Applied and Industrial Microbiology

Applied and  Industrial Microbiology The metabolic activities of microorganisms shape much of our environment, and microbial reactions are essential to life on Earth. Bacteria and fungi in particular are capable of many metabolic processes that are useful to humans. In this chapter, we examine how the vast metabolic capability of microbes is harnessed to serve human needs. (Chapter 8 discusses some aspects of recombinant DNA technology that give organisms enhanced or new functions to make them more valuable in industry, agriculture, or medicine.) Applied microbiology, the commercial use of microorganisms, encompasses two distinct fields: food microbiology, which includes the use of microorganisms in food production and the prevention of food spoilage and food-related illnesses, and industrial microbiology, which involves both the application of microbes to industrial manufacturing processes and solutions to environmental, health, and agricultural problems. We begin by considering food microbiology. Bread Food Microbiology Fermented Vegetables The Roles of Microorganisms in Food Production LE A R N I N G O U TC O M E 25.1 Describe how microbial metabolism can be manipulated for food production. Biochemists use the word fermentation to refer to the partial oxidation of sugars to release energy using organic molecules as electron acceptors (see Chapter 5). In food microbiology, however, fermentation may refer to any desirable changes that occur to a food or beverage as a result of microbial growth. In contrast, spoilage denotes unwanted change to a food that occurs from undesirable metabolic reactions, the growth of pathogens, or the presence of unwanted microorganisms. Fermentative microbes naturally occur on grains, fruits, and vegetables. In antiquity, people relied on these naturally occurring microbes to produce fermented foods and drinks. However, the same microbes are not always present on a food from harvest to harvest, yielding varying results. Most modern commercial food and beverage production relies on starter M25_BAUM7206_05_SE_C25.indd 778 Saccharomyces cerevisiae (sak-ă-rō-mī′sēz se-ri-vis′ē-ī) metabolizes sugars to leaven1 bread. Bakers add the yeast to flour, salt, and other ingredients to make dough, which is kneaded to introduce oxygen. The dough rises when metabolic reactions of yeast release CO2 , producing expanding pockets within the dough. Ethanol produced by fermentation evaporates during baking. Sourdough bread is made using starter cultures consisting of yeast and lactic acid bacteria. Lactic acid produced by the bacteria gives sourdough its characteristic taste. People around the world ferment many types of vegetables. Most of these vegetable products are the result of the actions of lactic acid bacteria, such as Streptococcus (strep-tō-kok′ŭs), Leuconostoc (loo′kō-nos-tŏk), Lactobacillus (lak′tō-bă-sil′ŭs), or Lactococcus (lak-tō-kok′ŭs), which specifically produce lactic acid during fermentation. Lactic acid acidifies the food and produces a “sour” flavor, as in sourdough bread. Food products derived from the fermentation of cabbage include Korean kimchi (kim-chē′) and German sauerkraut. Soy sauce is made by the fermentation of soybeans and wheat by lactobacilli, yeast (S. cerevisiae), and the fungus Aspergillus oryzae (as-per-jil′ŭs o′ri-zī). Chocolate is derived from the fermentation of cacao seeds. Coffee production relies on natural fermentation to release the coffee bean from the outer layers of the coffee berry so that the bean can then be dried and roasted. Pickles are another common fermented food. People often equate “pickles” with cucumbers, but other foods, such as beets and eggs, can also be pickled. Pickling refers to the process of preserving or flavoring foods with brine (saturated salt water) or acid. Microbial fermentation can be the source of the acid— as it is with dill pickles. Various spices can be added to the pickling solution to enhance taste. Because pickled foods are acidic, few pathogenic microorganisms survive, making pickling an excellent preservation method. Fermented foods are not made for human consumption alone. Silage, a product used as animal feed on many farms, is made by the natural fermentation of potatoes, corn, grass, grain stalks, or other types of green foliage. The vegetation is cut up and stored in silos that are closed to air. Under such moist, anaerobic conditions, the vegetation in the silos ferments, producing many organic compounds that make the silage aromatic and tasty to livestock and more easily digested by them. Some farmers produce silage from baled hay and grasses wrapped in plastic. ­ Microorganisms are involved in producing many of our favorite foods and beverages, from bread to wine to yogurt. Indeed, fermented foods are among the oldest foods known and are culturally very diverse. The characteristic flavors, aromas, and consistencies of such foods result from the presence of acids or sugars made by microbes during fermentation. Besides conferring taste and aroma, microbial metabolism also acts as a preservative, destroys many pathogenic microbes and toxins, and, in some cases, adds nutritional value in the form of vitamins or other nutrients. In the following sections, we explore how our knowledge of microbial growth and metabolism enables us to use microbes in food production and to control microbial activity that results in food spoilage. cultures composed of known microorganisms that perform specific fermentations consistently. In addition to an initial starter culture, secondary cultures may be added to further modify the flavor or aroma of foods. For example, the same starter culture is used to initiate formation of both Swiss cheese and blue cheese; the two cheeses differ because different secondary cultures are used in their production. 1From Latin levare, meaning “to raise.” 11/23/16 3:44 AM       Food Microbiology Fermented Meat Products Unless dried, cured, smoked, or fermented in some way, meat has a tendency to spoil rapidly. The combination of fermentation and drying or smoking has been used for centuries to preserve meats, particularly pork and beef. Dry sausages, such as salami and pepperoni, are made by grinding meat, mixing in various spices and starter cultures, allowing the mixture to ferment in the cold, and then stuffing it into casings. Fermented fish—common in Asian countries—are prepared by grinding fish in brine. Naturally occurring microbes ferment the mixture. The solids are removed and pressed to form a fish paste, while the liquid portion is drained off and mixed with flavoring agents to form fish sauce. Pasteurization kills unwanted microorganisms. Addition of starter bacterial culture Coagulation of milk proteins (curd formation) Fermented Dairy Products Disposal of liquid whey as waste product Production of unripened cheeses Cutting of curds ­ ­ Milk fermentation relies, for the most part, on the activities of lactic acid bacteria. Milk in an udder is sterile, but because milking introduces microorganisms, we pasteurize “raw” milk. The metabolic products of starter cultures give fermented milk products their characteristic textures and aromas. Buttermilk is made from fat-free milk (skim milk) by adding a starter culture of Lactococcus lactis (lak′tis) subspecies cremoris (kre-mōr′is) and Leuconostoc citrovorum (sit-rō-vō′rum). Yogurt production utilizes starter cultures of Streptococcus thermophilus (ther-mo′fil-us) and Lactobacillus bulgaricus (bulgă′ri-kŭs). Yogurt manufacturers mix pasteurized milk, milk solids, sweeteners, and other ingredients to a uniform consistency and then add the starter culture. They ferment the mix at 43°C and then cool it to stop microbial activity. They may add flavors prior to packaging. The microorganisms are left alive and may have beneficial effects; they are so-called probiotics; that is, live microorganisms administered to improve health and prevent disease. Cheeses can be hard or soft and mild or sharp. Regardless of the end product, the cheese-making process typically begins with pasteurized milk (FIGURE 25.1). One pound of cheese requires about five gallons of milk. Cultures of Lactococcus lactis coagulate protein in milk to form curds (solids) and whey (liquids). For cheeses sold as soon as they are made (called unripened cheeses, such as cottage cheese), the curd is removed, cut into small pieces, and packaged. Alternatively, the curdling process can be accelerated by the addition of the enzyme rennin, a type of protease (protein-digesting enzyme). Rennin was originally obtained from the stomach of cud-chewing animals, but now most of it is produced by the genetically modified fungus Aspergillus. Other cheeses (so-called ripened cheeses) are aged until they have the desired texture or taste. Hard cheeses, such as Parmesan and cheddar, are pressed until little water remains, while soft cheeses retain enough moisture to make them spreadable. Once curds are pressed, a cheese is aged for months to years while continued microbial activity imparts characteristic smells and tastes. Cheese producers utilize numerous different microorganisms and their enzymes along with various ripening regimens and culture conditions to make numerous, distinct types of cheeses. The average American ate 34 pounds of cheese in 2014, a clear sign of the size and importance of this microbe-dependent industry. ­ ­ ▲ FIGURE 25.1 The cheese-making process. The commercial production of all cheeses begins with the pasteurization of milk; the wide variety of cheeses available on the market results from the nature of subsequent processing steps. Why do cheeses that are aged longer have more acidic, “sharper” tastes? Figure 25.1 Fermentation during aging results in continued production of acid; the longer the cheese is aged, the more acid produced and the “sharper” the taste. M25_BAUM7206_05_SE_C25.indd 779 Production of ripened cheeses through pressing, addition of secondary microbial cultures, and aging (ripening) Products of Alcoholic Fermentation Alcoholic fermentation is the process by which various microorganisms convert simple sugars such as glucose into ethanol (drinking alcohol) and carbon dioxide (CO2 ). These two products of fermentation take on different importance for different foods and beverages. As is the case for the production of fermented dairy products, manufacturers use specific starter cultures in the large-scale commercial applications of alcohol fermentation. In the next sections we consider the role of fermentation in the production of wine, spirits, beer, sake, and vinegar. 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C Wine and Spirits Generally, wine production proceeds as follows 1 (FIGURE 25.2): 2  Preparation of must. Winemakers crush the fruit and remove the stems to form must (fruit solids and juices). They make red wines from the entire must of dark grapes, whereas they make white wines using only the juice of either dark or light grapes. Weather conditions and soil composition affect the sugar content of fruit, which in turn greatly affects the quality of the wine. Preparation of must by stemming and crushing of grapes (or other fruit) Addition of starter culture of yeast and bacteria Fermentation. A thin film of bacteria and yeast naturally covers grapes and other fruit, and in some wineries these natural microbes produce the wine. However, in most commercial operations, vintners add sulfur dioxide (SO2 ) to retard growth of naturally occurring bacteria and then add starter cultures of yeast and bacteria to ensure that wine of a consistent quality is generated from year to year. Saccharomyces ferments sugar in fruit juice to alcohol to make wine. Leuconostoc removes acids naturally present in grapes. 