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In Queso Emergency

RESEARCH SPOTLIGHT: Using Lactose Oxidase as a natural antimicrobial to inhibit growth of Listeria monocytogenes in Queso Fresco

This research spotlight highlights the work of master’s student Brenna Flynn. Brenna is investigating whether a naturally derived enzyme, lactose oxidase, can inhibit the growth of Listeria monocytogenes in queso fresco. Listeriosis is a high-risk infection caused by the consumption of L. monocytogenes which predominantly affects those with compromised immune systems, pregnant women, and the elderly. It also has a high lethality rate which makes it a main cause for concern in food safety.

Listeria monocytogenes colonies on MOX plates

L. monocytogenes is a problem for the dairy industry because it commonly contaminates products after pasteurization. Many dairy products are ready-to-eat (RTE), meaning that there’s no further kill step after pasteurization to reduce Listeria counts before the consumer eats the product! In addition, queso fresco has a near-neutral pH with a high water content, making it an ideal environment for pathogens to thrive if the cheese becomes contaminated. In ARG, we are trying to find an effective control method to reduce or eliminate L. monocytogenes in queso fresco to mitigate future L. monocytogenes outbreaks and keep consumers safe!

Preparation of lab-scale queso fresco

With increased consumer demand for clean-label ingredients, enzymatic antimicrobials can be an option for producers. Lactose oxidase is a naturally derived enzyme that oxidizes the sugar in milk, lactose, into lactobionic acid, while reducing oxygen into hydrogen peroxide. This reaction is an activator of a native antimicrobial system in milk. Hydrogen peroxide itself is also a widely used antimicrobial that is generally recognized as safe at low concentrations in food. In previous studies, hydrogen peroxide has shown to be an effective inhibitor of L. monocytogenes in queso fresco.

Brenna has been making lab-scale queso fresco to test if lactose oxidase has antimicrobial properties. She has run multiple experiments, using lactose oxidase as both an additive to the milk during the cheesemaking process and as a topical application. She then inoculates the cheese with L. monocytogenes to determine if lactose oxidase has an antimicrobial effect.

So far, her experiments have shown that lactose oxidase is effective at inhibiting the growth of L. monocytogenes! Lactose oxidase could potentially be used as an antimicrobial in the dairy industry to keep consumers everywhere safe!

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No Need to Cry Over Spoiled Milk!

RESEARCH SPOTLIGHT: IDing of food spoilage organisms for dairy manufacturers

This research spotlight highlights the work that Lab Technician Tim DeMarsh is doing to help prevent spoilage microbes from causing issues in the dairy industry. Contamination with spoilage microbes can result in food waste and loss of revenue for dairy producers. Remediating spoilage issues can be difficult, especially when producers don’t know which microbe is responsible. It’s much easier to create solutions when manufacturers can specifically target the organism that is causing the problem. Producers can send their contaminated food samples to the Alcaine Research Group where Tim identifies the offending species. Using this information, the manufacturer can develop strategies to mitigate this specific species and prevent future instances of spoilage.

To identify the organism, it must first be isolated from the food sample. The microbe is grown up on a petri dish for isolation and then its cells are split open, exposing its DNA for extraction. Tim then uses a technique called Polymerase Chain Reaction (a.k.a. PCR; the same process used for COVID-19 tests!) to make billions of copies of the organism’s DNA. The PCR product is loaded into an agarose gel, a scientific version of your favorite Jell-O dessert. Then, the gel is exposed to an electrical field. Since DNA is a negatively charged molecule, it migrates through the gel to the positively charged side, creating bands of DNA.

To make these bands really stand out, the gel is stained with ethidium bromide, a chemical that binds to DNA and fluoresces under UV light. When the gel is exposed to UV light, colorful bands of DNA aggregates are visible, verifying that Tim’s PCR process was successful. He now has billions of copies of the pesky spoilage microbe’s DNA! The DNA is purified and sent to another lab at Cornell that sequences the DNA for him, creating a genetic fingerprint for the organism.

When Tim receives the DNA sequence data, it must be cleaned up a little, and then he compares the sequence to an NCBI database that contains over 1.3 billion unique DNA sequences. The database identifies close matches to the DNA sequence and provides names of organisms that are similar. If the fidelity of the match is 99% or higher, he can be relatively confident that his organism matches the organism in the database, and he has found the name of the species causing the spoilage issues!

The name of the microbe is then given to the dairy processor, and they can create a targeted approach to remediating the issue. This service provided by the Alcaine Research Group helps manufacturers ensure that they are producing high-quality, spoilage free products that consumers everywhere will enjoy!

  • Agarose Gel
  • The group cleaning up the DNA sequence
  • Sequence in the database
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Got Milk?

