Respiratory syncytial virus (RSV) infects small foci of respiratory epithelial cells via infected droplets. Infection induces expression of type I and III interferons (IFNs) and proinflammatory cytokines, the balance of which may restrict viral replication and affect disease severity. We explored this balance by infecting two respiratory epithelial cell lines with low doses of recombinant RSV expressing green fluorescent protein (rgRSV). A549 cells were highly permissive, whereas BEAS-2B cells restricted infection to individual cells or small foci. After infection, A549 cells expressed higher levels of IFN-β-, IFN-λ-, and NF-κB-inducible proinflammatory cytokines. In contrast, BEAS-2B cells expressed higher levels of antiviral interferon-stimulated genes, pattern recognition receptors, and other signaling intermediaries constitutively and after infection. Transcriptome analysis revealed that constitutive expression of antiviral and proinflammatory genes predicted responses by each cell line. These two cell lines provide a model for elucidating critical mediators of local control of viral infection in respiratory epithelial cells. Airway epithelium is both the primary target of and the first defense against respiratory syncytial virus (RSV). Whether RSV replicates and spreads to adjacent epithelial cells depends on the quality of their innate immune responses. A549 and BEAS-2B are alveolar and bronchial epithelial cell lines, respectively, that are often used to study RSV infection. We show that A549 cells are permissive to RSV infection and express genes characteristic of a proinflammatory response. In contrast, BEAS-2B cells restrict infection and express genes characteristic of an antiviral response associated with expression of type I and III interferons. Transcriptome analysis of constitutive gene expression revealed patterns that may predict the response of each cell line to infection. This study suggests that restrictive and permissive cell lines may provide a model for identifying critical mediators of local control of infection and stresses the importance of the constitutive antiviral state for the response to viral challenge.

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Background Necrotizing soft tissue infections (NSTI) are characterized by fulminant tissue destruction and high mortality rates. Primary treatment is surgical debridement and antibiotics; hyperbaric oxygen therapy (HBOT) may be considered as adjuvant therapy. HBOT for NSTI remains controversial given the lack of quality evidence. HBOT is frequently used for NSTI at our institution; however, the early COVID-19 pandemic forced a significant decrease in its utilization. This practice change created an unusually high proportion of NSTI patients who did not receive HBOT at our institution and thus provided an opportunity to compare outcomes amongst NSTI patients who received HBOT against those who did not. Methods We performed a retrospective cohort study of patients aged 18 years and older admitted with the diagnosis of NSTI between January 2018 and December 2020, a period that included the first 6 months of the pandemic (Figure 1). We collected data on patient demographics, comorbidities, infection location, pathogen, severity of illness (APACHE II score), whether HBOT was provided and number of dives. Outcomes included 90-day mortality, amputations, inpatient antibiotic days, ventilator days, and hospital length of stay. Figure 1Proportion of NSTI patients admitted to R Adams Cowley Shock Trauma Center who received HBOT between January 2018 through December 2020. Proportions were averaged over six month blocks. Results 253 patients were included of whom 143 (56%) received HBOT and 110 (43.3%) did not. Baseline characteristics were similar between the groups except for Wound Surface Area (WSA) and APACHE II score (Table 1). Length of stay was significantly longer in HBOT patients (15.3 days vs 11.7 days p=0.005, Table 2). Fewer HBOT patients had amputations (7.5% vs 21% p=0.007) and fewer died within 90 days of admission (6.3% vs 14.5% p=0.029). When stratifying patients by APACHE II score, HBOT patients with APACHE II scores > 18 had significantly lower mortality than non-HBOT patients (9.7% vs 32.4% p=0.035). Conclusion In this population of NSTI patients who underwent surgical debridement, HBOT was associated with lower incidence of amputation, and lower mortality. Patients with APACHE II scores >18 who received HBOT had lower mortality than those who did not. A prospective study should further evaluate the impact of HBOT on mortality and amputations. Disclosures All Authors: No reported disclosures.

The host innate response contributes to the severity of disease caused by Respiratory Syncytial Virus (RSV). The primary target of RSV is the airway epithelium. The host becomes infected by inhaling droplets containing viral particles, which then infect small foci of cells. Infected cells then express types I and III interferons (IFN) and pro-inflammatory cytokines to alert neighboring epithelial and myeloid cells of danger. Despite their inherent differences, A549 (carcinoma, type II alveolar epithelium) and BEAS-2B (transformed bronchial epithelium) cells are often used interchangeably to study RSV-epithelial interactions. We compared these two cell lines by infecting them with low MOIs of RSV expressing GFP (rgRSV). We measured expression of types I and III IFNs, interferon stimulated genes (ISGs), pro-inflammatory cytokines and signaling intermediaries. We found that BEAS-2B cells contained rgRSV within foci of ~10–15 cells, but all A549 cells were infected by 48h. Both cell lines highly expressed IFN-β, IFN-λ1 and -λ2, but expression was greater in the A549s. Despite lower levels of IFN expression, BEAS-2Bs expressed higher levels of most classic antiviral ISGs (ISG15, MX1, PKR) and critical TLRs, RLRs and IFN receptors. In contrast, A549 cells expressed higher levels of NF-kB associated genes (CCL2, CCL5, CXCL8). Our data suggests that a balance between expression of NF-kB genes and ISGs may determine local control of RSV infection by respiratory epithelial cells.