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Pulmonary Function and Persistent Clinical Symptoms in Children and Their Parents 12 Months After Mild SARS-CoV-2 Infection

Background Pulmonary involvement is the leading cause of morbidity and mortality after severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. Long-term impairment has been reported in adults with severe infection. However, most infections cause only mild symptoms or are even asymptomatic, especially in children. There is insufficient evidence regarding pulmonary outcome measures in mild SARS-CoV-2. The objectives of this study were to determine spirometry parameters after SARS-CoV-2 infection and correlate those with reported persisting symptoms in children, adolescents, and adults. Methods Data on clinical symptoms during acute infection as well as SARS-CoV-2 serology results were recorded. Twelve months after infection, spirometry was performed and information on persisting symptoms was collected using a structured questionnaire. 182 participants (108 SARS-CoV-2 positive) from 48 families were included; 53 children (< 14 years), 34 adolescents and young adults (14–25 years), and 95 adults. Results Spirometry values did not significantly differ between the particular subgroups of the cohort (adults, adolescents, children; infected and non-infected individuals). Adults reported more symptoms during acute infection as well more persisting fatigue (29.7% of participants), reduced physical resilience (34.4%), and dyspnea (25.0%) 12 months after infection than adolescents (fatigue 26.7%, reduced physical resilience 20%, and 0% dyspnea) and children (4%, 0%, 0%, respectively). There was no correlation between persistent subjective symptoms and spirometry results. Discussion Children and adolescents are less affected than adults by acute SARS-CoV-2 as well as by post-infection persistent symptoms. Spirometry was not able to demonstrate any differences between healthy individuals and participants who had suffered from mild SARS-CoV-2 12 months after the infection.


INTRODUCTION
Since the end of 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread globally and caused more than 370 million infections worldwide to date. Over 5.6 million deaths have been reported due to the new coronavirus disease  in this global pandemic [COVID-19 dashboard 1 , (1)]. Morbidity and mortality have significantly impacted health care providers and communities. However, most infected individuals show a mild clinical course and many are asymptomatic (2,3). Children typically have less severe disease and are more often asymptomatic than adults (4)(5)(6).
Persistent dyspnea and cough have been reported even months after hospital discharge (16,17). Adult survivors of severe COVID-19 show restrictive and, less frequently, obstructive changes in lung function tests (18). A reduced carbon-monoxide (CO) diffusion capacity as well as a reduced 6-min walking distance, and oxygen desaturation on exercise have been reported 4 months after COVID-19 (19). Even in mild cases without hospitalization, persistent pulmonary symptoms, such as dyspnea and reduced exercise capacity, and reduced pulmonary function test results have been reported 2 months after infection (20).
Little is known about the differences in pulmonary function tests and subjective persistent symptoms in children, adolescents and adults more than just a few months after mild SARS-CoV-2 infection. We prospectively assessed households with SARS-CoV-2 infected and non-infected parents and their children and performed spirometry around 12 months after infection. We correlated infection status and spirometry parameters with reported acute symptoms during infection and persistent chronic symptoms.

Study Design
We conducted a single-center, cross-sectional study to assess pulmonary function after mild SARS-CoV-2 infection in children (< 14 years), adolescents (14-25 years), and adults (> 25 years), and correlated results with SARS-CoV-2 serology and reported symptoms. This study is part of the COVID-19 BaWü prospective household study, where families with at least one SARS-CoV-2 PCR-confirmed or seropositive index case 1 https://coronavirus.jhu.edu/map.html of SARS-CoV-2 were included. Cohort data on intrafamilial transmission dynamics of SARS-CoV-2 (21) and long term serologic responses after SARS-CoV-2 infection (22) have been published. Participants were recruited and first examined (SARS-CoV-2 serology, clinical data collection) between April and August 2020 (T1) at Freiburg, Tübingen, Heidelberg and Ulm University Medical Centers in Baden-Württemberg, Germany. Second visit (T2) with blood drawing took place between February and May 2021. Voluntary recruitment of participants took place during the first wave of the SARS-CoV-2 pandemic between May and August 2020 via local health authorities and by the means of in-hospital databases of families with at least one laboratory-confirmed SARS-CoV-2 infection. Only participants recruited at Ulm University were subjected to spirometry measurement at T2 (approx. 12 months after SARS-CoV-2 infection, the time-point was set by the part of the study focusing on serology) and are part of this study. Participants or their parents gave written consent to participate in the study.

