Culture-Based Virus Isolation To Evaluate Potential Infectivity of Clinical Specimens Tested for COVID-19.

Real-time reverse transcription-PCR (RT-PCR) is currently the most sensitive method to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that causes coronavirus disease 2019 (COVID-19). However, the correlation between detectable viral RNA and culturable virus in clinical specimens remains unclear. Here, we performed virus culture for 60 specimens that were confirmed to be positive for SARS-CoV-2 RNA by real-time RT-PCR. The virus could be successfully isolated from 12 throat and nine nasopharyngeal swabs and two sputum specimens. The lowest copy number required for virus isolation was determined to be 5.4, 6.0, and 5.7 log10 genome copies/ml sample for detecting the nsp12, E, and N genes, respectively. We further examined the correlation of genome copy number and virus isolation in different regions of the viral genome, demonstrating that culturable specimens are characterized by high copy numbers with a linear correlation observed between copy numbers of amplicons targeting structural and nonstructural regions. Overall, these results indicate that in addition to the copy number, the integrity of the viral genome should be considered when evaluating the infectivity of clinical SARS-CoV-2 specimens.

Macrolide-resistant Mycoplasma pneumoniae in children in Taiwan.

The aim of this study was to estimate the prevalence of macrolide-resistant Mycoplasma pneumoniae in Taiwan and to compare the clinical courses of pediatric patients with macrolide-resistant (MR) M. pneumoniae and macrolide-susceptible (MS) M. pneumoniae infection. Patients were among the children admitted to Chang Gung Children's Hospital with mycoplasmal pneumonia between February and December 2011. Detection for macrolide resistance was performed after informed consent was obtained. We retrospectively reviewed medical records and compared the clinical courses of two groups of patients of 73 children enrolled into our study. The rate of macrolide resistance in M. pneumoniae was 12.3 %. Longer hospital stay was observed in the MR patients than MS patients [median, 7 days vs. 5 days (P = 0.019)]. Clinical features or radiographic or laboratory findings are not helpful to differentiate MR from MS mycoplasmal pneumonia. Early diagnosis of MR mycoplasmal pneumonia is crucial for the best management of these patients and obviates the need for extensive etiological searches of these nonresponding cases.

Impact of patient characteristics on innate immune responses and inflammasome activation in ex vivo human lung tissues infected with influenza A virus.

Background: Influenza A virus (IAV) infection poses a persistent global health challenge, necessitating a nuanced grasp of host immune responses for optimal interventions. While the interplay between aging, immunosenescence, and IAV is recognized as key in severe lower respiratory tract infections, the role of specific patient attributes in shaping innate immune reactions and inflammasome activity during IAV infection remains under-investigated. In this study, we utilized an ex vivo infection model of human lung tissues with H3N2 IAV to discern relationships among patient demographics, IAV nucleoprotein (NP) expression, toll-like receptor (TLR) profiles, PD-1/PD-L1 markers, and cytokine production. Methods: Our cohort consisted of thirty adult patients who underwent video-assisted thoracoscopic surgery during 2018-2019. Post-surgical lung tissues were exposed to H3N2 IAV for ex vivo infections, and the ensuing immune responses were profiled using flow cytometry. Results: We observed pronounced IAV activity within lung cells, as indicated by marked NP upregulation in both epithelial cells (P = 0.022) and macrophages (P = 0.003) in the IAV-exposed group relative to controls. Notably, interleukin-2 levels correlated with variations in TLR1 expression on epithelial cells and PD-L1 markers on macrophages. Age emerged as a modulating factor, dampening innate immune reactions, as evidenced by reduced interleukin-2 and interferon-γ concentrations (both adjusted P < 0.05). Intriguingly, a subset of participants with pronounced tumor necrosis factor-alpha post-mock infection (Cluster 1) showed attenuated cytokine responses in contrast to their counterparts in Cluster 2 and Cluster 3 (all adjusted P < 0.05). Individuals in Cluster 2, characterized by a low post-mock infection NP expression in macrophages, exhibited reduced variations in both NP and TLR1-3 expressions on these cells and a decreased variation in interleukin-2 secretion in comparison to their Cluster 3 counterparts, who were identified by their elevated NP macrophage expression (all adjusted P < 0.05). Conclusion: Our work elucidates the multifaceted interplay of patient factors, innate immunity, and inflammasome responses in lung tissues subjected to ex vivo H3N2 IAV exposure, reflecting real-world lower respiratory tract infections. While these findings provide a foundation for tailored therapeutic strategies, supplementary studies are requisite for thorough validation and refinement.

Transcriptome Differences in Normal Human Bronchial Epithelial Cells in Response to Influenza A pdmH1N1 or H7N9 Virus Infection.

Avian influenza A (H7N9) virus infections frequently lead to acute respiratory distress syndrome and death in humans. The emergence of H7N9 virus infections is a serious public health threat. To identify virus-host interaction differences between the highly virulent H7N9 and pandemic influenza H1N1 (pdmH1N1), RNA sequencing was performed of normal human bronchial epithelial (NHBE) cells infected with either virus. The transcriptomic analysis of host cellular responses to viral infection enables the identification of potential cellular factors related to infection. Significantly different gene expression patterns were found between pdmH1N1- and H7N9-infected NHBE cells. In addition, the H7N9 virus infection induced strong immune responses, while cellular repair mechanisms were inhibited. The differential expression of specific factors observed between avian H7N9 and pdmH1N1 influenza virus strains can account for variations in disease pathogenicity. These findings provide a framework for future studies examining the molecular mechanisms underlying the pathogenicity of avian H7N9 virus.