3 Clarification. Filtration or settling removes solids—a process called clarification. 1 4 Aging. Wine is aged in wooden barrels. Continued yeast metabolism and the leaching of compounds from the wood add taste and aroma. 1 5 Bottling. The wine is then bottled and distributed.  2 Aging of wine 5 Bottling of wine ▲ FIGURE 25.2 The wine-making process. ed wine is produced by fermenting must (fruit solids and juice) from dark grape varieties, whereas white wine results from the fermentation of juice alone from either dark or light grape varieties.  Mashing. Malt and additional sources of carbohydrate called adjuncts (such as starch, sugar, rice, sorghum, or corn) are combined with warm water, allowing enzymes released during malting to generate more sugars. The sugary liquid is called wort.  1 Malting. Barley is moistened and germinated, a process that produces catabolic enzymes that convert starch into sugars, primarily maltose. Drying halts germination, and the barley is crushed to produce malt. 4 R Beer and Sake Beer, the most commonly consumed alcoholic beverage in the world, is made from barley in the following steps (FIGURE 25.3): 1 Clarification of wine  Dry wines are those in which all the sugar has fermented, whereas sweet or dessert wines retain some sugar. Most table wines have a final alcohol content of 10% to 12%, but fortified wines have an alcohol content of about 20% because distilled spirits (discussed next) are added. Sparkling wine is produced by the addition of sugar and a second fermentation that produces CO2 inside the bottle. Distilled spirits are made in a process similar to wine making in that Saccharomyces ferments fruits, grains, or vegetables. The difference is that during the aging process, the alcohol is concentrated by distillation. In this process, the liquid is heated to drive off the alcohol, which is then condensed and added back. The net result is an increase in the alcohol content (100proof spirits are 50% alcohol). Brandies are made from fruit juices, whiskeys are made from cereal grains, and vodka is often made from potatoes. 1 Fermentation of must (crushed fruit) or of juice alone into wine 3  1 2 M25_BAUM7206_05_SE_C25.indd 780 1 3 Preparation for fermentation. The brewer removes the spent grain from the wort and adds hops—the dried flower-bearing parts of the vinelike hops plant. The wort mixture is boiled to halt enzymatic activity, extract flavors from the hops, kill most microorganisms, and concentrate the mixture. The remains of the hops are removed after boiling.  1 1  1 11/23/16 3:44 AM       Food Microbiology Barley is moistened and germinated, producing enzymes that convert starch into sugars. Barley is then dried to halt germination, and crushed to produce malt. 2 Mashing malt and adjuncts with warm water allows enzymatic activity to generate more sugars. Solids are removed to Mashing kettle produce wort. Addition of hops for flavoring 3 Cooking of wort halts enzymatic activity, extracts flavor from hops, and kills the microorganisms present. Removal of hops Addition of yeast culture 5 Aging. The beer is aged, pasteurized or filtered, and bottled or canned. Beer has a considerably lower alcohol content (about 4%) than either wine or distilled spirits.  1 1 Sake (rice beer) is made from cooked rice in which the starch is first converted to sugar (by the fungus Aspergillus oryzae) and the sugar then is fermented by Saccharomyces. Sake has an alcohol content of about 14%. Vinegar Vinegar2 is produced when ethanol resulting from the fermentation of a fruit, grain, or vegetable is oxidized to acetic acid. Acetic acid bacteria such as Acetobacter (a-sē′tō-bak-ter) or Gluconobacter (gloo-kon′ō-bak-ter) perform this secondary metabolic conversion, generating about 4% acetic acid. Different flavors of vinegar are derived from different starting materials. Cider vinegar, for example, is made from apples. TABLE 25.1 on p. 782 summarizes types of fermented foods along with the organisms involved in their production. The Causes and Prevention of Food Spoilage LE A R N I N G O UTC O M E S 25.2 Describe how food characteristics and the presence of microorganisms in food can lead to food spoilage. 25.3 List several methods for preventing food spoilage. 4 Wort ferments into beer. Whereas many chemical reactions involving food are desirable because they enhance taste, aroma, or preservation, other reactions are not. Spoilage of food involves adverse changes in nutritive value, taste, or appearance. Spoilage leads not only to economic loss but also potentially to illness and even death. In this section, we examine the causes of food spoilage before considering ways to prevent it. Causes of Food Spoilage 5 Aging, filtering or pasteurization, and bottling finish the process. ▲ FIGURE 25.3 The beer-brewing process. The type of beer produced (ale, lager) is determined largely by the type of yeast used (top fermenting or bottom fermenting, respectively). 4 Fermentation. A starter culture of Saccharomyces is added to the wort. Most beers—called lagers—are made with the bottom-fermenting yeast S. carls­ bergensis (karlz-bur-jen′sis). Other beers—called ales—are produced with the top-fermenting yeast S. cerevisiae (so called because the yeast floats in the vat).  1 M25_BAUM7206_05_SE_C25.indd 781 Foods may spoil because of inherent properties of the food itself. Such intrinsic factors include nutrient content, water activity, pH, and physical structure of the food as well as the competitive activities of microbial populations. Alternatively, food spoilage results from extrinsic factors that have nothing to do with the food itself but instead with the way it is processed or handled. Intrinsic Factors in Food Spoilage The nutritional composition of food determines both the types of microbes present and whether they will grow. Some foods contain natural antimicrobial agents, such as the benzoic acid in cranberries, and thus are not prone to spoilage. In contrast, fortified foods—those enriched in vitamins and minerals to improve the health of humans— may inadvertently facilitate growth of microorganisms by providing more nutrients. 2From French vinaigre, meaning “sour wine.” 11/23/16 3:44 AM TABLE 25.1 p er 25    Applied and Industrial Microbiology t ha       C Some Fermented Foods and the Microorganisms Used in Their Production Food Starting Material Representative Culture Microorganisms Cabbage Various lactic acid bacteria Cucumbers, peppers, beets Various lactic acid bacteria Soybeans and wheat Aspergillus oryzae and Lactobacillus spp. Fermented Vegetables P Sauerkraut/kimchi ickle Soy sauce R Miso ice and soybeans or rice and other grains A. oryzae, Lactobacillus spp., and Torulopsis etchellsii ork, beef, chicken Various lactic acid bacteria Fermented Meat Products P Dry salami Fish sauce/paste Ground fish Various naturally occurring bacteria Fermented Dairy Products Milks P Buttermilk P Yogurt asteurized skim milk Lactococcus lactis subspecies cremoris and Leuconostoc citrovorum asteurized skim milk Streptococcus thermophilus and Lactobacillus bulgaricus asteurized milk L. lactis, including subspecies cremoris Cheeses P Cottage cheese Milk curd Starter culture as in cottage cheese without further additions Soft cheese (e.g., Camembert)a Milk curd Starter culture as in cottage cheese plus Penicillium camemberti Mold-ripened (e.g., oquefort) Milk curd Starter culture as in cottage cheese plus Penicillium roqueforti Corn, grains, vegetation Various naturally occurring bacteria Wine Grapes Saccharomyces cerevisiae Distilled spirits Fruits, vegetables, grains S. cerevisiae Beer Barley S. cerevisiae or S. carlsbergensis Sake Cooked rice A. oryzae and Saccharomyces spp. Vinegar Fruits, vegetables, grains S. cerevisiae and Acetobacter or Gluconobacter Bread Flour, salt, etc. S. cerevisiae R Hard cheese (e.g., cheddar) Animal Feed Silage Alcoholic Fermentations aIn addition to being a soft cheese, Camembert is also a mold-ripened cheese. Water activity refers to water that is not bound physically by solutes or to surfaces and is thus available to microbes. The water activity of pure water is set at 1.0, and most microbes require environments with a water activity of at least 0.90. Moist foods with water activities near 0.90, such as fresh meat, support microbial growth. Dry foods (e.g., uncooked dry pasta) with water activities near zero do not support microbial growth. Food processors reduce water activity by drying foods or by adding salts or sugars. Thus, even though jam is moist, its sugar content is very high, and its water activity is too low to support much microbial growth. Acidity of food can either be an intrinsic chemical property of the food, as in the case of citrus fruit, or result from fermentation, as with dill pickles. In either case, a pH below 5.0 typically reduces microbial growth except by a few molds or lactic acid bacteria. A pH closer to neutral (7.0) supports a wider range of microbial growth. Since the pH of most foods is close to neutral, pH is generally not a deterrent to spoilage microorganisms. Physical structure is a visible intrinsic factor. Rinds or thick skins usually protect fruits and vegetables, and eggs have shells. These coverings are dry and nutritionally poor, and thus M25_BAUM7206_05_SE_C25.indd 782 they support little microbial growth. If the outer covering is broken or cut, however, microbes can reach the moist interior of the food and cause it to rot. Ground meat has more surface area and more oxygen and may have bacteria mixed within it during the grinding process. Thus, ground meat supports microbial growth and spoils faster than whole cuts of meat. In uncut meat, the largest volume of the meat is anaerobic and not exposed to microorganisms. To some extent, microbial competition can also be an intrinsic factor in food spoilage, especially in fermented foods. Fermented foods are populated with large numbers of fermentative bacteria but few pathogens because the latter do not readily grow in the environment produced by the fermenters. Furthermore, pathogens that require the rich nutrient levels found in the human body are not typically capable of growing on “lower-nutrient” foods. TABLE 25.2 summarizes the effects of intrinsic factors on food spoilage. Extrinsic Factors in Food Spoilage Extrinsic factors, including how food is processed, handled, and stored, also govern food 11/23/16 3:44 AM       Food Microbiology TABLE 25.2 Factors Affecting Food Spoilage Foods at Greatest Risk of Spoilage Food at Least Risk of Spoilage Intrinsic Factors Nutritional Composition Chemically rich or fortified foods (steak, bread, milk) Chemically limited foods (flour, cereals, grains) Water Activity Moist foods (meat, milk) Dry foods or those with low water activity (dried pasta, jam) pH Foods with neutral pH (bread) Foods with low pH (orange juice, pickles) Physical Structure Foods