Biotherapeutics: Pre, pro and synbiotics in small animals (Proceedings)

The concept that microbes might be beneficial, rather than simply detrimental, to health, enjoys a long history of anectodal evidence. Among the oldest observations is the Old Testament recording of Abraham ingesting sour milk (Genesis 18:8). The purported health benefits of microbial products are varied and often substantiated by testimonials rather than science. However, increasing, hypothesis driven research is generating evidence based examples of therapeutic benefits of biotherapy, indicating that credence might be reasonably given to selected products for selected indications.The term "probiotic" was originally coined in 1965 to refer to substances secreted by one microorganism which stimulates the growth of another. The term has been modified at least 5 times, and continues to vary with the author. A reasonable working definition, as proposed by the WHO definition works best: Live microorganisms which when administered in adequate amounts confer a health benefit on the host (WHO (World Health Organization), 2006) In contrast to probiotics, prebiotics refer to non-digestible food ingredients (including dietary fiber) that beneficially effect the bacterial population. They differ from other fermentable carbohydrates in that they interact with selective microorganisms.. As such, organisms presumably associated with health benefits will be stimulated, shifting the composition of the intestinal microbiota toward beneficial organisms. For example, fructo-oligosaccharides and inulin, transgalactosylated oligosaccharides and soybean oligosaccharides selectively promote the growth of bifidobacteria. Metabonomics is the scientific discipline that studies compounds formed from prebiotics. Synbiotics contain both pre and probiotics, with "syn" implying a synergistic effect of the prebiotic on the probiotic portion of the combination products. Note that the prebiotic portion should have demonstrated positive effects on the specific probiotic component of the combination product should "match". Understanding the impact of probiotics is incumbent upon a description of the normal gastrointestinal microflora. The total body microbiota (human) includes 1012 to 1012 bacteria, outnumber other cells by approximately 10 fold; 30% of dry fecal matter represents microbes. The number of species in the gastrointestinal tract (again in humans) is estimated to be between 400 and 500. Only a fraction are conducive to culture; based on 16S rRNA sequencing, some of these unculturable microbes are closely related to those already identified whereas others represent totally new species. Their role is critical, protecting against invasion of pathogenic strains of bacteria, facilitating normal bowel smooth muscle function, supporting digestion of certain foods, and contributing to nutrition through the production of vitamins (vitamin K; vitamin B in some species) and other nutrients (eg, short-chain fatty acids).. Microbes impact the maturation and maintence of the intestinal immune system, influence cell proliferation and facilitate energy salvage (eg, through conversion of nutrients to short chain fatty acids). The catalytic potential of the microbiota may contribute to (or detract from) health through production of beneficial (or detrimental) metabolites. Among the organisms most commonly cited as beneficial are Lactobacillus and Bifidobacterium. Beneficial metabolites include the short chain fatty acid carbohydrate product butyrate (energy source, anti-inflammatory, immunosuppressant). Benefits of microbial fermentation of plant products include the production of enterolignans associated with estrogenic and antioxidant effects. Oxalobacter formingenes transforms oxalates; its absence increases the risk of oxalate stones in humans. Bacterial byproducts have a number of adverse effects. Protein byproducts, produced primarily in the distal colon, include ammonia and amines resulting from deaminations; these products are associated with pro-carcinogenic effects (eg, nitrosamines). Cysteine and methionine degradation yields sulfides, which inhibit colonic use of butyrate. Anaerobic colonic fermentation of aromatic amino acids tyrosine to phenols and tryptophan to indoles, and their subsequent metabolism, yields several procarcinogenic compounds. Bacterial deconjugation and dehydroxylation of bile acids contribute to their enterohepatic circulation; other compounds conjugated by compounds such as taurine, glycine, sulfate may likewise be recycled. Many of these determinental compounds play a role in hepatic encephalopathy as well.

