The sous-vide cooking method inspired an idea to bring promising technology from the laboratory into the barn. Researchers at Purdue University have successfully developed an on-site bovine respiratory disease test that can provide results within an hour.
The team of researchers has been steadily advancing point-of-care technology to address this disease, which is the most common and costly disease affecting cattle in the world.
Mohit Verma, assistant professor of agriculture and bioengineering who led the research, said: “We want to see if this technology is strong enough for farms and how messy we might become.” “We are not overly cautious about cleanliness. , Because we want the test to be easy to use. Respiratory diseases can spread quickly from animal to animal and can be devastating. Rapid diagnosis will lead to correct treatment and reduce unnecessary antibiotic use."
He said the team also added an easy-to-read color change from red to yellow to indicate test results. A paper detailing this work was published in the journal Veterinary Research.
"We have been working hard to improve our testing so that it is out of the laboratory and into the hands of farmers and veterinarians, and it works very well in the field," Verma said. "One of the keys to this progress is to use a sous-vide water bath to keep the temperature needed for its work at around 149 degrees Fahrenheit. My brother is doing sous vide cooking, and this idea just emerged. It's easy to bring to the farm. , Fill with water and test."
He said that bovine respiratory disease, or BRD, accounts for half of all cattle deaths due to disease in North America, causing US$900 million in losses to the beef industry each year. There are many strains of bacteria and viruses that can cause this disease, so it is difficult to treat it effectively.
"Some bacteria that cause BRD have become resistant to certain antibiotics," Verma said. "Unfortunately, because standard tests can take several days to provide results, farmers need to treat cattle before they know the pathogen. This can lead to the use of ineffective antibiotics or overuse of antibiotics."
The technology created by the Verma team can identify three strains of the first four bacteria that cause BRD: Pasteurella multocida, Mannheimia haemolyticus, and Haemophilus somnifera.
The nasal swab collects the required sample, and then puts the swab into a vial containing the corresponding primers and reagents, which were developed by the team and used as a biosensor for bacteria. The vial and its contents (called a test) are heated in a water bath to perform a chemical reaction. If there are bacteria designed to detect, the detection will change the color.
This technique tests DNA from bacteria and uses a nucleic acid amplification method called loop-mediated isothermal amplification or LAMP. When bacterial DNA is present, LAMP amplifies it. As the level of nucleic acid increases, it will change the pH of the test, causing a color change.
Verma said the advantage of LAMP over other methods is that it does not require sample extraction and processing, which can be lengthy and expensive, and can produce results within an hour. Its results match the results of polymerase chain reaction or PCR, and the test time is 60%-100%.
Ana Pascual-Garrigos participated in the project as an undergraduate student majoring in biochemistry and was the first author of the paper. Other members of the research team and co-authors of the paper include Jennifer Koziol from Purdue University School of Veterinary Medicine; Aaron Ault from Purdue University Elmore School of Home Electronics and Computer Engineering; and Timothy Johnson and Jon from Purdue University Animal Science Department Schoonmaker; Deepti Pillai from the Department of Comparative Pathobiology, Purdue University; and Murali Kannan Maruthamuthu, Josiah Levi Davidson, and Grigorii Rudakov from the Department of Agriculture and Bioengineering, Purdue University.
"It is exciting to work on a product that can greatly affect the cattle industry," said Pascual-Garrigos, who is now studying for a PhD. At Cambridge University. "This research shows that it is possible to have more immediate care solutions worldwide."
Verma and his team advanced the technology to this stage with a USD 1 million grant from USDA-NIFA. In a parallel project, Verma is using the same technology to conduct a saliva-based test for COVID-19. BRD technology is part of his start-up company Krishi Inc. The startup received $100,000 from the Purdue Ag-Celerator Fund earlier this year.
The next step in their pursuit of putting the technology in the hands of veterinarians and farmers is to develop paper test strips. These bands can include multiple tests, so that one band can identify the presence of several different pathogens. They also plan to apply the same method to detect other infectious diseases in cattle and pigs, and investigate the potential for detecting food contamination.
"This platform is very versatile," Verma said. "We just need to change the matrix we use-develop new primers and tests-for different pathogens. We are working hard to apply our technology to solve other health problems, and we believe it has the ability to quickly detect new viruses to help prevent global epidemics. Disease potential."
Verma's team's preliminary research to obtain a USD 1 million USDA-NIFA grant was funded by the initiated Agricultural Science and Economic Development Extension (AgSEED) grant. AgSEED was established through Crossroads funding from the Indiana State Legislature to cultivate the state's leadership in plant and animal agriculture and rural development. Verma also received seed funding for the Disease Diagnosis Inventors Challenge, which was created by the Institute of Inflammation, Immunology, and Infectious Diseases of Purdue University in collaboration with the Department of Comparative Pathobiology, which provided funding for the implementation of the pilot project; Indiana State Institute of Clinical and Translational Science; and Indiana State Analytical Science and Engineering Alliance.
Writer: Elizabeth K. Gardner; 765-441-2024; ekgardner@purdue.edu
Source: Mohit Verma: msverma@purdue.edu
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On-farm colorimetric detection of Pasteurella multocida, Mannheimia haemolyticus and Haemophilus narcolepsy in rough bovine nose samples
Ana Pascual-Garrigos, Murali Kannan Maruthamuthu, Aaron Ault, Josiah Levi Davidson, Grigorii Rudakov, Deepti Pillai, Jennifer Koziol, Jon P. Schoonmaker, Timothy Johnson, and Mohit S. Verma
Link: On-farm colorimetric detection of Pasteurella multocida, Mannheimia haemolyticus and Haemophilus narcolepsy in rough bovine nose samples | Veterinary research | Full text (biomedcentral.com)
This work modified the loop-mediated isothermal amplification (LAMP) test to detect bovine respiratory disease (BRD) bacterial pathogens Pasteurella multocida, Mannheimia hemolyticus, and lethargy in a colorimetric format on the farm Haemophilus. BRD poses a significant health and economic burden worldwide, partly because of the challenges involved in identifying the pathogens that cause the disease. Methods such as polymerase chain reaction (PCR) have the potential to identify pathogenic agents, but they require laboratory equipment and a large amount of sample processing, which makes the process lengthy and expensive. To overcome this limitation, LAMP allows accurate detection of pathogens in unprocessed samples by the naked eye, making it possible to make faster and more accurate diagnoses on the farm. The test developed here provides 66.7-100% analytical sensitivity and 100% analytical specificity (using man-made samples), and when tested on five bulls on the feedlot, the consistency with PCR results reached 60-100% . Using consumer-grade water baths can be implemented on the farm by taking nasal swabs from cattle and providing colorimetric results within 60 minutes. This test has the potential to provide cattle farmers and veterinarians with a quick fencing diagnosis.
Maureen Manier, Head of Department, mmanier@purdue.edu
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