High-Throughput Urinary Testing: More Data, More Problems?

If a urinary tract infection (UTI) has ever ruined your day, you’re not alone. This uncomfortable condition is one of the most common infectious illnesses in the United States, responsible for more than ten million office visits and nearly $4 billion in costs each year. UTIs are usually diagnosed based on the results of three different types of test:

1.     Dipstick test: a prepared test strip provides basic information about the urine, including things like pH and the presence of blood cells or nitrites (a byproduct of some bacterial infections). The dipstick is fast and inexpensive but not very sensitive, leading to both false negative and false positive results.

2.     Microscopy: examining urine under a microscope to check for the presence of red and white blood cells or bacteria directly. This is a relatively quick and inexpensive test but is also prone to false negatives and false positives.

3.     Urine culture: currently the gold standard of UTI diagnosis. For this test, a sample of urine is plated on materials that will allow bacteria to thrive and then it is observed for 48 hours to see if any bacterial colonies will grow. Culturing urine samples allows the lab to identify the species of bacteria responsible for an infection and test antibiotics to determine the best treatment. Growing cells in culture can be difficult, however, particularly if bacteria live together cooperatively in a growth called a biofilm which is common in the urinary tract. This makes false negative results a problem for urine culture.

Staphylococcus aureus bacteria bound by a biofilm. These growths make bacteria more difficult to detect with urine culture. Photo credit Rodney M. Donlan, Ph.D and Janice Carr, 2005.

Recently, new technology has been introduced into UTI diagnosis that may eventually change the way in which infections are identified. Polymerase chain reaction (PCR) allows the lab to test urine samples for several bacterial genes to identify infections without the need to grow bacteria directly in culture. PCR is more sensitive than urinary culture and may be able to provide evidence of bacteria that are more difficult to identify by culture, like biofilm infections. More sensitive still is next-generation sequencing (NGS).In this technique, rather than testing for a few specific bacterial genes that may be present in the urine sample, the lab can identify every single bit of genetic material in the urine. This allows the lab to identify every organism that is living in the urinary tract, and test for the presence of genes that confer antibiotic resistance to suggest a more targeted antibiotic therapy. These methods are poised to make UTI diagnosis and treatment more precise and less prone to false negatives.

How next generation sequencing works. Photo credit to Jenny Cham, based on a lecture by Alan Pittman, Ph.D.

However, these more sensitive tests do have limitations. By identifying many more species of bacteria than urine culture, NGS may detect species that are present in urine but not responsible for urinary symptoms. In a comparison of NGS and urine culture, although NGS and culture identified the same bacteria in UTI patients 96% of the time, NGS also identified many species of bacteria in healthy control participants. Why were these healthy patients testing positive for bacteria? Although historically the urinary tract has been described as a sterile environment, modern evidence suggests that the urinary tract has its own microbiome, where multiple species of microbes live without causing human illness. This research is still in its infancy, and until the microbiome of the bladder has been thoroughly described it will be difficult for doctors to determine whether a species identified via NGS is an infectious organism or a normal part of the patient’s urinary tract. Another disadvantage is cost: these new tests are also much more expensive and not often covered by insurance, and not all doctors are willing to use the results to make treatment decisions. 

So, are these new tests worth it? That’s a question only the patient and their doctor can answer. For now, these more sophisticated techniques are best used as a companion to traditional urinary culture, not a replacement, until clinical standards are established. It might be helpful to think of NGS as similar to “23andMe for your bladder”- the information is interesting, and some of it might be useful for your medical provider, but for now it’s mostly medical trivia. Here’s a summary of the test options:

TEST SPEED COST DRUG
TESTING
FALSE
NEGATIVES
FALSE
POSITIVES
Dipstick Instant $
Microscopy Instant $
Culture 2 days $$ Uncommon
PCR 1 day $$$ Uncommon Uncommon
NGS 4 days $$$$ Uncommon

Peer-edited by Kathryn Weatherford and Chiung-Wei Huang.



Fever; good for more than just a day home from school

https://pixabay.com/en/cold-ill-fever-thermometer-1972619/

“Mom, I think I have a fever,” was the sure fire way to stay home from school as a young child.  One such instance, my mom put her hand to my forehead and told me to go get ready for school. I walked away wondering how she could possibly know I didn’t have a fever without using a thermometer. I decided it could only be one thing: special mom powers.

