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Background: The aim of this study was to describe the prevalence of persistent symptoms of COVID in hospitalized pediatric population one year after admission compared to a control group. Method(s): Prospective observational study conducted in 2 hospitals. We included patients aged 0-18 years hospitalized for acute COVID-19 more than a year ago and controls, matched by age and sex, hospitalized for causes other than COVID-19, and who had never COVID-19 at recruitment or during the follow-up. Families were contacted and a standardized survey was conducted. Persistent COVID/disease was defined as the presence of symptoms with onset in the first 3 months after COVID-19 and with persistence for more than 2 months. Result(s): 50 cases and 46 controls were analyzed, 58.3% male, 36% <5 years. Families were interviewed a median of 1.89 years (interquartile range;1.25-2.07) after hospitalization. The definition of persistent COVID-19/disease was met in 34% of cases vs. 37% of controls (p=0.767). Symptoms persisted >=11 months in 24% (12/50) of cases vs. 13% (6/46) of controls (p=0.182), with no differences by age group. The most frequent symptoms at 1 year in cases were fatigue (8%), headache (6%), poor appetite (6%), abdominal pain (6%) and variations in heart rate (6%). In controls, persistent symptoms were mostly abdominal pain (6%) and poor appetite (6%). The number of readmissions was 11/50 (22%) and 6/46 (13%) (p=0.267), respectively. On emotional/behavioral items, 16/50 (32%) of cases reported that their emotional state was worse or much worse than before admission, compared to 16/46 (34.7%). No risk factors associated with the development of persistent symptoms were found, except the length of hospital admission (p=0.043). Conclusion(s): In this study, the prevalence of persistent symptoms was not different in patients with and without COVID-19. 1-year persistence was higher in COVID-19 cases but did not reach significance. Persistence correlated with length of hospitalization.
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Background: Little is understood about which comorbidities are associated with severe outcomes in children hospitalized with acute COVID-19. Some confusion lies especially for cancer or diabetes. Method(s): Data from 2 multicenter prospective cohort studies of hospitalized children (aged 0-18 years) with confirmed SARS-CoV-2 in Spain and Colombia were combined for this analysis. Data were obtained from 116 hospitals. Outcome was classified as (in decreasing order of severity): death, mechanical ventilation (MV), pediatric intensive care unit (PICU) admission, high flow/CPAP, oxygen therapy with nasal prong (NP) and hospitalization without respiratory support. Risk factors for severity, adjusting for age and gender, were identified using multinominal logistic regression and a backwards selection process. Result(s): A total of 1,753 patients were included, 734 (41.8%) in Spain and 1,019 (58.1%) in Colombia. The most frequent comorbidities were asthma (9.0%), chronic neurological disorder (NRL) (7.4%), immunosuppressive medication (7.2%), malignant neoplasms (5.4%) and chronic lung disease (not asthma) (CLD) (4.5%). Comorbidities associated with the different endpoints are summarized in Figure 1. Asthma was associated with a significantly increased risk of death (OR: 4.17;95%CI 1.34-12.97), MV (OR: 7.94 (3.59-17.56)), PICU admission (OR: 3.37 (1.91- 5.96)), high flow/CPAP (OR: 6.65 (2.69-16.46)), and NP (OR: 3.85 (2.57-5.77)) compared to hospitalization without respiratory support. NRL was associated with increased risk of death (OR: 7.34 (3.01-17.90)), MV (OR: 3.07 (1.20-7.82)) and high flow/CPAP (OR: 4.36 (1.68-11.29)). CLD was associated with increased risk of death (OR: 6.22 [2.28-16.94]) and NP (OR: 3.1 (1.74-5.58)) and in addition, chronic cardiac disease was associated with increased risk of MV (OR: 5.21 (1.76-15.41)) and PICU (OR: 2.78 (1.27-6.08)). Risks of death (OR: 4.49 (2.03-9.05)), MV (OR: 2.97 (1.52-5.81)), PICU (OR: 4.27 (2.89-6.33)), and NP (OR: 4.67 (3.64-5.99)) were higher in the Colombia Cohort. Conclusion(s): Asthma, chronic neurological, cardiac and lung disease;and belonging to the Colombia cohort were consistently associated with multiple severe outcomes of COVID-19. Cancer and diabetes association with selected endpoints rather than with most endpoints may be more related to the baseline disease than with the actual COVID-19.
