MCB 100 Introductory Microbiology Spring 2019 CHAPTER 26

1. MCB 100 Introductory Microbiology Spring 2019 CHAPTER 26 APPLIED AND ENVIRONMENTAL MICROBIOLOGY Food Microbiology Industrial Microbiology Environmental Microbiology

2. Food Preservation – Canning *Commercial Sterilization vs. Biological Sterilization Biological sterilization means eliminating all living organisms from a given sample or environment. Commercial sterilization means freedom from Clostridium botulinum. The goal of commercial canning procedures is to ensure that a food product is free of the endospores from Clostridium botulinum, not necessarily free of all microbial life. Clostridium botulinumis important because: - it produces an extremely potent neurotoxin - grows anaerobically at room temperature - is fairly common in the soil - It makes endospores that are very resistant to heat A process that kills C. botulinumendospores kills most microbes. (Thermophillic bacteria that are more heat resistant than C. botulinum are usually not a problem in food preservation because they don't grow at room temperature.)

3. Botulism Which ONE of the following statements about Clostridium botulinum is FALSE? A. C. botulinum produces an extremely potent neurotoxin. B. C. botulinum grows at room temperature in the absence of non-combined oxygen (O2). C. C. botulinum is fairly common in the soil. D. C. botulinum endospores are killed very quickly by exposure to heat, 10 minutes at 63oC will kill them all. E. A process that kills Clostridium botulinum endospores will kill most microorganisms.

4. Botulism Which ONE of the following statements about Clostridium botulinum is FALSE? A. C. botulinum produces an extremely potent neurotoxin. B. C. botulinum grows at room temperature in the absence of non-combined oxygen (O2). C. C. botulinum is fairly common in the soil. D. C. botulinum endospores are killed very quickly by exposure to heat, 10 minutes at 63oC will kill them all. E. A process that kills Clostridium botulinum endospores will kill most microorganisms.

5. Food Preservation - Chemical Preservatives Various chemicals that prevent the growth of microorganisms can be added to foods as long as these chemicals: - pose no danger to the consumer and - do not damage the taste of the product. High concentrations of salt and sugar can inhibit microbial growth by reducing the water activity of the food. Some herbs naturally contain substances that inhibit the growth of bacteria including: garlic, cloves, cinnamon and oregano.

6. Pasteurization* Pasteurization is a form of heat treatment that kills certain pathogens or food spoilage organisms. The object of Pasteurization is to prevent the transmission of disease causing germs or to prevent the rapid spoilage of product without damaging the product by overcooking it. Pasteurization is not the same as sterilization in that it does not kill all of the microorganisms in the product. Temperatures below 100oC are used. For example: Milk is Pasteurized by heating to 63oC for 30 minutes or 72oC for at least 15 seconds. This treatment is lethal for Salmonella and Mycobacterium tuberculosis and thus prevents transmission of salmonellosis and tuberculosis in milk. Pasteurization is used to prolong shelf life of beer and to kill disease causing germs in fruit juices. In wine production Pasteurization of the grape juice kills undesirable microorganisms before the fermentation step. The combination of Pasteurizing the juice then inoculating it with a starter culture helps to insure the consistent production of good wine.

7. Pasteurization The application of a minimal amount of heat to a product to kill harmful microorganisms. Pasteurization is not the same as sterilization. Sufficient heat to kill all microorganisms in milk, juice, beer and other products ruins the taste and nutritional value of the product. Procedures that can be used in the pasteurization of milk 1. LTH - low temperature holding Milk is heated to 62.8oC for 30 minutes. (This is also called LTLT for low temperature, long time.) 2. HT - high temperature Milk is heated to 72oC for at least 15 seconds. (This is also called flash pasteurization.) 3. UHT - ultra high temperature Milk is heated to 149oC for a few seconds. 149oC is 300oF, which is well above boiling. (This treatment sterilizes the milk, so no refrigeration of the product is needed until after the package is opened. Disadvantages of UHT pasteurization are: some loss of nutrients, taste is affected and it is slightly more expensive than other techniques.)

