Post-COVID syndrome (PCS) develops in individuals following SARS-CoV-2 infection, typically three months after the onset of COVID-19, with symptoms persisting for at least two months that cannot be explained by an alternative diagnosis [1].

Previous SARS and MERS epidemics left recovered patients with persistent fatigue, chronic dyspnea, and a reported decline in quality of life. A constellation of such diverse clinical symptoms, collectively termed PCS, has been described in a significant proportion of patients who recovered from SARS-CoV-2-induced COVID-19, despite the cessation of viral replication [2]. It is estimated that at least 10% of COVID-19 survivors develop PCS [3].

In the Russian Federation, as of July 1, 2021, enhanced clinical follow-up protocols for COVID-19 survivors were implemented, enabling comprehensive screening for potential complications. In order to facilitate monitoring, treatment adjustment, and mitigation of risk factors, including those associated with prior COVID-19, enhanced follow-up examinations are recommended not only for patients who experienced severe disease but also for all individuals reporting post-COVID sequelae, regardless of initial disease severity. Through this program, patients can identify coronavirus-related health issues at the earliest stage and implement timely interventions [4]. The immune system plays a critical role in preventing potential complications. Therefore, it is of particular interest to evaluate alterations in innate and adaptive immunity in patients with moderate COVID-19 six months after recovery and their contribution to PCS development.

The study was aimed at investigating immunological parameters in patients recovered from moderate COVID-19 during the post-COVID period.

The study included 60 individuals discharged from Mono-Infectious Hospital No. 1 of the Semashko City Clinical Hospital in Rostov-on-Don, where they had been treated with a confirmed diagnosis of “Moderate COVID-19 coronavirus infection, complicated by bilateral interstitial pneumonia”. The mean age of the patients was 54.3 ± 15.6 years, with 29 men and 31 women. Age differences between the two gender groups were statistically insignificant (p > 0.05). A comparison group of 20 age-matched healthy volunteers was included. All participants provided written informed consent. The clinical study was conducted in accordance with the World Medical Association’s Declaration of Helsinki “Ethical Principles for Medical Research Involving Human Subjects” and the Clinical Practice Guidelines of the Russian Federation approved by Order No. 266 of the Ministry of Health of the Russian Federation (June 19, 2003).

Clinical and laboratory parameters were assessed one and six months after hospital discharge. All subjects underwent complete blood count (CBC), including erythrocyte and hemoglobin levels, total leukocyte count, and leukocyte differential. Biochemical analysis included C-reactive protein (CRP), ALT, AST, albumin, urea, creatinine, glucose, LDH, ferritin, total protein, lactate, amylase, and bilirubin. Lymphocyte surface markers were evaluated using flow cytometry. CD3, CD4, CD8 T-cell subsets, CD19 B-cell, and CD16 natural killer cells were quantified using a Cytomics FC 500 flow cytometer (Beckman Coulter, USA). Serum levels of IgA, IgM, IgG, interferon-gamma, and IL-6 and IL-10 cytokines were measured by enzyme-linked immunosorbent assay (ELISA). Statistical analysis was performed using STATISTICA 10 (StatSoft Inc., USA). Descriptive statistics for quantitative variables are presented as median and interquartile range (IQR; 25th–75th percentiles), denoted as Me [LQ; UQ]. Group comparisons were made using the nonparametric Mann-Whitney U-test, with statistical significance set at p < 0.05.

Patients discharged from the mono-infectious hospital with a diagnosis of “Confirmed moderate COVID-19 coronavirus infection, complicated by bilateral polysegmental interstitial pneumonia, respiratory failure (RF) grade 0–I,” had an average hospital stay of 12.85 ± 2.75 days.

One month after discharge, the majority of patients reported pronounced asthenia, including weakness, reduced appetite, and decreased work capacity (65% of patients remained under outpatient follow-up with a general practitioner). A significant proportion of the examined individuals (36 patients, 60%) complained of palpitations and blood pressure fluctuations. Forty-seven patients (78%) reported exertional dyspnea and cough (mostly dry and unproductive). Twelve patients (20%) experienced intermittent episodes of dizziness and paresthesia.

