Research questions
‘Impact COVID-19 on the EMR’ examined the prevalence of SARS-CoV-2 antibodies and its determinants in the EMR. How does the prevalence of SARS-CoV-2 antibodies evolve over time? What are the sociodemographic, biological and behavioural determinants?
Design
‘Impact of COVID-19 on the EMR’ is a longitudinal study of prevalence of SARS-CoV-2 antibodies. To assess the presence of SARS-CoV-2 antibodies, blood samples are collected by a self-finger prick. To investigate the determinants of SARS-CoV-2 antibodies, sociodemographic and behavioural data such as compliance with measures, vaccination status/attitude, acquired infection(s), cross-country mobility, social contacts and overall wellbeing is collected through an online questionnaire. Data collection took place twice, from April to July 2021 and September to November 2021 in each sub region in the EMR.
Setting
The Meuse-Rhine Euroregion covers the border area between Belgium, the Netherlands and Germany. The sub regions are the Belgian provinces Limburg and Liège (including the German-speaking Community), South Zuid Limburg (NL), and the German City Region Aachen and Districts of Düren and Heinsberg. In 2021 the EMR counted around four million inhabitants (7).
Project consortium
The project consortium consists of the Public Health Service South Limburg (NL), Sciensano (Belgian institute of public health, BE), Maastricht University Medical Centre (NL), and foundation euPrevent (NL).
SAMPLING FOR BASELINE PARTICIPATION
Study population
The study focused on the registered general adult population living in private households. In Germany it was however not possible to identify the latter criterion completely. The sampling frame consisted of all persons registered in the latest version of the National Register (NR) in Belgium, the Basic Registration of Persons (BRP) in The Netherlands and the Registration Offices (Einwohnermeldeämter) in Germany.
Sample size
The sample size was set at 3,500 in each country, that in turn was distributed among the sub regions according to the size of the adult population. The included adult population was classified according to age group and gender. The study population was divided into eight age groups: 18-29, 30-39, 40-49, 50-59, 60-69, 70-79, 80-89 and ≥ 90. After ordering, a stepwise selection was applied, where the step size in each sub region was based on the total eligible population of that sub region divided by its sample size, starting from an ad-random number. During stepwise sampling, for each initially selected individual, 2 substitutes were selected that match in age group and gender. In Germany, the inhabitants of District Heinsberg and City Region Aachen were only classified by gender and an ad random sample was taken.
Opt-out
In South Limburg (NL), an opt-out procedure was required by the Dutch Ethical Committee. This procedure entailed that an announcement letter about the study was sent to each individual from the sample, informing them on the study and giving the option to disclose refusal of participation. Those who did not wish to participate, had 12 days to opt-out online: via the link in the information letter, by mailing the enclosed return card, or by calling the call center set up for this project. For the Belgian and German sub regions, this procedure was not required and for cost-effective motives therefor not applied.
Substitution
Participation in this study implied performing a self-fingerpick test and completing an online questionnaire. Given the uncertainty of the response rate, the project consortium opted for substitution. Substitution of non-participants can help achieve the predetermined net sample, both in size as in composition (8). For each initial selected case, two reserve cases with an identical socio-demographic characteristics are selected. The selection probability of the substitute cases is conditional. This contradicts the idea of equal selection chances. Nevertheless, the probability of being an initial selected case is equal of being selected as a substitute case.
FOLLOW-UP
Between three to five months after receiving an invitation to participate in the study, all invitees that participated – filling in the questionnaire and/or sending a blood sample and/or signing the informed consent - were invited for a follow-up round of data collection.
MEASUREMENTS AT BASELINE AND AT FOLLOW-UP
SARS-CoV-2 antibody testing
The Euroimmun Anti-SARS-CoV-2 QuantiVac ELISA (IgG) test and Euroimmun Anti-SARS-CoV-2 NCP ELISA (IgG) test (EUROIMMUN Medizinische Labordiagnostika AG, Lübeck, Germany) were used to determine antibody responses to SARS-CoV-2 using the Virion/Serion Immunomat (Virion/Serion, Würzburg, Germany) (8,9).