without rinds, skins, or shells; ground meat Foods with rinds, skins, or shells; intact foods Microbial Competition Foods that lack natural microbe populations (ground beef ) Foods with resident microbial populations (pickles) Extrinsic Factors Unprocessed foods (raw milk, fruit) Amount of Preservatives Foods without preservatives (meats, natural foods) Foods with either naturally occurring or added preservatives (garlic, spices, sulfur dioxide) Storage Temperature Foods left in warm conditions Foods kept cold Storage Packaging Foods stored exposed Foods wrapped or sealed P Degree of Processing rocessed foods (pasteurized foods) spoilage. Microorganisms can enter food in a variety of ways; some are introduced accidentally during harvesting from soil or contaminated water, and commercial food processing introduces others. Mechanical vectors, such as flies, also deposit microbes onto food. However, most microbial contaminants leading to spoilage can be traced to improper handling by consumers. Methods to prevent the introduction of microbes into food or to limit their growth will be presented shortly. Classifying Foods in Terms of Potential for Spoilage Prevention of Food Spoilage Food spoilage can begin during production, processing, packaging, or handling. Spoilage results in significant economic losses not only to producers—in the form of lost productivity, food recalls, and even the loss of jobs resulting from the shutdown of unsafe food processing facilities—but also to consumers in the form of medical expenses and time lost from work. In the next sections, we examine some of the many techniques used to prevent food spoilage: food processing methods, the use of preservatives, and attention to temperature and other storage conditions. Food Processing Methods Industrial canning is a major food packaging methodology for preserving foods (FIGURE 25.4). After food is prepared by washing, sorting, and processing, it M25_BAUM7206_05_SE_C25.indd 783 ▲ FIGURE 25.4 Industrial canning. Food is processed, put into cans or jars, and heated under pressure for a length of time and a temperature sufficient to kill most microbes, including endospore-forming bacteria. Why isn’t the worker wearing a mask and gloves while handling open cans of food? Figure 25.4 The food is about to be steam sterilized, so any introduced microbes will be killed. Based on the previous considerations, foods can be grouped into three broad categories, depending on their likelihood of spoilage. Perishable foods, such as milk, tend to be nutrient rich, moist, and unprotected by coverings. They need to be kept cold, or they spoil relatively quickly (within days). Semi­ perishable foods, such as tomato sauce, can be stored in sealed containers for months without spoiling as long as they are not opened. Once opened, however, they may spoil within several weeks. Nonperishable foods are usually dry foods, such as pasta, or canned goods that can be stored almost indefinitely without spoiling. Nonperishable foods are typically either nutritionally poor, dried, fermented, or preserved (discussed shortly). is packaged into cans or jars and subjected to high heat (115°C, 239°F) under pressure for a given amount of time, depending on the food. This temperature is not that of a sterilizing autoclave in a microbiology laboratory, so the food must be heated longer. (Autoclave temperature [121°C, 250°F] would severely change the food’s taste, texture, and nutritional content.) The heating is followed by rapid cooling. The heat kills vegetative mesophilic bacteria and destroys endospores formed by Bacillus (ba-sil′lŭs) and Clostridium (klos-trid′ē-ŭm). Although canning results in the elimination of most contaminating microorganisms, it does not sterilize food; hyperthermophilic microbes remain, but because they cannot grow at room temperature, they do not pose a threat. Spoilage ca n occ ur if t he food is u nder processed or if mesophilic microbes contaminate the can during 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C cooling or thereafter. The two most frequent contaminants found in canned goods are Clostridium spp. and coliforms, organisms which grow well anaerobically in low-acid foods. Contamination with the resistant endospores of Clostridium is especially problematic because if the cans or jars are not heated sufficiently, the endospores germinate, grow, and release toxins inside the sealed container, which can result in botulism. Pasteurization is less rigorous than canning and is used primarily with beer, wine, and dairy products because it does not degrade the flavor of these more delicate foods. By heating foods only enough to kill mesophilic, non-endospore-forming microbes (including most pathogens), pasteurization lowers the overall number of microbes, but because some microbes survive, pasteurized foods will spoil without refrigeration. Moisture is essential for microbial growth; therefore, desiccation (drying) is an excellent food preservation technique. Drying greatly reduces or eliminates microbial growth, but it does not kill bacterial endospores. Although some fruits are still dried by being left in the sun, today most commercially dried foods are processed in ovens or heated drums that evaporate the water from the foods. Lyophilization (lī-of′i-li-zā′shŭn), or freeze-drying, involves freezing foods and then using a vacuum to draw off the ice crystals. Freeze-dried foods such as soups and sauces can be reconstituted by mixing with water. Gamma radiation, produced primarily by the isotope cobalt-60, penetrates fruits, vegetables, and meats, including fish, poultry, and beef, to cause irreparable and fatal damage to the DNA of microbes. Such ionizing irradiation is controversial because some consumers believe erroneously that irradiated foods become radioactive, are less nutritious, or contain toxins. Nevertheless, because irradiation can achieve complete sterilization, it is used routinely to preserve some foods, such as spices. Ultraviolet (UV) light is not ionizing and does not penetrate very far. It is used to treat packing and cooling water used in industrial canning and to treat work surfaces and utensils in industrial meat processing plants. In aseptic packaging, paper or plastic containers that cannot withstand the rigors of canning or pasteurization are sterilized with hot peroxide solutions, UV light, or superheated steam. Then a conventionally processed food is added to the aseptic package, which is sealed without the need for additional processing. The Use of Preservatives Humans have preserved foods with salt or sugar throughout history. Both chemicals draw water by osmosis out of foods and microbes alike, killing microbes on the food and retarding the growth of any subsequent microbial contaminants. Bacon is an example of a high-salt food; jellies are examples of high-sugar foods. Whereas salt and sugar act by removing water, some natural preservatives actively inhibit microbial enzymes or disrupt cytoplasmic membranes. For example, garlic contains allicin, which inhibits enzyme function. Benzoic acid, produced naturally by cranberries, also interferes with enzymatic function. Cloves, cinnamon, oregano, and thyme (and, to some degree, sage and rosemary) produce oils that interfere with the functions of membranes of microorganisms. M25_BAUM7206_05_SE_C25.indd 784 As we have seen, fermentation preserves some foods by producing an acidic environment that is inhospitable to most microbes. For other foods such as meats, the use of wood smoke during the drying process introduces growth inhibitors that help preserve the food. Other naturally occurring and synthetic chemicals can be purposely added to foods as preservatives. Acceptable preservatives are harmless and do not alter the taste or appearance of the food to any great extent. Organic acids, such as benzoic acid, sorbic acid, and propionic acid, are commonly used in beverages, dressings, baked goods, and a variety of other foods. Gases, such as sulfur dioxide and ethylene oxide, are used to preserve dried fruit, spices, and nuts. All such chemical preservatives inhibit some aspect of microbial metabolism, but many do not actually kill microbes; in other words, they are germistatic rather than germicidal. Some chemicals work better against bacteria than against fungi and vice versa. Benzoic acid, for example, has a largely antifungal function and does not affect the growth of many bacteria. Attention to Temperature During Processing and Storage In general, higher temperatures are desirable during food processing and preparation to prevent food spoilage, whereas lower temperatures are desirable for food storage. High temperatures, such as those of pasteurization, canning, or cooking, kill potential pathogens because proteins and enzymes become irreversibly denatured. However, heat does not inactivate many toxins; for example, botulism toxin may remain in cooked foods even when the bacteria that produced it are dead. Unlike heat, cold rarely kills microbes but instead merely retards their growth by slowing metabolism. Even freezing fails to kill all microorganisms and may only lower the level of microbial contamination enough to reduce the likelihood of food poisoning after frozen foods have been thawed. To prevent food spoilage, foods should be prepared at sufficiently high temperatures and then stored under conditions that do not facilitate microbial growth. Foods should be properly stored in appropriate containers, cold foods should be kept cold, foods should be cooked thoroughly, and leftovers should be refrigerated or frozen to reduce spoilage. Wrapping leftovers or putting them in containers enables foods to be stored away from exposure to air or contact that could result in contamination. One microbe for which cold storage does not suppress growth is Listeria monocytogenes (lis-tēr′ē-ă mo-nō-sī-tah′je-nēz), the causative agent of listeriosis and a common environmental bacterium. This bacterium is prevalent in certain dairy products, such as soft cheeses, and grows quite well under refrigeration; therefore, it is best to prevent its entry into foods. Foodborne Illnesses LE A R N I N G O UTC O M E 25.4 Discuss the basic types of illnesses caused by food spoil- age or food contamination, and describe how they can be avoided. Spoiled food, if consumed, can result in illness from toxins produced by the microbes or from the microbes themselves, but 11/23/16 3:44 AM       Industrial Microbiology not all foodborne illnesses result from actual food spoilage. Foodborne illnesses may also result from the consumption of harmful microbes or their products in food. Foodborne illnesses (food poisoning) can be divided into two types: food infections, caused by the consumption of living microorganisms, and food intoxications, caused by the consumption of microbial toxins instead of the microbes themselves. Typical signs and symptoms of food poisoning are generally the same regardless of the cause and include nausea, vomiting, diarrhea, fever, fatigue, and muscle cramps. Symptoms occur within 2–48 hours after ingestion, and the effects of the illness can linger for days. Most outbreaks of food poisoning are common­source epidemics, meaning that a single food source is responsible for many individual cases of illness. The Centers for Disease