Spirometry
Spirometry was performed with the Easy On-PC spirometer (ndd Medical Technologies, Andover, MA, United States) after [23] standards and was evaluated with the EasyOne Connect platform (ndd Medical Technologies, Andover, MA, United States). Z-scores for forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1), FVC/FEV1, forced expiratory flow (FEF) at 75% of FVC (FEF 75%) as well as FEF25-75 were recorded (24) and are reported as FVCz, FEV1z, FVC/FEV1z, FEF75z, and FEF25-75z, respectively. Individuals who were not able to perform spirometry of adequate quality according to ATS/ERS guidelines were not included in the study, for example children < 4 years as they are not able to perform spirometry maneuvers sufficiently. Participants were grouped according to their age as children [age 4-14 years, adolescents/young adults ≥ 14-25 years, and adults (i.e., the parents of the participating children/adolescents)].

Serology and Clinical Data
SARS-CoV-2 seropositivity at T1 (approx. 4 months after infection with the virus) was used as a proxy of previous SARS-CoV-2 infection. Serological response was measured using EuroImmun-Anti-SARS-CoV-2 ELISA for anti-Spike-1 IgG, Siemens Healthineers SARS-CoV-2 for anti-RBD (receptor-binding domain) IgG and Roche Elecsys for anti-NCP (nucleocapsid-protein) IgM and IgG. Seropositivity was defined as at least any two of the three SARS-CoV-2 assays being positive [Details in Renk et al. (22)]. In the subcohort that performed spirometry no participants were hospitalized, therefore all SARS-CoV-2 infections were classified as mild or asymptomatic, depending on reported clinical data.
At T1 a questionnaire comprising demographical data, and clinical symptoms as fever, cough, diarrhea, and dysgeusia around the time of SARS-CoV-2 infection was filled out by participants or their parents (for children < 14 years of age). At T2, participants completed an online questionnaire [via REDCap© 2 (25,26)] that addressed a range of symptoms and health outcomes. In the present study, we focus on questions regarding three self-perceived subjective symptoms relevant to pulmonary involvement (fatigue, reduced physical resilience and dyspnea), their perceived severity and duration, and the extent to which they limited daily activities, all in comparison before the SARS-CoV-2 infection. Persistent symptoms were classified as symptoms still present at T2. The cohort reported here is a sub-cohort of a larger, multi-center cohort and post-/long-COVID symptoms are reported elsewhere for the whole cohort (27

Spirometry
There were no statistically significant differences in z-scores for FVC, FEV-1, FVC/FEV-1, FEF-25-75, and FEF 75 between SARS-CoV-2 seropositive and seronegative participants (Figure 1 and Supplementary Table 1) in the whole cohort or in the subgroups of children, adolescents and adults. Mean spirometry parameters were within the normal ranges in all subgroups. Patient-reported smoker status, asthma, and pollen allergies were not associated with reduced spirometry parameters.

