Lab Report PDF

Preview

Summary

Lab Report Number 1...

Description

Paige Menard Bio 282 Rachel Larsen

Paige Menard 9/20/19 Rachel Larsen Plate Count Experiment Lab Report Microbes Growing In Adverse Environments

Paige Menard Bio 282 Rachel Larsen Microbes Growing In Adverse Environments Abstract: Salt is known for its antimicrobial aspects, such as protecting fruit from microbes. Sugar is also said to do the same, but for this experiments purpose we only saw the attraction of microbes. Salt can be used to help protect our fresh fruits and other products from harmful pathogens, that could be detrimental to our bodies. The goal of this experiment was to see if salt or sugar would have a more abundant amount of microbes present. We hypothesized that the salt would have fewer microbes present than the sugar, but the results showed that sugar actually had lower counts. Instead of making an inhabitable environment for microbes with the salt, we made an environment where they could thrive. The second experiment was able to show that with the varying amount of salt, it does minimize the chances for microbes to make a home. Salt is able to preserve fruits and other produce from microbes, but must have the right amount to make an unlivable environment.

Introduction: Bacteria can live in varying environments, and it is important to note that they can live on all of the different foods we eat. Salmonella is the most common bacterial pathogen that have been isolated from a wide variety of fresh produce (FDA, 2019). Before you eat any fruits or vegetables that you buy from anywhere, you should always rinse them with water. They are susceptible of containing harmful germs, such as Salmonella, E.coli, and Listeria, which can make not only you, but your family as well sick. (CDC, 2019). Vegetables and fruits are an essential part of our diet because of the nutrients, but as we discussed previously they can sometimes be the host to a large microbe population. Foodborne pathogens can contaminate vegetables and fruits at any stage of their production process with a potential for human infection such as, growth, harvest, and preparation. Through the use of salt on foods, it not only prevents spoilage of such foods, but also it is important in not allowing the growth of food borne pathogens such as Salmonella or Clostridium when applied with the right amount (Espocitio, 1996). The simplest way that salt inhibits is through osmosis, or dehydrations. This results in the reduction of water activity, which is free water molecules in

Paige Menard Bio 282 Rachel Larsen food that is overall necessary for the survival and for the growth of microbes. Microorganisms that do not have access to enough water will not grow as fast as the ones with plenty of access to water (Pelagia Research Library, 2015). The counterpart of salt can also be used in preserving foods, which is sugar. Sugar has been used for generations in food preparation, preservation of fruits such as apples, pears, and jams. Foods that have a .95 water activity will have enough water activity to support microbial life such as bacteria. Since sugar makes a juice, it attracts more bacteria since there is more water activity if you use too much sugar (FDA, 2014). With sugar it is added to food, and it binds with the water in the fruits and vegetables, and this gives the microorganisms less places to grow. During osmosis small amounts of fruit aid is removed with water as well. This is because sugar makes the cell membranes semi-permeable, allowing water to be released faster than sugar. This is apparent when you make Strawberry shortcake, the water is extracted from strawberries to make a juice. Large amounts of sugar are used in preserving food, that can also be used to inhibit microbial activity (Chavan, 2012). In this study, we observed what microbes did when sugar and salt were added. We predicted with salt on one strawberry there will be less microbes/bacteria on the strawberry. We also predicted that the sugar strawberry would attract more microbes. With the first experiment we saw a higher number of microbes on the salt strawberry, then the sugar strawberry, and then as we suspected the control had less than both of the strawberries. This then turned into another experiment that we were interested in. The next experiment was to have one control strawberry, one strawberry with 1 teaspoon of salt, and one strawberry with half a teaspoon of salt. We were more interested in seeing the change in microbes with more salt on them, and see how that would inhibit the microbial growth. Ally, one of the lab partners, claimed that the half a teaspoon had way more salt on it then the first round. So we are sure that this will have better results. With these changes, we predicted that the strawberry with 1 tablespoon will have the least amount of bacteria present. Salt is able to preserve foods better than sugar, because it cannot do the same when it is prepared the same way.

