Encephalitis

The clinical laboratories (Innsbruck, Mayo Clinic, Oxford, and Sydney; centers 1–4) sent the following groups of coded serum samples and clinical information to the Institute for Quality Assurance (IfQ; Lübeck, Germany):

Phase I: 89 coded samples sent to centers 1–4 and center 5 (Euroimmun) for testing (figure 1)

MOG-IgG clearly positive: 39 blinded samples from all laboratories with a previously determined clearly positive MOG-Ab serostatus (high titers or fluorescence-activated cell sorting [FACS] binding ratios, supplementary methods, table e-2, links.lww.com/NXI/A189), all of them diagnosed with inflammatory demyelinating diseases known to be associated with MOG-IgG (such as ADEM, aquaporin-4 [AQP4] antibody–negative neuromyelitis optica spectrum disorder (NMOSD), optic neuritis, myelitis, and other demyelinating diseases).

MOG-IgG clearly negative (negative or very low titers or FACS binding ratios, supplementary methods, table e-2, links.lww.com/NXI/A189): 40 blinded samples from all laboratories with a previously determined clearly negative MOG-Ab serostatus. Eighteen of the 40 samples were from people who also presented with clinically overlapping features such as optic neuritis, myelitis, ADEM, or encephalitis. The other samples were from controls (7 from people with MS, 5 from people with other neurologic diseases, and 10 from healthy controls).

Ten technical controls (humanized monoclonal MOG-Ab 8-18-C5,^{30 (link)} 5 samples IgG1, and 5 samples IgM (kappa) in different dilutions, but of unknown IgG or IgM concentration, contributed by center 5.

Phase II: 100 coded samples sent to 5 centers for testing (18 repeat and 82 new, figure 1)

Nine positive and 9 negative samples from phase I were sent out a second time to assess interassay variations.

Thirty healthy blood donors were contributed by the IfQ. No clinical information was available, and samples were not pretested for antibodies against MOG or other autoantigens.

MOG-IgG low/borderline positive: 39 blinded samples from all laboratories with a previously determined low positive MOG-IgG serostatus (just above the individual cutoff values, supplementary methods, table e-2, links.lww.com/NXI/A189). Thirty-six of these samples were from people with inflammatory demyelinating diseases associated with MOG-IgG and 3 were from patients with MS.

MOG-IgG borderline negative: 13 blinded samples from all laboratories with a previously determined borderline negative MOG-IgG serostatus (just below the individual cutoff values, supplementary methods, table e-2, links.lww.com/NXI/A189). Five of these samples were from patients with inflammatory demyelinating diseases associated with MOG-IgG and 8 were from controls (3 from people with MS and 5 from people with other neurologic diseases).

All patients in our study consented to the use of their medical records for research purposes. The study was approved by the Institutional Review Board of Mayo Clinic, Rochester, MN (No. 08-007846).
Serum samples from 394 patients and controls were tested: 91 patients were classified as having a MOG-IgG–like clinical phenotype and included ADEM (28), AQP4-IgG seronegative NMO (27, fulfilling Wingerchuk diagnostic criteria for NMO, either 1999 or 2006 [excluding antibody status]), optic neuritis (21 single, 2 relapsing), or longitudinally extensive transverse myelitis (10 single, 3 recurrent). The control samples were collected from patients with MS (244, selected from the Olmsted County MS population-based cohort), hypergammaglobulinemia (42), and other (17, encephalitis, glioma, Creutzfeldt-Jakob disease, glaucoma). Sensitivity was calculated as the percentage of positives cases within the MOG-IgG–like clinical phenotype cohort. Specificity was calculated as the percentage of positive cases in the MS cohort and those with other neurologic presentations inconsistent with an MOG-related clinical phenotype. Positive predictive value (PPV) was calculated as the percentage of positive test results in patients with MOG-IgG–like clinical phenotypes of all positive test results and estimates the reliability of a positive test result. In contrast, the negative predictive value is the percentage of negative test results in patients without an MOG-IgG–like clinical phenotype of all negative test results and is an estimate of how reliably a negative test result rules out the disease. This study was approved by the Mayo Clinic Institutional Review Board.
All samples were stored at −80°C at the Mayo Clinic central laboratory. They were divided into aliquots and provided frozen as coded samples to the 3 neuroimmunology laboratories: Mayo Clinic; Oxford, UK; and Euroimmun, Germany. All samples were tested by investigators blinded to the clinical information. Methodologies of the 3 assays are shown in table 1, and staining of cells considered positive and negative by all 3 assays is illustrated in the figure