 Comparative analysis of CBC parameters one month post-recovery/discharge revealed no significant differences compared to healthy donors. Evaluation of biochemical markers showed statistically significant differences only in elevated CRP levels (10.4 [ 14.7; 8.5] vs. 2.8 [ 1.2; 4.7]) and LDH (338 [ 240; 429] vs. 84 [ 14; 218]). A significant increase was also observed in ALT levels (60.5 [ 41.2; 108.5] vs. 23.0 [ 19.8; 56.8]), likely due to hepatotoxic medications administered both during hospitalization and after discharge (e.g., anticoagulant therapy).

Assessment of innate and humoral immunity one month post-recovery demonstrated a statistically significant increase in the relative and absolute counts of cellular immune components: CD3+, CD4+ T-lymphocytes, and CD8+ T-lymphocytes. In contrast, the humoral arm showed a marked reduction in B-lymphocytes, while IgG and IgA levels were elevated (Table 1).

Six months after COVID-19 recovery, 46 patients (76.7%) continued to report significant asthenia. Sleep disturbances (insomnia, nightmares) were present in 21 patients (35%), intermittent headaches in 15 patients (25%), and persistent cough in 18 patients (30%). Half of the cohort (29 patients, 48.3%) complained of new-onset tachycardia and elevated blood pressure, which had not been present prior to the infection.

For screening anxiety and depression in study participants, the Hospital Anxiety and Depression Scale (HADS) was used. This is a “subjective” assessment tool. Among the surveyed patients, 26 individuals (43.3%) scored 8–10 points, indicating subclinically significant anxiety/depression, while 34 patients (60%) scored 0–7 points (normal range).

Comparative analysis of CBC parameters six months post-recovery/hospital discharge revealed a statistically significant decrease in leukocyte levels compared to healthy donors. All other CBC parameters showed no significant differences (Table 2).

Evaluation of biochemical markers demonstrated minor but significant elevations in predictors of severe disease progression, such as CRP and LDH, compared to the control group (Table 3). No other biochemical parameters exhibited significant differences.

Six months post-infection, patients continued to show a statistically significant increase in relative and absolute counts of T-lymphocytes, persistent imbalance in humoral immunity, with reduced absolute and relative B-lymphocyte counts and elevated IgA levels (Table 4).

Comparison of immunological parameters at one and six months post-infection revealed a trend toward normalization of cellular immunity, characterized by decreased relative counts of total T-lymphocytes, CD4+ T-lymphocytes, and both relative/absolute CD8+ T-lymphocyte levels. In humoral immunity, increased B-lymphocyte counts and normalization of serum immunoglobulin levels were observed (Table 5).

The severity of COVID-19 can be explained by its multi-organ involvement [5]. Current evidence suggests that COVID-19 severity results from both the direct cytopathic effects of the virus and tissue damage caused by the immune response to viral infection. Cohort study findings at six months post-acute infection indicate that recovered patients primarily experienced fatigue/muscle weakness, sleep disturbances, and anxiety/depression [6]. These results highlight the critical need to investigate the pathogenesis of long COVID and develop effective interventions to mitigate its risks. COVID-19 patients exhibit lymphopenia and significant depletion of CD8+ T-lymphocytes and NK cells. The prior research has documented immunological alterations in patients with moderate COVID-19 during the acute phase, correlating with varying degrees of lung involvement. Key findings included reduced T-lymphocyte counts, dysimmunoglobulinemia, and elevated IL-6 levels in patients with more extensive pulmonary lesions [7]. These SARS-CoV-2-induced immune response alterations lead to adaptive immunity suppression: diminished proliferative and antiviral activity, culminating in profound immunosuppression [8]. Even six months post-infection, monitoring these changes remains essential for patient rehabilitation.

Analysis of cellular and humoral immunity parameters during the post-COVID period revealed the following results: at one month, cellular immunity alongside humoral imbalance (dysimmunoglobulinemia) was pronouncedly activated; at six months, T-lymphocyte maturation activity was persistently declining and trending toward normalization. Absolute/relative B-lymphocyte counts were significantly reduced while IgA levels were elevated, necessitating ongoing immunological surveillance.

COVID-19, even after the cessation of viral replication in the human body, induces a range of symptoms collectively referred to as PCS. Analysis of laboratory parameters revealed that following moderate COVID-19, general clinical and biochemical markers returned to reference values within six months. However, a distinctive feature of this disease is the persistence of immune status alterations even after an extended period. These include sustained activation of T-lymphocyte maturation processes and suppression of the humoral immune response accompanied by dysimmunoglobulinemia. Clinically, these changes may manifest as increased susceptibility to respiratory viral infections, prolonged disease course, and the necessity for continued immunological monitoring.