key: cord-0764782-aeo7tcyt
authors: Rabi, Firas A.; Al Zoubi, Mazhar S.; Al‐Iede, Montaha Mohammed; Kasasbeh, Ghena; Badran, Eman Farouk
title: Coronaviruses in Children: A Review of Potential Mechanisms of Childhood Protection
date: 2020-11-28
journal: Acta Paediatr
DOI: 10.1111/apa.15691
sha: eaea37163c721dcf8bb06024ad5c8705eabf5055
doc_id: 764782
cord_uid: aeo7tcyt

AIM: The 2019 coronavirus disease (COVID‐19) has spread worldwide and the number of cases continues to rise exponentially. Epidemiologic reports indicate that severity of illness increases with age. However, the reasons behind the relative protection of children and infants are unclear. Whether the rationale is host‐related or virus‐dependent is important to determine since the latter could change with viral mutations. We review factors that could affect the susceptibility of children to the novel coronavirus. METHODS: We search publications indexed on PUBMED. RESULTS: Descriptions of the pathophysiology of current and previous coronavirus infections suggest several viral targets and immunomodulatory pathways affecting the severity of illness. There is limited evidence to suggest age‐variability of viral cell receptors and transmembrane co‐factors required for coronavirus entry and replication. However, the ensuing cytokine storm and the effect of higher melatonin in children are age‐dependent and could explain decreased disease variability in children. CONCLUSION: We believe that current evidence suggests host factors can play a role in disease severity in children and thus may remain protective despite potential virus mutation in the future. However, we recognize and discuss avenues of future research that can further illuminate the reasons children are protected from severe COVID‐19 illness.

The coronavirus disease of 2019 (COVID-19) is currently causing a devastating global pandemic, threatening to overburden healthcare facilities worldwide. Initial impressions indicate the likelihood of pediatric infection is significantly less than the age distribution would suggest, and conjecture is that children younger than 10 years old have some physiologic or inherent resistance.

Understanding the apparent pediatric resistance to COVID-19 could predict whether it will change with future SARS-CoV-2 mutations.

The global prevalence of COVID-19 in children remains unknown. The lack of estimates for pediatric COVID-19 diagnosis can be largely attributed to the unprecedented global health challenge that has limited the universal availability of testing to diagnose asymptomatic carriers. The age distribution of COVID-19 cases in South Korea, one of the first countries to implement mass testing, shows that only 8.0% of patients were below 19 years of age, and the main risk factor for severe illness and death was increased age 1 .

While it remains to be seen if children are as vulnerable to SARS-CoV-2 infection as adults, the disease appears to be less severe in the pediatric and neonatal age group 1,2 . We review the clinical presentation of COVID-19 in the pediatric age group and search for possible reasons behind the decreased severity in children and neonates.

Coronaviruses (CoVs) are zoonotic positive-strand RNA viruses that are subclassified into four genera: alpha and beta CoVs (most likely from bats and rodents), and delta and gamma CoVs (most likely from avian species) 3 . Due to their propensity for rapid mutation and recombination, crossover infection from natural hosts to humans is possible. Seven human CoVs (HCoVs) have been identified to date -four of which cause usually mild, upper respiratory symptoms and three responsible for acute respiratory distress syndrome (ARDS) pandemics with high morbidity and mortality.

In 1965, alpha HCoV-229E and beta HCoV-OC43 were isolated from adults with common colds 4 . These viruses were considered to play a negligible role in pediatrics since they appeared to cause only mild upper respiratory symptoms in children, unless they infected premature infants or children with chronic underlying disease in whom they could cause severe lower respiratory tract illness 4 . HCoV-OC43 and HCoV-229E are considered inconsequential pathogens that are endemic globally and are responsible for ~1.5% and ~0.5%, respectively, of respiratory illnesses requiring hospitalization, generally causing respiratory distress only when there is a coinfection with another virus 4 .

In 2004, HCoV-NL63 was isolated from a seven-month-old infant with bronchiolitis 5 . In studies of children hospitalized with respiratory tract infections, HCoV-NL63 has been reported to cause between 1.2 to 9.3% of respiratory disease in children, with the variability attributed to severity of illness in the study population, or the collection season of respiratory specimens 6 .

In 2005, a 71-year-old man presented in Hong Kong with pneumonia after returning from Shenzhen, China. After testing negative for influenza and other known coronaviruses at the time, he was eventually diagnosed with a new coronavirus, termed HCoV-HKU1 7 . In one study of 11, 399 hospitalized pediatric patients aged 14 years and younger, HCoV-HKU1 was detected in 38 (0.3%), half of which were coinfected with another pathogen, such as influenza A, Mycoplasma pneumoniae, or respiratory syncytial virus 8 .

