Term Paper MICRB 201H MOOOOOve Over Microbes: Psychrotrophs in Milk Introduction

Alaina Weinheimer Dr. Porter

Got milk? Dairy products are a large part of Americans diet. From the cheese on a slice of pizza to the milk added to cereal, the average American consumes about 604 pounds of milk every year (1). With high-tech sterilization and pasteurization techniques used by farmers and refrigerating these products at home, one would think that the cream Nana adds to her coffee in the morning is scotch-free of contaminants. Think again. A plethora of microbes and their byproducts are adapted to withstand the seemingly rigorous preservation processes. Not all of these microbes and by-products are pathogenic, but those that are can cause serious gastrointestinal problems. According to one study, about 32.5% of raw bulk milk contains pathogens (2). Not all pasteurization and sterilization processes are perfect, so there is a considerable chance that these pathogens could reach your dinner table. The masters at surviving the processes milk goes through are the psychrotrophs (3). Whether it be contamination by the microbe itself or a product of the psychrotrophs, no glass of milk is entirely safe. The three most common pathogenic psychrotrophs in milk are L. monocytogenes, Y. enteroclitica, and B. cereus. Although, these pathogens are no reason to stop drinking milk, they are definitely something to consider while at the grocery store (Im cereus!) Milk Sterilization Techniques Bacteria have no problem entering the milk. Mother Nature knows no boundaries. There is a significant amount of bacteria in raw milk. The ruminant intestinal tract of cow is considered a reservoir of bacteria. It is speculated that the bacteria enter the ruminant of a cow

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

via fecal contamination of the cows food and water. These microbes amplify in the gut of a cow and typically exit via the feces, which can lead to fecal contamination elsewhere. Though uncommon, some foodborne pathogens cause mastitis, resulting in the excretion of the bacteria into the milk. Usually the pathogens concentration from mastitis is so dilute that its presence is not even detectable in milk samples. For the most part, though, milk is contaminated by contact with contaminated equipment, such as milk cans (4). In short, there are many avenues for microbes to come into contact with raw milk. The first real obstacle psychrotrophs have to face is heat. To reduce the number pathogens and spoilage-causing microbes, milk is often pasteurized. Pasteurization involves heating milk to a temperature that kills bacteria without causing the milk to boil. The heat is intended to denature proteins of microbes in the milk, which kills the microbes. There are different types of pasteurization. For small quantities of milk, batch pasteurization is used. In this case, milk is heated to 63C for thirty minutes. When pasteurizing large quantities of milk, a plate heat exchanger may be used, allowing the milk to be heated at higher temperatures and cooled more quickly. This type of pasteurization is most commonly used for milk that is sold at the grocery store. It is called High Temperature Short Time (HTST) pasteurization. The name explains it all. More specifically, the high temperature that the milk is heated to is around 72C and the short time is fifteen seconds. As a rule, the higher the temperature the milk is heated, the shorter the heating time. After the fifteen seconds, the milk is cooled to under 10C (5). At 72C, it is presumed that the microbial count is reduced without burning the milk (3). The taste of milk is attributed to its chemical properties. At higher temperatures, the chemical

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

properties of milk are altered, causing a taste unpleasant to most Americans (9). These techniques greatly reduce microbial load but pasteurization never completely rids the milk of microbes. There are sterilization techniques, such as Ultra High Temperature treatment, which is used to annihilate the microbial population in milk. The milk is put under pressure and heated at 140C for five seconds (5). At 140C, it would be expected that the milk would lose its fresh taste and a considerable amount of vitamins would be destroyed. However, the pressure reduces this destructiveness. Scientists at Oregon State University are finding that the higher the pressure applied to the milk, the lower the temperature required to kill the bacteria. High pressure techniques could become the most common milk process in the future (9). For the most part, in order for microbes to contaminate milk at home, they must endure the heat of pasteurizing techniques. The second obstacle bacteria face is cooling. Milk is preserved by refrigeration. At low temperatures, it is thought the growth of remaining bacteria is slowed or completely halted. Temperatures below 7C are thought to be the appropriate temperature of refrigeration (6). Refrigerated milk usually expires after a week (7). On top of heating, microbes must survive cooling to reach a consumers body. Pasteurization Survival To overcome these obstacles of heating and cooling, microbes must have special adaptatoins. Many pathogens are mesophilic, growing optimally between 10 - 50C, mainly because the human bodys temperature hovers around 37C (10). Mesophilic bacteria are generally wiped out during pasteurization because they cannot survive the heat, and their growth