The Euroimmun Anti-SARS-CoV-2 QuantiVac ELISA (IgG) detects IgG antibodies binding the S1 antigen of SARS-CoV-2 in a quantitative manner using a 6-point calibration curve. The S1 antigen contains the immunologically crucial receptor binding domain (RBD), which is a key target antigen for virus neutralisation. The total amount of IgG antibodies is expressed as relative units per milliliter (RU/ml).
The Euroimmun Anti-SARS-CoV-2 NCP ELISA (IgG) is a semi quantitative test which detects IgG antibodies directed against the nucleocapsid protein. Whereas S directed assays detect both post natural infection and vaccine-induced antibodies, nucleoprotein assays only detect antibodies post natural infection.
Both tests were performed using the manufactures instructions. The resulting extinction is calculated to a ratio (extinction of sample/extinction of calibrator) in case of the Euroimmun Anti-SARS-CoV-2 NCP ELISA (IgG) or is plotted against a 6-point calibration curve in case of the Euroimmun Anti-SARS-CoV-2 QuantiVac ELISA (IgG) in order to determine the relative units per milliliter (RU/ml).
For the Euroimmun Anti-SARS-CoV-2 QuantiVac ELISA (IgG) results were interpreted as followed:
Value (RU/ml)
|
Interpretation
|
<8 RU/ml
|
negative
|
≥ 8 to < 11 RU/ml
|
borderline
|
≥ 11 RU/ml
|
positive
|
In this study, borderline results were considered positive.
Self-finger prick
Serological data were collected twice with the same cohort to assess the prevalence of SARS-CoV-2 (infection and vaccine induced) antibodies and the changes over time in the EMR. Using a collection kit distributed by mail, participants collected circa 20 drops of capillary blood by self-finger pricking with a safety lancet into a BD Microtainer® K2EDTA collection tube. The addition of ethylenediaminetetraacetic acid in the test tube prevents clotting of blood, thereby enabling the analysis of the sample even days after collection.
At home self-finger pricking makes participation in the study easily accessible and allows the collection of large numbers of samples for the two defined time periods. The self-finger prick method with following laboratory testing showed to be feasible in a small pilot. The testing procedure and the retrieved amount of blood was validated with 7 finger prick samples from hospital colleagues who were known positive for SARS-CoV-2. Test procedure and numbers of antibody titers were similar as the number of antibody titers of 7 samples obtained by regular venous blood collection.
Home-sampling increases participation and is a cost-effective alternative for clinic-testing (10). Furthermore, the self-finger prick method offers the advantage of minimizing unnecessary movements of individuals to central blood collection locations, thus not overburden health care staff for whom work load is already high due to the continuing epidemic (11). Nevertheless, self-finger pricking might be difficult for motoric impaired participants or participants who find it difficult to collect a blood sample from themselves.
Questionnaires
In addition to the serological data of two timeframes within the same cohort, accompanying questionnaires will provide insight into the determinants of SARS-CoV-2 antibodies as well as the impact of the corona pandemic and measures taken in the EMR on mobility, behavior and wellbeing. Both questionnaires were available in Dutch, German and French. Participants could choose in which language the questionnaire was completed. Seven topics were included in both questionnaires: infection prevention measures (compliance, support, opinion on usefulness and level of difficulty to adhere), mobility (changes in mobility, cross border visits), social network, COVID-19 infection(s) and symptoms, vaccination, general health and socio-demographics. In the second questionnaire one topic on traveling during the summer was added (see appendix Table A1 for complete content of the questionnaires).
Consent
An informed consent was to be filled in and signed by participants of this study. Participants in all participating countries were asked to consent: to participate in the study, for keeping personal data for a longer period of time to use for future research in the field of infectious diseases. In the Netherlands and Germany participants were asked consent to be contacted for future research. In Belgium participants were asked to be re-contacted for the follow-up round of the study.
In the Netherlands, the study was submitted to the medical ethics review committee of Maastricht University Hospital and Maastricht University (METC azM/UM) and was assessed as not requiring a WMO. For Germany, no ethical assessment of the project was needed. The project had to be approved by the heads of Gesundheitsamt Heinsberg, Düren and Aachen, and the in-house lawyers of Kreisverwaltung Heinsberg, Düren and Städteregion Aachen. In Belgium, the study was approved by the Medical Ethics Committee of the University Hospital Ghent and the University of Ghent (BC-09754).