Prill et al showed that the four strains of HCoV were detected equally in inpatient children and control (asymptomatic) children and those who had been admitted were more likely to have been coinfected with other viruses 9 . Symptomatic children whose only detectable respiratory virus is a CoV were more likely to be under the age of 3, have underlying heart disease, or other chronic illness 10, 11 .

These results suggest that these four HCoVs are endemic in pediatrics.

Three other HCoVs have been responsible for three large scale epidemics with alarming morbidity and mortality: severe acute respiratory syndrome coronavirus (SARS-CoV-1), the Middle

This article is protected by copyright. All rights reserved East respiratory syndrome coronavirus (MERS-CoV) and, most recently, SARS-CoV-2 that is responsible for COVID-19 3 .

In 2002, SARS-CoV-1 was identified as the causative agent behind a life-threatening pneumonia and respiratory failure outbreak that was -until then -the most pathogenic human coronavirus identified 12 . A highly contagious CoV, it had a high mortality rate in the adult population (9.6 to 16.7%) but children younger than 12 years old ran a less aggressive clinical course and no deaths were reported in children and adolescents 13 . Whereas adults followed a "triphasic" clinical course, the majority of pediatric patients appeared to follow a "biphasic" pattern 13 In late 2019, SARS-CoV-2 was first isolated from a cluster of pneumonia cases in Wuhan City, initiating a pandemic that, as of October 28, 2020 (date of this writing), has been diagnosed in every country in the world, infected more than 44 million of people with a case fatality rate as high as ~2.6% with no signs of slowed spread 18 . By far the largest coronavirus pandemic to date, it has overwhelmed healthcare facilities and, in some cases, patients died from ARDS simply due to a shortage in respiratory support devices 19 . Lacking any effective treatments for COVID-19, the threat to global health security has necessitated the implementation of drastic quarantine measures in Accepted Article multiple countries: the Tokyo 2020 Olympics were delayed for the first time since World War 2;

Saudi Arabia has banned international visitors from making the annual religious pilgrimage (Hajj) of over 2 million Muslims for the first time since the 1918 Spanish flu pandemic; and international travel was essentially brought to a halt [20] [21] [22] .

The homological characterization of the receptor-restricting area of SARS-CoV-2 has been shown to be similar to that of SARS-CoV-1, suggesting that the pathogenesis of COVID-19 infection may be comparable to that of SARS 3 . Despite similarities between SARS-CoV-1 and SARS-CoV-2, the latter is causing a pandemic of a much greater proportion mainly due to two reasons. First, asymptomatic transmission occurs with SARS-CoV-2 whereas there have been no reported instances of transmission of SARS-CoV-1 before the onset of symptoms. Second, a much larger proportion of SARS-CoV-2 patients have relatively mild disease, increasing the difficulty of proactive containment.

While it is currently too early to accurately determine, the case fatality rate of COVID-19 is between 0.3% and 11.4% 23 . However, despite the rapid spread of the disease, COVID-19 remains relatively mild in children, with a hospitalization rate per 100,000 of 8.0 in children younger than 18 years versus 164.5 in adults 24 . The reason for the decreased severity of CoV infections in children as compared to adults remains unknown. Unlike other viruses such as influenza, it appears that younger children are more likely to recover from SARS-CoV-1 and -2 and MERS 2 . It is noteworthy that younger children are often more prone to severe respiratory infections caused by almost all other respiratory viruses.

The RNA genome of SARS-CoV-2 contains ~30,000 nucleotides and codes for at least four main structural proteins: envelope (E), membrane, nucleocapsid, and spike (S) proteins which facilitate entry into cells (Figure 1 

This article is protected by copyright. All rights reserved balance between ACE-1 and ACE-2 levels has been implicated in several diseases including cardiovascular disease, diabetes, hypertension, and acute respiratory distress syndrome 27 .

The ACE-2 protein is expressed in many tissues but primarily on the surface of type 2 alveolar epithelial cells in normal lungs 28 Upon inhalation of SARS-CoV-2, the S-protein binds to ACE-2 and creates a S-protein-ACE-2 complex (Figure 2A ). The affinity with which SARS-CoV-2 binds to ACE-2 is even higher than that of SARS-CoV-1 30 . The S-protein-ACE-2 complex is then proteolytically processed by type 2 transmembrane serine protease (TMPRSS2) leading to cleavage of ACE-2 and activation of the spike protein ( Figure 2B ) 27 , similar to the mechanism employed by influenza and human metapneumovirus, thus facilitating viral entry, through endocytosis, into the target cell. It has been suggested that cells in which ACE-2 and TMPRSS2 are simultaneously present are most susceptible to entry by SARS-

The endocytosed viral particles are actively maintained in an acidic environment, and the endosomal pH plays a key role in SARS-CoV-2 processing and replication. Hydroxychloroquine, an antimalaria drug, has the ability to increase endosomal pH and was postulated as a potential prophylactic medication to prevent SARS-CoV-2 infection. However, recent studies have shown that there is no clinical benefit of hydroxychloroquine in the prevention or treatment of COVID-19 32 .

