How does it actually work? How does water activity control microbial growth? We’re going to talk about its mode of action, how it actually does that. Here in this slide, I have little beautiful ovals that are going to represent our microorganism. In here, we have – interior to the microbe – water activity of 0.95, and the environment that that microorganism is in is at 0.90. We know from thermodynamics, if we have a difference in water activity, we have a difference in energy level that it wants to go from high to low to even that out to equilibrate, so the water inside that microbe is going to want to leave. It does, and it moves out into the environment. What happens inside that cell is that trigger pressure is lost. That starts to stress that microorganism out. In response, the microorganism is going to try to stop that. How it does it is it’s going to try to equilibrate its own energies and water activities with the environment. You can see in the next little pathway there that the microbe tries to adapt by altering its membrane to reduce its water activity to maintain that trigger pressure. How it’ll do it is either they’ll produce or transport in small solutes to reduce the water activity. This could be amino acids, polyols, sugar, something like that. It’s trying to compensate for that loss of water out of it. You can see that it is able to drop it a bit, so it’s now at 0.93, but it’s still not matching the environment of 0.90.
In the next little section here, we see that is not able to do any more. That’s as low as this microorganism can do. If it’s unable to reach that equilibration with water activity to its surroundings, then that microbe will remain in what we call a lag phase, where there’s no growth or it’ll begin to sporulate and go dormant. It’s just in stasis right now. It can’t do anything. It can’t grow. The water is not available for it to take in to start reproducing. It will stay like that until the environment changes. If it’s back in an environment that’s 0.93 or above, then it would be able to start growing, but at this point right now, it is in stasis.
In the 1950s, W. Scott did some experiments on this idea of water activity in the different bacterium. What he did is he took different types of food and inoculated them with different bacteria. Here’s a list of the bacterium that he put in there, specific strains, and then he observed what happened. Did it grow? Did it not?
You’ll see the whole top part of this table here is based on Staph aureus. At various levels, we have inhibition, and we also have growth in this toxin that’s formed. That’s the dangerous part of the microorganism and produces that toxin which is what makes us sick. I wanted to point out that precooked bacon towards the middle there, you can see at 0.86 that there is growth for that strain, but at 0.84 there is not. You’ll see that it’s inhibited. That’s a really small difference in water activity, but it illustrates – you can also see as you go up that list – that from anything above 0.85 or above is the difference between growth or inhibition. For staph, that’s true that if you can lower your water activity to below 0.85, you will not have any staph. As a matter of fact, you’ll also find out as we go along here that nothing, no pathogenic bacteria, will be able to grow. Staph is the hardest. It’s the most hardy. It’s the most adaptive of these pathogenic bacteria.
What I did also want to point out is he put this in milk, cream filling, eggs, meat, cheese, beef, bacon, all of these different things are inoculated with staph, but it didn’t matter. The water activity limit stayed the same. That cut off is poor bacteria or bacterium, but not food matrix related, which is really important. You can use this in any of the industries.
Here’s the list where those lie for each one of them. We can see at the top here we have botulism, E. coli is listed here, there’s salmonella, Listeria, and then at the very bottom, you can see that Staph aureus, the aerobic version of that, is that 0.86. There are no more pathogenic bacteria that will grow below that level. If you have ever seen that – about the 0.85 – for water activity, that is why, because nothing else can adapt to a water activity lower than that. Staph is the hardiest, the most adaptive as the case may be.
This is a table we have for not only those pathogenic bacteria but also molds and spoilage, and where no microbial growth is. I wanted to point those out. Also, you can see where foods that are generally in that similar range to those microorganisms. We have at 0.85 and up: that’s where all the potentially hazardous foods lie, above that level. If we go down to 0.7, so between 0.85 and 0.7, that’s where you’ll find the yeast and molds will be, and those are our spoilage ones, but there aren’t any spoilage molds below 0.7. You see this funny little section with the osmophilic yeast right there, and there are a few molds but they don’t produce spoilage. Then under below 0.6, you’ll get no microbial growth at all. Nothing will grow below that.
