndrome and colon cancer. The binding of probiotic microbe may trigger receptors to initiate signals that result in the synthesis of cytokines (pro-inflammatory proteins released by immune cells), and thus enhancing immune function. As extensively studied probiotic bacterium, Lactobacillus GG (LGG) colonizes the GIT in 1-3 days after consumption, and thus, reduces infections like Clostridium difficile, pediatric diarrhea, traveler’s diarrhea and antibiotic-associated diarrhea. The intestinal immunity is also enhanced by LGG due to increase in the number of immunoglobulin cells in the mucosa. This might be useful against the causative agents of inflammatory bowel disease and ulcerative colities. Probiotic microbes might even serve as efficient delivery vehicles to deliver drugs, vitamins or vaccines. Lactobacillus lactis has been genetically modified to secrete the anti-inflammatory molecule interleukin-10 to reduce colitis in mice¹⁴.

Who Benefits Most from Probiotics?¹⁵

The people who benefit most from probiotics are those who are most susceptible to infections, those who have disorders involving diarrhea, those people taking antibiotics, and pregnant and nursing mothers, and people undergoing radiation treatments of the pelvis or abdomen^{16, 17}. These people will see the greatest difference in their health by using probiotics regularly.

Ideally, everyone would be consuming probiotics or a fermented food which contains beneficial organisms. When you stop consuming probiotics it takes just a short time before all of the bacteria from the probiotic are gone from your system. No probiotic has been found which permanently colonizes the intestinal tract.

Future Research In The Probiotics Area¹⁸

• Determine the physiological role, mechanisms of action, and extent of influence of probiotics in human health using human feeding studies. Studies on high-risk human populations for colon cancer or cancer recurrence would be a possible target for some studies.
• Validate biomarkers used for assessing probiotic function. Testing of predictions based on biomarker studies with actual results in human clinical evaluations is needed. Biomarker validation in the areas of immune system, cancer, and gut micro-ecology is especially important. Once validated, biomarkers will be useful tools to assess dose-dependence and strain-specific responses. Biomarkers commonly used to select strains for probiotic use (adherence in tissue cultures, cholesterol assimilation, competitive exclusion of pathogens in tissue culture, inhibitor [bacteriocins, organic acids] production, lactase activity) have never been tested in controlled studies to determine if mutants without the characteristic perform any differently in clinical evaluations.
• Assess effects of probiotics on populations and activity of gut microbes. The application of gene based methods holds much promise in this field.
• Determine the role of probiotics as part of a whole food compared to isolated component.
• Improve reliability and ease of taxonomic classification of probiotic bacteria. Improve strain performance and activity.
• Conduct studies with consumers to understand how best to communicate the concept of probiotics and to determine favorable probiotic formats.
• Conduct research to improve product formats, consumer acceptance, stability, and efficacy of probiotic-containing products.

References:

1. Metchnikoff, E. 1907. Essais optimistes. Paris . The prolongation of life. Optimistic studies. Translated and edited by P. Chalmers Mitchell. London : Heinemann, 1907.
2. Vaughan, RB. 1965. The romantic rationalist: A study of Elie Metchnikoff. Med Hist., 9:201-15.
3. Tissier, H. 1900. Recherchers sur la flora intestinale normale et pathologique du nourisson. Thesis, University of Paris , Paris , France .
4. Nissle, A. 1918. Die antagonistische Behandlung chronischer Darmstörungen mit Colibakterien. Med Klin 1918, 2:29-30.
5. Cheplin, H.A., and L.F. Rettger, 1920. Studies on the transformation of the intestinal flora, with special reference to the implantation of Bacillus acidophilus, II. Feeding experiments of man. Proc Natl Acad Sci U S A. 1920 December; 6(12): 704–705.
6. Rettger, L.F., W.N. Levy, L. Weinstein, and J.E. Weiss. 1935. Lactobacillus acidophilus and its therapeutic application. Yale University Press, New Haven .
7. Fuller, R. 1989. Probiotics in man and animals, J. Appl. Bacteroil., 66:365-378.
8. Tannock, G.W. 2003. Probiotics: Time for a dose of realism. Curr. Intest. Microbiol. 4:33-42.
9. http://nccam.nih.gov/health/probiotics
10. Parvinder Kaur and T. Satyanarayana*, “Probiotics: A Beneficial Health Option”, Everyman’s Science VOL. XXXIX NO. 4, October— November’04, pp-224-229
11. Conway, L., Asia Pacific J. Clin. Nutr., 5, 10-14, 1996.
12. Teitelbaum J.E. and Walker W.A., Annu., Rev. Nutr. 22, 107-138, 2002.
13. Kopp-Hoolihan, L. J. Amcr Diet. Assoc., 101 , 229-238, 2001.
14. Abott, A., Nature, 427, 284-286, 2004.
15. http://www.hacres.com/diet/articles/Probiotics
16. Delia P, Sansotta G, Donato V, Messina G, Frosina P, Pergolizzi S, et al. Prevention of radiation-induced diarrhea with the use of VSL#3, a new high-potency probiotic preparation. Am J Gastroenterol 2002;97:2150-2.
17. Urbancsek H, Kazar T, Mezes I, Neumann K. Results of a double-blind, randomized study to evaluate the efficacy and safety of Antibiophilus in patients with radiation-induced diarrhoea. Eur J Gastroenterol Hepatol 2001;13:391-6.
18. The Institute of food technologists, Food technology, Vol. 53, no. 11, November 1999.
19. Chukeatirote, E., Songklanakarin, J. Sci. Technol., 2003, 25(2) : 275-282.
20. G.C. O'Sullivan*, P. Kelly, S. O'Halloran, C. Collins, J.K. Collins, C. Dunne and F. Shanahan, Current Pharmaceutical Design , 2005, 11, pp-3-10.

About Authors:

Vikash Kumar
SBMN Institute of Pharmaceutical Sciences & Research, Asthal Bohar, Rohtak
Mobile- 09896439451, H. No 78, Type-II, M. D. U. Campus Rohtak (Haryana).
E-mail: vikasruhilo1@gmail.com, vikasruhil01@rediffmail.com

Balvinder Singh
SBMN Institute of Pharmaceutical Sciences & Research, Asthal Bohar, Rohtak