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

is slowed during refrigeration because refrigeration temperatures are generally below their growth range. The professionals at surviving the conditions of milk processing are the psychrotrophs. Psychrotrophs are microbes that grow optimally at room temperatures, 20 35C, but are capable of growing at refrigeration temperatures (6). One would presume that psychrotrophs would be annihilated by pasteurization temperatures, well above 35C, but they manage to be the leaders in milk contamination. The methods to their madness include the excretion of enzymes that can endure the pasteurization temperatures and the formation of spores (3). The strategy of contaminating milk by enzyme excretion is usually used by rod-shaped Gram-negative bacteria. There are different types of heat-resistant enzymes that produce different effects on the quality of milk. These enzymes include proteases and lipases. Proteases deteriorate milk quality by breaking down proteins in the milk. Pseudomonas, Achromobacter, and Aeromonas species commonly excrete the proteases in milk that cause this to happen (3). Pseudomonas species also often produce lipases in milk. Lipases degrade the fat in milk. Casein micelles are often the target of lipases. Naturally, milk contains both proteases and lipases by itself, but the bacterial excretion contributes to the degradation of the quality of milk (11). In short, rod-shaped Gram-negative bacteria contaminate milk indirectly by excreting heat resistant enzymes that degrade the quality of milk. Gram-positive take the second approach to surviving milk preservation processes. These bacteria form spores during pasteurization of milk at high temperatures. The spores allow the bacteria to survive the heat. After pasteurization, when the milk reaches a more favorable

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

temperature for cell growth, the cell is then activated and the bacteria then germinates as a regular, vegetative cell. Increasing pasteurization temperatures have been linked to an increase in spore forming bacteria (11). Although, Gram-positive bacteria are less common in milk than Gram-negative bacteria, they are the leading cause of milk spoilage in good hygiene farms. The genii that form spores during milk pasteurization are Bacillus and Clostoridium. Bacillus species mainly cause the spoilage in milk. Bacillus cereus is the most common species isolated from milk. This species causes the milk to taste bitter and may contain an enterotoxin. Some Grampositive bacteria use the same tactic as the Gram-negative bacteria when it comes to milk contamination. For instance, Gram-positive Enterococcus species produce protesases (3). In summary, Gram-positive bacteria survive pasteurization and preservation processes by endospore formation and heat-resistant enzyme excretion, enabling the bacteria it contaminate the milk you put on your Lucky Charms. Pathogenic Party-poopers (no pun intended) Although some psychrotrophs can survive pasteurization and proliferate in ones milk glass, the main cause of foodborne illness via pasteurized milk consumption is postpasteurization contamination. Residual water on cooling tanks provides a brewing pot of psychrotrophs. Other sources of contamination include the transfer of the milk to jugs and athome cross-contamination (8). Not all bacteria that grow in milk are pathogenic; however there are a considerable variety of pathogenic bacteria that cause foodborne illnesses. The three most common pathogenic psychrotrophs in milk include Listeria monocytogenes, Yersinia enterocolitica, and Bacillus cereus. To avoid infection by Y. enterocolitica, pack your bags and

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

head South. This bacteria is most common in milk in the northern part of the northern hemisphere, and cases of infection by Y. enterocolitica has been increasing in the past fifteen years. Y. enterocolitica causes a type gastroenteritis called yersiniosis. Symptoms of yersiniosis include abdominal pain, headaches, fever, and of course, diarrhea. Although Y. enterocolitica contains an enterotoxin, this bacteria causes damage by invasiveness. While youre heading South to get away from Y. enteroclitica, also avoid California, where the first major outbreak of foodborne illness caused by L. monocytogenes occurred (6). L. monocytogenes typically contaminates milk when the milk makes contact with wet areas of dairy plants, but the bacteria can also entire raw milk via mastitis. Even if the milk is pasteurized, L. monocytogenes has been found to survive minimal HTST (2). Not surprisingly due to its name, Listeria monocytogenes causes the disease called listeriosis. Listeriosis causes symptoms similar yersiniosis. They are both gastrointestinal diseases. People of different age groups and pregnancy statuses are affected by listeriosis differently, but the typical adult experiences fever, muscle aches, and diarrhea (13). Unlike Y. enterocolitica and L. monocytogenes, B. cereus is unavoidable by train, plane, or automobile. (Neither are the other two pathogens, but their prevalence is more concentrated in certain areas.) Psychrotrophic and mesophilic strains of the spore-forming B. cereus exist (6). Both strains are capable of producing toxins that harm milk consumers. One of these toxins causes diarrhea, and the other toxin causes both diarrhea and vomiting. B. cereus can excrete these toxins at low temperatures, temperatures as low as your refrigerator. In fact, B. cereus has been isolated from unopened cartons of milk that had been refrigerated at 10C before the expiration date. Milk isnt the only dairy product B. cereus chills out in (no pun intended).

