Researching urinary extracellular vesicles

15 October 2024

Veterinary student Nicole Cardoza reports on the BSAVA PetSavers research project entitled Urinary extracellular vesicles in the pathogenesis of canine urinary tract infection and asymptomatic bacteriuria.

As a third-year veterinary student at the University of Cambridge on the scout for summer research project opportunities, I came across the BSAVA PetSavers research project led by Dr Tim Williams at the Queen’s Veterinary School Hospital. Such an opportunity just couldn’t pass me by, especially working with a project leader as amazing as Tim!

What are urinary extracellular vesicles and why are they important?

Urinary extracellular vesicles were a foreign concept at first, but through reading literature published by Dr Williams, I soon familiarized myself with this exciting field of research. Acting as a snapshot in time, urinary extracellular vesicles are shed by cells lining the urinary tract, containing a representative sample of proteins and biological molecules that were present at the time of its release from its origin cell. Tim and I were interested in researching whether or not these urinary extracellular vesicles possessed any antimicrobial properties as this could have implications for future research into the pathophysiology of ascending urinary tract infections.

Our research began by acquiring free-catch urine samples from canine patients at the Queen’s Veterinary School Hospital. A challenge we initially faced was collecting samples that were representative of normal, healthy canine urine. The selection criteria for our urine samples included a normal pH and urine specific gravity, low protein content and an absence of haemoglobin and bacteria; to examine whether our samples met these criteria we completed routine urine analysis and sediment exams.

However, as one may expect, many of our unwell canine patients at the hospital didn’t provide the normal healthy urine samples we desired so unfortunately many samples had to be discarded at this early stage of our research.

What did our findings show and how are they clinically relevant?

In the lab, we isolated our urinary extracellular vesicles (uEVs) by differential ultracentrifugation and used nanoparticle analysis to identify and quantify them. We incubated them with luminescent E. coli bacteria (BL21) to observe whether the presence of uEVs impeded the growth of bacterial colonies (Figure 1). Using a paired t-test, the area under these bacterial growth curves was compared to that of a control group in which E. coli BL21 were incubated in PBS without the addition of urinary extracellular vesicles.

Figure 1

To our excitement, we identified a spectrum of antimicrobial potency in our uEV samples. Fourteen of our 23 canine urinary extracellular vesicle preparations did indeed demonstrate significant antimicrobial activity, often at physiologically relevant concentrations.

Our findings are especially promising for future research into the field of canine urinary extracellular vesicles and their role in the prevention of ascending urinary tract infections. Further research could investigate whether there is any relationship between the antimicrobial activity of canine urinary extracellular vesicles and the incidence of ascending urinary tract infections. Our data from urinary extracellular vesicles isolated from normal healthy urine could be used as a baseline to compare the antimicrobial activity of uEVs isolated from patients who have a history of recurrent urinary tract infections or subclinical bacteriuria. To confirm whether urinary extracellular vesicles are the effectors of antimicrobial activity in canine urine, live imaging studies and further manipulation of urinary extracellular vesicles could reveal the mechanism of action behind the antimicrobial properties of uEVs and support the hypothesis of uEVs playing a defensive function against urinary tract infections.

What have I learned from this research project?

This research has been a tremendous opportunity for me to pilot my scientific career in veterinary clinical research and has introduced me to the wonderful clinical pathology team at the Queen’s Veterinary School Hospital at Cambridge as well as the welcoming staff at the Karet laboratory in the Cambridge Institute for Medical Research. The opportunity to present my research (for the first time no less!) at BSAVA Congress in 2023 was a tremendous introduction to the world of conferences and scientific research. It was a challenging feat but I’m extremely grateful to have had the opportunity to work with Dr Tim Williams on this fascinating project. Student presentations from BSAVA Congress 2023 can be viewed here.

A great deal of gratitude and thanks goes to BSAVA PetSavers for making this research possible through their funding. Not only did their contributions create this scientific opportunity for me, but because of their funding I was able to receive financial support over the summer while I completed my research. For any future students interested in completing a research project I would strongly recommend that they leap at any opportunity to work with BSAVA PetSavers!

About the authors

Nicole Cardoza is a penultimate year Veterinary Medicine student at Girton College, University of Cambridge. She completed this project in the summer of her third year. Nicole’s interests lie with small animals, particularly taking a keen interest in pathology and diagnostic imaging. She’s thrilled to have been involved in this project and can’t wait to kickstart her endeavours in veterinary research

Tim Williams qualified from the University of Cambridge, and following a year working in small animal practice, he undertook a PhD in the Feline Research Group at the Royal Veterinary College, London. He subsequently joined the Department of Veterinary Medicine, University of Cambridge, first as Senior Clinical Training Scholar (resident) and subsequently Associate Professor in Clinical Pathology. He is a Diplomate of the European College of Veterinary Clinical Pathology and Fellow of the Royal College of Pathologists.

Supporting more BSAVA PetSavers research

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