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Wednesday, July 16, 2014

How Next-Generation Sequencing Rapidly Diagnosed a CNS Infection

By Dan Hurley

A rapidly progressing case of meningoencephalitis in a 14-year-old boy was diagnosed as neuroleptospirosis in 48 hours with a new, relatively inexpensive genetic sequencing system that, neurologists say, promises to transform the way such cases are routinely handled.

Although only a handful of academic centers presently have access to both the rapid sequencing technology and the bioinformatics capabilities necessary to decipher its output, a wider rollout is expected over the next few years.

In the meantime, the neurologist who served as first author of the new report told Neurology Today that clinicians faced with similarly serious, unknown CNS infections should contact his team.

“We can accept samples from external folks,” said Michael R. Wilson, MD, an assistant professor of neurology at the University of California, San Francisco (UCSF), where he works in the genomics laboratory of study coauthor Joseph DeRisi, PhD. “We’re very interested in seeing more cases like this.” (Interested neurologists may e-mail him, he said, at

            As reported first in the June 4 online edition of New England Journal of Medicine, the patient, under the care of an immunologist for severe combined immunodeficiency, presented three times to a Wisconsin medical facility over the course of four months “with fever and headache that progressed to hydrocephalus and status epilepticus necessitating a medically induced coma.”

The cause of the boy’s illness — as is evident in more than half the cases of meningoencephalitis — remained undiagnosed despite extensive clinical testing, including MRI, and biopsy of the right frontal lobe. In addition to his routine monthly IV immune globulin therapy, intravenous glucocorticoids were administered for possible neurosarcoidosis, but his clinical status only worsened.

UCSF researchers were asked to help diagnose the case by a pediatrician at the University of Wisconsin School of Medicine, James Gern, MD, who was collaborating with them on an unrelated study.

Serum and CSF were sent overnight, and then fed into an Illumina MiSeq instrument. One of the latest iterations of so-called “next generation” sequencing devices, it generated 8,187,737 sequences.

Then, in just 96 minutes, those sequences were analyzed with a bioinformatics pipeline developed at UCSF, comparing them with reference sequences in the National Center for Biotechnology Information databases of bacteria, viruses, fungi, and parasites.

            In all, less than 48 hours after the samples had reached UCSF, 475 sequences corresponding to the leptospira genome were identified in the patient’s CSF but not in the serum, with no other convincing hits to other bacteria or viruses.

Without waiting for the identification to be confirmed by targeted PCR — performed soon after by two independent laboratories — the Wisconsin team decided to begin treatment for neuroleptospirosis. Glucocorticoids were tapered, and antibiotic coverage was narrowed to high-dose IV penicillin. The patient recovered over the next seven days, and was discharged to inpatient rehabilitation another 14 days later. He returned home after 11 days of rehabilitation close to his premorbid functional status.

Check back tomorrow afternoon for the full discussion of these findings and their implications from outside experts in the July 17 issue of Neurology Today. Read our previous stories on next-generation sequencing here: