The prevalence of TTV infection in children with ARI was high in our study, reaching 38% of cases. It was associated with an infection by another respiratory virus in practically all cases. The children with detection of TTV in NPA more frequently had pneumonia on radiography, and a clinical course with fever and frequent admission to the PICU. Our results appear to indicate that TTV infection is a marker of impaired immune response and a facilitator of respiratory infections and greater severity. The study of the immune response of our patients has shown that TTV+ children have a higher abundance of the filaggrin protein in NPA, with a lower gene expression. This result could mean that there is an alteration of the epithelial barrier in the TTV+ group, which is associated with liberation of this molecule to the supernatant of their NPA. Filaggrin is a protein involved in the integrity of the epithelial barrier and could play a role in allowing or favoring these respiratory infections22.
Chronic TTV infections have been reported in healthy individuals, and colonization is considered to begin very early in life, possibly transplacental in some cases, or in the family environment by the fecal-oral route23,24. Colonization increases with age. Although the prevalence of TTV in blood samples has been more often studied than in respiratory secretions, the respiratory route is considered to be a frequent path of dissemination and contagion15. The role of TTV in pediatric respiratory infections is poorly understood. Maggi F et al15, in the most important study in this regard, found that the presence of TTV was associated with bronchopneumonia and other respiratory viral infections, similarly to our study. They hypothesized that coinfection could increase the severity of other respiratory infections. Our study supports this hypothesis, and we observed that our TTV+ patients had coinfection with other CRVs and a more severe clinical outcome.
To study the respiratory secretions of these children in terms of their immune response is tempting, to better understand the pathophysiology of this association. The TTV+ patients had higher expression of TLR3 and IL33 in the NPA. Although they were not statistically significant results, this lack of significance might be due to the small sample analyzed; however, it suggests that in the group infected by TTV there is a higher detection of genes upregulated by viral infection. The increased expression of TLR3 and IL-33 in NPA has been previously described by our group, primarily in children with bronchiolitis, and this was the main diagnosis in our cohort25. They have also been overexpressed in animal models of asthma exacerbation26. Some studies have shown that viruses like RSV or HRV induce IL-33 synthesis by damaging epithelium in pulmonary diseases27.
The most important finding is that the frequency of TTV+ children that expressed filaggrin mRNA was lower than in the TTV-, and that the filaggrin protein was nevertheless increased in NPA from the TTV+. Filaggrin is a protein involved in the integrity of the epithelial barrier and its mutations have been associated with atopic dermatitis or ichthyosis28,29. In our TTV+ patients, filaggrin is likely underexpressed, and the increase of filaggrin in NPA is part of the ongoing repair mechanism to maintain barrier function, triggered to respond to the epithelial damage caused by viral infection. Filaggrin release from keratohyalin granules into the keratinocyte cytoplasm is a main event in the cornification process, and it is critical for skin barrier function. The increase of filaggrin observed in the supernatant might be explained by damage to the cellular integrity and the consequent liberation of intracellular filaggrin to the extracellular medium30.
Whether the presence of TTV in respiratory samples is a marker of a deficient immune response or is a consequence of it is difficult to elucidate. We cannot rule out that the previous presence of chronic TTV infection predisposes patients to infection by other CRVs and to a greater severity.
On the other hand, the association of altered filaggrin with the possible development of asthma, as well as increased Th-2 immune response in our patients who clinically had a diagnosis of bronchiolitis or recurrent wheezing, suggest that the presence of TTV infection could have a role in respiratory diseases. Several studies have suggested that TTV plays a role in the development and/or exacerbation of respiratory diseases in childhood such as asthma31. It has been postulated that TTV has a role in respiratory dysfunction, either alone or synergistically with other viruses, and could act as an enhancer of inflammation systemically or at specific body sites, such as the upper and lower airways32.
A limitation of our study is that it was performed during a pandemic. Thus, and as has been previously mentioned, the sample size was small, and although plausible, it does not allow for firm conclusions to be drawn about the immune response. However, it provides a comprehensive view of TTV respiratory infection, which has been little studied so far, in a homogeneous population of infants.
In conclusion, TTV infection is common in children with viral respiratory infections and could be associated with pneumonia and greater severity, as well as an alteration in the epithelial barrier due to low filaggrin gene expression. Larger prospective studies will be able to unravel whether TTV favors respiratory viral infections or is a marker of impaired immune response.