Control and Prevention (CDC) estimates that about 48 million cases of food poisoning occur in the United States each year. Of these, about 128,000 people require hospitalization and 3000 die. Researchers identify the microbes involved in only about 14 million of the total cases. The U.S. Department of Agriculture estimates the economic cost of food poisoning—due to loss of productivity, medical expenses, and death—at roughly $5 billion to $10 billion per year. More than 250 different foodborne diseases have been described. TABLE 25.3 lists 10 common causes of foodborne illness and their sources. Except for the protozoan Toxoplasma gondii (tok-sō-plaz′mă gon′dē-ē), all are bacterial agents. A few fungi (Aspergillus and Penicillium [pen-i-sil′ē-ŭm]) and viruses (e.g., hepatitis A, noroviruses) also cause food poisoning. M Why e ell T Even though microbes are naturally present in raw milk, raw milk is not generally used for the production of cheeses. Why not? Industrial Microbiology The potential uses of microorganisms for producing valuable compounds, as environmental sensors, and in the genetic modification of plants and animals makes industrial microbiology one of the more important fields of study within the microbiological sciences. In the sections that follow, we will examine the use of microbes in industrial fermentation, in the production of several industrial products, and in the treatment of water and wastewater. The Roles of Microbes in Industrial Fermentations LE A R N I N G O UTC O M E 25.5 Describe the role of genetically manipulated microor- ganisms in industrial and agricultural processes and the basics of industrial-scale fermentation. In industry, the word fermentation is used differently from its use in food microbiology or in the study of metabolism. Indus­ trial fermentations involve the large-scale growth of particular microbes for producing beneficial compounds, such as amino acids and vitamins. Temperature, aeration, and pH are all regulated to maintain optimal microbial growth conditions. Generally, industrial fermentations start with the cheapest growth medium available, often the waste product from another process (such as whey from cheese production). Because of its scale, industrial fermentation is performed in huge vats that can be readily filled, emptied, and sterilized (FIGURE 25.5). Vats are typically made from stainless steel so that they can be cleaned and sterilized more easily. There are two types of industrial fermentation. In batch pro­ duction, organisms ferment their substrate until it is exhausted, TABLE 25.3 Most Common Bacterial and Protozoan Agents of Foodborne Illnesses Campylobacter jejuni Affected Food Products R Organism aw and undercooked meats; raw milk; untreated water Comments Most common cause of diarrhea of all foodborne agents Escherichia coli O157:H7 Meat; raw milk Listeria monocytogenes Dairy products; raw and undercooked meats; seafood; produce Common in soils and water, contamination from these sources occurs easily; grows at refrigerator temperature Salmonella spp. aw and undercooked eggs; meat; dairy products; fruits and vegetables Second most common cause of foodborne illness in the United States Shigella spp. Salads; milk and other dairy products; water Third most common cause of foodborne illness in the United States Staphylococcus aureus Cooked high-protein foods Toxoplasma gondii Meat (pork in particular) Yersinia enterocolitica M25_BAUM7206_05_SE_C25.indd 785 R P Vibrio vulnificus P P P P Home-prepared foods R Clostridium botulinum roduces a neurotoxin roduces an enterotoxin roduces a potent toxin that is not destroyed by cooking arasitic protozoan aw and undercooked seafood Causes primary septicemia (bacteria in the blood) ork; dairy products; produce Causes generalized diarrhea and severe cramping that mimics appendicitis; grows at refrigerator temperature 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C additives and supplements, dyes, plastics, and fuels. Recombinant organisms add to this diversity by producing pharmaceuticals, such as human insulin, which are not normally manufactured by microbial fermentations. In the sections that follow we consider a variety of industrial products produced by microbes, including enzymes, alternative fuels, pharmaceuticals, pesticides, agricultural products, biosensors, and bioreporters. Enzymes and Other Industrial Products ▲ FIGURE 25.5 Fermentation vats. Such large containers are used for growing microorganisms in the vast quantities needed for the largescale production of many industrial, agricultural, and medical products. Why are industrial fermentation vats made of stainless steel instead of wood? Figure 25.5 Stainless steel can be cleaned more easily to avoid contamination of the product being produced. and then the end product is harvested all at once. In continuous flow production, the vat, also called a bioreactor, is continuously fed new medium while wastes and product are continuously removed. For this setup to work, the organism must secrete its product into the surrounding medium. Industrial products are produced as either primary or secondary metabolites of the microorganisms. Primary metabo­ lites, such as ethanol, are produced during active growth and metabolism because they are either required for reproduction or by-products of active metabolism. Secondary metabolites, such as penicillin, are produced after the culture has moved from the log phase of growth and entered the stationary phase (see Chapter 6, Figure 6.20), during which time the substances produced are not immediately needed for growth. Recombinant DNA techniques are used for the production of recombinant microorganisms. (Chapter 8 examines these techniques and some of their uses.) Most genetically modified organisms used in industry have been specifically designed to produce a stable, high-yield output of desirable chemicals or to perform certain novel functions. Industrial Products of Microorganisms LE A R N I N G O U TC O M E 25.6 List some of the various commercial products produced by microorganisms. Microorganisms, particularly bacteria, metabolically produce many industrially useful chemicals, including enzymes, food M25_BAUM7206_05_SE_C25.indd 786 Enzymes are among the more important products made by microbes. Most are naturally occurring enzymes for which industrial uses have been devised. For example, amylase, produced by Aspergillus oryzae, is used as a spot remover. Pectinase, obtained from species of Clostridium, enzymatically releases cellulose fibers from flax, which are then made into linen. Proteases from a variety of microbes are used in meat tenderizers, spot removers, and cheese production. Streptokinases and hyaluronidase are used in medicine to dissolve blood clots and enhance the absorption of injected fluids, respectively. Microbes also make the enzymatic tools of recombinant DNA technology, including restriction enzymes, ligases, and polymerases. Some products made naturally by microorganisms are useful to humans as food additives and food supplements. Food additives generally enhance a food in some way, such as by improving color or taste, whereas food supplements make up for nutritional deficiencies. Amino acids and vitamins are two important microbial products used as supplements. Vitamins are added directly to foods or sold as vitamin tablets. Amino acids are either sold in tablet or powder form or combined to make new compounds, such as the sweetener aspartame (made from the amino acids phenylalanine and aspartic acid). Other organic acids, such as citric acid, gluconic acid, and acetic acid (vinegar), are also microbially produced to be used in food manufacturing. Citric acid is used as an antioxidant in foods, and gluconic acid is used medically to facilitate calcium uptake. Other industrial products made by microbes include dyes and cellulose fibers used in woven fabrics. Microbially produced biodegradable plastics can replace nonbiodegradable, petroleum-based plastics. This is possible because many bacteria produce a carbon-based storage molecule called polyhydroxyalkanoate (PHA), a polymer with a structure similar to petroleum-based plastics (see Beneficial Microbes: Plastics Made Perfect? on p. 74). Alternative Fuels Photosynthetic microorganisms use the energy in sunlight to convert CO2 into compounds that can be used as fuels—so-called biofuels. Other microorganisms convert biomass (organic materials such as plants or animal wastes) into renewable biofuels, including ethanol, methane, and hydrogen. Ethanol, which is made during alcoholic fermentation and is the simplest alternative biofuel to synthesize, can be mixed 11/23/16 3:44 AM       Industrial Microbiology TABLE 25.4 Pharmaceuticals Foremost among the pharmaceutical substances that microorganisms produce are antimicrobial drugs. About 6000 antimicrobial substances have been described since penicillin was first produced during World War II. Of these, 100 or so have current medical applications. The bacteria Streptomyces (streptō-mī′sēz) and Bacillus and the fungus Penicillium synthesize the majority of useful antimicrobials, though recombinant DNA techniques enable the modification of other microbes to produce new versions of old drugs. In addition to antimicrobials, genetically modified microbes are producing hormones and other cell regulators. TABLE 25.4 lists some products of recombinant DNA technology used in medicine. Pesticides and Agricultural Products Farmers use microbes and their products in a variety of agricultural applications, particularly with regard to crop management. Bacillus thuringiensis (thur-in-jē-en′sis) is one of the more widely used organisms because during sporulation it produces Bt toxin, which, when digested by the caterpillars of such pests as gypsy moths, diamondback moths, and tomato hornworms, destroys the lining of the insect’s gut wall, eventually killing it. Bt toxin isolated from the bacterium can be spread as a dust on plants, but with recombinant DNA technology the Bt gene can be added to a plant’s genome. In this way plants such as corn, soybeans, and cotton protect themselves by manufacturing their own Bt toxin. There is little evidence of insect resistance to this Bt toxin despite the fact that large tracts of farmland are devoted to raising Bttoxin-expressing plants. Some people are leery of the prospect of transgenic foods derived from these plants, whereas others contend that the genetic manipulation of crops is crucial if we are to continue to feed the world’s population. In any case, one thing is clear: The genetic manipulation of plants is big business. ­ with gasoline to make gasohol, which is used in existing cars. Currently, in the United States, food crops such as corn are fermented to ethanol, but production of alternative fuel from nonfood crops or crop wastes—for example, inedible switchgrass or corn stalks—may use less water, fertilizer, and food supplies. Much research currently is focused on developing microbes to generate a more complex alcohol—butanol, which contains more energy per mass than ethanol, does not absorb water from the atmosphere like ethanol, and can be burned in most engines designed for gasoline without modification. All microorganisms synthesize hydrocarbons as part of their normal metabolism, but