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Background: mRNA vaccines elicit a durable humoral response to SARS-CoV-2 in adults, whereas evidence in children is lacking. This study aimed to evaluate the early and long-term immunological response after the BNT162b2 vaccine in children with or without a previous SARS-CoV-2 infection. Method(s): In a multicenter, prospective, observational study we profiled the immune response to the BNT162b2 vaccine in children aged 5-11 years attending the Pediatric Departments at the University of Padua and Bambino Gesu Children's Hospital in Rome (Italy). Forty-four healthy children (HC), 20 immune compromised (IC), and 18 children who previously developed MIS-C (MIS-C) were included in the study. Blood samples were collected pre-, 1, and 6 months after a 2-doses vaccination schedule. Neutralizing antibodies (NAbs) and anti-S-RBD IgG titers were analyzed through Plaque Reduction Neutralization Test (PRNT) and chemiluminescent immune-enzymatic assay (CLIA), respectively. B and T cell phenotypes were analyzed by flow cytometry. Geometric mean titers (GMTs) and 95% confidence intervals and median and interquartile range (IQR) of variables were evaluated according to pre-existing confirmed COVID-19. Result(s): Eighty-two children were studied;60 with a molecular-documented previous COVID-19 (Group A) and 22 without previous infection defined as the absence of antigen-specific antibodies before the vaccination (Group B). Overall, in Group A we observed higher NAbs GMTs, anti-S-RBD titers, and T- and B-reg cells than in Group A, at both 1 and 6 mo after vaccination (table);Nabs against the parental virus resulted to be greater in Group A than in Group B by a factor of 18 and 11, at 1 and 6 mo after vaccination, respectively. Both Groups recorded a decrease in antibody titers of approximately 50-70% between 1 and 6 months. A significant difference for Omicron NAbs (p=0.02) and anti-S-RBD (p=0.07) titers decay was observed between Group A and B;in contrast, Parental NAbs titers appeared to have similar trends in the 2 groups (p=0.47). Comparable antibody titers at 1 and 6 mo. (p=0.37) were detected across the three categories of HC, IC, and MIS-C (table). Conclusion(s): mRNA vaccination triggers a higher humoral response in children with a previous history of COVID-19, regardless of the immune deficiency or previous MIS-C, at least up to 6 mo, providing insight into boosting preexisting immunity with mRNA vaccines.
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BACKGROUND: In May 2020, the ACCESS (The vACCine covid-19 monitoring readinESS) project was launched to prepare real-world monitoring of COVID-19 vaccines. Within this project, this study aimed to generate background incidence rates of 41 adverse events of special interest (AESI) to contextualize potential safety signals detected following administration of COVID-19 vaccines. METHODS: A dynamic cohort study was conducted using a distributed data network of 10 healthcare databases from 7 European countries (Italy, Spain, Denmark, The Netherlands, Germany, France and United Kingdom) over the period 2017 to 2020. A common protocol (EUPAS37273), common data model, and common analytics programs were applied for syntactic, semantic and analytical harmonization. Incidence rates (IR) for each AESI and each database were calculated by age and sex by dividing the number of incident cases by the total person-time at risk. Age-standardized rates were pooled using random effect models according to the provenance of the events. FINDINGS: A total number of 63,456,074 individuals were included in the study, contributing to 211.7 million person-years. A clear age pattern was observed for most AESIs, rates also varied by provenance of disease diagnosis (primary care, specialist care). Thrombosis with thrombocytopenia rates were extremely low ranging from 0.06 to 4.53/100,000 person-years for cerebral venous sinus thrombosis (CVST) with thrombocytopenia (TP) and mixed venous and arterial thrombosis with TP, respectively. INTERPRETATION: Given the nature of the AESIs and the setting (general practitioners or hospital-based databases or both), background rates from databases that show the highest level of completeness (primary care and specialist care) should be preferred, others can be used for sensitivity. The study was designed to ensure representativeness to the European population and generalizability of the background incidence rates. FUNDING: The project has received support from the European Medicines Agency under the Framework service contract nr EMA/2018/28/PE.