Parents and doctors have used fevers to monitor infection for decades; no special mom powers after all. Before the discovery of antibiotics, doctors used high fevers to treat various diseases and medical conditions. Even today, increasing body temperature is a used to treat cancer, in combination with other treatment options. Despite fevers being used to treat infection and cancer for decades, it was only recently that doctors and scientists begun to understand how fever treatment works. A recent study by Lin and colleagues suggests fever may help immune cells, called T cells, find their way to the infected part of the body.

T cells help fight off many different types of infections caused by bacteria, viruses, and fungi, collectively referred to as microbes. T cells are part of the adaptive immune system, meaning each T cell is trained to fight one specific invader. Therefore, during infection, the correct T cell (i.e. the one trained to fight that particular infection) needs to receive the message that there is an invader and then travel to the location of invader in the body.

Adhesin molecules on blood vessels act as a Velcro hook, while integrins on T cells act as the Velcro loop.

Adapted from a photo by Hadley Paul Garland

When you get an infection, your body quickly recognizes there is a foreign invader; a microbe that has lost its way and entered enemy territory (your body). Once an invader is identified, your cells begin making molecules to activate the body’s innate immune system to help fight the foreign invader. Some of these molecules can act like messages that travel all over the body to alert other cells of danger or travel to the brain to cause fever. Another subset of these molecules act as bread crumbs to lead immune cells to the infection battlefield or place where the invader was identified. The first to arrive on the battlefield are innate immune cells. Innate immune cells, in combination with other parts of the innate immune response such as fever, and mucus, use their broad arsenal to try to quickly disarm the foreign invader. However, the meanest of microbes can endure the hottest of fevers, escape the stickiest of mucus, and hinder the most brutal attacks by innate immune cells. In the event the innate immune system cannot fight off the invader, messengers are sent to find the right T cell for the fight. Once the T cell trained to fight the infection is called upon, the T cell travels through the bloodstream, the highway for immune cells during infection, following chemokine breadcrumbs to the infection battlefield.

While traveling on the bloodstream highway, T cells put proteins called integrins on their surface – think of them as the loop part of Velcro (Image 2). Cells that make up the wall of the blood vessel near the battlefield put proteins called adhesion molecules on their surface – these act as the hook part of Velcro. When the T cell is traveling past the battlefield, adhesion molecules on the blood vessel wall will bind the integrins and catch the T cells, like a Velcro tether. This ensures the T cells do not miss the battlefield location (Image 3).


T cells traveling through the bloodstream are caught near the site of the infection by binding of T cell integrins to blood vessel adhesin molecules. Clustering of integrins strengthens the bond between the T cell and the blood vessel to ensure the T cell does not pass the infection location on its journey.

Interestingly, fevers can help with this tethering process to get T cells to the right place at the right time. Fevers increase the number of adhesion molecules lining the blood vessel wall at the location of the infection. This increases the chance T cells will be “caught” as they travel through the bloodstream. The study by Lin and colleagues shows fevers increase the activity of integrins, making them extra sticky. T cells sense the increased body temperature during fever and begin making more heat shock proteins, which are proteins that help protect cells from fevers. It turns out heat shock proteins also bring integrins together as clusters and clusters of integrins are stickier than a single integrin. Going back to our Velcro analogy, imagine the hook side of the Velcro trying to grab onto one small piece of the loop. This bond is not going to be very strong. However, if the Velcro hooks attach to multiple small loop pieces, this bond will be much stronger. Further, clustering of integrins on the cell surface acts as an blinking exit sign on the bloodstream highway, signaling to the T cell to leave the bloodstream, by going through the blood vessel wall, to get to the infection battlefield. Once on the other side of the blood vessel, the T cell can begin to fight the microbial intruder.

Fevers are often thought of as a simple consequence of illness without much thought to their actual purpose. However, the studies of Lin and others show inducing fever is an important and intentional process carried out by the body to improve the immune system’s ability to fight infection. We will never know what a human immune system without fever looks like, but one thing is for certain, without warm foreheads there would be a lot more children at home watching cartoons.

Peer edited by Laetitia Meyrueix and Samantha Stadmiller.

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