Paige Menard Bio 282 Rachel Larsen

Methods: For both of the experiments three strawberries were purchased. For one of the experiments just salt was used, and for the other experiment sugar and salt were used. For the first batch of strawberries salt was put on one, sugar was put on one, and one was a control. For the second experiment varying amounts of salt was used, one strawberry had ½ a teaspoon of salt, the other strawberry had 1 tablespoon of salt, and the other strawberry was the control. The second experiment was specified amounts, while the first experiment was randomized of how much salt and sugar was put on. The strawberries were completed around 5:30 PM and sat in plastic sandwich bags overnight, with some air. For both experiments the strawberries were purchased at Hannafords, one set of strawberries was your average plastic container strawberry, but for the second experiment there was Canadian strawberries that come in quartz. The procedures followed for the rest of the testing is found in the lab manual on pages 89 and 90 (Larsen, 2019).

Results: The goal of this study was to see what kind of conditions that microbes can survive in, those environments being salt and sugar environments. Our initial hypothesis was that we would see less microbes on the salt strawberry and we would see more microbes on the sugar strawberry, because it draws out moisture from strawberries. Therefore it makes a better living environment for the strawberries, so we expected to see more microbes on the sugar strawberry than on the salt strawberry. To our surprise what we saw on our plates was clearly different than what we had originally hypothesized. In figure one you can see that the salt strawberry has a much higher CFU/g’s, than that of the sugar strawberry. The average for the salt strawberry was 31,000 CFU/g’s, while the sugar strawberries average was 10,766.667 CFU/g’s. You can also see this information in table 1. The different plate counts for the salt strawberry were as follows, plate 1 had 32 colonies, plate 2 had 3 colonies, and plate 3 did not make sense to include. 32 colonies for plate 1 made sense, but for plate 2 it showed 3 colonies which was on the low spectrum, so it is

Paige Menard Bio 282 Rachel Larsen not as reliable. The counts from the plates followed the trend of 10-fold dilutions. Figure one you can see the sugar standard deviation from the error bars is much more than the salt strawberry. We believed the amount of sugar and salt that we applied to each played an important role on why our hypothesis was wrong for the first experiment. We simply just used a dipping and roll method for the salt and sugar application the first time. Since we did not specify the amount of sugar and salt for each strawberry, we did not put enough of either to really see huge results that we were hoping to see. Because of this experiment, we planned a second experiment to further investigate if salt would lyse the cells. For this experiment we bought more strawberries, they were different than the first experiments, and were Canadian strawberries. The second experiment had one control strawberry, one strawberry with a ½ teaspoon of salt, and one strawberry with 1 teaspoon of salt. We hypothesized that the strawberry with the most salt, 1 teaspoon will have the least amount of bacteria due to its antimicrobial properties. With the less salt comes more ability for the bacteria to access the strawberry. As you can see in figure 2 our hypothesis was correct. Figure 2 shows that the control strawberries CFU/g was undoubtedly more than both the 1 teaspoon and ½ teaspoon. The control strawberry had a CFU/g of 30,000,000, while the strawberry with ½ a teaspoon of salt had 29,433 CFU/g, while the strawberry with a teaspoon of salt on it had an average of 12,250 CFU/g. The one teaspoon went from 135 to 11 to 0 to 0, which is not a perfect 10-fold dilution, but is close. The ½ teaspoon went from 203, to 18, to 5, and to 1, again, these are not the perfect 10-fold dilutions, but it still shows a downward trend of the microbes on each plate. You can see in table 1 and table 2, the difference in the CFU/g’s for the control strawberries, and how drastic the difference is. The amount of salt we used for the second experiment was able to show that the amount of salt you use does matter, and that the more salt you use the less likely it is for microbes to be able to grow.