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Background: Clinical manifestations of children's coronavirus disease-2019 (COVID-19) were initially considered less severe compared with adult patients. However, there is now increasing evidence of a "long-tail" of COVID-19 related symptoms lasting for several months after recovery from the acute infection. Long COVID-19-related symptoms and mechanisms are poorly characterized and understood, with several phenotypes reported, often driven by long-term tissue damage (such as lung, heart and brain) and pathological inflammation due to viral persistence and/or immune deregulation. Purpose(s): The objective of this study was to evaluate atrio-ventricular mechanics, by means of two-dimensional speckle-tracking echocardiography, in previously healthy children recovered from asymptomatic or mildly symptomatic severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in a long-term followup. Method(s): We analysed a cohort of 157 paediatric patients, mean age 7 +/- 4 years, who had a confirmed diagnosis of SARS-CoV-2 infection and were asymptomatic or mildly symptomatic for COVID-19. Patients underwent standard transthoracic echocardiogram and speckle tracking echocardiographic study 148 +/- 68 days after diagnosis. One hundred seven age, sex, and body surface area comparable healthy subjects were used as control group. Result(s): Left ventricular ejection fraction was within normal limits in postCOVID-19 cases and CTRL with no significant differences between the two groups (postCOVID-19: 65.6 +/- 4% vs CTRL: 65.0 +/- 5%, p = 0.182).Left ventricular (LV) global longitudinal strain (postCOVID-19: -20.5 +/- 2.9%;CTRL: -21.8 +/- 1.7%;p < 0.001) was significantly reduced in cases compared with CTRLs. An amount of 11 (7%) postCOVID-19 cases showed impaired GLS values < -17% and 95 subjects (60%) presented with a strain lower than -16% in more than 2 segments. These subjects did not show any difference regarding symptoms or serological findings. Moreover, GLS was significantly reduced in children with disease's onset during the second wave of COVID-19 pandemic, compared with those during the first wave (second wave: -20.2 +/- 2.6%;first wave: -21.2 +/- 3.4%;p = 0.048). Finally, peak left atrial systolic strain was within the normal range in the postCOVID-19 group with no significant differences compared to CTRL (postCOVID-19: 49.1 +/- 12%;CTRL: 49.5 +/- 18%). Conclusion(s): SARS-CoV-2 infection may affect left ventricular deformation in children despite an asymptomatic or only mildly symptomatic acute illness. Our data show an amount of 60% of children, recovering from asymptomatic or mildly symptomatic COVID-19, with still mild subclinical systolic cardiac impairment in the midand long-term follow-up after the infection. This subtle impairment was seen to be worse in children recovering from the second wave of COVID-19 compared to the first one. A follow-up is needed to verify the reversibility of these alterations and their impact on long-term outcomes.
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Background: Clinical manifestations of children's coronavirus disease-2019 (COVID-19) were initially considered less severe compared with adult patients. However, there is now increasing evidence of a “long-tail” of COVID-19 related symptoms lasting for several months after recovery from the acute infection. Long COVID-19-related symptoms and mechanisms are poorly characterized and understood, with several phenotypes reported, often driven by long-term tissue damage (such as lung, heart and brain) and pathological inflammation due to viral persistence and/or immune deregulation. Purpose: The objective of this study was to evaluate atrio-ventricular mechanics, by means of two-dimensional speckle-tracking echocardiography, in previously healthy children recovered from asymptomatic or mildly symptomatic severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in a long-term follow-up. Methods: We analysed a cohort of 157 paediatric patients, mean age 7±4 years, who had a confirmed diagnosis of SARS-CoV-2 infection and were asymptomatic or mildly symptomatic for COVID-19. Patients underwent standard transthoracic echocardiogram and speckle tracking echocardiographic study 148±68 days after diagnosis. One hundred seven age, sex, and body surface area comparable healthy subjects were used as control group. Results: Left ventricular ejection fraction was within normal limits in postCOVID-19 cases and CTRL with no significant differences between the two groups (postCOVID-19: 65.6±4% vs CTRL: 65.0±5%, p=0.182).Left ventricular (LV) global longitudinal strain (postCOVID-19: −20.5±2.9%;CTRL: −21.8±1.7%;p<0.001) was significantly reduced in cases compared with CTRLs. An amount of 11 (7%) postCOVID-19 cases showed impaired GLS values < −17% and 95 subjects (60%) presented with a strain lower than −16% in more than 2 segments. These subjects did not show any difference regarding symptoms or serological findings. Moreover, GLS was significantly reduced in children with disease's onset during the second wave of COVID-19 pandemic, compared with those during the first wave (second wave: −20.2±2.6%;first wave: −21.2±3.4%;p=0.048). Finally, peak left atrial systolic strain was within the normal range in the postCOVID-19 group with no significant differences compared to CTRL (postCOVID-19: 49.1±12%;CTRL: 49.5±18%). Conclusions: SARS-CoV-2 infection may affect left ventricular deformation in children despite an asymptomatic or only mildly symptomatic acute illness. Our data show an amount of 60% of children, recovering from asymptomatic or mildly symptomatic COVID-19, with still mild subclinical systolic cardiac impairment in the mid- and long-term follow-up after the infection. This subtle impairment was seen to be worse in children recovering from the second wave of COVID-19 compared to the first one.A follow-up is needed to verify the reversibility of these alterations and their impact on long-term outcomes. Funding Acknowledgement: Type of funding sources: None.