Viral entry initiates the self-replication of the SARS-CoV-2 RNA and the virus spreads, presumably in the mucosal epithelium of the upper respiratory tract initially. Further multiplication occurs in the lower respiratory tract due to the abundance of expression of ACE-2 in type 2 alveolar cells 33 . When the majority of these cells are infected by SARS-CoV-2, a number of pathological processes occur that define ARDS. Pneumocyte destruction, decreased surfactant production, viremia-

This article is protected by copyright. All rights reserved induced cytokine storm, and impaired vascular permeability lead to diffuse alveolar damage and ineffective gas exchange 34 . In addition, destruction of ACE-2 causes a dysfunction of the reninangiotensin-system, further exacerbating the pulmonary edema and hypoxemia 35 .

It is apparent that several elements of the pathogenesis of COVID-19 can alter the severity of illness (Figure 3) . The age-dependence of any of these factors may help explain the decreased clinical severity seen in children with COVID-19.

There are many ways by which the pediatric immune system differs from that of adults. A discussion of the general decline of immune function known to be associated with aging, immunosenescence, is beyond the scope of this paper, and has been reviewed well by Aiello et al 36 .

Adults are more likely to have co-morbid conditions that weaken their immune system such as diabetes, hypertension, or cardiovascular conditions. In addition, adults are more likely to have a hyper-immune response that may cause destruction of healthy cells alongside the infected cells. The known factors necessary for SARS-CoV-2 infection and development of ARDS are reviewed below.

Children tend to have fewer outdoor activities, particularly in the winter months. SARS-CoV-1 and SARS-CoV-2 both started in China in the winter seasons of 2002 and 2019, respectively. In addition, children undertake less international travel, further decreasing their exposure risk, hence the fact that most children diagnosed with SARS, MERS, and COVID-19 were household contacts of adult cases 37, 38 . If this were the primary factor leading to decreased severity of CoVs in children, then one would expect other winter respiratory diseases to similarly spare children. However, we see quite the opposite for influenza, for example, where the highest burden of hospitalization is in the 5 to 17- year age group 39 . Moreover, as the SARS-CoV-2 pandemic continued into the spring and summer months of 2020, it became apparent that the rate of infection in children is similar to that in adults 38 .

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ACE-2 is the receptor for SARS-CoV-1, HCOV-NL63, and SARS-CoV-2. Expression of ACE-2 in the airway epithelium is dynamic and correlated with cellular differentiation 33 . The factors modifying the expression of ACE-2 are unknown to date. Studies have shown either no agedependent differences in ACE-2 levels in alveolar fluid of patients with ARDS 34 , or decreased ACE-2 expression with age 27, 40 . In one study assessing ACE-2 receptor expression in tissues of 224 patients with lung cancer, there were no significant disparities in ACE-2 gene expression between racial groups (Asian vs. Caucasian), age groups (older or younger than 60 years old), or gender groups (male vs females) 41 . However, smoking and angiotensin-receptor blockers (ARBs) have both been shown to upregulate ACE-2 gene expression 42 . Since children are less likely to have either exposure, that could at least partially explain the COVID-19 age discrepancy. Further epidemiologic data is required to determine whether nonsmoking adults not taking ARBs are as equally protected from COVID-19.

Muus et al. 43 analyzed the expression of ACE-2 in 164 donors. Although the number of children was small, the authors did note that ACE-2 expression in children was lower than that in adults 43 . It would be reasonable to surmise that if ACE-2 expression is reduced in children, then SARS-CoV-2 would be less likely to cause infection at any given viral load.

The TMPRSS2 gene, located at human chromosome 21q22.3, encodes an androgen-regulated type II transmembrane serine protease that is expressed in many tissues, including the prostate and lungs 44 . The physiologic role of TMPRSS2 is currently unknown and TMPRSS2 null mice have no reported abnormalities, but it has been implicated in prostate carcinogenesis 45, 46 . However, studies have shown that the interaction of TMPRSS2 with the S-protein-ACE-2 complex is essential for the spread of several viruses, including coronaviruses [47] [48] [49] . Since it appears that TMPRSS2 is dispensable for normal physiology, it thus constitutes an attractive antiviral target. Camostat mesylate, a drug approved for pancreatitis in Japan, is a TMPRSS2 inhibitor that blocks SARS-CoV-2 cell entry in vitro 49 . Clinical trials as a therapeutic agent for COVID-19 are underway 50 .