8. Milk Microbiology Milk contains nutrients that can support the growth of many microorganisms. MILK CONTENTS - Protein (Proteins are a good carbon and nitrogen source. The predominant protein in milk is called casein, there are also immunoglobulins.) - Carbohydrates (Milk sugar, lactose, is a disaccharide: glucose+galactose.) - Lipids (butterfat) - Vitamins (Milk is naturally a source of B- vitamins, A and D may be added.) - Minerals (especially calcium) - Trace elements - water, - pH of about 7, - isotonic salt concentration Sources of microorganisms that may contaminate milk 1. Feed and water that the cows get 2. Bits of soil, dust, dirt and manure from the barn 3. Milking equipment may be heavily contaminated if not disinfected properly. 4. There are bacteria on the hide and udders of the cow. 5. Humans that are handling the milk may introduce microorganisms.

9. Steps taken to fight microbial contamination of milk 1. The barn and milking equipment is kept as clean as possible, hoses, tanks, and cows’ udders are regularly washed with disinfectant solution. This reduces the number of bacteria in the milk. 2. The milk is immediately cooled to 4oC to reduce the growth of the bacteria that do get into the milk. The milk is kept refrigerated continually, except during pasteurization. 3. The milk is pasteurized. This is a process of heating milk enough to kill pathogenic bacteria. It's not enough heat to sterilize the milk, since that would ruin its taste, but it reduces the number of bacteria in the milk and rids it of pathogens. 4. Milk is given an expiration date that accounts for the growth of microorganisms at refrigerator temperatures.

10. Cheese and Yogurt Production Milk sugar is converted to lactic acid by bacterial fermentation. Reduced pH causes precipitation of the protein casein to form curds. Cheese making begins with pasteurized milk to eliminate undesirable microorganisms. Starter cultures are then added to the milk: For yogurt the bacteria used includes: Streptococcus thermophilus and Lactobacillus bulgaricus The incubation temperature is warm. (43oC or 110 oF) For cheese, the starter culture used depends on the desired product. Several species of lactic acid bacteria are used, most commonly: Lactococcus lactis and Lactococcus cremoris (30oC) For ripened cheeses, the curds are pressed to remove water and aged to allow the microorganisms to continue producing flavorful compounds. Sharp cheddar cheese differs from mild cheddar because it is aged longer. Penicillium molds may be used to ripen soft cheeses.

11. Beer ProductionThe yeast Saccharomyces cerevisiae makes alcohol when fermenting sugars. Beer is a grain product. Most of the sugar found in grain is in the form of starch. Yeast don’t have the enzyme needed to break down starch.

12. Beer Production* Summary: Sugar from barley is converted to ethanol by yeast (fermentation). Problem One – get sugar from barley (yeast don’t break down starch) Malting: Barley is moistened and allowed to germinate. Enzymes in the seeds begin the process of breaking down starch to simple sugars. After a few days the barley is dried and ground to a powder (malt). Mashing: Malt and adjuncts (other grains) are mixed with water to dissolve the sugars. Enzymes from the barley seeds continue to convert amylose to maltose. The sugary liquid is called the wort. Problem Two – stop bacteria from ruining the beer Problem Three – fermentation products tend to taste sour, too sour Preparation for fermentation: Spent grain is removed from the sugary liquid (wort). Hops are added to the wort and the mixture is boiled. Fermentation: The wort is cooled to room temperature and yeast is added. The culture is incubated for a while with air bubbled through the liquid to allow the yeast to grow rapidly. Air flow is shut off and the yeast must shift to anaerobic metabolism (ethanol fermentation). Aging: a low temperature aging period is used in lager production.

14. Water Treatment* Potable Water = Safe Drinking Water - Free of dangerous pathogenic microorganisms - Free of dangerous concentrations of harmful chemicals Drinking Water and Pathogenic Microorganisms There are over 100 different bacteria, viruses and protozoa that cause infectious human diseases that may be transmitted in contaminated water. (Fecal-Oral cycle pathogens) Most of the pathogens found in water are associated with fecal contamination.

15. Potable water is: A. free of all chemicals. B. loaded with Salmonella and coliform bacteria. C. contaminated with high levels of organic matter D. as salty as water in the ocean. E. safe to drink.

16. Potable water is: A. free of all chemicals. B. loaded with Salmonella and coliform bacteria. C. contaminated with high levels of organic matter D. as salty as water in the ocean.E. safe to drink.