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Background and Aim: Clinical manifestations of children's corona-virus disease-2019 (COVID-19) are generally considered less severe compared with adult patients. The objective of this study was to evaluate cardiac involvement in healthy children with asymptomatic or mildly symptomatic severe acute respiratory syn-drome coronavirus-2 (SARS-CoV-2) infection. Method(s): We analysed a cohort of 210 paediatric patients, mean age 7 +/- 4 years, who had a confirmed diagnosis of SARS-CoV-2 infection and were asymptomatic or mildly symptomatic for COVID-19. Patients underwent standard transthoracic echocardiogram and speckle tracking echocardiographic study 138 +/- 65 days after diagnosis. Seventy-two age, sex, and body sur-face area comparable healthy subjects were used as control group. Result(s): Left ventricular ejection fraction was within normal limits but significantly lower in the cases group compared to controls (62 +/- 4% vs. 65 +/- 5%;P = 0.012). Left ventricular (LV) global longi-tudinal strain (-20,91 +/- 2,83 % vs.-22,73 +/- 2,51 %;P lt;0.001) was significantly reduced in cases compared with CTRLs. Regional LV strain analysis showed a significant reduction of the LV mid-wall segments strain among cases compared to controls. Furthermore, in the cases group, there were 25% of subjects with a regional peak systolic strain below-16% (-2.5 Z score in our healthy cohort) in at least two segments. These subjects did not show any difference regarding symptoms or serological findings. Conclusion(s): SARS-CoV-2 infection may affect left ventricular deformation in 26% of children despite an asymptomatic or only mildly symptomatic acute illness. A follow-up is needed to verify the reversibility of these alterations and their impact on long-term outcomes.
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Background: Understanding the long-term kinetics of the immune response against SARS-CoV-2 infection is crucial in guiding public health policies and optimizing of vaccination strategies. While it is known that SARS-CoV-2 specific antibodies may persist in adults 12 months after infection, data are lacking in the pediatric population. We herein describe the long-term immune response in children following SARS-CoV-2 infection. Methods: Single-centre, prospective observational study analyzing family clusters of COVID-19 attending the Pediatric Department, University of Padua (Italy). Confirmed COVID-19 infection was defined by positive SARS-CoV-2 PCR and/or IgG serology. All patients with confirmed infection at enrolment underwent serological follow-up at 1-4, 5-10, and >10 months after infection. Plasma was analyzed to quantify anti-SARS-CoV-2 S-RBD IgG, by chemiluminescent immunoassay, performed on MAGLUMI™2000 Plus (Snibe Diagnostics). IgG title >4.3 kBAU/L was considered positive. Results: Among 902 subjects (252 COVID-19 family clusters), 698 had confirmed COVID-19, including 352 children/older siblings aged 8.6 ±5.1 years, and 346 parents aged 42.5 ±7.1 years;of those, 96.5% cases had asymptomatic/mild COVID-19. Children showed significantly higher S-RBD IgG titers than older subjects across all follow-up time points, with an overall mean S-RBD IgG titer <3 years of age five-fold higher than adults (282.3 [139-516.6] kBAU/L vs 56.7 [24.6-136.9] kBAU/L, p<0.001) (Table). The longitudinal analysis of 60 subjects sampled at least twice during follow-up demonstrated the persistence of antibodies up to 10 months from infection in all age classes. Subjects >6 years of age showed a significant progressive decline of the S-RBD IgG titer from the first serological follow-up. While, in younger children antibodies remained stable at 5-10 months of follow-up (p=0.0625), with a subsequent significant decline afterwards (p<0.001). Conclusion: In our unique family cluster cohort, we confirmed the different kinetics of the COVID-19 humoral response across several age groups of asymptomatic/mild COVID-19 cases in our family-cluster cohort. Children presented with higher S-RBD IgG titer at every time point up to 10 months of follow-up. Children less than 3 years demonstrated a more intense long-term resilience of their immune response, which started to decline significantly only after ten months from infection.