Water activity is a critical parameter for compliance, and it can be used to justify a limited microbial testing which is very important. The FDA has it in their definition of potentially hazardous food. They also have it with FSMA, the Food Safety Modernization Act, both in the HARPC, which is a risk based approach, and in HACCP as a critical control point. We have the 21CFR 110 for good manufacturing practices. USDA also has that as a critical point and good manufacturing practices, and pharma, you can also find that in USP 1121, and also the new one that’s coming, but it won’t be out until next year, 922 as well. The last one, if you’re familiar with ICH, it’s part of the decision tree for assessing hazards. This is the International Conference on Harmonization. Now, these are just a few places where you can find water activities specifically listed as a critical control point, or it mentions how it can be used to justify the limited microbial testing, but this is not an exhaustive list. You can still find it more. This is just a taste.
Now, I’d like to talk about common food pathogens. We’re going to go through them one by one. There’s two differentiations between the pathogens. We have the foodborne intoxication: those are caused by actually ingesting a toxin. The toxin is produced in the food, and then you ingest it, and then you get very sick. There are some examples of what would be foodborne intoxication. Then the second type is foodborne infection. These are caused by ingesting the pathogenic microorganism, and then it gets into your GI tract and then it starts to grow. Intoxication is formed in the food, and infection is formed, essentially, in the gut. That toxin is formed in your gut.
Let’s start with staph aureus. This one is facultative. If you remember, that means that it can grow in both situations. It can grow without air, without oxygen or with oxygen. This is a concern for pharmaceutical companies because they have creams and things, and you’ve got people who are immune compromised. Staph is always a concern for them. It can be destroyed by heat treatment and nearly all sanitizing agents, which is very good. It has the lowest water activity limit, so it’s at 0.85, the lowest of all the pathogens. Sources: we find it on your skin and sores, hair, in your nasal passages, in your nose. On food, you can have that with hand contact with food. Then that food does not require any additional cooking. You have a lot of cross contamination issues with that in salads, filled bakery goods and sandwiches. The interesting thing is if you find staph aureus on food processing equipment, it is generally an indication of poor sanitation. This one can be taken care of quite easily if you’re careful with cleaning and prep and try to minimize that cross contamination issue.
Next, we’re going to talk about botulism. It’s anaerobic. It will not grow in a pH below 4.6. It just needs three minutes of boiling to destroy. Its water activity limit is a little higher at 0.94. Where you’re going to find botulism is nature, soil, water, plants. In foods, it’s improperly canned foods, especially the low acid foods, so beets, green beans, baked potatoes wrapped in foil. In smoked fish you have the herb infused oil where you have the herbs that will contain the botulism on there, and then it’s infused in oil, and now you’ve got an anaerobic condition. Honey can cause children, more specifically infants, to have infant botulism, so that’s why they have the recommendation to not feed your child honey until they’re a year old. Botulism, it’s unusual in that it’s anaerobic. Some of them are not, but that’s where it can be a real difficulty because once you remove the air and you have a higher pH, if you don’t do the retort, then you can have botulism be an issue.
Salmonella – salmonella is the number one for most reported cases. This is now into food infection, where you have to ingest it and then it grows in your gut. It’s more common in the summer months, and that is because it’s warmer weather and we have more active animal life. It is also facultative. We’ve seen that before, where it can go either in an oxygen rich or oxygen depleted environment. It’s also killed by cooking and pasteurization, and the water activity limit for this one is 0.95. The sources are contamination by feces, contaminated drinking water, person to person contact. In foods, we have inadequately cooked poultry and poultry products, eggs and egg products, raw fruits, vegetables, unpasteurized milk and milk products like raw milk cheese, which there’s a recall for that as well, flour which there is also for that as well for salmonella right now, and peanut butter, which we’ve seen in the past. For flour, how that could be contaminated is it actually could be contaminated at the processing facility if it’s not kept clean or there was something else there to infect it with salmonella, or it could be in the field. You could actually be growing the wheat and if, say the fertilizer was contaminated with salmonella or something like that, then it can even be on the grain before it’s even come into process. For peanut butter, most commonly, there’s birds around the processing plant that can contaminate with salmonella. Generally, salmonella is killed during roasting, but if the exposure happens after the roasting or as again introduced after the roasting, then that salmonella is viable. For salmonella, the important thing to know is that it won’t proliferate at a lower water activity, below 0.85. As a matter of fact, we call this – peanut butter and flour are part of it – the low moisture your food group, so they generally have a low water activity, quite a bit lower than 0.85. If they are contaminated with salmonella at that point, they’re harmless. They can’t grow, right? We already talked about that, but the problem is is they’re additives to other things. When you add flour to make a batter or a peanut butter into something, now you’ve introduced it to a high water activity environment, and they will start to grow. That’s where the problems start happening.
