Antibody-secreting Cells, Childhood Pneumonia, Diagnosis, Enzyme-linked Immunospot (ELISpot), Mycoplasma Pneumonia
Conditions
Brief summary
To investigate the Mycoplasma pneumoniae-specific circulating antibody-secreting cell (ASC) response and Mycoplasma pneumoniae-specific interferon (INF)-γ-secreting T cell response, along with polymerase chain reaction (PCR) and serology, in a cohort of children with community-acquired pneumonia (CAP) and controls.
Interventions
DIAGNOSTIC_TESTEnzyme-linked immunospot (ELISpot) assay [Blood]
The ASC ELISpot will be developed based on the improved methods recently described \[Nat Protoc 2013;8:1073-87\]. This protocol allows rapid (6-8 h) detection of specific ASCs in small volumes (1-2 ml) of blood. M. pneumoniae protein P1 (50 μl/ml) will be used as antigen. The optimal concentration of coating antigen will be assessed in advance in two-fold serial dilutions for clear spot definition. The M. pneumoniae-specific T cell ELISpot will be developed based on methods recently described \[Nat Protoc 2009;4:461-9\].
Sponsors
University Children's Hospital, Zurich
Study design
Observational model
COHORT
Time perspective
PROSPECTIVE
Eligibility
Sex/Gender
ALL
Age
3 Years to No maximum
Healthy volunteers
Yes
Inclusion criteria
CAP cohort: * Children of age 3 to 18 years; * In- and outpatients; * Clinically diagnosed community-acquired pneumonia (CAP); Healthy control cohort: \- Healthy asymptomatic children of age 3 to 18 years undergoing an elective surgical procedure; Family control cohort: \- Family members of index CAP patients.
Exclusion criteria
* Hospital-acquired pneumonia; * Immunodeficiencies; * Chronic lung disorders.
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Change in numbers of M. pneumoniae-specific ASCs and M. pneumoniae-specific INF-γ-secreting T cells in blood from inclusion (day 0) to 1-month follow-up (day 28) | At day 0 (inclusion, disease presentation) and at day 28 (follow-up, disease resolution) | Enzyme-linked immunospot (ELISpot) assay and flow cytometry |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Change in M. pneumoniae DNA levels in respiratory samples from inclusion (day 0) to 1-month follow-up (day 28) | At day 0 (inclusion, disease presentation) and at day 28 (follow-up, disease resolution) | PCR |
| Change in total and M. pneumoniae-specific antibody levels (immunoglobulin (Ig)G, IgM, IgA) from inclusion (day 0) to 1-month follow-up (day 28) | At day 0 (inclusion, disease presentation) and at day 28 (follow-up, disease resolution) | Enzyme-linked immunosorbent assay (ELISA) |
| Outcome of community-acquired pneumonia (CAP) assessed by clinical assessment of body temperature (°C) and respiratory rate (per minute) at 1-month follow-up (day 28) | At day 28 (follow-up) | Clinical assessment of body temperature (°C) and respiratory rate (per minute), with worse outcome defined as body temperature more than 38.5°C and respiratory rate according to age more than 40/min for 3 years, more than 34/min for 4-5 years, more than 30/min for 6-12 years, and more than 16/min for 13-18 years. |
Outcome results
None listed