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Introduction: Evidence suggests that, compared with adult patients, clinical manifestations of children's COVID-19 may be less severe. However, multiple reports have raised concern about the so called pediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2 (PIMS-TS) which resembles other inflammatory conditions (i.e. Kawasaki disease, toxic shock). Patients affected by PIMS-TS showed cardiac involvement with myocardial injury, reduced left ventricle systolic function and coronary artery abnormalities, and in some cases, need for inotropes/ vasopressors and extracorporeal life support (ECLS). Little is known regarding cardiac involvement in pediatric patients with SARS-CoV-2 infection and none or only mild symptoms of disease. Methods: We analyzed 52 pediatric patients (29males, 56%) with diagnosis of SARS-CoV-2 infection based on either PCR analysis of nasopharingeal swab (NPS), or serological finding of IgG on blood sample and asymptomatic (23%) or only mildly symptomatic (77%) for COVID-19. Patients underwent transthoracic echocardiogram (TTE) after a median time of 3.6 months from diagnosis and negative NPS for SARS-CoV-2. Offline analysis with GE EchoPAC software to measure global longitudinal strain (GLS) of the LV using 2D speckle tracking imaging. Therefore, we compared the results with an age-matched group of 32 controls (18males, 56%). Results: Cases and controls were similar regarding age and gender. LV biplane EF was significantly lower in the cases group, although still in the normal range (62.4±4.1% vs. 65.2±5.5%, p=0.012). TAPSE and LV-GLS were comparable between the two groups. GLS analysis showed significant strain reduction of the LV midwall segments and of the basal anterior, posterior and septal inferior segments among cases compared to controls. On the other hand, apical segments showed higher deformation in cases compared to controls. Furthermore, in the case group there were 14 subjects (27%) with a strain below 16% (mean value minus 2.5 SD) in at least 2 segments. Conclusions: SARS-CoV-2 infection may affect LV deformation in asymptomatic or only mildly symptomatic children, showing a peculiar pattern with lower longitudinal strain in all mid-wall segments of LV compared to control subjects. The clinical significance of this findings is unclear and follow-up is needed to verify the reversibility of this alterations.
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Background and Aim: Salivary SARS-CoV-2 Ab determination could be suitable for monitoring the viral spread and vaccination efficacy, especially in pediatric patients. We investigated N/S1-RBD IgG antibody levels in salivary samples of infectious-naïve vaccinated subjects and of COVID-19 patients, further comparing levels with serum anti-SARS-CoV-2 S-RBD IgG. Methods: A total of 72 subjects were enrolled at the Padova University Hospital: 36 COVID-19 patients and 36 health care workers (HCW), who underwent a complete vaccination campaign with BNT162b2 (BioNTech/Pfizer). All collected a salivary sample, using Salivette (Sarstedt, Nümbrecht Germany). For 9 HCW, salivary samples were collected at three different times within the same day (before breakfast, at 10 am, and after lunch). A serum sample was also collected for all individuals. Time post symptoms onset or time from the first vaccine were also recorded. Salivary COVID-19 N/S1 RBD (sal-IgG) ELISA (RayBiotech, GA, USA) and anti-SARS-CoV-2 S-RBD IgG Ab (ser-IgG) (Snibe Diagnostics, Shenzhen, China) were used for determining IgG Ab. Results: Subjects' mean age (±sd) was 35.8±18.2 yrs. Age significantly differed (p<0.001) from COVID-19 patients [29.7±17.3 yrs] and HCW [47.1±12.9 yrs]. Positive sal-IgG were found in 70/72 (97.2%) samples;in sera, 71/72 (98.6%) samples were positive to ser-IgG. The sal-IgG median levels differed from COVID-19 to vaccinated HCW, being in salivary samples 0.21 kAU/L and 0.8 kAU/L (p =0.030), respectively;median levels for ser-IgG in COVID-19 and vaccinated HCW were 135 kBAU/L and 940 kBAU/L, respectively (p<0.001). Salivary IgG levels were not influenced by time post-symptom onset or time post-vaccination, both on vaccinated HCW (rho= -0.147, p=0.402) and COVID-19 subjects (rho=0.0267, p=0.986). Ser-IgG levels was not influenced by the time post-symptom onset for COVID-19 subjects (rho=0.102, p=0.419), while a strong significant correlation was found with time post-vaccination in HCW (rho=-0.6292, p<0.001). Sal-IgG levels were notinfluenced by the daytime of collection (rho=0.148, p=0.373). Passing-Bablok regressions showed that sar- IgG and ser-IgG comparability was assessable only when ser-IgG values were divided by 1000, being slope and intercept 0.068 (95%CI: 0.069-0.341) and 0.221 (95%CI:- 0.097 to 0.786), respectively. Conclusions: Salivary IgG is efficiently detectable both in COVID-19 and in vaccinated individuals and analyses appeared to be not influenced by the daytime of collection. The analyses performed showed that, overall, sal-IgG were lower than ser-IgG, and thus comparability with serum levels needs to be better explored.