Paige Menard Bio 282 Rachel Larsen

Figure 1 35000 30000

CFU/G

25000 20000 15000 10000 5000 0

Salt Strawberry

Control

Sugar

Food Counts

Figure 1: Salt and Strawberry Experiment 1. This figure shows the microbes growing on strawberries in different conditions, those being a regular condition, one in salt, and one in sugar.

Paige Menard Bio 282 Rachel Larsen Strawberries CFU/g averages Experiment 1 Salt 31,000 Control 7,333 Sugar 10,766.667 Figure 2: Salt Strawberry Experiment 2. This figure shows the microbes growing in different environments, one again growing in a regular environment, one in ½ teaspoon of salt, and one in 1 teaspoon of salt.

Table 1: Average CFU/g’s for experiment one. This table shows the amount of microbes for each of the given environments.

Strawberries CFU/g averages Experiment 2 Salt ½ Tsp 29,433 Control 30,000,000 Salt 1 Tsp 12,250 Table 2: Average CFU/g’s for experiment two. This table shows the amount of microbes for each of the given environments for the second experiment.

Discussion: Through this study I was able to learn about preserving fresh fruit and vegetables, and also learn that with the right amount of salt or sugar, it can control the amount of microbes that can be seen. Salt is able to lyse the microbes when the right amount is applied, like we have seen through the graphs, and the charts. But sugar on the other hand wasn’t able to do the same, maybe if it was in a different environment it would have been more controlled. Since sugar makes the water activity within the food come out, it usually makes for a great environment for microbes to take over. But if we were to do a third experiment, it would be cool to see how sugar reacts with a fruit or vegetable in a scenario where it thrives, and is able to have a sealed space with no air, and it where it could preserve the fruit or vegetable with a minimization of microbes. In our findings for the first experiment it was much different than what we hypothesized. Through the graphs you can see that salt microbes were much more present, than that of the sugar strawberry. The CFU/g on the salt strawberry was 31,000, while the sugar strawberries average CFU/g was 10,766.667. We believe that because the colonies were more apparent for the

Paige Menard Bio 282 Rachel Larsen salt strawberry that we did not apply enough salt, and it was not enough salt to lyse the bacteria. Instead of killing the bacteria with the salt, we created a condition where they love to live, in moisture. Although we did not see our hypothesize quite work, we were able to see sugar attract microbes, we saw this through the CFU/G being higher than that of the control strawberry, which you can see in table 2. Since we were not able to see a perfect 10-fold dilution, it could have also been to human error. One thing that we could have done differently for our experiments was use the same package of strawberries, so we could have consistent results for the control strawberry. I believe since we used a different package strawberries that made our control strawberry have a significant difference to that of the first experiment. We purchased both sets of strawberries at Hannaford’s supermarket, but one of the packages was your average strawberry package, but the second experiment strawberries were quart style strawberries. These strawberries were right as you walked into the store, and were susceptible to air and were not sealed. This could be the jump in the controls from 31,000,000 in the second experiment, to 7,333 CFU/g’s for the first experiment. Another area where we could account for some of our error bars is human error. Since we were not able to count some of our plates it could be not as accurate, and we also could have made mistakes counting the strawberry colonies since so many did have an abundance of microbes present. Another area where we could have seen error is because for the second experiment we could only count one of the control strawberry plates, and that was the 3rd plate. We really couldn’t get an accurate average, because we couldn’t count any of the plates except for that one. Although the main goal of our study was to see the difference between the amount of microbes in a salt and a sugar environment. We did learn a little about how each of them attract or kill bacteria. But what we really learned is the importance that salt brings to the table, and it’s preservation properties. Salmonella is a pathogen that is accountable for 70 to 80% of the pathogen outbreaks in China. Salmonella is commonly related to meat, but lately it is associated with the fresh fruits and vegetables that most of us eat on a daily basis (Wang, 2018). Salmonella affects both meat and fresh produce, making this a situation that salt could help with, with its antimicrobial functions. Salt has been known for hypertension, obesity, and overall not the