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

Chocolate milk, ice cream, and infant formula all provide a welcoming home for this pathogen. Whats even scarier is that not much B. cereus is required to cause infection. About 1 X 107 CFU/mL of B. cereus will make a milk drinker or ice cream lover take several trips to the toilet (3). The psychrotrophic pathogens that reside in pasteurized milk are terrifying enough to make one return their carton back to their fridge. Though most of us dont want to live in an igloo, neither do mesophilic bacteria, but like Eskimos, some mesophilic bacteria make it work in cold climes. One mesophilic bacteria that has strains capable of growing in temperatures as low as 6.9C is E. coli (6). A particularly dangerous strain of E. coli is STEC, Shiga Toxin-Producing E. coli. STEC is extremely potent; infection requires only 1 100 CFU/mL. The Shiga toxin that is produced is linked to causing non-bloody diarrhea, diarrhea-associated hemorrhagic colitis (inflammation of the colon), hemolytic uremic syndrome (HUS, reduction of red blood cell count and acute kidney failure) and thrombotic thrombocytopenic purpura (blood clot formation in small blood vessels) (2). None of these diseases are pleasant. In fact, most can cause death. The ability of mesophilic pathogens, such as STEC, to grow in cold temperatures is alarming. Prevelance of Pathogens Considering all of the pathogens that can be swimming in your bowl of Raisin Bran, drinking milk does not seem appealing, even when the milk is pasteurized. Milk lovers can rest at ease, though. These pathogens are not prevalent enough to make the milk industry shut down. L. monocytogenes is hardly common in raw milk, let alone pasteurized milk. The prevalence of L. monocytogenes in bulk milk has been reported to be 1 12% (2). Y. enteroclitica also

Term Paper MICRB 201H

Alaina Weinheimer Dr. Porter

appears rarely in milk. In the testing of samples of milk from 183 different farms in Finland (which is considered to have the highest quality hygienic practices), only 7.7% of the samples contained Y. enteroclitica (14). Disease caused by Y. enteroclitica only occurs in one out of every 100,000 people each year (16). Even less common in milk than L. monocytogenes and Y. enteroclitica is STEC (thankfully!). The prevalence of STEC in milk is 0.8% - 3.8%. On the other end of the spectrum, the presence of B. cereus in bacteria is a little alarming. In 20.8% of the milk samples that were taken in the study of Finnish milk, the scientists detected B. cereus. However, the concentration of B. cereus in the milk samples hovered around only 1 CFU/mL, which is 1 X 107 CFU/mL less than the necessary concentration to cause infection (14). The only major outbreaks of B. cereus reported occurred in 1988 and 1989 in Canada. Thirty-six people became ill from drinking milkshakes from a fastfood joint in Ontario 1988. The 1989 incident took place in a school in Quebec, where seventy-four students were infected by milk that contained B. cereus. The contamination was attributed to temperature abuse and poor stock rotation (15). Overall, though, the prevalence of these pathogens in milk is hardly any reason to avoid the dairy aisle at Giant. Regarding the future, the presence of pathogens in milk has been decreasing over years due to the reduction in post-pasteurization contamination. This means that the main concern of milk contamination will center on the psychrotrophs that have features well-adapted to the pasteurization processes, such as Bacillus cereus strains (3). Better pasteurization techniques are emerging that can kill more bacteria with less heat. These techniques involve the application of high pressure at a temperature lower than HTST pasteurization (9). As for now, milk is still safe enough to drink. The industry is merely improving.

Term Paper MICRB 201H Conclusion

Alaina Weinheimer Dr. Porter

Microbes are everywhere. Despite the current rigorous pasteurization techniques used by farmers, bacteria still manage to contaminate the milk in your fridge. These bacteria mainly include psychrotrophs, which either produce endospores to survive pasteurization or enzymes that are heat-resistant to pasteurization. Some mesophilic bacteria have been found to grow in refrigerated milk, including the extremely pathogenic Shiga Toxin-Producing E. coli., STEC. The diseases the psychrotrophs cause mainly affect the gastrointestinal system, but the mesophilic STEC affects blood cells. Regardless of infection location, these pathogens are hardly prevalent enough to render the abstinence of dairy products. In fact, pasteurization techniques are currently improving, making milk taste fresher while killing more microbes. The presence of microbes in milk is definitely something to consider, but no reason to not drink milk. So go on, rock that milk mustache!