RESEARCH SPOTLIGHT: Fermenting surplus skim milk into a value-added product

This week, we are highlighting the work of master’s student Lucas Wise. Lucas is focusing on repurposing surplus skim milk into a value-added product. High prevalence of lactose intolerance in the world and increased demand for plant-based milk alternatives has caused a decline in skim milk consumption. For producers, this is resulting in financial loss and environmental damage from increasing skim milk waste. The goal of his project is to develop technologies to derive added value compounds, ingredients, and products from skim milk via the fermentation of lactose.

While traditional brewer’s yeast, Saccharomyces cerevisiae, can’t ferment lactose, alternative yeasts can externally break down lactose into its two smaller subunits, glucose and galactose. Non-Saccharomyces yeasts can also selectively ferment the glucose, leaving galactose behind to be recovered!

Lactose cleavage into its two monomers, glucose and galactose

Galactose is a low glycemic index sugar, meaning that it doesn’t cause a huge spike in blood sugar levels after its ingestion. Galactose can be used to make healthier snacks and beverages, serve as a pre-cursor to other “rare” sweeteners like tagatose, and can be used in pharmaceuticals and health-promoting galactooligosaccharrides, which are prebiotics.

The anaerobic fermentation of lactose also produces ethanol, the ingredient in all of our favorite drinks!

The fermentation process can leverage the lactose in discarded skim milk and provide an alternative method of upcycling this dairy waste, easing the burden of waste on dairy producers. Companies could avoid major financial losses and reduce their environmental impact because the skim milk surplus gets repurposed.

Currently, Lucas is in the screening stage, using a variety of yeast species to determine the best process performance parameters and improve the fermentation stability and sensory properties of the product. This will be followed by an optimization seeking to maximize ethanol and galactose yields from the fermentation. This work could potentially be used to create a stand-alone fermented beverage!

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Holey Cow!

RESEARCH SPOTLIGHT: Late-blowing defect in cheese caused by Clostridium tyrobutyricum

This week, we will be highlighting a project that PhD Student Margarita Valdiviezo is working on with assistance from one of our lab techs, Dana. They are working with an interesting organism that has long acted as a pest to cheese producers- Clostridium tyrobutyricum. C. tyrobutyricum is a spore forming bacteria; this means that it is able to survive pasteurization temperatures that kill off most other unwanted microbes. Once conditions are favorable, and vegetative cells begin to grow in the cheese from the spores. Butyric acid and gas are produced as a byproduct of their metabolism. This causes a defect knowing as “late-blowing”. As you can see in the photo, this causes cracks and bubbles to form in the cheese that are not desirable.

Samples were received from a cheese producer to isolate and confirm the identity of C. tyrobutyricum, as well as to develop a sporulation protocol that Margarita will be able to apply to her research. Margarita plans to develop biotechnological strategies that will reduce spore forming bacteria in organic dairy products.

Margarita and Dana are still in the process of attempting sporulation. The process has been as follows: a sample of the cheese was digested in a stomacher and plated. Then, an overnight tube was made with an isolated colony from the plate. The following day, 1mL of that tube was transferred to a biphasic media bottle and allowed to incubate. All of this took place in an anaerobic chamber, as C. tyrobutyricum is a very oxygen sensitive organism and can only grow in anaerobic environments. The next step in this process is to confirm the presence of spores using a phase-contrast microscope.

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What’s Shakin’?

RESEARCH SPOTLIGHT: Fermentation of Dairy Waste Stream into Value-Added Product

For our first research spotlight, we are highlighting some of the novel dairy fermentation work going on in ARG. Marie Lawton and Kate Jencarelli have been aerobically fermenting a synthetic media supplemented with lactose to emulate dairy effluents. Acid whey, whey permeate, and milk permeate are by-products of dairy processing. They’re generally low in solids but have high levels of the milk sugar lactose remaining.

These by-products are typically waste and must be treated and disposed or they are used as animal feed. But what if we were able to convert this dairy waste into something great?

How can we upcycle dairy waste and create a value-added product?

Using the tools of microbiology, we can convert the milk sugars into something of value and help dairy processors turn their waste into a functional product.

With the rise in consumer trends for functional beverages, products like kombucha with live cultures have increased in popularity. Why couldn’t we make a kombucha like product out of these dairy effluents?

Acetic acid is the compound that gives kombucha its signature flavor, and dairy effluents are also packed with vitamins and minerals. Fermentation of these dairy effluents could break into a new niche in the market, and help dairy producers everywhere recycle their waste into a new profitable product!

Anaerobic fermentation of these dairy effluents is also being investigated in our lab. This process produces ethanol and could be another means of upcycling dairy waste. Research is ongoing to determine how to optimize acetic acid and ethanol production from the dairy waste stream.

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Fermenting Ideas for Dairy Culture

We are a lab in the Department of Food Science at Cornell University in Ithaca, New York. We have a passion for the the world of dairy science and fermentations. Whether it’s chasing phage tails, teaching yogurt, or fermenting something wild…we are all about trying something new.

We hope you enjoy following along with the research and adventures we undertake. Please reach out if you have any questions, ideas, or just want to say hello!

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