**6. Discussion**

This retrospective study in LTRs with COVID-19 showed lung function decline after COVID-19 in most patients with moderate COVID-19. In most patients with mild COVID-19 evolution, the lung function evolution was not affected. Patients who showed lung function decline after COVID-19 in the first month did not recover in the following year, and in this group of patients, most showed a further lung function deterioration. After 12 months, 10 patients (56%) showed a decreased FEV1 as compared to pre-COVID FEV1 measurements, with a FEV1 range of −4.98% to −40.4%. In four patients (*n* = 22%), the FEV1 decrease was 5–10%, three patients (*n* = 17%) lost ≥ 10–20%, and three patients (*n* = 17%) lost ≥ 20%. Most patients did not develop DSA even one year post-COVID.

Although CLAD-BOS was frequently diagnosed after moderate COVID-19, we had no patients with CLAD-RAS or a mixed phenotype. These results were in line with a retrospective multicenter study that collected data from three Dutch transplant centers and included 74 LTRs that showed a significantly lower lung function that remained significantly lower compared to the pre-COVID-19 values [23].

The so-called wild type of the SARS-CoV-2 was first demonstrated in China at the end of 2019. Relevant virus mutations were the Alpha variant (B.1.1.7, first demonstrated in the United Kingdom in September 2020), followed by the Beta variant (mutation E484K, first seen in South Africa in May 2020), the Gamma variant (P.1, initially detected in Brazil, in November 2020), the Delta variant (B.1.617.2, initially detected in India in May 2021), and the Omicron variant (November 2021).

Although a genotyping PCR was not initially performed at our hospital, the abovedescribed patients were studied between 1 February 2020 and 1 March 2021 and probably mainly suffered from the wild type, Alpha variant, Beta variant, or the Gamma variant of SARS-CoV-2 based on the predominant strains detected in this period.

The numbers of affected patients at that time (beginning of the pandemic) were relatively small. The second wave of the pandemic in Switzerland was in October 2020 [24]. At the beginning of the pandemic's spread in Switzerland, specifically in March 2020, there were only 3000 COVID-19 patients diagnosed despite widespread testing, but this rapidly increased to over 500,000 in January 2021 [25].

At the time of this study, patients had not yet received the vaccinations because they were not yet available. The approval of the first COVID-19 mRNA vaccine, called BNT162b2 (Pfizer BioNTech) [26], was on 19 December 2020, followed by the COVID mRNA vaccine by Moderna, which was approved on 12 January 2021 in Switzerland [27]. Moreover, at this time there were no clear guidelines on how to deal with immunosuppression in LTRs with COVID-19 and vaccination uptake was slightly delayed due to prioritization of elderly persons at the beginning of the vaccine roll-out.

We now know that vaccination in lung transplant recipients is a key strategy that reduces the risk of severe COVID-19 and hospitalization [28]. Vaccination is also now considered as an indirect treatment in the prevention of CLAD BOS in LTRs due to SARS-CoV-2 infection. Unfortunately, two problems in vaccination of LTRs have become evident. The first problem is that LTRs have a blunted humoral and cellular immune response after COVID-19 vaccination [29]. The second is a shorter duration of the protective effects of the vaccine [29–32].

A weaker immune response was demonstrated in immunosuppressed transplant recipients who received the trivalent influenza vaccine; these patients showed significantly lower antibody titers [33]. Another study also showed a weaker response in immunosuppressed heart transplant recipients after pneumococcal vaccination [34]. In LTRs, the standard therapy is a triple immunosuppression in which most patients receive a combination of a calcineurin inhibitor (CNI), a mycophenolate derivative such as mycophenolate mofetil (MMF), and a corticosteroid (typically prednisone or prednisolone) [35]. All types of immunosuppressive drugs have different mechanisms of action that when combined will severely blunt the immune response. CNI blocks T-cell activation and proliferation, MMF impairs the proliferation of B and T lymphocytes and increases apoptosis, while corticosteroids mainly affect T lymphocytes by impairing their development, survival, activation, and migration [36].

Our study had some obvious limitations, namely the small number of patients, the single-center experience, and the retrospective design of the study. The data should therefore be interpreted with caution; for firm conclusions, further studies are needed.

In other viral infections in LTRs, acute rejection and chronic lung allograft dysfunction are well-known complications as well [37]. Allograft dysfunction is not only caused by direct effects of viral replication, but also by immunologically mediated lung injury [37]. The exact mechanisms are only partially understood.

In conclusion, this study suggested a potential relationship between SARS-CoV-2 infection and CLAD. More specifically, our hypothesis was that the risk of the development of CLAD-BOS was higher in the LTRs with moderate COVID-19 compared to those with mild COVID-19. Moreover, the decline in FEV1 could already be seen as soon as 1 month after COVID, with an additional FEV1 deterioration in the following months.

In LTRs, emphasis on prevention of COVID-19 by minimizing exposure and widespread use of vaccinations is certainly warranted because the increased severity of SARS-CoV-2 infections appears to increase the risk of CLAD development. The exact role of vaccination in LTRs requires further studies that include long-term follow-up data on FEV1 evolutions for different clinical situations because the current serological data show a suboptimal antibody response in these immunosuppressed patients. Thus, the protective effects regarding COVID-19 severity and disease course are not well studied to date.

**Author Contributions:** Conceptualization, R.H. and M.M.S.; methodology, R.H. and M.M.S.; formal analysis, R.H. and M.M.S.; data curation, R.H.; writing—original draft preparation, R.H.; writing review and editing, R.H. and M.M.S.; supervision, M.M.S. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Institutional Review Board Statement:** Swiss Medical Ethics Committee (Swissethics No. 2021-00293).

**Informed Consent Statement:** Informed consent was obtained from all subjects involved in the study.

**Data Availability Statement:** All the data are available from the corresponding author upon reasonable request.

**Acknowledgments:** We would like to thank Julian Stuart for reading and correction of the English language.

**Conflicts of Interest:** The authors declare no conflict of interest.