This article is protected by copyright. All rights reserved If ACE-2 receptor expression is not age-dependent, then TMPRSS2 expression may explain the pediatric reduced susceptibility to COVID-19. Cells expressing both ACE-2 and TMPRSS2 appear to rare in children 43 , although this study had a limited number of children. However, a PubMed search for "TMPRSS2" and "children" or "age" showed no other relevant results, and this avenue of research may be ripe for exploration.

In the early stages of CoV infection, neutrophils and activated epithelial and dendritic cells express a cluster of pro-inflammatory cytokines and other proteins such as IL-1β, IL-2R, IL-6, IL-8, IL-10, TNFα, C-reactive protein, ferritin, D-dimer, and procalcitonin 51 . This suggests that the overproduction of cytokines and chemokines ("cytokine storm") is associated with disease severity.

Previous animal studies on SARS-CoV-1 showed that peak disease severity coincided with maximal immune dysregulation rather than levels of peak viremia, indicating that the cytokine storm was more responsible for disease severity than the virus itself 52 . The cytokine storm in neonates is blunted since neonates are known to have reduced neutrophil number and function, and have been shown to have lower production of pulmonary inflammatory mediators during ARDS 53 . While neonatal neutropenia can predispose neonates to increased infection, the development of ARDS may be diminished, and there is clinical evidence that mortality from ARDS increases with age 53 .

Melatonin is a hormone produced by the pineal gland that has gained acceptance as a natural sleep aid in adults, but also has a variety of favorable biological and therapeutic activities. Melatonin exerts immunomodulatory effects through various pathways. 54 It inhibits nuclear factor kappa-B (NF-κB) activation, which plays a role in the activation of ARDS. It also stimulates nuclear factor erythroid 2-related factor 2 (Nrf2) which plays a role in the protection of various organs during oxidative stress. Melatonin also upregulates superoxide dismutase and downregulates nitric oxide synthase, exerting an anti-oxidative effect that may help attenuate the damage from high-oxygen therapy in those with severe ARDS. In addition, melatonin stimulates the production of natural killer cells and T and B lymphocytes, and animal studies have shown it augments the antigen-presenting

This article is protected by copyright. All rights reserved function of macrophages. There are no reports yet of the use of melatonin in the treatment of COVID-19, but it has been proposed as a potential adjuvant medication and theoretically shows promise 54 .

All mammals, including humans, produce melatonin from serotonin in response to retinal light-dark perception cycles and plays an important role in sleep organization 55 . Neonates develop photorhythmic production of melatonin by 9 weeks of age, and infants and children with a welldeveloped sleep cycle have a higher amplitude melatonin production cycle 56 . In fact, neonatal exposure to irregular light patterns can impair melatonin production in premature infants or infants who required neonatal intensive care, such as those with congenital heart disease, has been associated with poor outcome including sudden infant death syndrome 57 . The production of melatonin is known to decline with age due to a variety of reasons, and melatonin supplementation has been suggested to boost immune function in the elderly 58 .

Melatonin deficiency is related to suppressed immunocompetence and the increased levels of melatonin in children compared to adults may explain the age discrepancy seen in oxidative stress seen in ARDS 59,60 .

Several explanations have been proposed for why children differ from adults in SARS-CoV-2 infection rate and COVID-19 clinical severity, including potential protective factors in children or exacerbating factors in adults. We provide the first review of the main mechanisms of coronavirus infection and the age-variability of various components of SARS-CoV-2 infection and clinical severity. Reduced likelihood of exposure to SARS-CoV-2 was a preliminary theory but developing epidemiologic data suggests that children are at least as likely as adults to contract the virus. Reduced quantity or maturity of ACE-2 receptors or TMPRSS2 in children have been postulated, but current data are insufficient to support this theory. However, reduced cytokine storms in children and protection afforded by greater levels of immunomodulatory melatonin in the young can help explain the reduced COVID-19 severity.

Based on currently published data, we cannot conclusively determine that the childhood protection seen in COVID-19 is exclusively host dependent and future viral mutations may change the Accepted Article current age distribution of clinical severity. It is possible that children are simply less likely to develop severe coronavirus infection than adults due to biological factors that we described, or others that are yet to be defined. We believe there are multiple avenues to be explored to further explain the protective features of young age.

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