Term Paper MICRB 201H Literature Citation

Alaina Weinheimer Dr. Porter

1. "Consumption - US." Dairy Statistics. Wisconsin Cheese. Web. 15 Apr 2013. 2. Oliver, S. P., and R. A. Almeida. "Foodborne Pathogens, Mastitis, Milk Quality, and Dairy Food
Safety."National Mastitis Council. National Mastitis Council, 2005. Web. 15 Apr 2013. <http://nmconline.org/articles/MilkQualFoodSafety.pdf>.

3. Champagne, Claude P., Richard R. Laing, Denis Roy, and Assanta Mafu Akier. "Psychrotrophs
in Dairy Products: Their Effects and Their Control." Critical Reviews in Food Science and Nutrition. 34.1 (1994): 1-30. Web. 15 Apr. 2013.

4. Oliver, S. P., B. M. Jayarao, and R. A. Almeida. "Foodborne Pathogens in Milk and the Dairy
Farm Environment: Food Safety and Public Health Implications." Foodborne Pathogens and Disease. 2.2 (2005): n. page. Web. 15 Apr. 2013.

5. "Milk Microbiology." International Livestock Research Institute. International Livestock

Research Institute. Web. 15 Apr 2013. <http://www.ilri.org/InfoServ/Webpub/fulldocs/ilca_manual4/Microbiology.htm

6. "Extended Shelf Life Refrigerated Foods: Microbiological Quality and Safety." IFT. International
Food Technologies, 1 Feb 1998. Web. 15 Apr 2013. <http://www.ift.org/KnowledgeCenter/Read-IFT-Publications/Science-Reports/Scientific-Status-Summaries/ExtendedShelf-Life-Refrigerated-Foods.asp&xgt;.

7. "Shelf Life of Milk." EatByDate. EatByDate. Web. 15 Apr 2013.

<http://www.eatbydate.com/dairy/milk/milk-shelf-life-expiration-date/>.

8. Walter de Santana, Elsa Helena, Vanerlia Beloti, et al. "Milk Contamination in Different Points
of the Dairy Process. ii) mesophilic, psychrotrophic and proteolytic microorganisms." Semina: Cincias Agrrias, Londrina. 25.4 (2004): 349-358. Web. 15 Apr. 2013.

9. "Pressure processing retains fresh milk taste, extends shelf life." Oregon State University

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Alaina Weinheimer Dr. Porter

Extension Service. Oregon State University, 13 Dec 2006. Web. 15 Apr 2013. <http://extension.oregonstate.edu/news/release/2006/12/pressure-processing-retainsfresh-milk-taste-extends-shelf-life>.

10. "Mesophiles." ORACLEThinkQuest. ORACLEThinkQuest. Web. 15 Apr 2013.

<http://library.thinkquest.org/CR0212089/meso.htm>.

11. "Milk Enzymes." MilkFacts. MilkFacts. Web. 15 Apr 2013. <http://www.milkfacts.info/Milk

Composition/Enzymes.htm>.

12. "Psychrotrophic Spore-Forming Bacteria in Raw and Pasteurized Milk." Dairy Food Science
Notes. Cornell University, 7 Jul 2010. Web. 15 Apr 2013. <http://foodscience.cornell.edu/cals/foodsci/extension/upload/CU-DFScience-NotesBacteria-Psychrotophic-Spores-12-10.pdf>.

13. "Listeria." Centers for Disease Control and Prevention. Centers for Disease Control and
Prevention, 7 Jan 2013. Web. 15 Apr 2013. <http://www.cdc.gov/listeria/definition.html>.

14. Ruusunen, M., M. Salonen, et al. "Pathogenic Bacteria in Finnish Bulk Tank Milk." Dept. of
Food Hygiene and Environmental Health. 10.2 (2013): 99-106. Web. 15 Apr. 2013.

15. "Foodborne Illness Outbreaks." Milk Safety Notes. BC Center for Disease Control, 28 Jun 2002.
Web. 15 Apr 2013. <http://www.bccdc.ca/NR/rdonlyres/34B36D22-D767-4140-B03235FE8AAD409F/0/Outbreak_Bacillus_Milk.pdf>.

16. "Yersinia enterocolitica." Centers for Disease Control and Prevention. Centers for Disease
Control and Prevention, 25 Oct 2005. Web. 15 Apr 2013. <http://www.cdc.gov/ncidod/dbmd/diseaseinfo/yersinia_g.htm>.