Discussion

Very few prospective studies were performed for bacterial/fungal complications of viral CAP in hospitalized adults. In this present study, the incidence, outcome, and risk factors of bacterial/fungal complications were investigated on prospective analysis of two consecutive years’ data from patients hospitalized with viral pneumonia. Using comprehensive microbiological tests, our data indicated that 34.5% of viral CAP-associated hospitalizations had evidence of bacterial/fungal superinfection. Apart from illness severity and clinical outcome, the incidence and types of bacterial/fungal infections with non-influenza respiratory viruses were also analogous to that associated with influenza, except for patients with SARS-CoV-2 infection. The high rate of superinfection complicated the clinical outcome of patients with viral pneumonia, while patients with superinfection presented significantly more in-hospital mortality than those without (22.7% vs 2.4%), especially when they came from non-ICU wards. In the follow-up period, the incidence of death within 120 days after admission was significantly higher in patients with bacterial/fungal superinfection than in those without. Furthermore, we found that PaO2/FiO2< 300, BUN ≥ 7.1 mmol/L, leukocytosis, and lymphocytopenia were independent risk factors of superinfection.
With the development of multiplex PCR assays for viruses, recent etiology studies have demonstrated an elevated proportion of viral infection (varies from 6% to 58%) and a declined detection of bacterial infection (7.8-24.8%)[2, 18, 32]. Flu was the first ranking viral pathogen, which was in line with other studies [5, 18]. Non-influenza viruses constituted exceeding one-quarter of the patients with viral pneumonia. Mixed viral infection was diagnosed in approximately one out of ten patients, indicating that virus co-infection may be the common phenomenon in viral pneumonia cases, and multiplexing of molecular assays could facilitate the identification of those patients [33]. In contrast to secondary bacterial infections, these mixed viral infections were not associated with a more severe disease course. Most studies published to date in viral CAP have included only one influenza season [5, 14]. The present consecutive 2-year prospective study demonstrated a regular seasonality profile in viral pneumonia, with a peak time in the period of November to March. In accordance with previous studies, we found comparable clinical characteristics among influenza, non-influenza, and mixed viral infection groups [18, 34]. Yet, patients with influenza had a higher rate of antiviral medication, and so perhaps the shorter length of hospital stay.
In our cohort, the severity of diseases, complications, and clinical outcomes of non-influenza pneumonia was analogous to that of influenza pneumonia, which were in line with previous studies [18, 35]. Accumulating evidence reported the notorious complication of bacterial/fungal superinfection followed by influenza infection [9], however, there were few clinical data on whether the risk of bacterial/fungal superinfection of non-influenza pneumonia was similar to that of influenza pneumonia. Our data demonstrated that 34.5% of patients had bacterial/fungal superinfection (ranging from 38%-50% by different viruses), except for SARS-CoV-2 infection. It was consistent with reports in previous studies patients with respiratory tract viral illness [15]. There was limited data on superinfection among patients with SARS-CoV-2 infection [36]. In our study, 5% of patients with SARS-CoV-2 had bacterial superinfection, which was relatively low compared to other respiratory viral infections. The immune system lacks the preexistence of a clonally expanded population of antigen-specific lymphocytes during the first year of the SARS-CoV-2 pandemic, the new virus may trigger an exacerbated inflammatory response that makes superinfection difficult to occur [37]. Notably, there were no significant differences in the rate of bacterial/fungal infection by a viral pathogen (χ2 =11.490, P = 0.244). This finding is original, since previous works that studied bacterial/fungal complications of viral CAP patients did not make any comparison between influenza and non-influenza viruses. These findings suggested equal attention should be given to pneumonia caused by non-influenza viruses.
Influenza infection may facilitate the nasopharyngeal colonization of opportunistic pathogens (such as A. baumannii , K. pneumoniae , Pseudomonas species or C. striatum ) that can gain better access to the lower respiratory tract by chemotaxis and cell motility genes and cause severe secondary infection [38] [39] [40]. It has reported that influenza-infected patients co-infected with P. aeruginosa and Aspergillus spp. were correlated with a significant mortality [37]. Among antibiotic-resistant microorganisms from patients in ICU, infection with K. pneumonia ,A. baumannii, and S. aureus was independently associated with a higher risk of death vs infection with another microorganism [41-44]. In the present study, the most frequent pathogen identified in the two years was P.aeruginosa andK. pneumonia , followed byA. baumannii and H. influenza , which was similar to the trend reported in a multicenter study [14]. Of note, fungal infection were documented 9 (9/255, 3.5%) patients. Up to 75% ofS. aureus were methicillin-resistant, 53% of K. pneumoniaresistant to β-lactam antibiotics, including third-generation cephalosporins and carbapenems, 33% of MDR P. aeruginosa and 31% of A. baumannii resistant to carbapenem, which may explain the higher mortality of patients with superinfection.