DISCUSSION
This study reports spirometry outcomes and persistent subjective symptoms 12 months after mild SARS-CoV-2 infection in children, adolescents and adults. We have found no differences in spirometry parameters between individuals with history of SARS-CoV-2 infection and healthy controls. Only a subgroup of participants who experienced cough during acute infection showed a lower FEF 25-75z compared to those without cough. However, the results still lie within the normal range and are likely not clinically relevant. Additionally, this study found no evidence for a correlation between reported persistent symptoms as fatigue, reduced physical resilience, and dyspnea, and spirometry values. Pathological pulmonary function tests, especially reduced CO-diffusion capacity, have been reported to persist even after discharge of SARS-CoV-2 infected patients with moderate or severe pulmonary involvement (17,(29)(30)(31). Spirometry parameters 2-3 months after discharged have been reported to be reduced but a normalization of all values with the exception of FVC has been demonstrated at 6 months (32)(33)(34). Fewer data exist regarding patients with milder course who were not treated as inpatients. One study showed that patients who were still symptomatic 2 months after first symptom onset did show reduced FEV-1 and vital capacity as well as impaired CO-diffusion capacity and elevated airway resistance (20). A Swiss study found no changes in spirometry in milder cases 4 months after SARS-CoV-2 infection (19). Regarding children and adolescents, one study reported no changes in spirometry or CO-diffusion capacity 2 months after SARS-CoV-2 infection in seven children (35) and one pre-print reports no abnormalities in spirometry or multiple breath washout in 73 children and adolescent 2.6 months after infection (36).
The cohort reported here is unique due to its composition of family members of all age groups. Even though direct comparison with other studies is difficult, it is reassuring that 12 months after mild or asymptomatic SARS-CoV-2 (the longest interval compared with the other published studies) no objective changes can be found in spirometry in all three age groups.
In this study, correlation of clinical symptoms with spirometry showed only mild reduction in FEF25-75z in adults who had reported cough during acute infection. Some previous studies have reported mild reduction of spirometry parameters (20,45) in symptomatic patients, others did not (36,46). All studies show spirometry parameters in the normal ranges-as does the study reported here. Therefore, mild symptomatic SARS-CoV-2 infection does not lead to measurable changes in spirometry in children, adolescents, and adults. Whether cough in our cohort during acute infection is an indication of more pulmonary involvement and therefore more severe disease or that underlying conditions as airway hyperreactivity or asthma, unknown to the patients and therefore not reported, cannot be definitively clarified with this study. All changes in spirometry found in this study are subtle and spirometry results are still in the normal ranges and therefore most likely not of clinical relevance.
In the cohort reported here almost 25% of adults but no children or adolescents reported dyspnea 12 months after SARS-CoV-2 infection and more than 30% of adults but only 7.1% of adolescents and no children reported reduced physical resilience. Prevalence of persisting subjective symptoms after SARS-CoV-2 infection is low in children (47) whereas, depending on the study, around 30% of adults, who had not been hospitalized, reported ongoing symptoms between 3 weeks and 9 months after infection (30,(48)(49)(50). Pulmonary symptoms as cough mainly resolved within weeks after infection (16) but dyspnea may persist several months up to 1 year in adults (16,17). Adolescents, especially teenage girls, seem to suffer more frequently from long-term sequelae (27,51) but children do not (27,52). The data reported here are in concordance with the published literature and support the evidence that long-term sequelae after SARS-CoV-2 infection in children are rare. Some authors have suggested a possible link to dysfunctional breathing (for example inducible laryngeal obstruction) after SARS-CoV-2 infection (36), a condition that is well known to occur after other (infectious) triggers (53). Further studies are needed to elucidate this possible pathological link.
The major strength of this study is that it reports clinical and lung function data in multiple age groups (children, adolescents and adults) after the most common clinical scenario (asymptomatic or mild SARS-CoV-2 infection).
Limitations of the study are the single-center design and the lack of longitudinal spirometry data. Additionally, a more detailed evaluation including CO-diffusion capacity and exercise testing would have been desirable. The evaluated subgroups are small, especially the adolescent group. This group does include teenagers and young adults and is quite heterogeneous. It was decided to keep the three age groups as they best represent the structure of participating families. The data on symptoms during the acute phase of SARS-CoV-2 infection as well as on the persistent symptoms on follow-up have to be interpreted with caution. It is based on self-reporting that may be biased.

CONCLUSION
Our data show that mild SARS-CoV-2 infection in a household cohort including children, adolescents and adults did not cause persistent, clinically relevant, functional pulmonary abnormalities detectable with spirometry. Self-perceived persistent symptoms as fatigue, reduced physical resilience and dyspnea were more common in adults, less in adolescents and rare in children. The presence of symptoms was not associated with reduced spirometry parameters. More sensitive pulmonary function tests (e.g., spiroergometry, CO-diffusion), would be needed to evaluate subtle pulmonary changes in detail. Despite this reassuring finding, a pulmonary follow-up should be offered to all patients after SARS-CoV-2 infection with persistent respiratory symptoms, since dysfunctional breathing might possibly be the underlying cause in some patients.

DATA AVAILABILITY STATEMENT
Data are available upon reasonable request from the corresponding author.

ETHICS STATEMENT
The studies involving human participants were reviewed and approved by Ulm University Ethics Committee (Permit No. 152/20). Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.

AUTHOR CONTRIBUTIONS
SB designed the study, performed the spirometry, analyzed the data, and wrote the first draft of the manuscript. DF performed the spirometry and analyzed the data. MH analyzed the data and prepared the graphs. BM gave critical input to statistical analysis and analyzed the data. MZ coordinated the study and curated data. AH and PF designed online questionnaire and analyzed the data. RE, HR, and MS analyzed the data and gave valuable input to the manuscript. K-MD procured funding and provided key resources. AJ conceived the study, analyzed the data, and performed the spirometry. All authors critically revised the manuscript and agreed to the final version of the manuscript.

FUNDING
This work is part of the COVID-19 BaWü study and was financially supported within the framework of the special funding line in the state of Baden-Württemberg for COVID-19 research to the University of Ulm, Germany. The funders had no role in study design, data collection, data analysis or the decision to publish.