Paulo Bispo, PhD, launched his clinical research training in his home city of Sao Paulo, Brazil at the ripe age of 17. Now, he’s collaborating with Sao Paulo universities to improve the diagnosis and untangle hidden causes of infectious uveitis and endophthalmitis, serious eye disorders that cause inflammation in the back portions of the eye and are leading causes of blindness worldwide.
Bispo’s pilot research, which is funded through our Five Senses: Input and Response opportunity, illustrates an interesting case study in the interplay between “bench” and “bedside” in clinical and translational medicine. In this case, a challenging clinical problem — the complex diagnosis of intraocular inflammatory diseases, where around half of all cases are of unknown origin — is driving the basic-science discovery of novel or elusive pathogens in order to diagnose and treat these infections more precisely. A key part of the project is to develop an affordable portable genomic device that can be used to support diagnosis and treatment in places lacking laboratory infrastructure.
You are investigating new methods for diagnosing serious intraocular infections, including uveitis and endophthalmitis. What’s wrong with our current methods?
The challenge with the diagnosis of intraocular infections is that there are so many causes. The methods we use right now are very time-consuming and not sensitive enough.
“The challenge with the diagnosis of intraocular infections is that there are so many causes. The methods we use right now are very time-consuming and not sensitive enough.”
In addition to a wide range of infectious pathogens, in some cases, autoimmune dysfunction and trauma can also cause intraocular inflammation. The physician needs to investigate a long list of possible causes based on the clinical presentation to begin testing for infectious pathogens. Some types of cancer can mimic the signs and symptoms of uveitis for example. If the cause is suspected to be infectious, this might point to another long list of pathogens from all four types of microbes–viruses, bacteria, parasites, and fungi–as the cause.
Testing for certain pathogens requires intraocular fluid, which we are able to extract only in minute quantities, often just a drop or two. With 40 or more possible pathogens, how are you going to test them all?
What if we could instead look for all the microbes at the same time using a method that is sensitive, fast, and requires just one drop to potentially detect all the microbes that could be associated with infection? That’s our aim with this pilot study.
You’re using an advanced genome-sequencing technique to blanket-screen for pathogens in that drop of intraocular fluid. Why this method?
Approximately half of uveitis and endophthalmitis cases are treated without knowing the cause in part because we don’t know all the associated pathogens, or because the sensitivity of the cultures we have to detect known causative agents is very poor. In our patient population, around 50 to 60% of cases are negative after extensive laboratory testing. What is causing the infection in those cases?
Just think about that: Half of these patients are going to be treated using a trial-and-error approach for the whole course of their disease based solely on their physicians’ educated guesses. Sometimes physicians are very good at guessing, and sometimes not. The problem isn’t the physician or the physician’s training; the problem is the multifactorial nature of the infection.
In the method we are using, which we call unbiased sequencing, we extract the DNA from the intraocular fluid and sequence bits of DNA in the sample — the pathogen as well as the human DNA. We start from a place where we say: Let’s see what is there.
Using unbiased methods, we can start to develop all-in-one tests for detection of all possible etiologies using only one assay, and also potentially start to find new causes of infection. Not only will that help improve diagnosis in the short term, but in the future we can develop better diagnostic methods and treatments based on new understandings of the pathogenesis.
You are collaborating with researchers in Brazil. How will that contribute to this work?
I earned my PhD from the Federal University of Sao Paulo, and I will now be collaborating with researchers from both the Federal and State Universities of Sao Paulo on this work. This will help increase the number of patients we can reach, as the teaching hospitals from these Universities serve more than 22 million people who live in the Sao Paulo metro area.
Even more importantly, this collaboration helps to increase the diversity of DNA samples we’re processing. In Brazil, the causes of infection are different than here. Toxoplasma is more common there, for example, as is tuberculosis and some tropical diseases that are uncommon here, including Zika, chikungunya, and dengue. This diversity will help us validate the test to be as universal as is possible.
“Knowing that I’m doing work that is highly translational, work that can improve the way we treat and diagnose patients and improve their quality of life – these are strong personal motivations to wake up every day and return to the lab.”
One of the things I really want to do in the future is to bring these technologies to places where people don’t have access to complex laboratory infrastructure. The technology we’re developing is very small, and it’s cheaper to use than other next-generation sequencing methods. We can run this test with minimal infrastructure. I think the portability of this technology is an attractive advantage that could be particularly useful in areas that are considered medical deserts, especially in the Global South.
The goal is to have an affordable genomic-based test that could be performed, for example, in rural areas that don’t have a strong laboratory infrastructure, or any infrastructure.
How does the pilot funding support this research?
The pilot has been fundamental to help me establish the minimal infrastructure needed in the lab, to purchase the devices and the reagents that we need, and to start to develop and optimize the protocol. It enables us to commence proof-of-principle studies as the basis for larger clinical validations.
If we find new etiologies that are interesting, we could potentially also have projects investigating the pathogenesis of infection. Our future work will hinge on what we discover through this pilot research.
What inspires you on your research path?
I started my training in biomedical sciences and translational research at a young age because I’ve always loved medicine, and specifically wanted to do clinical and translational work. I’ve always loved being in the lab, working on research that will potentially benefit patients. Almost two decades later, I’m still doing this. It’s where I’m most happy.
I’m working a lot right now. Since I’m junior faculty, I’ve been writing many grants over the past few years, as well as writing papers and generating data. For me, that’s very energizing.