This is actually a big concern. How do you inoculate for salmonella on flour without changing the properties of that ingredient, well, flour or peanut butter or something else? There are studies done, Dr. Bradley Marks was actually working on getting a database for the Michigan State University on low moisture foods and salmonella if you were interested in looking at that or contributing to that. There’s also some articles. It’s definitely a big concern of how we can combat salmonella and E. coli as well, salmonella in particular, to mitigate those problems so we don’t find that when people start using them in their own food products that they get contaminated.
Now, Listeria. Listeria is also facultative. It can grow in refrigerated temperatures, which can be a real problem and unusual for most of our microbes that we’re talking about today. It can also be killed by cooking and pasteurization, and it has a water activity limit of 0.92, so a little bit lower, a little more hardy than some of the other ones we’ve seen. Sources for this are soil, water and animals carrying that bacterium. In foods, we see it in uncooked meats like raw hamburger, vegetables, unpasteurized milk and cheese, and cooked or processed foods. Certain soft cheeses, we talked about there was one for Heinz for last year, processed and ready to eat meats and smoked seafood. This is also been a factor for hotdogs, so a similar thing: if you have a low oxygen environment, refrigerate it. Listeria can grow if it’s present in that food. The thing about Listeria which is good to point out, is that at least 90% of people who get Listeria infections are in a high risk group like pregnant women, older adults, people with weakened immune systems. Healthy children and adults occasionally do get infected with Listeria, but they rarely become seriously ill, which is important to point out.
Next, we’re going to talk about E. coli, also facultative. Most strains of E. coli are actually harmless and important in the digestional tract. It actually is used quite a bit in the pharmaceutical industry to make other drugs and things. It’s very effective that way. However, this particular strain that we’re talking about here is the nasty one that does make people sick. It can be cooked and pasteurized to kill it. Some packagers of lettuce can use or have used a chlorine wash that is partially effective so it does help. It’s not a for sure kill to that E .coli, so it’s always important to cook your food well, to wash your raw ingredients well, to refrigerate and or defrost correctly, and watch for cross-contamination. E. coli is relatively dangerous and that you can have a low infectious dose but it’s relatively difficult to kill. We’ll see this in intestines of birds and animals. In food, we have ground beef, raw milk and milk products, raw fruit and vegetables like the greens, like the lettuce. Cross-contamination is a real issue. We’ve seen that a lot in the news. Last year, if you remember, all of the lettuce out of Arizona last April had E. coli and they had to throw all of that away. I’m not sure they even found what was causing that. I was looking recently and I didn’t see if they’d actually identified where it was. Right now, like I mentioned before, Aurora packing company, they have a recall for E. coli as well.
Bacillus cereus is anaerobic. It multiplies very quickly at room temperature, and its incubation time is minutes to hours, but it doesn’t last very long. This is mostly confused with the 24-hour stomach flu. If you’ve come down with a bug that only lasts a day, you probably got this guy, and gave yourself some nice food poisoning. The sources for this are mammals, shellfish, and contaminated water. In foods, it’s raw and undercooked poultry, raw milk and milk products, and for starchy foods like rice, sauces, soups, they’ve been left in the danger zone for more than two hours, and now, this can grow and give people food poisoning.
Campylobacter is the last one we’re going to talk about today. It’s microaerophilic, where it has to have a low oxygen, less than atmospheric. It also can be killed by cooking and pasteurization. It is the number one cause of bacterial diarrhea. This is also known as traveler’s diarrhea. If you ever get this when you’ve been traveling abroad, this is probably what you’ve been infected with. The problem with it is that it can cause bigger issues in the future like IBS, GBS, or arthritis, but the thing is it’s generally isolated to a person or a group, so it’s not extremely widespread. Its water activity limit is extremely high at 0.99, highest out of all of them. You’re gonna find this in the gut of mammals, shellfish, and contaminated water. In foods, we have raw, undercooked poultry and raw milk and milk products.
If we know that water activity, if we can reduce that, will change what microbes we’re going to grow, how do you actually do that? How can you formulate for water activity? First, you can dehydrate the product. That is the ancient way of preserving food is just to dry it out. You lower the water activity. Microbes can’t grow when it stays for a long time. You can have edible films or coatings that would be to prevent or limit the moisture migration.
Moisture migration is caused by differences in water activities. If you can limit that shift in moisture change from one component to the next, then you can limit where that can grow.