Longitudinal Household Assessment of Respiratory Illness in Children and Parents During the COVID-19 Pandemic

Key Points Question What is the incidence of SARS-CoV-2 infection in children detected when removing testing barriers, and how do symptoms and severity compare with the burden of other respiratory illnesses? Findings In this cohort study of 1209 children and adults from 307 households in the Netherlands, when comparing the incidence (per person-year) of SARS-CoV-2 infection in children (0.22 at age <12 years and 0.21 at age 12-17 years) with that of adults (0.27), no significant difference was found. Unlike the SARS-CoV-2 disease burden in adults, the burden in children with SARS-CoV-2–positive respiratory illness episodes was similar to that of children with SARS-CoV-2–negative episodes. Meaning This study found that SARS-CoV-2 symptoms in children were not more severe than symptoms from other common respiratory illnesses, which could be an important consideration in pediatric COVID-19 vaccine recommendations.

SARS-CoV-2 virus. Follow-up household sampling schemes were stratified based on presence of SARS-CoV-2 in the initial NTS outbreak samples. The full outbreak sampling protocols for SARS-CoV-2 positive and negative outbreaks are summarized in supplement Table 1. This includes: 1) Dried Blood Spot (DBS) by selffinger-prick at the start of the outbreak study and ten days after completion of the outbreak period (convalescent sample); 2) additional NTS and saliva for symptomatic subjects throughout the outbreak period as well as 3) frequent saliva and fecal sampling for SARS-CoV-2 positive outbreaks.
In the extended follow-up, the outbreak study entry criteria were slightly adapted; immediate NTS results were only available for the person(s) who reported respiratory symptoms (index case). If SARS-CoV-2 positive, the outbreak study was initiated.
All samples were temporarily stored in the participant's home freezer, collected by research assistants from home, and subsequently transported to the laboratory on dry ice and stored at -80 degrees Celsius (°C) until further analysis.
Monitoring of respiratory symptoms was temporarily intensified using daily symptom diaries for each household member, supplemented with daily symptom severity scores and disease questionnaires for those episodes meeting the case definition for Acute Respiratory Illness (ARI). The following symptoms were included in the diary; nasal congestion/runny nose, cough, headaches, sore throat, fever, shortness of breath, cold shivers, muscle aches, loss of taste or smell, fatigue, vomiting and diarrhea. The outbreak study lasted at least 21 days or until 21 days after the last ARI onset in a household member.
For study procedures and data collection we used a custom made study app (COVapp), compatible with Apple and Android systems, developed by the University Medical Center Utrecht (UMCU) in collaboration with YourResearch Holding BV. The study app was used to collect four types of self-reported data: (1) baseline questionnaire data, (2) reporting of new onset of respiratory symptoms or fever, (3) registration of all virological and serological specimens collected through participant self-sampling, (4) symptom diaries and disease questionnaires in case of an outbreak. All data entered in the App by the participants were stored in an online secured database and could be accessed and navigated in real-time by study personnel using an online portal with authorized login.

Laboratory analyses
NTS were analysed using the SARS-CoV-2 RT-PCR based on the presence of the E-gene, [1]  DBS were transported on dry ice to the laboratory, where they were stored at -80˚C. Two bloodspots of 3/16" of serum were punched out of the filter paper and were incubated in 200 μl Blotto-blockingbuffer containing 5% Surfact-Amps 20 (ThermoFisher Scientific Inc., Rockford, USA) overnight at 4°C to release serum from DBS for a test dilution of approximately 1 in 40. 90 μl of the eluted serum was used and subsequently tested for the presence of IgG antibodies reactive with the SARS-CoV-2 spike trimer, S1 and SARS-CoV-2 N antigens in a protein microarray, essentially as described previously. 2 Signal exceeding 45,000 fluorescent units were considered positive.
Sequencing of RT-PCR NTS was performed using an in house designed amplicon-based approach as described before. [3][4][5] Several modifications were made to the protocol as primer concentrations were increased from 0.125 to 1 pmol for specific primer pairs. In addition modified primers were added to the original primer set to match current circulating variants. Primer sequences and concentrations are available upon request. AMPure XP beads