Next-Gen Sequencing for Diagnosing Brain Infections

June 22, 2016
Next-Gen Sequencing for Diagnosing Brain Infections
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Adding to the ever expanding list of molecular diagnostic uses for next-generation sequencing, investigators at John Hopkins University report that they were able to diagnose or rule out suspected brain infections using the advanced genetic sequencing technique on brain tissue samples.

The findings from this study, which were recently published in Neurology: Neuroimmunology & Neuroinflammation through an article entitled “Next-generation sequencing in neuropathologic diagnosis of infections of the nervous system” demonstrates that using computers to rapidly analyze huge amounts of genetic and biological information could be a cost-effective addition to a pathology lab's toolkit. This is a much needed diagnostic advancement since more than 50 percent of inflammatory disorders and infections of the brain go undiagnosed—often leading to treatments for individual symptoms that can do more harm than good.

"By incorporating modern genetic sequencing techniques into pathology diagnostics, we were able to investigate the potential presence of infection in 10 subjects and found appropriate explanations of clinical problems in eight out of 10 patient cases examined in this study," explained senior study author Carlos Pardo-Villamizar, M.D., associate professor of neurology at the Johns Hopkins University School of Medicine. "We hope to develop this technique further as a way to bring the diagnosis rate of inflammatory brain disorders and infections closer to 100 percent so we can treat patients more effectively."

In the current study, the Johns Hopkins researchers identified ten patients at—six males and four females between the ages of 16 and 68—with clinical signs of a brain infection, such as fever and rapid onset of neurological symptoms, like weakness in limbs, partial paralysis numbness, headache, or seizures. Each patient had a biopsy of a brain lesion that first identified by MRI. 

Using commercially available next-generation sequencing technologies that read millions of pieces of DNA at once, the investigators sequenced DNA from the brain tissue for each person and compared the results to a database containing human and nonhuman DNA sequences, 2,817 bacterial genomes, 4,383 viral genomes and 26 single-cell pathogen genomes. After subtracting out the human DNA results, they ranked the top three other species found in each sample that were thought to be potential infectious organisms.

In three of the patients, the research team definitively identified the cause of individual infections as Mycobacterium tuberculosis, JC virus, and Epstein-Barr virus.

"Identifying organisms involved in brain infections is critical for establishing adequate treatments and, in the case of tuberculosis or viral infections, is a major challenge in clinical practice," Dr. Pardo-Villamizar stated.

Additionally, the scientists reported that next-gen sequencing revealed possible but unspecified infections in two other patients. In one patient, the researchers identified several bacteria species in the biopsy related to Delftia acidovorans, a rare, rod-shaped bacillus that physicians treated with antibiotics. In the other patient, they identified a relatively high number of sequences from Lactococcus, a normally noninfectious bacterium.

However, the research team reanalyzed the sequence data a year later and found the sequences matched up to a newly emerging pathogen, Elizabethkingia—a genome that was unavailable during the original analysis. The bacterium, found in soil, has been linked mostly to newborn meningitis or inflammation of the membrane covering the brain.

"One of the limitations of using this kind of diagnostic tool is that we can only identify the pathogens for which we have the genetic sequences available," noted lead study author Steven Salzberg, Ph.D., professor of biomedical engineering at Johns Hopkins. "As we continue to sequence the genomes of more organisms, the tool will become steadily more powerful." The cost for genetic sequencing using the latest technology is less than $500 per sample and takes a couple of days. The comparison of the DNA results to other species takes less than two hours.

Intriguingly, in five of the patients, the sequencing didn't point to a specific pathogen as a cause. In those cases, the negative findings helped clinicians to understand better the diseases those patients were experiencing—allowing them to avoid treating patients with drugs that may do harm.

"If someone has an infection, then you don't necessarily want to give them steroids to reduce inflammation. You'll make the infection worse by knocking down the immune system," Dr. Pardo-Villamizar remarked. "Having a correct diagnosis is the only way to ensure the best, individualized precision treatment."

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