ABSTRACT
Background: Evidence suggests that, compared with adult patients, clinical manifestations of children's COVID-19 may be less severe. However, multiple reports have raised concern about the so called pediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2 (PIMS-TS) which resembles other inflammatory conditions (i.e. Kawasaki disease, toxic shock). Patients affected by PIMS-TS showed cardiac involvement with myocardial injury, reduced left ventricle systolic function and coronary artery abnormalities, and in some cases, need for inotropes/vasopressors and extracorporeal life support (ECLS). Little is known regarding cardiac involvement in pediatric patients with SARS-CoV-2 infection and none or only mild symptoms of disease. Methods: We analyzed 52 pediatric patients (29males, 56%) with diagnosis of SARS-CoV-2 infection based on either PCR analysis of nasopharingeal swab (NPS), or serological finding of IgG on blood sample and asymptomatic (23%) or only mildly symptomatic (77%) for COVID-19. Patients underwent transthoracic echocardiogram (TTE) after a median time of 3.6 months from diagnosis and negative NPS for SARS-CoV-2. Offline analysis with GE EchoPAC software to measure global longitudinal strain (GLS) of the LV using 2D speckle tracking imaging. Therefore, we compared the results with an age-matched group of 32 controls (18males, 56%). Results: Cases and controls were similar regarding age and gender. LV biplane EF was significantly lower in the cases group, although still in the normal range (62.4±4.1% vs. 65.2±5.5%, p=0.012). TAPSE and LV-GLS were comparable between the two groups. GLS analysis showed significant strain reduction of the LV mid-wall segments and of the basal anterior, posterior and septal inferior segments among cases compared to controls. On the other hand, apical segments showed higher deformation in cases compared to controls. Furthermore, in the case group there were 14 subjects (27%) with a strain below 16% (mean value minus 2.5 SD) in at least 2 segments. Conclusion: SARS-CoV-2 infection may affect LV deformation in asymptomatic or only mildly symptomatic children, showing a peculiar pattern with lower longitudinal strain in all mid-wall segments of LV compared to control subjects. The clinical significance of this findings is unclear and follow-up is needed to verify the reversibility of this alterations.
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Background: Further knowledge on adaptive immunity to SARS-CoV-2 (CoV-2) in children is needed in order to define possible immunization strategies and reconsider pandemic control measures. We analyzed anti-CoV-2 antibodies (Ab) and their neutralizing activity (PRNT), alongside antigen (Ag) specific cellular response, in relation to virus load in nasopharyngeal swabs. Methods: We analysed 42 CoV-2 patients at 7 days after symptoms onset. CoV-2 viral load (VL) was measured by RT-PCR and digital droplet PCR on longitudinal samples of nasopharyngeal swabs (NP). Virus infectivity (FFU) was tested by virus focus forming assay. CoV-2 antibodies were investigated by Diasorin (CoV-2 Ab) and neutralization assay (PRNT). CoV-2-specific CD4-CD40L+ T-cells and Spike specific B-cells were analysed by flow cytometry. Plasma proteomic profiling was measured by 2 Olink panels. We calculated the area under the curve (AUC) of the viral load from NP collected every 48 hours up to undetectable VL. Mann-Whitney was used to compare means in individuals with neutralizing activity (PRNT+) or not (PRNT-);linear regression was used to evaluate the associations between virus load and infectivity over time. Principal component analysis (PCA) was used to analyse proteomic data. Results: Higher VL was found in seronegative patients expressed in terms of both CoV-2 Ab (p=0.003) and PRNT (p=0.0007). Similarly, lower FFU was associated with higher CoV-2 Ab (p=0.003;rho=-0.67) and PRNT (p=0.023;rho=-0.46). Further, the AUC of the viral load in NP showed an inverse correlation with CoV-2 Ab (p=0.031;rho=-0.54). Development of humoral response was associated with the presence of CoV-2 specific IgD-CD27+ B cells, with a higher frequency of CoV-2 specific B cells found in seropositive compared to seronegative (p=0.001). Besides, individuals developing neutralizing Ab had higher frequency CD4-CD40L+ T-cells compared to PRNT- (p=0.03). The plasma proteome confirmed the association between cellular and humoral CoV-2 immunity, with PRNT+ showing higher viral signal transduction molecules (SLAMF1, CD244, CLEC4G). Conclusion: This work provides a virological and immunological characterization of SARS-CoV-2 infected children presenting a differential Abmediated neutralizing activity. It demonstrates that children with neutralizing antibodies present reduced viral load, faster virus clearance and lower in vitro infectivity. These data provide information that can drive vaccination endpoints and quarantine measures policies.