This retrospective cross-sectional study was conducted in Germany among hospital-based physicians. The study was granted an exemption determination from a central Institutional Review Board (IRB) in the United States prior to starting data collection. No personal identifiable information was captured during the course of the study. Prior to participating, physicians provided their informed consent to proceed with the study.
The study was conducted in two main phases (Fig. 1). A qualitative phase was initially conducted between July 13, 2020 and August 13, 2020 in which 12 physicians were interviewed to assess how TBE is diagnosed and managed in real-world practice, as well as to examine the feasibility of questions to be included in the quantitative phase. The quantitative phase, which was conducted between October 14, 2020 and May 7, 2021, consisted of two parts, a screening and a chart review survey.Fig. 1

Study schematic

To be eligible to participate in either phase of the study, physicians must have reported (1) being, as their primary specialty, an emergency room (ER) specialist, an intensive care unit (ICU) physician (i.e., medical, neurological, or pediatric ICU), an infectious disease specialist, a neurologist, or a pediatrician, (2) being in clinical practice for ≥ 3 years, and (3) spending ≥ 60% of their time in clinical practice. Qualitative phase participants must have also reported (1) working in a hospital-based setting ≥ 70% of the time and (2) managing ≥ 2 patients with meningitis, encephalitis, or non-specific CNS symptoms per year and prescribing or ordering testing for some patients. Quantitative phase participants must have also reported (1) working in a hospital-based setting ≥ 50% of the time and (2) managing ≥ 5 patients with meningitis, encephalitis, or non-specific CNS symptoms per year and prescribing or ordering testing for these patients.
In the quantitative phase, physicians (N = 500) were first screened in order to identify up to 200 who met the eligibility criteria for the chart review survey, with an approximately even split of ER specialists, ICU physicians, infectious disease specialists, neurologists, and pediatricians/neuro-pediatricians, and to gauge the caseload of patients with meningitis, encephalitis, or myelitis symptoms among the physician specialties of interest.
Prior to chart review survey data collection, the survey instrument was piloted with a convenience sample of 5 physicians who met the eligibility criteria described above to verify that the questions were appropriate and sufficiently clear to respondents and that the required data points were easy to collect (to reduce the amount of potential missing data). For the chart review, physicians completed a 50-min cross-sectional web-based survey, including a minimum of 2–3 retrospective case report forms (CRFs) for patients who had presented with meningitis and those who presented with encephalitis. The chart review survey collected profile information about the physician (e.g., gender, age, specialty, practice setting, etc.) and about their clinical practice (e.g., number of patients with meningitis, encephalitis, and myelitis seen in the past year with etiology, diagnostic testing performed, etc.).

For the qualitative interviews, results were summarized with means (continuous data) or counts and percentages (categorical data), as well as with illustrative verbatim quotes. For the chart review survey, sample characteristics variables were reported as counts and percentages. The count and percentage of patients who received a TBE test and who did not receive a TBE test were also reported. The TBE positive rate was computed as the number of patients who had a positive TBE test divided by the number of patients who received a TBE test and then multiplied by 100 to convert to a percentage value. TBE testing rate and TBE positive rate were computed among the subsets of patients who experienced different types of symptoms for the aggregate patient sample, tick bite (tick bite, no tick bite, and don’t know), seasonality (admitted during tick season1and not admitted during tick season), headache (headache and no headache), fever groups (no fever, fever > 38 °C to 39 °C, and high fever > 39 °C), clinical manifestations and flu-like symptoms (yes or no). Chi-square tests were used to examine whether the distribution of TBE positive rate varied across the five manifestations of interest (i.e., meningitis only, encephalitis only, myelitis only, a combination of meningitis, encephalitis, and/or myelitis, and non-specific neurological symptoms) for the aggregate sample, for patients who presented with headache, and for those who presented without headache. p-values < 0.05, two-tailed, were considered statistically significant.