FOOD MICROBIOLOGY: THE BASICS AND THE DETAILS OF CHEESE PRODUCTION
(August 2003)
Food spoilage has been an important problem throughout human history. Finding ways to overcome this problem was crucial as communities became larger and individuals no longer grew their own food. Some kind of system was needed to maintain the nutrient content of various food stuffs for long periods of time and prevent them from rotting and becoming inedible.
Early solutions to food spoilage
Food spoilage is caused by the growth of microorganisms, primarily bacteria and fungi, that convert nutrients into energy which they use for their own growth. Depletion of the nutrient content of food as well as the secretion of byproducts from this biochemical process are two things which contribute to the spoilage of food rendering it inedible. Since ancient times, humans have used many methods to extend the shelf life of food although not always understanding how these processes worked. Salting and drying are two very simple techniques that prevent rotting; both make the food an inhospitable environment for microorganisms. Canning is another technique first developed in the late 18th century by Nicholas Appert, a French confectioner, who, after 15 years of research, realized that if food is sufficiently heated and then sealed in an air tight container it will not spoil. Here the heating of food, kills all residual microorganisms present in the food and immediate sealing prevents the reentry of other contaminanting organisms. Napoleon immediately put this discovery to work in his armed forces and awarded Appert a prize of 12,000 francs for his discovery. Later, an Englishman, Peter Durand, took the process one step further and developed a method of sealing food into unbreakable tin containers. This was perfected by Bryan Dorkin and John Hall, who set up the first commercial canning factory in England in 1813. In 1859, Louis Pasteur definitively showed that microorganisms were responsible for food spoilage for the first time. This discovery led to the coining of the term “pasteurization” to describe the process where liquids with the potential to spoil (milk in particular) are heated for preservation.
Fermentation
In some cases, the growth of microorganisms in food can be put to good use for the production and preservation of various types of food. Fermentation is arguably the earliest example of biotechnology and refers to the metabolic process by which microbes produce energy in the absence of oxygen and other terminal electron acceptors in the electron transport chain such as fumarate or nitrate. In ancient times, it was considered as a way to both preserve food and to retain nutritional value. It was probably accidentally discovered in ancient Egypt when dough, made from ground up wheat and rye, was left for a period of time before cooking. In contrast to dough that was immediately cooked, it was observed that the aged dough expanded in size and when cooked produced tastier, lighter bread. The process was not completely reproducible: sometimes the uncooked dough yielded good bread and other times it did not. However if small amounts of good dough was added to the next batch, the bread was again tasty. The Romans went onto improve and perfect this process and popularized this sort of bread throughout the European continent. The discovery of fermentation in Egypt also led to the first production of wine and alcohol. All these discoveries were largely phenomenological and it would be another 3000 years before the exact cause of fermentation was uncovered. It was Louis Pasteur, again, in 1857 who was able to demonstrate that alcohol can be produced by yeast when grown in particular conditions. This discovery revolutionized the modern food industry: for the first time the agent of fermentation was identified and could be used commercially.
Industrial processes using fermentation
Fermentation by bacteria, yeast and mold is key to the production of fermented foods. Fermenting yeast produces the alcohol in beer and wine. In fact, the smell of fresh baked bread and rising dough can be attributed to alcohol produced from yeast. Fermentation is used to make many ethnic foods such as sauerkraut and miso. Soy sauce is produced by fermenting Aspergillus ortzae, a fungus, growing on soy beans. Erwinia dissolvens, another type of bacteria, is essential for coffee bean production; it is used to soften and remove the outer husk of beans. Finally, fermentation of milk produces most dairy products. Without microbes, we would not be able to eat many types of different food that we enjoy today. Table 1 shows example of several foods that are produced through fermentation with specific organisms.
The biochemical process
All organisms need energy to grow. This energy comes from the reduction of adenosine triphosphate (ATP) into adenosine diphosphate (ADP) and results in the release of energy and a phosphate group. In this way ATP serves as a storage molecule of energy which can be used by the cell. But where does the ATP come from? Cells get their ATP from the controlled chemical breakdown of glucose to form two molecules of pyruvate. This process requires two molecules of ATP but results in the release of four molecules or a net gain of two molecules of ATP. This process is referred to as glycolysis and is illustrated in Figure 1. Once pyruvate is formed, it can be processed in several different ways. Mammalian cells usually process pyruvate by putting it into the tricarboxylic or Kreb’s cycle. In the presence of oxygen, oxidative phosphorylation produces more ATP from the byproducts of the Kreb’s cycle reactions. This is referred to as aerobic respiration. However when oxygen is limiting, other processes must be used in order to deal with pyruvate. This is done through anaerobic respiration or fermentation and involves the breakdown of pyruvate into simpler compounds. Two of the most important fermentation processes which are used on an industrial scale are ethanol or lactic acid fermentation. This is illustrated in Figure 1.
