1. Microbial Growth In Cows, Humans, & Other Animals

2. Ruminant Microbiota Cows, sheep, goats, etc. evolved to forage on grasses. Rumen precedes stomach. In developed countries, cows are fed grain (esp. corn) to speed up growth. The normal microbiota are used to grass (high fiber, low starch).

3. Grass-fed Microbiota Fibrobacter, Ruminococcus, Eubacterium, and other genera reside in the rumen. They possess the enzymes to digest the fiber into monosaccharides Animal cells do not Many of these bacteria are pH-sensitive Russell & Rycklik (2001). “Factors that Alter Rumen Microbial Ecology.” Science 292: 1119-22.

5. Switching to Grain Promotes new types of fermentation. –Acetic & butyric replaced by lactic. Acids build up in rumen, lowering the pH. –~7.0 lowered to 4.6. Metabolism of fiber is compromised.

6. Altered Bacterial Profile Growth of new, often pathogenic, bacteria is promoted. Also pathogenic in humans! –E.coli O157:H7, Campylobacter. Antibiotics in animal feed. –Bacteria evolve resistance.

7. Environmental Conditions pH Temperature Osmolarity Oxygen levels Nutrients Toxins

8. Oxygen Obligate aerobes – Mycobacterium Facultative anaerobes – E.coli, enterics Obligate anaerobes – Clostridium Aerotolerant anaerobes – Lactobacillus, Streptococcus pyogenes Microaerophiles – Micrococcus luteus

9. (a)Obligate aerobes; (b) obligate anaerobes; (c) facultative (an)aerobes; (d) microaerophiles; (e) aerotolerant anaerobes See also table 6.1

10. Detoxifying Oxygen Superoxide Dismutase (SOD), Catalase, Peroxidase Superoxide free radicals: O 2 - O 2 - + O 2 - + 2H +  H 2 O 2 + O 2 (SOD) 2 H 2 O 2  2 H 2 O + O 2 (catalase) H 2 O 2 + NADH + H +  2 H 2 O + NAD + (peroxidase)

11. Catalase Test

12. Detoxifying Oxygen Obligate anaerobes lack all 3 enzymes Obligate aerobes always produce these enzymes Facultative anaerobes do so under aerobic conditions Aerotolerant anaerobes can produce SOD Microaerophiles are sensitive to high O 2

13. The Human Gut

14. Genera & Species in your Gut Lactobacillus spp. Bifidobacterium spp. Bacteroides fragilis Bacteroides oralis Bacteroides melaninogenicus Clostridium perfringens Clostridium septicum Clostridium tetani Pseudomonas aeruginosa Escherichia coli Enterococcus faecalis Salmonella enteritidis Klebsiella pneumoniae Enterobacter aerogenes Proteus mirabilis Staphylococcus aureus Streptococcus spp. Lactococcus lactis

16. Human Intestinal Microbiota 90 – 95% are obligate anaerobes Bifidobacterium, Clostridium, Eubacterium, Fusobacterium, Peptococcus, Peptostreptococcus and Bacteroides Up to 10% are facultative/aerotolerant anaerobes Lactobacillus, Escherichia coli, Klebsiella, Streptococcus, Staphylococcus and Bacillus Pseudomonas – only known obligate aerobe 10 11 – 10 12 CFUs per ml ~1.2 kg

17. Lactobacillus and Bifidobacterium: –control pH. –Produce bacteriocins. –Take up space. –Interact with WBCs. –Interfere with quorum sensing. Intestinal microbiota produce many vitamins and digest certain carbs.

18. Scientific Findings? Lactobacillus interferes with pathogens. Probiotic treatments can alleviate symptoms of infant diarrhea. The Microbiome.

19. Conversations with the Host Stimulate production of mucus. Interact with the immune system. Interfere with activities of pathogens. Xavier & Podolsky (2000). “How to Get Along – Friendly Microbes in a Hostile World.” Science 289: 1483-4. Neish et al. (2000). “Prokaryotic Regulation of Epithelial Responses by Inhibition of I  B- Ubiquitination.” Science 289: 1560-3.

21. Newer Experiments Germ-free mice. –Gnotobiotic studies. Single organism studies. Microbiome transplants.

22. “Germ-Free” Animals Decreased resistance to infection. Chronic inflammation. Used for gnotobiotic studies.

23. “Germ-Free” Mice

25. The role of the immune system in governing host-microbe interactions in the intestine. Brown et al. (2013). Nature Immunology 14, 660–667.

26. Bacteroides fragilis Gram stain Example of a single organism study.

28. Bacteroides fragilis Polysaccharide A (PSA) regulates immune response. B. fragilis eases symptoms of ulcerative colitis (induced by infection) in mice. So does purified PSA.

29. Obesity Study Fecal transplantation from human to mice. Two groups: obese adults and lean adults. Effects on mouse weight gain. Ridaura et al. (2013). Gut Microbiota from Twins Discordant for Obesity Modulate Metabolism in Mice. Science 6 September 2013: Vol. 341 no. 6150.

31. More Connections Autism. Autoimmunity. Allergies. Diabetes. What else will be discovered?

32. Probiotics Foods with live microbes that promote good health. Yogurt, kefir, raw sauerkraut, miso, kombucha, and more. [Pasteurized products – no live beneficial organisms.] OTC pills – freeze-dried, live microbes.

34. Probiotic Organisms Many Lactobacillus species Bifidobacterium Eubacterium Normal residents of the human GI tract.

35. Probiotics Ingestion may lead to persistent or transient infection. Either result can help immune system. Probiotic may secrete something that affects immune regulation.

36. How do they get there? The fetus is possibly sterile. Vaginal births: initial colonization (vagina has similar microbes to the intestine). –Vaginal fluids contain prebiotics. Caesarian births: altered microbial profile.

38. Developing Microbiota Breast-feeding. –Breast milk contains probiotics and prebiotics. –Primarily Bifidobacterium. Solid foods: alters microbial profile. Development affected by antibiotic use, infections, diet, and more. http://www.connecticutcenterforhealth.com/baby.html http://www.pdrhealth.com/drug_info/nmdrugprofiles/nutsupdrugs/p ro_0034.shtml