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Background: SARS-CoV-2 infected children are often asymptomatic or paucisymptomatic compared to adults. The immune response plays a pivotal role in dictating the clinical outcome in infected adults, but it is still poorly investigated in the pediatric population. Methods: Fifty-seven family clusters of SARS-CoV-2, attending the Department for Women's and Children's Health (University of Padova), were enrolled between March and September 2020, for a total 209 subjects. SARS-CoV-2 infection was confirmed in 155 patients (SARS+: 93 ≥15 years [group A];34 children ≥6-15 years [group B];28 children <6 years [group C]) by virus molecular detection and/or serology. In 41 available samples, measurement of SARS-CoV-2 levels (VL) was performed by an in-house quantitative One-Step ddPCR method. A blood sample was obtained at a median [IQR] of 2.8 [2.1-3.7] months after baseline (symptom's onset and/or first positive virus detection). Neutralizing antibodies (Nabs) were detected by a Plaque Reduction Neutralization Test (PRNT). Activated (CD8+CD38+HLA-DR+) and regulatory T cells (T-regs;CD4+Foxp3+CD127-CD25+) were analyzed by flow cytometry. Results: VL did not differ by age (18507 [326-339315], 6723 [3427-114587], and 21106 [162-152500] copies/5μl, in group A, B and C, respectively;overall, p=0.955). Group C had the highest PRNT titer compared to the other groups (overall, p<0.0001). Activated CD8 and regulatory T cells were significantly higher in SARS+ than in SARS- subjects (p<0.001). CD8 activated cells were significantly higher in group A compared to the other groups (p<0.001;Figure a), and were inversely correlated with PRNT titer (group A: rs=-0.527, p<0.0001;B: rs=-0.494 p=0.003;C: rs=-0.547 p<0.0001;Figure b). Conversely, T-regs were significantly higher in group C compared to the others (p<0.001;Figure c), and were positively correlated with PRNT values in children (group C: rs=0.662 p=0.0001, B: rs =0.532 p=0.001;A: rs =0.160, p=0.125;Figure d). Conclusion: Levels of SARS-CoV-2 did not differ among age classes, but adults displayed a higher T cell activation and a lower production of anti-SARS Nabs than children. Conversely, younger infected children had the highest production of anti-SARS Nabs and the lowest non-specific T cell activation, most likely due to their higher expression of regulatory T cells.
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Background: Recent evidences suggest that SARS-CoV-2 neutralizing antibodies (Nabs) may persist over time, however lack of knowledge still regards the pediatric population. Methods: A single-centre, prospective observational study evaluated family clusters of COVID-19 attending the Pediatric Department, University Hospital of Padua (Italy). Confirmed COVID-19 was defined by positive SARS-CoV-2 molecular detection and/or serology;patients/family symptom's and virological positivity were considered to define the infection onset (baseline). Blood samples were analyzed in pair to detect Nabs through Plaque Reduction Neutralization Test (PRNT), and IgG through chemiluminescent immuneenzymatic assay (CLIA) MAGLUMI™ 2000 Plus;IgG >1.1 kAU/L and/or PRNT≥1:10 were considered positive. SARS-CoV-2 viral load (VL) was quantified by multiplex quantitative assay based on One-Step RT-ddPCR. Geometric mean titers (GMT) and 95% Confidence Intervals of IgG/PRNT were evaluated, stratified by age and time from baseline to sample collection. Trends over time of immune-virological response were assessed. P-value <0.05 was considered statistically significant. Results: Among 213 subjects (57 families) evaluated, 155 had confirmed COVID-19 including 73 (47%) children/older siblings of 8 years median age (IQR 4-13) and 82 (53%) parents aged 42 years (IQR 34-46);93.5% had asymptomatic/mild COVID-19. From the cumulative analysis of 194 blood samples, Nabs persisted over a median period of 95 days (IQR, 67-133) from baseline. Children showed significantly higher NAbs than older subjects, with children <3 years ranging from a 4-fold difference at 1-2 months to 8.8-fold difference at 3-6 months after baseline, compared to adults (table). The longitudinal assessment of 42 subjects sampled at 60 days (SD+/-9.9) and at 150 days (SD+/-24.2) showed a 2-fold increase in NAbs in children <6 years (PRNT 144, 95% C.I. 74.42-277.94 versus 303, 95% C.I. 196.43-468.57) and a substantial stability in Nabs among older subjects. CLIA IgG significantly decreased over time for all age classes, becoming negative in 13/42 subjects (31%), compared to 1/42 subjects detected by PRNT. Among 32 individuals checked for VL within 4 days from baseline, VL directly correlated with PRNT titers in subjects >15 years (Pearson Coefficient =0.70, p=0,0349) but not in pediatric cases. Conclusion: Asymptomatic/mild COVID-19 disease triggers in children a superior and persistent humoral response compared to adults.