TECHNICAL AND LOGISTIC REQUIREMENTS DEVELOPED TO CONDUCT THE STUDY
Online tool
An online tool was developed by an external software developer to fulfill multiple needs in this cross-border study. The online tool functioned as an intersection where all components of the cross-border study were connected.
For compliance with GDPR regulations ID-numbers were generated and could be downloaded via the tool to be linked to the invitees in each country. From the 10 digit identification number, the following could be identified: sub region, gender, age group, ranking; initial sample, first or second substitute.
Employees who composed the test kits used the online tool to scan and connect the ID-numbers to a blood tube containing a random 4 digit number.
Invitees received the announcement letter or invitation letter with their personal ID-number and the link to the online tool where multiple actions could be taken: filling in the questionnaire, consulting the blood results, retrieving information on the study, and interpretation of test results. Dutch invitees could fill in their ID-number to opt-out for the study and the researchers processed the return cards in the online tool to register the opt-outs.
To process the informed consents, researchers and/or employees scanned the ID-number in the online tool, together with the items of consent given by the participant.
The result of the SARS-CoV-2 antibody tests were determined by the lab and noted in an Excel file together with the blood tube code. This file was periodically delivered to the software developer to be imported and consulted by the researchers and participants in the online tool.
Via a helpdesk page, employees could assist participants with no computer (skills) or difficulties with filling in the questionnaire. At the end of filling in the questionnaire, participants choose a pin code to consult the antibody test result three weeks after sending the blood sample. If participants forgot this code, employees could reset the pin code.
At any moment, the researchers could download a file with the overview of the ID-numbers and information about opt-out, completion of questionnaire, availability of blood results, and informed consents. At the end of the study, the anonymized dataset combining data from the questionnaire and the antibody test could be downloaded by the researchers and used for analysis.
Call center
For the data collection phase, a call center was set up by Public Health Service South Limburg (NL) in all three languages. Call center employees received training from the involved researchers of the Public Health Service South Limburg on the online tool. Invitees to the study could call the call center for more information, to request the antibody result, to solve technical problems, to assist with the online questionnaire.
Call center employees logged the questions in a logbook anonymously, only mentioning the ID-number of invitees to ensure privacy. This way, the researchers could monitor the number and type of questions. Alternatively, participants could send an email to the researchers via the email address mentioned in the letters or on the website. Researchers answered the emails in the appropriate language.
Fieldwork
Invitees were sent an invitation letter in the official language of the sub region with an information brochure, a link and ID-code for filling in an online questionnaire, an informed consent form accompanied by a home-sampling kit including a pretested instruction with pictures on how to use a finger prick to sample some droplets of blood in a micro tube, two finger prick lancets (one extra in case of failure), a micro tube (BD Microtainer® K2EDTA collection tube), plaster, gauze bandage, plastic safety bag and a pre-stamped envelope for return to the laboratory.
In both rounds and in all sub regions, one week after sending the invitation a general reminder letter was sent whether or not invitees participated. The documents were uniform in the three countries, except for some variations that are specific to the data collection procedures, information on local authorities and logistical characteristics.
The fieldwork procedures were carried out in each country by the responsible project partners. Variations in the fieldwork procedure are due to local factors and national legislations in conducting scientific research. In weekly consultations, the most important decisions were taken in consultation with all partners in order to achieve a parallel approach, as much as possible.
The laboratory for analysis of the blood samples was located in the Netherlands. Participants from Germany and Belgium could not send blood samples straight to the laboratory, seeing that personal information (informed consent) could not cross the border. For that reason, the Ministry of the German Speaking Community in Belgium served as collection point for Belgian participants: informed consents were scanned in the online tool by employees of the Ministry of the German Speaking Community and send to Sciensano, while the blood samples were picked up by a courier to the laboratory. In Germany, a similar procedure was followed where the Public Health Service Düren was used as the collection point.