Paige Menard Bio 282 Rachel Larsen greatest for your health. But if we want to change the amount of sodium on our food, and in it, it will not change the number of organisms on food, but it may affect their growth, and if they stay living or not (Stringer, 2005). These pathogens such as Salmonella can continue to grow within our bodies, but with sodium in our stomachs, they tend to lyse the cells, ending up with their deaths (Stringer, 2015). Sodium acts to help prevent spoilage, and it is able to cause some microbe cells to go into a “osmotic shock,” and this helps the water leave the cell (Institute of Medicine, 2010). Through this experiment we have been able to learn astounding information about salt, and it’s antimicrobial properties, but we also have been able to learn through these articles that salt can help with the preservation of all foods, not just strawberries. If we were to do another experiment, I would like to do an experiment where we see how sugar is able to preserve food. Since in our previous experiments we learned how it could attract more microbes, it would be cool to do an experiment where sugar is used to preserve food. Since we mostly focused on salt, and in result we had to leave air in the bags. But if we were to do a sealed environment, where we know that sugar is able to preserve, since it helps preserve jams, and other sealed foods.

References:

Paige Menard Bio 282 Rachel Larsen CDC. (2019, June 6). Fruit and Vegetable Safety | Food Safety | CDC. Retrieved from https://www.cdc.gov/foodsafety/communication/steps-healthy-fruits-veggies.html Chavan, U. D., & Amarowicz, R. (2012). Osmotic Dehydration Process for Preservation of Fruits and Vegetables. Journal of Food Research, 1(2), 202–209. doi: 10.5539/jfr.v1n2p202 FDA. (2019, April 12). Bacteria and Viruses. Retrieved from https://www.foodsafety.gov/foodpoisoning/bacteria-and-viruses FDA. (2014, August 27). Water Activity (aw) in Foods. Retrieved from https://www.fda.gov/inspections-compliance-enforcement-and-criminalinvestigations/inspection-technical-guides/water-activity-aw-foods He, C., Esposito, C., Phillips, C., Zalups, R. K., Henderson, D. A., Striker, G. E., & Striker, L. J. (1996). Dissociation of glomerular hypertrophy, cell proliferation, and glomerulosclerosis in mouse strains heterozygous for a mutation (Os) which induces a 50% reduction in nephron number. The Journal of clinical investigation, 97(5), 1242–1249. doi:10.1172/JCI118539 Institute of Medicine (US) Committee on Strategies to Reduce Sodium Intake; Henney JE, Taylor CL, Boon CS, editors. Strategies to Reduce Sodium Intake in the United States. Washington (DC): National Academies Press (US); 2010. 4, Preservation and Physical Property Roles of Sodium in Foods. Available from: https://www.ncbi.nlm.nih.gov/books/NBK50952/ Larsen, RA ed. 2019. BIO 282 Microbiology Lab Manual. University Readers, San Diego, CA. Pelagia Research Library. Food Spoilage: Microorganisms and their prevention. (2015). Retrieved from https://pdfs.semanticscholar.org/3214/30ca38867d803d31b086cd672a98db38d2d4.pdf Stringer, S. C., & Pin, C. (2005). Microbial risks associated with salt reduction in certain foods and alternative options for preservation.Microbial risks associated with salt reduction in certain foods and alternative options for preservation.Institute of Food Research. Retrieved from https://acmsf.food.gov.uk/sites/default/files/mnt/drupal_data/sources/files/multimedia/pdfs/acm7 40a.pdf Wang, W., Zhou, Y., Xiao, X., Yang, G., Wang, Q., Wei, W., … Yang, H. (2018). Behavior of Salmonella Typhimurium on Fresh Strawberries Under Different Storage Temperatures and Wash Treatments. Frontiers in microbiology, 9, 2091. doi:10.3389/fmicb.2018.02091...