only some microbes produce hydrocarbons that could be useful fuels. The colonial alga Botryococcus braunii (bot′rē-ō-kok′ŭs brow′nē-ē), for example, produces hydrocarbons that account for 30% of its dry weight. One day, this alga may be an important source of fuel, but the technology to harvest such hydrocarbons does not yet exist. One exception is the harvesting of methane gas, which can be collected and piped through natural gas lines (natural gas is mostly methane) to be used for cooking and heating. The largest potential source of methane is from landfills, where methanogens anaerobically convert wastes into methane via anaerobic respiration. Some communities already use methane from landfills to produce energy. Methane can also be used as a fuel in properly equipped cars. Although a variety of microbes release hydrogen as part of their normal metabolism, the hydrogen-producing metabolic pathway does not cost effectively produce fuel. An economically efficient method for the production of hydrogen fuel using sunlight and photosynthetic microbes would have widespread applications and enormous appeal, but much research is needed to develop technologies to produce hydrogen as a viable energy source. Biofuels may play an important role in the future of the energy industry, possibly serving as a renewable replacement for fossil fuels. Some Products of Recombinant DNA Technology Used in Medicine Modified Cell Uses of Product Interferons Escherichia coli, Saccharomyces cerevisiae To treat cancer, multiple sclerosis, chronic granulomatous disease, hepatitis, and warts Interleukins E. coli To enhance immunity Tumor necrosis factor E. coli In cancer therapy Mammalian cell culture To stimulate red blood cell formation; to treat anemia Tissue plasminogen activator Mammalian cell culture To dissolve blood clots Human insulin E. coli For diabetes therapy Taxol E. coli In ovarian cancer therapy Factor VIII Mammalian cell culture In hemophilia therapy Macrophage colony stimulating factor E. coli, S. cerevisiae To stimulate bone marrow to produce more white blood cells; to counteract side effects of cancer treatment E Product rythropoietin To ease childbirth E. coli To correct childhood deficiency of growth hormone Hepatitis B vaccine Carried on a plasmid of S. cerevisiae To stimulate immunity against hepatitis B virus R E. coli Human growth hormone elaxin M25_BAUM7206_05_SE_C25.indd 787 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C Pseudomonas syringae (soo-dō-mō′nas sēr′in-jī) is another example of a bacterium with agricultural applications. This bacterium produces a protein that serves in the formation of ice crystals, but the protein is not essential to the survival of the organism, and scientists can remove its gene. When sprayed on crops such as strawberries, the strain lacking the gene, known as P. syringae ice - (“ice-minus”), inhibits the formation of ice, protecting the plants from freeze damage. TABLE 25.5 lists selected industrial products produced by microorganisms and some of their uses. Biosensors and Bioreporters Among the relatively new applications of microorganisms to solve environmental problems are biosensors and bioreporters. Biosensors are devices that combine bacteria or microbial products (such as enzymes) with electronic measuring devices to detect other bacteria, bacterial products, or chemical compounds in the environment. For example, one such biosensor measures blood glucose levels in diabetic patients by using the enzyme glucose oxidase to convert glucose into an electrical signal. Bioreporters are somewhat simpler sensors that are composed of microbes (again usually bacteria) with innate signaling capabilities, such as the ability to glow in the presence of biological or chemical compounds. Currently, biosensors and bioreporters are used to detect the presence of environmental pollutants (e.g., petroleum) and to monitor efforts to remove harmful substances; they may also be useful in detecting bioterrorist attacks. Because bacteria Water Treatment LE A R N I N G O UTC O M E S 25.7 Contrast water contamination and water pollution. 25.8 Describe two waterborne illnesses. 25.9 Explain how water for drinking and wastewater are treated to make them safe and usable. Water contamination refers to the presence of anything in the water. Contaminants can be natural or artificial; they can be physical, as in the case of silt; chemical, as in the case of dissolved minerals; or biological, as in the case of amoebas. Water pollution, in contrast, generally refers to something added to water by human activity beyond the capacity of natural processes to get rid of it. So, all water pollution is contaminating, but not all contaminants are pollutants. Water treatment facilities process water to remove harmful contaminants and pollution. Water Pollution Many pollutants are not readily visible. Although physical and chemical pollutants are important, of greater concern is biological pollution of water with human pathogens such as might enter water in sewage. Biological pollutants can cause significant human diseases, which can be prevented only through water treatment. ­ TABLE 25.5 Selected Industrial Products Produced by Microorganisms are very sensitive to their environments, they can detect compounds in very small amounts. Biosensors and bioreporters could serve as early warning systems to give officials more time to respond by quickly detecting the metabolic waste products of weaponized biological agents. Products Made Use of Product Enzymes Spot removers Streptokinase Breakdown of blood clots estriction enzymes, ligases, polymerases Molecular biology, recombinant DNA technology R Amylase, proteases Food Additives/Supplements Amino acids, vitamins Health supplements Citric acid Antioxidant Sorbic acid, lysozyme Food preservatives Other Industrial Products P Indigo lastics Dye used in manufacturing clothes Biodegradable substitutes for petroleum-based plastics thanol Methane Hydrogen, hydrocarbons Used in gasohol Burned to generate heat and electricity P E Alternative Fuels otential fuels Pesticides and Agricultural Products Bt toxin M25_BAUM7206_05_SE_C25.indd 788 Insecticide Waterborne Illnesses Consuming contaminated water, either as drinking water or in water added to foods, can result in a variety of bacterial, viral, or protozoan diseases. No known fungal water contaminants cause diseases. Each year, contaminated drinking water results in roughly 3 billion to 5 billion episodes of diarrheal disease worldwide, including over 3 million deaths among children ages five years or younger. Intoxication can also occur from the presence of microbial toxins in the water. Water treatment removes most waterborne pathogens, so waterborne illnesses are generally rare in the United States as compared to countries with inadequate water treatment facilities. Outbreaks that do occur in the United States are point­ source infections, in which a single source of contaminated water leads to illness in individuals that consume the water. In some marine environments, massive overgrowth of dinoflagellates, such as Gonyaulax (gon-ē-aw′laks), chemically pollutes water with their toxins. These overgrowths are sometimes called red tides because of the color that might be imparted to the water by the huge number of dinoflagellates. Their toxins can be absorbed and concentrated by shellfish, and human intoxication results from consumption of these mollusks rather 11/23/16 3:44 AM       Industrial Microbiology Disease Bacteria Campylobacter jejuni Acute gastroenteritis Escherichia coli Acute gastroenteritis Salmonella spp. Salmonellosis, typhoid fever Shigella spp. Shigellosis (bacterial dysentery) Vibrio cholerae Cholera Viruses Hepatitis A virus Infectious hepatitis Norovirus Acute gastroenteritis P oliovirus oliomyelitis Eukaryotic Parasites Cryptosporidium parvum Cryptosporidiosis Entamoeba histolytica Amebic dysentery Giardia intestinalis Giardiasis Schistosoma spp. Schistosomiasis Toxin Producers Gonyaulax (saxitoxin) aralytic shellfish poisoning E E GING DIS AS CAS STUDY E M ER E Potable (pō′tăbl) water is water that is considered safe to drink, but the term potable does not imply that the water is devoid of all microorganisms and chemicals. Rather, it implies that the levels of microorganisms or chemicals in the water are low enough that they are not a health concern. Water that is not potable is polluted; that is, it contains organisms or chemicals in excess of acceptable values. The permissible levels of microbes and chemicals in potable water vary from state to state. Nationally, the U.S. Environmental Protection Agency (EPA) requires that drinking Organism P Treatment of Drinking Water TABLE 25.6 Selected Waterborne Agents and the Diseases They Cause P than from water consumption, so such intoxication is not a true waterborne illness. TABLE 25.6 lists some common human waterborne pathogens and toxins. Emerging Disease Case Study: Attack in the Lake considers a pathogen that has recently acquired more attention. Clean water is vital for people and their activities. In the following sections, we consider the treatment of drinking water and the treatment of wastewater (sewage). In both cases, treatment is designed to remove microorganisms, chemicals, and other pollutants to prevent human illness. Attack in the Lake M25_BAUM7206_05_SE_C25.indd 789 P P Naegleria is an amoeba that lives in warm freshwater, where it feeds on bacteria, grows, and divides. It normally doesn’t bother people. But when people jump or dive into water where Naegleria makes its home, the amoeba can enter the nose, penetrate the nasal mucous membrane, and crawl up the olfactory nerves to the brain, where it causes primary amebic meningoencephalitis ( AM). AM results in part from the body’s efforts to ward off the infection with inflammation; the brain swells, producing headaches and other neurological symptoms. No antimicrobial drugs are known to kill Naegleria in the brain. Finally, the patient lapses into coma and usually dies within one to two weeks of infection; only three survivors have been verified in hundreds of cases. Scientists have not been able to determine why some people are infected while millions of others swim and play in warm lakes without consequence. pidemiologists have been unable to develop water safety standards to protect against Naegleria: they haven’t developed a method to accurately measure the number of amoebas in the water, determine the concentration of Naegleria required to initiate an infection, or design preventive measures other than avoiding recreation in warm lakes and wearing nose clips while swimming. Fortunately, AM is rare; only about 133 cases have occurred in the United States since 1962, though more cases are being reported now than in the past. For Callie and her family, this emerging disease is not rare enough. P E E ­ ­ ­ Callie was more than worried— she was terrified. Her 10-yearold son wasn’t acting normally. He was incoherent, and his eyes were rolling. It was as if he looked right through her, eyes unfocused and staring. How life can change. Just two weeks ago the family had been enjoying their summer vacation at the lake. veryone had been so happy—splashing, swimming, and diving, even though the lake had been warmer than normal. “Almost like a bathtub,” her husband had said. Still, it had been a grand week, the opposite of this, the second week since they returned. This week seemed to have stretched to an eternity. First, Caleb had complained of a headache, then he developed a fever, and on the third day the nausea and vomiting had started—he threw up 20 times by Callie’s estimation. However, she could handle these things; after all, the flu or a “stomach bug” was not that uncommon to a mother of five older children. But now, Caleb was unable to move his head. He’d had a seizure and had been hallucinating. What was happening? Would it never end? The doctors at the emergency room suspected bacterial meningitis and started an aggressive course of antibiotics; however, Caleb was not going to get better. Callie’s worry was about to turn to anguish. Her son’s brain was being consumed by Naegleria fowleri. 