Genomic analysis, immunomodulation and deep phenotyping of patients with nodding syndrome

Abstract

The aetiology of nodding syndrome remains unclear, and comprehensive genotyping and phenotyping data from patients remain sparse. Our objectives were to characterize the phenotype of patients with nodding syndrome, investigate potential contributors to disease aetiology, and evaluate response to immunotherapy. This cohort study investigated members of a single-family unit from Lamwo District, Uganda. The participants for this study were selected by the Ugandan Ministry of Health as representative for nodding syndrome and with a conducive family structure for genomic analyses. Of the eight family members who participated in the study at the National Institutes of Health (NIH) Clinical Center, three had nodding syndrome. The three affected patients were extensively evaluated with metagenomic sequencing for infectious pathogens, exome sequencing, spinal fluid immune analyses, neurometabolic and toxicology testing, continuous electroencephalography and neuroimaging. Five unaffected family members underwent a subset of testing for comparison. A distinctive interictal pattern of sleep-activated bursts of generalized and multifocal epileptiform discharges and slowing was observed in two patients. Brain imaging showed two patients had mild generalized cerebral atrophy, and both patients and unaffected family members had excessive metal deposition in the basal ganglia. Trace metal biochemical evaluation was normal. CSF was non-inflammatory and one patient had CSF-restricted oligoclonal bands. Onchocerca volvulus-specific antibodies were present in all patients and skin snips were negative for active onchocerciasis. Metagenomic sequencing of serum and CSF revealed hepatitis B virus in the serum of one patient. Vitamin B6 metabolites were borderline low in all family members and CSF pyridoxine metabolites were normal. Mitochondrial DNA testing was normal. Exome sequencing did not identify potentially causal candidate gene variants. Nodding syndrome is characterized by a distinctive pattern of sleep-activated epileptiform activity. The associated growth stunting may be due to hypothalamic dysfunction. Extensive testing years after disease onset did not clarify a causal aetiology. A trial of immunomodulation (plasmapheresis in two patients and intravenous immunoglobulin in one patient) was given without short-term effect, but longer-term follow-up was not possible to fully assess any benefit of this intervention.

Keywords: autoimmune; epilepsy; genetic; global health; infectious.

Figure 1

Pedigree of family unit examined at the NIH. The symbols with an asterisk indicate those individuals examined at the NIH. These include the patriarch and matriarch of the family, three unaffected daughters, two daughters with nodding syndrome and one grandson with nodding syndrome. Open symbols are unaffected individuals and shaded symbols are those with nodding syndrome. Circles indicate female, squares indicate males and the diamond icons indicate gender not specified. Numbers within symbol indicate the number of individuals represented by that shape and the letter ‘n’ indicates an unknown number, but more than one, of individuals being represented by that shape. Numbers outside the circle indicate age of the patient when seen at the NIH.

Figure 2

EEG recordings including sleep in patients with nodding syndrome. Patients underwent long-term EEG monitoring with a 27-channel digital EEG with time-locked video and single-lead EKG. The EEG electrodes were placed according to the internationally accepted 10–20 system of electrode placement. In addition, subtemporal F9, F10, T9, T10, P9 and P10 electrodes were placed for better localization. The EEG events were reviewed with at least three standard montages. (A) Interictal EEG performed on Patient 2 while awake (left) shows slow background activity with superimposed focal slowing and sharp waves in the temporal regions, at times becoming more diffuse. While asleep (right), generalized and multifocal spikes and polyspike and wave activity are present, frequently embedded within K complexes. (B) Interictal EEG performed on Patient 3 while awake (left) shows diffuse background slowing, at times with runs of semirhythmic or rhythmic theta activity. While asleep (right), findings similar to Patient 2 are observed.

Figure 3

Brain MRI findings in patients with nodding syndrome and an unaffected sibling. (A) Patient 1: (i) cerebral atrophy and mild colpocephaly; (ii) deposition of metal in globus pallidus more than typical for age; (iii) possible pituitary lesion. (B) Patient 2: (i) slight cerebral atrophy; (ii) deposition of metal in globus pallidus more than typical for age; (iii and iv) this signal change represents metal based upon the direction of the phase shift on SWI sequences. (C) Patient 3: (i) normal cerebral volume; (ii) deposition of metal in globus pallidus more than typical for age; (iii) subtle asymmetry of the hippocampi with left being smaller. (D) Unaffected sibling: (i) minimal bilateral colpocephaly; (i) deposition of metal more than typical for age in globus pallidus; (iii) deposition of metal in the substantia nigra and red nucleus; (iv) deposition of metal in the cerebellar dentate nuclei.

Figure 4

Immunomodulation results in decreases in antigen specific antibodies. (A) Serum antibodies to O. volvulus antigen-16 (Ov16) or (B) O. volvulus tropomyosin (OVTmy1) in two patients prior to (black bars) and after (grey bars) plasmapheresis. Total IgG antibodies specific for O. volvulus antigens are shown on the left and IgG4 antibodies are shown on the right. (C) Serum antibodies to OV16 (IgG and IgG4) and OVTmy1 (IgG and IgG4) in a patient prior to (black bars) and after (grey bars) administration of intravenous immunoglobulin (IVIg).