In CAP, viral-bacterial infection has been associated with a more complicated and worse outcome (e.g., a higher rate of hospital mortality or mechanical ventilation for > 7 days) than infections with viruses only, bacteria only, or no identified etiology [14, 45]. However, these conclusions are drawn from patients admitted to the ICU, few prospective studies were performed for the impact of bacterial/fungal complications on the overall population of patients with viral CAP. Our study clearly demonstrated that patients with bacterial/fungal superinfection had worse clinical outcomes as opposed to those without, whereas the case-fatality rate was found to be similar between patients with viral CAP and viral-bacterial CAP in patients from conventional wards in a recent study [16]. Such a gap probably owed to the older age and higher frequency of comorbidities of our cohort. Accumulating experimental data demonstrate that viral infection predisposes to bacterial superinfection by augmented bacterial adherence and dysregulation of the immune response [15, 46]. Unexpectedly, Martin-Loeches et al. [14] found the lack of association between appropriate antibiotic treatment and the outcome of viral-bacterial infection, partly underlining the major role of the immune response in the physiopathology of patients with severe influenza infection. In fact, respiratory viruses with both severe and mild disease courses such as influenza, RSV, and HRV may induce immunosuppression. Type I interferons productions are shown to increase after respiratory virus infection, which inhibits the phagocytosis of macrophages, decrease the recruitment of leukocyte to the lung, and increase levels of anti-inflammatory cytokines. For adaptive T cells, CD8+T cells are important in recovery from virulent influenza infections [47]. In our study, we found a profoundly higher leukocyte count, lower platelet count, and lymphocyte count. Of note, CD8+ T cells were decreased in the superinfection group indicating inadequate adaptive immune responses. These effects all may then contribute to the susceptibility toward various superinfections [48].
Given the incidence and worse outcome of superinfection in viral CAP, some predictors should be identified to determine the initiation of antimicrobial treatment. In patients with severe influenza pneumonia, PCT is shown to be a reasonably accurate marker for the detection of bacterial pneumonia. However, it might not be sufficient as a stand-alone marker for withholding antibiotic treatment [49, 50]. More complementary and simple-to-use markers are needed. Previously, some data suggest that the superinfection rate increased progressively with higher admission BUN levels among 12, 363 patients [51]. The current study confirmed that BUN ≥ 7.1 mmol/L was an important independent predictor of mixed viral-bacterial/fungal infection. We also found that hypoxemia might be a good predictor for superinfection, and reflect the state of alveolar ventilation dysfunction. Further research is needed to investigate the pathogenesis of hypoxemia in superinfection. Similar to study in adult patients with respiratory tract viral illness, leukocytosis was indecently associated with superinfection in patients with viral CAP. Respiratory virus-induced type I interferons mediate dysfunction of leukocytes that contributes to the increased susceptibility of various superinfections [48]. At the same time, we also found that lymphocytopenia were independent risk factors of superinfection. Animal model research indicated that lymphocyte deficiency would deactivate neutrophil and macrophages, resulting in impaired bacterial clearance [52]. Data from patients with influenza indicated that lymphopenia on admission was associated with the occurrence of nosocomial infection, which partially favored our result [10].
This study has some limitations. First, PCR detection of viruses in the NP swabs did not indicate the real pathogen of infection. However, low respiratory tract specimens for viral identification were obtained in 238 (93.3%) in our study, indicated that the causation was believable. Second, the diagnosis of bacterial infection was largely based on routine culture methods. The widely empirical use of antibiotics would result in underestimating the incidence of bacterial superinfection. Third, the nasopharyngeal microbiota may play a critical role in viral respiratory infection[39]. Because the background nasopharyngeal microbiota of these patients with viral CAP before hospitalization was unknown, it was hard to trace the isolated bacterial/fungal strains were due to nosocomial infection or the invading downward of existing colonizing strains [40]. Longitudinal studies are needed to address this issue.
In conclusion, superinfections may make up a significant proportion of total viral CAP cases, and the incidence and types of bacterial/fungal infections with non-influenza respiratory viruses were also analogous to that associated with influenza. These findings suggested equal attention should be given to pneumonia caused by non-influenza viruses. Mixed viral-bacterial/fungal infections were associated with a higher mortality rate than primary viral infection. An aggressive microbiological diagnostic approach should be initiated for those with a high risk of superinfection, and clinicians should consider the epidemiology of bacterial pathogens in this setting and consider empiric treatment for those who are critically ill.