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C Sedimentation and Flocculation 1 Removal of large and particulate materials (b) A water treatment facility Particulates Flocculation 2 Alum Removal of large and particulate materials Flocs settle Filtration 3 Microbial removal by one of three methods: Sand • Sand filtration • Activated charcoal • Membrane filtration Disinfection 4 Inactivation of remaining microbes by one of three methods: • Chlorine • Ozone • UV light Faucet Disinfection Release of treated water (a) The stages of water treatment ▲ FIGURE 25.6 The treatment of drinking water. (a) The stages of water treatment: sedimentation, flocculation, filtration, and disinfection. Why is it that chemical treatment cannot destroy most viruses? (b) A water treatment facility. Figure 25.6 Most chemicals are designed either to inhibit some aspect of active metabolism or to damage cellular structures; therefore, they do not damage acellular and nonmetabolizing viruses. 1 Sedimentation. During sedimentation, water is pumped into holding tanks, where particulate materials (sand, silt, and organic material) settle.  1 M25_BAUM7206_05_SE_C25.indd 790 2 Flocculation. The partially clarified water is then pumped into a secondary tank for flocculation, in which alum (aluminum potassium sulfate) added to the water joins with suspended particles and microorganisms to form large aggregates, called flocs (see Figure 11.23). The flocs also settle to the bottom of the tank. The water above the sediment is then pumped into a different tank for filtration. 1 3 Filtration. In this stage, the number of microbes is reduced by about 90% in one of several ways. One method uses sand and other materials to which  1  water have a count of zero coliform bacteria per 100 ml of water and that recreational waters have no more than 200 coliforms per 100 ml of water. Recall that coliforms are intestinal bacteria such as Escherichia coli (esh-ĕ-rik′ē-ă kō′lī). The presence of coliforms in water indicates fecal contamination and thus an increased likelihood that disease-causing microbes are present. The treatment of drinking water can be divided into four stages (FIGURE 25.6): 11/23/16 3:44 AM       Industrial Microbiology  ­ microbes adhere and form biofilms that trap and remove other microbes. Slow sand filters are composed of a 1-meter layer of fine sand or diatomaceous earth and are used in smaller cities or towns to process 3 million gallons per acre of filter per day. Large cities use rapid sand filters that contain larger particles and gravel and can process 200 million gallons per acre per day. Both types of filters are cleaned by back-flushing with water. Two other filtration methods are membrane filtration, which uses a filter with a pore size of 0.2 mm, and filtration with activated charcoal, which provides the added benefit of removing some organic chemicals from the water. Disinfection. In this stage, ozone, UV light, or chlorination is used to kill most microorganisms before the water is released for public consumption. Many European communities use ultraviolet light. Chlorine treatment is widely used in the United States in part because it is least expensive.  4 1 Chlorine gas, an oxidizing agent, is thought to kill bacteria, algae, fungi, and protozoa by denaturing their proteins within approximately 30 minutes of treatment. Chlorine levels must be constantly adjusted to reflect estimates of microbial load, the number of microbes in a unit of water—a higher load requires more chlorination. Chlorination does not kill all microbes: Most viruses are not inactivated by chlorine, and bacterial endospores and protozoan cysts are generally unaffected by any chemical treatment. Only filtration can completely remove all viruses, endospores, and cysts. ­ Water Quality Testing E. coli and other coliforms in water indicates a probability that pathogens are present as well. E. coli is consistently prevalent in human waste as long as (if not longer than) most pathogens and is easily detected. Several testing methods can be used to assess water quality. One testing method is the membrane filtration method (FIGURE 25.7a), which is simple to perform: A 100-ml water sample is poured through a fine membrane, which is then placed on selective media agar plates and incubated. Coliform colonies exhibit unique characteristics. The colonies are then counted and reported as number of colonies per 100 ml. This method gives a total coliform count. In another test, water samples are added to small bottles containing both ONPG (o-nitrophenyl-b-d-galactopyranoside) and MUG (4-methylumbelliferyl-b-d-glucuronide) as sole nutrients. Most coliforms produce b-galactosidase, an enzyme that reacts with ONPG to produce a yellow color, but the fecal coliform E. coli produces an additional enzyme, b-glucuronidase, which reacts with MUG to form a compound that fluoresces blue when exposed to long-wave UV light (FIGURE 25.7b). This test allows for the rapid detection of coliforms but gives an estimate for the number of bacteria rather than an actual count. Scientists now often use PCR-based tests instead because they are more specific and more rapid. The presence of viruses and particular bacterial strains cannot be determined with these tests; their presence must be confirmed by genetic “fingerprinting” techniques, in which water samples are collected and enriched to cultivate the organisms present. The DNA content of the enriched sample is then genetically screened for the identification of potential pathogens. Governments are currently reconsidering the use of coliform tests to indicate fecal contamination because some coliforms grow naturally on plants even when there is little fecal contamination, giving a false-positive result for fecal pollution. Regulators are considering replacing coliform tests with genetic assays that would specifically indicate the presence of E. coli. Water quality testing is a technique that uses the presence of certain indicator organisms to indicate the possible presence of pathogens in water. Because the majority of waterborne illnesses are caused by fecal contamination, the presence of (a) (b) M25_BAUM7206_05_SE_C25.indd 791 P P ▲ FIGURE 25.7 Two water quality tests. (a) Membrane filtration. The grid on the membrane makes it easier to count the colonies of fecal coliforms, which are green on this selective medium. (b) The ON G and MUG tests. The yellow color of the ON G bottle indicates the presence of coliforms, whereas the blue fluorescence in the MUG bottle indicates the presence of the coliform E. coli; the clear bottle is the negative control. 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t trickle filter system, which is similar to the slow sand filters used in treating drinking water but less effective in removing BOD than activated sludge systems. The biological activity in this phase reduces the BOD to 5% to 25% of the original. Most pathogenic microorganisms are also removed. Treatment of Wastewater Municipal Wastewater Treatment Today, people living in larger U.S. towns and cities are usually connected to municipal sewer systems—pipes that collect wastewater and deliver it to sewage treatment plants for processing. Traditional sewage treatment consists of four phases (FIGURE 25.8): 1 2 Primary treatment. Wastewater is pumped into settling tanks, where lightweight solids, grease, and floating particles are skimmed off and heavier materials settle onto the bottom as sludge. After alum is added as a flocculating agent, the sludge is removed, and the partially clarified water is further treated. Primary treatment removes 25% to 35% of the BOD in the water.  1 Secondary treatment. The water is aerated to facilitate the growth of aerobic microbes that oxidize dissolved organic chemicals to CO2 and H2 O. In an activated sludge system, aerated water is seeded with primary sludge containing a high concentration of metabolizing bacteria; flocculation also occurs during this step. Any remaining solid material settles and is added to the sludge from primary treatment. The combined sludge is pumped into anaerobic holding tanks. Some smaller communities accomplish secondary treatment using a  1 M25_BAUM7206_05_SE_C25.indd 792 1 3 Chemical treatment. Water from secondary treatment is disinfected, usually by chlorination, after which the wastewater is either released into rivers or the ocean or, in some states, used to irrigate crops and highway vegetation. Some communities remove nitrates, phosphates, and any remaining BOD or microorganisms from the water by passing it over fine sand filters and/or activated charcoal filters. Nitrate is converted to ammonia and discharged into the air (removes roughly 50% of the nitrogen content), whereas phosphorus is precipitated using lime or alum (removes 70% of the phosphorus content). Such tertiary treatment is generally used in environmentally sensitive areas or in areas where the only outlet for the water is a closed-lake system. 1 4 Sludge treatment. Sludge is digested anaerobically in three steps: First, anaerobic microbes ferment organic materials to produce CO2 and organic acids. Second, other microbes metabolize these organic acids to H2 , more CO2 , and simpler organic acids, such as acetic acid. Finally, the simpler organic acids, H2 , and CO2 are converted to methane gas. Any leftover sludge is then dried for use as landfill or fertilizer.  Sewage, or wastewater, is typically defined as any water that leaves homes or businesses after being used for washing or flushed from toilets. (Some municipalities also include industrial water and rainwater as wastewater.) Wastewater contains a variety of contaminants, including suspended solids, biodegradable and nonbiodegradable organic and inorganic compounds, toxic metals, and pathogens. The objective of wastewater treatment is to remove or reduce these contaminants to an acceptable level for releasing the water into a natural environment, using it for irrigation, or sending it to a water treatment facility to be repurposed as drinking water. At one time, “raw” (unprocessed) sewage was simply dumped into the nearest river or ocean; the idea was that wastes would be diluted to a point at which they would be harmless. Burgeoning populations and the realization that waterways were becoming increasingly polluted led to greater use of effective wastewater treatment processes. Because sewage is mostly water (less than 1% solids), most sewage treatment involves the removal of microorganisms. A key concept in the processing of wastewater is reducing biochemical oxygen demand (BOD), which is a measure of the amount of oxygen required by aerobic bacteria to fully metabolize organic wastes in water. This amount is proportional to the amount of waste in the water; the higher the concentration of degradable chemicals, the more oxygen is required to catabolize them, and the higher the BOD. Effective wastewater treatment reduces the BOD to levels too low to support microbial growth, thus reducing the likelihood that pathogens will survive. The following sections consider wastewater treatments of various types: the traditional sewage treatment used in municipal systems, treatments used in rural areas, a treatment used for agricultural wastes, and the use of artificial wetlands.  