The investigators propose a single center, prospective observational study in children with Lyme disease (LD), the Borrelia B-cell diagnostics (BRILLIANT) study, to assess the immune response against Borrelia burgdorferi (Bb) with the following main objectives:

1. Development of Bb-specific ASC ELISpot as a new test method for diagnosis of early LD.

   There is an urgent unmet clinical need for a better diagnostic tool for early LD, as the current standard two-tier testing has low sensitivity in recently infected patients and may show false positive results in recovered patients due to long-term persistence of antibodies against Bb. The measurement of Bb-specific ASC with the ELISpot assay my has the potential to overcome these issues and to improve diagnosis in early LD.

2. Extensive analysis of the immune response in LD. The immune response in LD is not well understood. Large-scale studies assessing the detailed immune cell subsets/phenotypes present in blood, CSF, or synovial fluid of LD patients with respective manifestations are lacking.

3. Isolation and characterization of causative Bb species. Existing literature suggests that Bb genospecies and/or genotypes may determine virulence and manifestations, but large-scale studies assessing Bb genospecies/genotypes in different manifestation of LD are lacking.

4. Collection of clinical data about symptoms, severity, routine laboratory and diagnostic test results, treatment, and outcome of LD.

5. Biobanking samples for analysis in the future.

Project population

Inclusion criteria: Children, 0-17 years of age, at University Children's Hospital Zurich:

• LD differential diagnosis cohort: Patients presenting at the ED with differential diagnosis of LD according to the treating physician.
• Control cohort: Previously healthy patients (HC) with routine blood investigations presenting at the ED or PID outpatient department

Exclusion criteria: Primary or secondary immunodeficiency.

Background:

Lyme disease (LD) is the most common tick born disease in Europe. It is caused by an infection with several genospecies of the spirochaetal bacteria Borrelia burgdorferi (Bb).

Although classical disease manifestations are well-known, the clinical presentation in children is often variable and inconclusive, which results in delayed diagnosis and treatment.

Methods/design:

The investigators are conducting an observational cohort study in children with LD. Study site is the University Children's Hospital Zurich. 502 patients will be enrolled. Children from 0-17 years of age presenting with signs and symptoms suspicious for LD are included in the study. Previously healthy children with routine blood investigation are enrolled as healthy controls. Patients will be excluded in cases of primary or secondary immunodeficiency.

Clinical and routine laboratory data regarding course and outcome, as well as venous blood samples are collected at first hospital contact and follow up visits (FUP). FUPs are scheduled at 28 days, 3 months and 6 months after hospital admission. Cerebrospinal fluid (CSF) and synovial fluid (SF) will be collected for the study only if sampling is indicated due to diagnostic or therapeutic reasons.

Primary objectives are to assess Bb-specific ASCs in blood using ELISpot assay, in order to develop new diagnostic tool for early LD. In addition, the investigators will examine immune response in patients with various LD manifestations using flow cytometry, ELISA assay, and ELISpot assay. Finally, the investigators will perform whole genome sequencing of causative Bb-species isolated from patients to investigate potential differences in virulence and associations with clinical presentations.

Discussion:

This single-centre, observational cohort study will improve the understanding of immunological response in LD in children. It will also provide new information about the virulence of distinct LD causing Bb-genospecies and will test a new approach in the diagnosis of early LD.

Inclusion Criteria:

• Patients presenting at the ED with differential diagnosis of LD according to the treating physician

Exclusion Criteria:

• Patients will be excluded in cases of primary or secondary immunodeficiency

Children from 0-17 years of age presenting with signs and symptoms suspicious for LD are included in the study. Previously healthy children with routine blood investigation are enrolled as healthy controls.

Data will be handled with uttermost discretion and is only accessible to authorized personnel who require the data to fulfil their duties within the scope of the research. On the CRFs and other project specific documents, participants are only identified by a unique participant number. Biological material will be stored in coded manner with unique participant numbers in restricted areas at the Children's Research Center and the IMM. Collaborators will not be granted access to the key or any personal patient information.

Project data are stored for at least 15 years. Biological data will be collected for 5 years with a start in 2023 until 2028 and destroyed after 10 years of study closure.

Access is authorized to personnel who require the data to fulfil their duties within the scope of the research.