Metabolomics of cerebrospinal fluid from humans treated for rabies

Abstract

Rabies is a rapidly progressive lyssavirus encephalitis that is statistically 100% fatal. There are no clinically effective antiviral drugs for rabies. An immunologically naïve teenager survived rabies in 2004 through improvised supportive care; since then, 5 additional survivors have been associated with use of the so-called Milwaukee Protocol (MP). The MP applies critical care focused on the altered metabolic and physiologic states associated with rabies. The aim of this study was to examine the metabolic profile of cerebrospinal fluid (CSF) from rabies patients during clinical progression of rabies encephalitis in survivors and nonsurvivors and to compare these samples with control CSF samples. Unsupervised clustering algorithms distinguished three stages of rabies disease and identified several metabolites that differentiated rabies survivors from those who subsequently died, in particular, metabolites related to energy metabolism and cell volume control. Moreover, for those patients who survived, the trajectory of their metabolic profile tracked toward the control profile and away from the rabies profile. NMR metabolomics of human rabies CSF provide new insights into the mechanisms of rabies pathogenesis, which may guide future therapy of this disease.

Figure 1

¹H NMR spectra obtained for CSF samples from controls, early stage rabies patients, and late stage (hospital day 19 and 20) rabies patients (survivor vs nonsurvivor).

Figure 2

Identification of three stages of Rabies pathogenesis. Partial least-squares discriminant analysis (PLS-DA) of ¹H NMR CSF metabolomic data comparing three stages of rabies (R²X = 0.30, R²Y = 0.77, Q² = 0.64). Disease stage groups were determined from metabolomic data using k-means cluster analysis. Open circles represent early stage disease, gray filled circles represent mid stage disease, and black filled circles represent late stage disease. Hotelling’s T² 95% confidence ellipse is indicated on the plot.

Figure 3

Tracking metabolic profiles of survivors and nonsurvivors. Partial least-squares discriminant analysis (PLS-DA) of ¹H NMR CSF metabolomic data comparing samples collected from rabies patients and controls (R²X = 0.34, R²Y = 0.87, Q² = 0.81). Open circles represent control samples; black filled circles represent rabies patient samples. Red filled circles track a nonsurviving (NS) patient from hospital day 10 to day 18, as disease progresses this patient diverges further from control samples. The green filled circles track a survivor (S) who changes from a diseased metabolic profile to reflect a normal metabolic profile as the patient recovers from rabies disease. Hotelling’s T² 95% confidence ellipse is indicated on the plot.