ha       C Nonmunicipal Wastewater Treatment Houses in rural areas, which typically are not connected to sewer lines, often use septic tanks, essentially the home equivalent of primary treatment (FIGURE 25.9). Sewage from the house enters a sealed concrete holding tank. Solids settle to the bottom, and the liquid flows from the top of the tank into an underground leach field that acts as a filter. Sludge in the tank and organic chemicals in the water are digested by microorganisms. However, because the tank is sealed, it must occasionally be pumped out to remove sludge buildup. Cesspools are similar to septic tanks except that they are not sealed. As wastes enter a system of porous concrete rings buried underground, the water is released into the surrounding soil; solid wastes accumulate at the bottom and are digested by anaerobic microbes. Treatment of Agricultural Wastes Farmers and ranchers often use oxidation lagoons to treat animal waste from livestock raised in feedlots—a penned area where animals are fattened for market. Oxidation lagoons accomplish the equivalent of primary and secondary sewage treatment. Wastes are pumped into deep lagoons and left to sit for up to three months, sludge settles to the bottom of the lagoon, and anaerobic microorganisms break down the sediment. The remaining liquid is pumped into shallow, secondary lagoons, where wave action aerates the water. Aerobic microorganisms, particularly algae, break down organic chemicals suspended in the water. Eventually, the microbes die, and the clarified water is released into 11/23/16 3:44 AM       Industrial Microbiology ◀ FIGURE 25.8 Traditional sewage treatment. (a) A municipal wastewater treatment facility. (b) The traditional sewage treatment process. Microbial digestion during secondary treatment removes most of the biochemical oxygen demand (BOD) before the water is chemically treated and released; dried sludge is recycled as landfill. (a) 1 Primary Treatment: Sedimentation Sewer line Floating contaminants are skimmed off and heavy ones settle as sludge 2 Water effluent Primary sludge (particulates + flocs) Primary sludge Secondary Treatment Activated sludge or trickle filter system promotes microbial degradation of organic material 3 Removal of 25–35% BOD Aerated sludge Chemical Treatment 4 Secondary sludge Removal of 75–95% BOD Sludge Treatment Anaerobic digestion of sludge Disinfection, often by chlorination, prior to release Methane burned off or trapped for fuel Dried sludge for landfill or agriculture (b) rivers or streams. One problem with oxidation lagoons is that they are open, which can be dangerous if floodwaters inundate the lagoons and spread largely untreated animal wastes over a wide area. Artificial Wetlands Since the 1970s, small planned communities and some factories have constructed artificial wetlands to treat wastewater. Wetlands use natural processes to break down wastes and to remove microorganisms and chemicals from M25_BAUM7206_05_SE_C25.indd 793 water before its final release. Individual septic tanks are not needed; instead, wastewater flows into successive ponds where microbial digestion occurs (FIGURE 25.10). The first pond in the series is aerated to allow aerobic digestion of wastes; anaerobic digestion occurs in the sludge at the bottom. The water then flows through marshland, where soil microbes further digest organic chemicals. A second pond, which is still and contains algae, removes additional organic material, and the water then passes through open meadowland, where grasses and plants 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C Sedimentation in septic tank Filtration in leach field House Sludge (must be pumped out eventually) Pipes beneath ground distribute water through leach field. ▲ FIGURE 25.9 A home septic system. After wastewater from the house enters the septic tank, solids settle out as sludge, and the effluent liquid is filtered by the soil in the leach fields. ◀ FIGURE 25.10 Wastewater treatment in an artificial wetland. The majority of the BOD is removed by microbial action in the first pond; natural filtration by plants and the soil removes pollutants and the rest of the BOD. Why aren’t artificial wetlands feasible for major metropolitan areas? Figure 25.10 Major metropolitan areas produce too much sewage and have too little available space for artificial wetlands. Houses and businesses of planned community release sewage Pond 1: Aeration • Aerobic digestion of wastes in water column Marsh Water • Soil microbes digest organics. Pond 2: Still water Water • Algae continue digestion of organics. Meadow Water • Grasses filter out pollutants. • Anaerobic digestion of wastes in sludge Water release into third pond or waterway M25_BAUM7206_05_SE_C25.indd 794 11/23/16 3:44 AM       Chapter Summary A Case of Dehydration? e ell M Why In a polluted lake, the microbes reproduced prolifically, died, and then sank to the bottom, “feeding” anaerobes in the sediment. Even though the surface water looks clear, why is it still unsafe to drink the lake water? Mateo immediately. In addition, local health authorities work with the CDC to identify the source of Mateo’s intoxication. Mateo remains on the ventilator for five months. After he can breathe independently, he is released into a nursing care facility for another three months. When Mateo returns home, he finds that he still tires easily, but ultimately he makes a complete recovery. The public health inspection reveals the source of infection to be marinated mushrooms from the restaurant where Mateo enjoyed dinner with his coworkers. Three other people were infected; all were successfully treated with antitoxin. 1. Why was Mateo’s botulism treated with antitoxin rather than antibiotics? 2. Propose one hypothesis for how the mushrooms became contaminated with Clostridium botulinum. P The initial examination at the center doesn’t immediately reveal a cause for Mateo’s symptoms. Blood and urine cultures are taken and come back normal. In the meantime, Mateo’s breathing has become more difficult, and he starts having weakness in his facial, arm, and leg muscles. Dr. Mitchell suspects a neurological disease and orders a brain scan and nerve conduction test. He also obtains a sample of cerebrospinal fluid (CSF). The brain scan and CSF are normal. In contrast, the nerve conduction test reveals decreased conductance (that is, an inability to generate proper action potentials). The results of the tests and Mateo’s symptoms prompt Dr. Mitchell to order another test for botulinum toxin, produced by Clostridium botulinum. Mateo’s condition continues to deteriorate— he must be put on a ventilator to assist his breathing, and his limbs are paralyzed. The results of the toxin test come back positive; Dr. Mitchell obtains antitoxin (antibodies against botulism toxin) from the Centers for Disease Control and revention (CDC) and begins treating T ­ trap pollutants. By the time the water reaches a final pond, most of the BOD and microorganisms have been removed, and the water can be released for recreational purposes or irrigation. One drawback of an artificial wetland is that it requires considerable space—an artificial wetland to serve a small community can cover 50 acres of land or more. CHAPTER SUMMARY Food Microbiology (pp. 778–785) 1. The commercial use of microorganisms is referred to as applied microbiology and includes two distinct fields: food microbiology and industrial microbiology. 4. Alcoholic fermentations, usually performed by yeasts, convert sugars to ethanol and carbon dioxide. Alcoholic fermentation is used in the production of wine, distilled spirits, beer, vinegar, and bread. 2. Food microbiology involves the use of microorganisms in food production and the prevention of foodborne illnesses. In this context, fermentations involve desirable changes to food; spoilage involves undesirable changes to food. 5. Intrinsic factors of food spoilage are properties of the food itself, such as moisture content and physical structure, that determine a food’s susceptibility to spoilage. Extrinsic factors of food spoilage include ways in which the food is handled. 3. Food fermentations involve the use of starter cultures—known organisms that carry out specific and reproducible fermentation reactions. For most of the great variety of fermented vegetables, meats, and dairy products, starter cultures of lactic acid bacteria are used. The acid produced results in “sour” flavors. Some of the live organisms remaining in the fermented foods are probiotics— they can have health benefits. 6. Industrial processes preserve food via canning, pasteurization, drying, freeze-drying (lyophilization), irradiation, and aseptic packaging techniques. Natural and artificial preservatives are added to some foods to inhibit microbial growth. In stores and at home, foods should be properly stored in appropriate containers, cold foods should be kept cold, foods should be cooked thoroughly, and leftovers should be refrigerated to reduce spoilage. M25_BAUM7206_05_SE_C25.indd 795 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C 7. Food poisoning is a general term that describes instances of food infections (illnesses due to the consumption of living microbes) or food intoxications (illnesses due to the consumption of microbial toxins). Food poisoning frequently follows poor food handling. Industrial Microbiology (pp. 785–795) 1. Industrial microbiology is concerned with the use of microorganisms for the production of commercially valuable materials. Such industrial fermentations synthesize desired products and can use genetically modified microbes. 2. Batch production is the growth of organisms followed by harvesting of the entire culture and its products. Continuous flow production involves the constant addition of nutrients to a culture and the removal of the products formed. Products may be either primary metabolites (produced during active growth) or secondary metabolites (produced during the stationary phase). 3. Microorganisms produce a variety of useful products, including enzymes, dyes, alternative fuels, plastics, pharmaceuticals, pesticides, biosensors, and bioreporters. Alternative fuels (biofuels) can be produced from products of photosynthesis or by fermentation of biomass into fuel. Biosensors combine microbes and electronics to detect microbial activity in the environment. Bioreporters use microbes alone as sensors. 4. Contaminated water has something in it. Contamination can be physical, chemical, or biological. Water pollution refers to human contamination of water. 5. Potable drinking water is derived via water treatment, which involves the removal of microbes and of organic and inorganic contaminants from water. Polluted water contains organisms or chemicals at unsafe levels. Some diseases, such as diarrhea or shellfish intoxication, can result from consumption of polluted water or of food harvested from polluted water. 7. Wastewater (sewage) refers to water used for washing or flushed from toilets. Wastewater treatment involves the removal of solids, organic chemicals, and microorganisms. Biochemical oxygen demand (BOD) is a measure of the amount of oxygen required to fully metabolize organic wastes. 8. Municipal wastewater treatment involves four phases. Primary treatment entails sedimentation of large materials (primary sludge) and flocculation. Secondary treatment involves sedimentation of secondary sludge, as well as the removal of microorganisms and organic material using activated sludge systems or trickle filters. In the third phase, effluent water is chemically treated (chlorinated) and released. In the fourth phase, primary and secondary sludge is digested and dried to produce landfill. Methane gas can also be recovered during the processing of sludge. 9. Septic tanks and cesspools are home equivalents of municipal wastewater treatment. After wastewater leaves the home, it is deposited in underground tanks. Sludge settles, and the water is released into the soil, where natural processes remove organic chemicals and microorganisms. 10. Oxidation lagoons are used by farmers and ranchers to process animal wastes. Waste is pumped into successive lagoons, where microorganisms digest wastes before the water is released into natural water systems. 11. In artificial wetlands—found in some planned communities and industrial sites—ponds, marshes, and meadowland remove organic compounds, chemicals, and microorganisms from sewage as the water moves through them. VI W E RE E QU STIONS FO R 6. Drinking water treatment involves four steps: sedimentation, flocculation, filtration, and disinfection. Sedimentation removes large materials. In flocculation, alum combines with suspended materials to make them precipitate. Filtration, using either slow or rapid sand filters, removes microorganisms and chemicals. Disinfection, usually chlorination, kills most microbes that remain after filtration. Water is potable following treatment if it has zero detected coliforms (indicator organisms) in 100 ml of water, as determined by one of several testing methods (membrane filtration, ONPG/MUG test). Answers to the Questions for Review (except Short Answer questions) begin on p. A-1. Multiple Choice 1. Food fermentations do all of the following except _____________. a. give foods a characteristic taste b. lower the risk of food spoilage c. sterilize foods d. increase the shelf life of the food 3. Which of the following lists foods in order, from perishable to nonperishable? a. dried pasta, cheese, fruit, uncooked ground beef b. dried pasta, fruit, uncooked ground beef, cheese c. uncooked ground beef, fruit, cheese, dried pasta d. uncooked ground beef, fruit, dried pasta, cheese 2. Commercially produced beers and wines are usually fermented with the aid of _____________. a. naturally occurring bacteria b. naturally occurring yeast c. specific cultured bacteria d. specific cultured yeast 4. Which of the following would be the best growth medium to use for industrial fermentations? a. corn b. synthetic medium made by hand c. whey from cheese production d. brewing mash M25_BAUM7206_05_SE_C25.indd 796 11/23/16 3:44 AM 5. Biodegradable plastics are made from which of the following microbial metabolites? a. sludge c. BOD b. PHA d. alum 6. Strains of the bacterium Pseudomonas syringae have been identified as being capable of _____________. a. producing plastics b. producing alternative fuels c. fermenting foods d. preventing ice formation 7. Which of the following is added during water or sewage treatment to promote flocculation? a. sludge c. BOD b. PHA d. alum       R Questions for eview Fill in the Blanks 1. Intrinsic factors affecting food spoilage are properties of __________________ rather than __________________. 2. Leaving foods out at room temperature __________________ the likelihood of food spoilage. 3. The two types of industrial fermentation equipment are designed for __________________ production or __________________ production. 4. Potable water is allowed to have __________________ coliforms per 100 ml of water tested. 5. A primary goal in wastewater treatment is to reduce the _________________________. 8. During chemical treatment of drinking water and wastewater, which of the following microbes is least likely to be inactivated or killed? a. algae c. fungal spores b. viruses d. bacteria 6. The presence of ________________________ in a water sample usually means pathogens have contaminated the source. 9. In which step is most of the organic content of sewage removed? a. primary treatment c. tertiary treatment b. secondary treatment d. sludge treatment 8. Compounds that can be used for energy and are made by microbes are called ____________________. 10. Industrial fermentation _____________. a. always involves alcohol production b. involves the large-scale production of any beneficial compound c. refers to the oxidation of sugars using organic electron acceptors d. is any desirable change to food by microbial metabolism 11. Lyophilization in food preservation is by _____________. a. cell lysis c. rapid heating b. gamma radiation d. freeze-drying 7. __________________ and ____________________ are the two types of food poisoning. 9. A __________________ is a device composed of microbes and electronics used to detect other microbes or their products. 10. __________________ is the amount of oxygen required by aerobic organisms to fully metabolize organic waste in water. Matching Match each term with its correct definition. A. Spoilage 2. _____ Refers to water that is fit to drink C. Coliforms 3. _____ Used in the processing of animal wastes; mimics primary and secondary wastewater treatment D. Pasteurization 4. _____ Water that is not bound by solutes F. Potable G. BOD 4. __________ Methane is a gas produced by microbial metabolism that can be used directly as a fuel source. 5. _____ Undesirable fermentation reactions in food, leading to poor taste, smell, or appearance 5. __________ The treatment of drinking water and sewage involves similar processes. 6. _____ Brief heating of foods during processing Indicate whether each of the following statements is true or false. Rewrite the underlined phrase to make a false statement true. 1. __________ The fermentation of dairy products relies on mixed acid fermentation.  2. __________ Sauerkraut production involves the alcoholic fermentation of cabbage.  Modified True/False   1. _____ Organisms whose presence in water indicates contamination from feces E. Secondary metabolites  3. __________ Pasteurization kills mesophilic microorganisms except endospore formers. B. Water activity  H. Oxidation lagoon  7. _____ Descriptor of the level of organic material present in wastewater  8. _____ Fermentative products produced by microorganisms during stationary phase M25_BAUM7206_05_SE_C25.indd 797 11/23/16 3:44 AM p er 25    Applied and Industrial Microbiology t ha       C VISUALIZE IT! 1. Label the steps in the cheese-making process. 1. _____________________ kills unwanted microbes. 2. Label the steps in the wine-making process. Removal of stems and crushing grapes to make 1. ______________________________ Addition of 2. _____________________ Addition of 2. _____________________ Coagulation of milk proteins (curd formation) Disposal of liquid whey as waste product Production of 3. _____________________ 3. ____________ Cutting of 4. _____________________ 4. ____________ Production of 5. _____________________ through 6. _____________________ 7. _____________________ and 8. _____________________ 5. ______________ 6. ____________ M25_BAUM7206_05_SE_C25.indd 798 11/23/16 3:44 AM       R Questions for eview R C ITICAL THINKING 1. Why does the application of recombinant DNA technology to food production enhance not only food quality but also food output? Given that it has the potential to feed more people, why are some people opposed to genetic modification of foods? 2. Given what you know about microbial nutrition and metabolism, explain why it is technically more difficult to achieve high yields of a secondary metabolite than of a primary metabolite. 3. Compare the types of alternative fuels that could be produced by microbes. Based on starting materials, which would provide the most renewable energy? ­ 4. One way that farmers are attempting to prevent the development of widespread insect resistance to Bt toxin is by planting non-Bt-producing crops in fields adjacent to Bt-containing crops. Insects can infest both fields, but the insects eating the non-Bt plants should survive at a higher rate than those in the Bt-producing field. Why would this prevent the dissemination of resistance to Bt toxin among insects? 5. Even though water and wastewater undergo essentially the same forms of treatment, treated wastewater usually is not put into the water system from which drinking water is derived. Why not? 6. Take a critical look at the garbage in all of your wastebaskets. How much of the material present could possibly be degraded M25_BAUM7206_05_SE_C25.indd 799 by microbes? How much could be recycled either at a recycling center or in compost? What would be left if the degradable or recyclable materials were removed? 7. Explain why sake—sometimes called rice wine—would be more accurately described as “rice beer.” 8. Inexpensive bulk wines and wines that have been left exposed to air for too long often acquire a “vinegary” smell or taste. Explain why this is so. 9. Describe the intrinsic and extrinsic factors that would contribute to the spoilage of butter, an apple, and a steak. 10. Suggest why neither the original food involved nor the organism responsible can ever be identified in more than 80% of food poisoning cases. 11. Which process would you expect to yield more product from the same-sized vats over the same amount of time: batch production or continuous flow production? Why? 12. Explain how biosensors might be used for water quality testing. Would biosensors identify the microorganisms present in water more accurately than an ONPG or MUG test? Is such specificity truly necessary? 11/23/16 3:44 AM M ING APP CONC EPT       Chapter 25   Applied and Industrial Microbiology Using the following terms, fill in the following concept map that describes microbial roles in food production. Soybeans and wheat Sugars in milk Malt from grains Saccharomyces cerevisiae (yeast) Fermentation Gluconobacter Lactobacillus bulgaricus Acetic acid in vinegar Alcohol in wine Bread to rise Microbial roles in food production include 1 of 2 by Sugars in bread dough Sugars in fruit juice by by 4 10 by by produces Aspergillus spp. and Lactobacillus spp. Alcohol in beer produces 5 3 or Streptococcus thermophilus causes produces 6 7 produces can be oxidized by Cheese Soy sauce 8 Acetobacter producing 9 M25_BAUM7206_05_SE_C25.indd 800 3/6/18 4:36 PM