17. General Municipal Water Purification 1) Start with the best water obtainable. - Deep Wells - Protected Reservoirs 2) Initial Disinfection – (Algicide if needed) 3) Flocculating Agent – Aluminum sulfate or alum is added to raw water to cause small particles and colloids to stick to each other forming heavier particles or flocs which settle to the bottom of the clarifying tank. 4) Clear water is run through a filter consisting of an activated charcoal bed and layers of sand. The activated charcoal helps to absorb organic contaminants. A biofilm called the zoogleal mass develops in the upper layers of the sand filter. This biofilm helps trap microorganisms and degrades organic matter. 5) The filtered water is disinfected again, usually with chlorine, ozone or UV-light. 6) Treated water is stored and pumped throughout the water system. Positive pressure prevents contamination of the water main in the event of a small leak.

18. Examples of fecal oral cycle pathogens • Bacteria • - Salmonella typhi - typhoid fever • - Shigella dysenteriae and strains of E.coli - dysentery • - Vibrio cholerae - cholera • - Campylobacter jejuni – gastroenteritis • Helicobacter pylori – gastric ulcers • Viruses • - Polio – paralysis and neural complications • - Hepatitis A – inflammation of the liver, • cirrhosis • - Rotoviruses - diarrhea • Protozoa • Giardia lamblia - dysentery • Entamoeba histolytica - dysentery • Balantidium coli - dysentery • Cryptosporidium parvum - dysentery

19. Indicator Organisms Tests for the presence of specific pathogens are very complicated and expensive. There are many different pathogens that may contaminate drinking water and produce a risk to humans. A test for a specific bacterium, such as Salmonella typhi, would do nothing to detect other types of disease causing microorganisms. It is very difficult to culture and detect viral or protozoan pathogens. Instead water is tested for the presence of indicator organisms. The presence of these bacteria in water is an indication that the water has been contaminated with fecal material. The presence of disease causing microorganisms in water is tightly linked to fecal contamination. Escherichia coli

20. Indicator Organisms – Examples* • Coliforms – small Gram negative bacteria that ferment lactose making acid & gas in 24 hrs., non-spore forming • coliform = similar to E. coli (Enterobacter aerogenes etc.) • - Enterococci (fecal Streptococci) • small Gram positive cocci found in the intestine • Characteristics of the coliform bacteria • - coliforms are always present in the large intestines • and fecal material of animals (mammals, reptiles, birds) • - in contaminated water, coliforms are present in larger numbers than pathogenic bacteria • - coliforms survive in water longer than pathogenic bacteria • - coliforms are easy to cultivate • - coliforms produce acid and gas from lactose in 24 hours • - lactose fermentation on certain selective media is a presumptive identification of a coliform

21. Modern Sewage Treatment Raw sewage is screened and pumped into a primary settling tank, particulates and flocs settle to the bottom forming the primary sludge. The effluent is piped into a secondary treatment that promotes the degradation of organic compounds by aerobic microorganisms. In an activated sludge system the water is mixed with some recycled sludge from some earlier batch of sewage. This sludge is an active biomass of microorganisms that grow on the nutrients in sewage. The water is kept well oxygenated. In a trickle filter system the effluent is passed over rocks that are covered with a biofilm. The microbial community in the mat degrades organic matter in the sewage. Much of the organic matter in the sewage is converted to carbon dioxide in this treatment. Microorganisms are allowed to settle out as flocs. This removes more organic matter from the water. Sludge from the primary settling tank and trickle filter tank is put into an anaerobic digestion tank. The Imhoff tank is an anaerobic environment, this means that the conditions are highly reduced. Bacteria and Archaea in the Imhoff tank break down organic matter and produce carbon dioxide and methane. Methane is burned off or used for fuel. Effluent is chlorinated before being released.

22. General Steps in Sewage Treatment* Primary treatment: removal of coarse solids using screening, skimmers, grinders and sedimentation in a settling tank provides mechanical removal of objects in the water. Secondary treatment: digestion of solid and dissolved nutrients in the sewage by anaerobic and aerobic microorganisms (in two or more steps). Chemical disinfection: chemicals are added that kill pathogenic bacteria and solids are disposed of. BOD - Biochemical Oxygen Demand. BOD is a measurement of the amount of oxygen that is used up by aerobic bacteria in a water sample in a five-day period. A high BOD is caused by a high concentration of organic matter while a low BOD indicates a low concentration of organic matter. The goal of sewage treatment is to reduce the BOD, i.e. degrade the organic matter that is contaminating the waste water.

23. Typical Steps in Wastewater Treatment

24. Microorganisms play essential roles in biogeochemical cycles A biogeochemical cycle is a description of the conversions of an element from one form to another by living organisms and non-biological chemical reactions in the environment. These transformations typically involve redox reactions.

26. Carbon Cycle