The number and diversity of microbes increases with age, and with location in the gastrointestinal tract. Peristalsis and low pH contribute to lower microbial counts in the stomach, duodenum, and jejunum because of the low pH and peristalsis.. Numbers and diversity increase in the ileum (104-8 cfu/ml) and colon. The environment of the colon markedly differs from that in the small intestine: the pH is higher (5-6), epithelial turnover is slower, the bacterial count is higher and bacterial growth is more rapid, redox potential is lower, and intestinal/bacterial by products contribute to a higher short-chain fatty acid concentration. Within the colon, bacterial growth and fermentation are more rapid proximally compared to distally; in general, carbohydrates are fermented more proximally and proteins, distally. Diet will have a profound impact on the microbiota, with non digestible foods serving as sources of energy and carbon. Major sources of food for microbes include resistant starches, dietary fibers such as cellulose, hemicellulose, pectin and inulin, and unabsorbed sugars and sugar alcohols. Proteins from the diet or host serve as substrates. Food also impact host gastrointestinal function and health and thus, indirectly impacts the microbiota. The gastrointestinal flora of both humans and dogs has been described and compared) and the microbiota of the cat has been described (Rastall 2004). As a carnivore, the gastrointestinal tract of the dog is smaller. Similarities between human and dog include the proportion of Gram negative to anaerobic organisms, the make-up of Gram negative isolates, and floral behavior in response to pro or prebiotics. Genera that are numerous in both the GI tract of restricted access dogs and humans include Lactobacillus, Bifidobacterium, Eubacterium, Bacteroides, and Peptostreptococcus.The major difference between dogs and humans appears to be the proportion of Bifidobacteria; however, a study examining floral changes in response to different housing environments demonstrated that the number of Bfidobacteria dramatically increase under conditions in which exposure to the environment is controlled.. Under such conditions, major differences between the two species among culturable bacteria are limited and include: 1. Bacterioids and Streptococcus as the most common isolates in the ilea and colon of the dogs; however, these isolates, while not the predominant isolate in humans, are present in a large proportion. 2. Fusobacterium is not as numerous in dogs compared to man. In the cat, Bifodobacteria appear to be even less numerous than in the dog. The use of pre and probiotics in dogs has been reviewed. Lactobacillus acidophilus DSM 13241 was fed (2X109 CFU/d) was studied in 15 healthy dogs decreased the number of culturable Clostridum sp and was associated with an increases in indices indicative of immunomodulation (incrased serum IgG, monocytes, decrease plasma nitric oxide). Prebiotics which have been studied include lactosucrose and fructo-oligosaccharides. Feeding 1.5 g lactusucrose/d for 2 weeks to healthy dogs (n=8) increased Bifidobacteria and decreased Clostrdium sp. Bifidobacteria and Lactobacillus also increased and Clostridium and Enterobacteracea decreased In cats receiving 0.75 g/day. A decrease in toxin levels and odor also was described for both dogs and cats. Fructo-oligosaccharides fed at a rate of 4 g/day to adult health dogs (n=20) increased Bifidobacteria and Lactobacillus and decreased Clostridium sp. Although both lactate and butyrate increased, ammonia, dimethylsulfide, and hydrogen sulfide also increased. Fructo-oligosaccharides at a rate of 0.75% in the diet of adult healthy cats for two weeks decrease Clostridium sp and E. coli. Rastall (2004) also described the formulation of a symbiotic containing Lactobacilli cutulred from one dog, following the tradition of probiotics being based on microbes isolated from the target species. Among the isolates cultured, L. mucosae, L. acidophilus, and L. reuteri, were subjected to prebiotic carbohydrates.

Different strains of probiotic bacteria are presumed to impart differential effects, with variability representing habitat preferences of the microbe, specific capabilities of the microorganism and differential enzymes. In the gastrointestinal tract, four microhabitats have been described: the lumen, the mucus layer of epithelial cells, the surface of epithelial cells, and the crypts of the ileum, cecum and colon (which generally are colonized with motile, spiral-shaped organisms). In contrast to probiotics, prebiotics refer to non-digestible food ingredients (including dietary fiber) that beneficially effect the bacterial population. As such, they differ from other fermentable carbohydrates in that they interact with selective microorganisms. As such, organisms presumably associated with health benefits will be stimulated, shifting the composition of the intestinal microbiota. For example, fructo-oligosaccharides and inulin, transgalactosylated oligosaccharides and soybean oligosaccharides selectively promote the growth of bifidobacteria. Metabonomics is the scientific discipline that studies compounds formed from prebiotics. Synbiotics contain both pre and probiotics, with "syn" implying a synergistic effect of the prebiotic on the probiotic portion of the combination products. Note that the prebiotic portion should have demonstrated positive effects on the specific probiotic component of the combination product should "match".The proposed mechanisms by which probiotics work are several fold. Known effects of commensal intestinal bacteria include maturation and maintenance of the immune system, gastrointestinal epithelial cell proliferation, energy capture, and through production of metabolites, either both beneficial and detrimental health effects. The natural microflora prevents invasion by foreign bacteria. The number of CFU ingested as probiotics is minor compared to the normal microbiota. However, they transit through regions of the gastrointestinal tract that are sparsely populated and as such, may transiently become the dominant microbe. Pre, probiotics and synbiotics purportedly impart a number of beneficial effects as has been and continues to be demonstrated using a variety of in vitro and in vivo models. Probiotics have been demonstrated to have an impact on a large diversity of potential health problems.. Included are gingival disease, gastric infections by Helicobacter pylorus (and thus medical conditions associated with infection), pancreatitis, antibiotic-induced diarrhea, (most promise included Lactobacillus GG, Lactobacillus sporogenes, and Saccharomyces boulardii), infectious Diarrhea, (but not C. difficile colitis), inflammatory bowel disease, liver Disease, bacterial translocation, urinary tract infection and oxalate urolithiasis; the roll in allergic diseases is evolving.

Probiotics and related compounds are not approved drugs and undergo no premarket approval process. As such, data supporting quality assurance, safety and efficacy for each product may not exist. Consumer Laboratories (www.consumerlab.com) has reviewed issues specifically related to quality assurance of a probiotic product. These include: 1. Labeling. Labels should list all types of bacteria or yeast, including genus and species. Labels generally list the number of colony forming units (CFU); generally 1 to 10 billion (109 to 1010) CFU are recommended (in humans) per day. 2. Viability of organisms, which may decrease during time of manufacture versus purchase due to exposure to heat, moisture and oxygen; 2.presence of contaminating (potentially pathogenic) organisms, including E. coli, Salmonella spp, Staphylococcus aureus and Pseudomonas aeruginosa (as per FDA requirements); and 3. the extent of enteric protection of selected organisms, including L. bulgaricus, S. thermophilus and Leuconostoc and Lactococcus sp. Organisms which generally do not need protection include most Lactobacillus, Bifidobacterium and Streptococcus, or organisms present as spores, including Bacillus and some Lactobacillus. Of 24 products (21 human, 3 pet) reviewed by Consumer Laboratories (October 2007), 5 (4 human, 1 pet) failed to contain the labeled amount of microbes, 6 (4 human, 2 pet) failed to provide at least 109CFU per serving (generally those that also failed to contain labeled amount) and 1 (pet) failed due to microbial contamination (with mold). Two of the pet and 4 of the human products did not include the number of CFU on the lab.