Dairy Fermentation
Milk is an excellent food source for humans and bacteria alike. It is full of vitamins, fats, minerals, nutrients and carbohydrates. It is rich in the protein casein which gives milk its characteristic white color. The most abundant carbohydrate is the disaccharide lactose, “milk sugar.” At room temperature, milk undergoes natural souring caused by lactic acid produced from fermentation of lactose by fermentative lactic acid bacteria. This accumulation of acid (H+ ions) decreases the pH of the milk and cause the casein to coagulate and curdle into curds and whey. Curds are large, white clumps of casein and other proteins. Whey is the yellow liquid that is left behind after the casein has formed curds. Thus, bacteria obtain nutrients from the milk, inadvertently curdle it and humans use it as the first step in making many dairy products.
The microbes important for dairy product manufacturing can be divided into two groups, primary and secondary microflora. Products undergoing fermentation by only primary microflora are called unripened and those processed by both primary and secondary microflora are called ripened. Primary microflora are fermentative lactic acid bacteria which cause the milk to curdle. During dairy product production, milk is first pasteurized to kill bacteria that cause unwanted spoilage of the milk and of the downstream milk products. Primary microflora consists of certain kinds of Lactococcus, Lactobacillus and Streptococcus that are intentionally added to pasteurized milk and grown at 30°C or 37°C (temperature depends on the bacteria added). Secondary microflora include several different types of bacteria (Leuconstoc, Lactobacillus, and Propionibacterium), yeasts and molds; they are only used for some types of surface ripened and mold ripened cheeses. The various combinations of microflora determine what milk product you will end up with.
Different unripened milk products are created by using various starting products and bacteria. For buttermilk production, Lactobacillus bulgaris (named for its country of discovery, Bulgaria) is added to skim milk to curdle it. Leuconostoc is then added to thicken it. Sour cream is made the same way except cream is used instead of skim milk. During yogurt production, dry milk protein is added to milk to concentrate the milk before addition of actively growing Streptococci and Lactobacilli. Butter is produced by curdling and slight souring from Streptococci growing in sweet cream. Leuconostoc is then added so it can synthesize diacetyl, a compound that gives butter its characteristic aroma and taste. The milk is then churned to aggregate the fat globules into solid butter.
Thus milk type and bacteria will determine the dairy product produced.
Cheese is an important product of fermentative lactic acid bacteria. Particularly in the past, cheese was valued for its long shelf life. Due to its reduced water content, and acidic pH, bacterial growth is severely inhibited. This causes cheese to spoil much more slowly than other milk products. Consequently, the art of cheese production has spread throughout Europe, each country manufacturing many different types of cheeses. Cheese production has three steps: curd formation, curd treatment and curd ripening.
1. Curd formation can use mare, ewe, cow or goat milk to produce “sour” or “sweet” curd. Sour curd is produced by fermentative lactic acid bacteria as mentioned above. Sweet curd is produced by adding an enzyme called renin instead of bacteria to curdle the milk. The curd is separated from the whey by draining. The curd can be used directly to make unripened cheeses such as ricotta or cottage cheese or can undergo further processing to make a ripened cheese.
2. Curd treatment consists of condensing and squeezing to form dense, hard curd. It is then molded into the desired shape, salted and mixed with different types of secondary microflora.
3. Secondary microflora ripen the cheese and will determine the final texture and aroma of each type of cheese. For hard ripened cheeses such as Cheddar, curds are further compressed and the bacteria particular for the cheese is added. The Cheddar is wrapped in wax or plastic to prevent contamination and then incubated to allow the bacteria to do its work. For soft ripened cheeses such as Camembert and Limburger, a microbe, usually mold, is added to the surface of the cheese that produces a protein-digesting enzyme. This enzyme breaks apart the curds and causes the cheese to become creamy and spreadable.
Many cities have long held traditions and nuances for producing a particular cheese i.e. the limestone caves in Roquefort, France which have constant heat and humidity that create unique and delightful cheeses. Figure 2 shows a schematic diagram of the cheese manufacturing process.
Thus, microbes can not only be harmful to society but also can be manipulated in a variety of ways for the benefit of society. Particularly in the preservation and production of food, microbes have proven to be useful and essential.
References
1. Alcamo, I. (2003). Microbes and Society. Missassauga, Ontario, Jones and Bartlett.
2. Doyle, M., L. Beuchat, et al., Eds. (1997). Food Microbiology: Fundamentals and Frontiers. Washington, DC, ASM Press.
3. Foster, E., F. Nelson, et al. (1957). Dairy Microbiology. Englewood Cliffs, Prentice Hall.
(Art by Fan Sozzi)