ABSTRACT
Background: SARS-CoV-2 (CoV-2) infected children often range from being paucysymptomatic to fully asymptomatic. The impact of this population on the epidemics due to their ability to transmit the virus and achieve protective immunity has been poorly defined. We explored CoV-2 infectivity potential and anti-CoV-2 cellular (CD8, NK and B) and humoral response in symptomatic (SY) and asymptomatic (AS) CoV-2 infected children, screened for a family member resulted infected. Methods: CoV-2 viral load was measured by RT-PCR and digital droplet PCR (ddPCR) on longitudinal samples of nasopharyngeal swabs in 9 AS and 33 SY (samples were paired according to symptoms'onset for SY and first family contact for AS). Virus infectivity was tested by Virus focus forming assay (FFA). CoV-2 antibodies were investigated by Diasorin (CoV-2 Ab) and Ab-mediated neutralization activity (PRNT) at diagnosis, (samples collected >5 days from symptoms onset in SY, or from first family contact in AS were excluded from this timepoint), and in the convalescent phase (CP) (10-14 days after infection). Cellular response was analyzed by flow cytometry: 1) Ag-specific B cells, by a S1+S2 CoV2-R-PE probe;2) Ag-specific CD8+T cells by ICAM+;3) natural-killer (NK) phenotype. Mann-Whitney was used for comparison;linear regression was used to evaluate the associations between virus load and infectivity. Results: AS showed lower viral load (p=0.004) and faster virus clearance (p=0.0002) compared to SY. Virus infectivity was associated with ddPCR (rho=0.66;p=0.002). ASY and SY showed similar levels of CoV-2 Ab and PRNT, at both diagnosis and at follow up. During the CP, the proportion of CoV-2 Ab negative was 33,3% for both groups and PRNT was negative in 16,6% and 15,7% of AS and SY respectively. Anti-CoV-2 cellular immunity was comparable between ASY and SY. Indeed Ag-specific B cells and CD8 T cells were detectable despite symptomatology and no major differences were found between the groups. Total NK frequency was similar between the groups, while a regulatory NK subset (CD56bright NK cells) was higher in AS compared to SY (p=0.01). Conclusion: These data show that AS have a lower infectivity potential compared to SY suggesting that mitigated restrictive measures or alternative screening may be considered for this population. In addition, these patients showed an intact ability to produce humoral and cellular CoV-2 specific responses hence contributing to achieve herd immunity as much as SY.
ABSTRACT
Background: Evidence suggests that clinical manifestations of children's COVID-19 may be less severe. However, it has been described the pediatric inflammatory multisystem syndrome temporally associated with SARS-CoV-2 (PIMS-TS) which resembles other inflammatory conditions (i.e. Kawasaki disease). Patients affected by PIMS-TS showed cardiac involvement with myocardial injury, reduced left ventricle systolic function and coronary artery abnormalities. Little is known regarding cardiac involvement in pediatric patients with asymptomatic or mildly symptomatic SARS-CoV-2 infection. Methods: We analyzed 23 pediatric patients (13males, 56%) with diagnosis of SARS-CoV-2 infection based on PCR analysis of nasopharin-geal swab (NPS), and asymptomatic or only mildly symptomatic for COVID-19. Patients underwent standard transthoracic echocardiogram (TTE) within 2-3 month from diagnosis and after negative NPS for SARS-CoV-2. We performed offline analysis with GE EchoPAC software to measure global longitudinal strain (GLS) of the LV using 2D speckle tracking imaging. Therefore, we compared the results with a matched group of 23 controls (13males, 56%). Results: Cases and controls were similar regarding age (5.9 ± 4.1years vs. 6.4 ± 4.4 years, p = 0.63), body surface area (0.98 ± 0.3m2 vs. 0.8 ± 0.4m2, p = 0.17), LV FS (37.9 ± 5.9% vs. 36.4 ± 8.3%, p = 0.74) and LV biplane EF (63.9 ± 5.2% vs. 66.4 ± 5.3%, p = 0.11). GLS analysis showed significant strain reduction of the LV mid-wall segments and of the basal anterior, posterior and septal inferior segments among cases compared to controls. Furthermore, in the case group there were 7 subjects (30%) with a strain below 16.5% in at least 3 segments. Conclusion: SARS-CoV-2 infection may affect LV deformation in asymptomatic or only mildly symptomatic children, showing a peculiar pattern with lower longitudinal strain in all mid-wall segments of LV compared to control subjects. The clinical significance of this findings is unclear and follow-up is needed to verify the reversibility of this alterations.