Human respiratory syncytial virus (RSV) is the most common cause of viral bronchiolitis and pneumonia in infants and children worldwide. Inflammation induced by RSV infection is responsible for its hallmark manifestation of bronchiolitis and pneumonia. The cellular debris created through lytic cell death of infected cells is a potent initiator of this inflammation. Macrophages are known to play a pivotal role in the early innate immune and inflammatory response to viral pathogens. However, the lytic cell death mechanisms associated with RSV infection in macrophages remains unknown. Two distinct mechanisms involved in lytic cell death are pyroptosis and necroptosis. Our studies revealed that RSV induces lytic cell death in macrophages via both of these mechanisms, specifically through the ASC (Apoptosis-associated speck like protein containing a caspase recruitment domain)-NLRP3 (nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3) inflammasome activation of both caspase-1 dependent pyroptosis and receptor-interacting serine/threonine-protein kinase 3 (RIPK3), as well as a mixed lineage kinase domain like pseudokinase (MLKL)-dependent necroptosis. In addition, we demonstrated an important role of reactive oxygen species (ROS) during lytic cell death of RSV-infected macrophages.

BackgroundSalmonella enterica serovar Enteritidis (S. Enteritidis) is a human and animal pathogen that causes gastroenteritis characterized by inflammatory diarrhea and occasionally an invasive systemic infection. Salmonella pathogenicity islands (SPIs) are horizontally acquired genomic segments known to contribute to Salmonella pathogenesis. The objective of the current study was to determine the contribution of SPI-13 to S. Enteritidis pathogenesis.MethodsWe deleted the entire SPI-13 (∆SPI-13) from the genome of S. Enteritidis CDC_2010K_0968 strain isolated from a human patient during the 2010 egg-associated outbreak in the US. The kinetics of infection of the wild-type parent and the ∆SPI-13 were compared in orally challenged day-old chickens and streptomycin pre-treated mice. The degree of intestinal inflammation and the survival of mutant strain within the avian (HD11) and murine (RAW264.7) macrophages were also determined.ResultsThe deletion of the SPI-13 resulted in impaired infection kinetics of S. Enteritidis in streptomycin pre-treated mice which was characterized by significantly lower (P < 0.05) viable counts in the ceca, liver and spleen, impaired ability to induce intestinal inflammation and reduced survival within murine macrophages. Conversely, there were no significant differences in the infection kinetics of ∆SPI-13 in day-old chickens in any of the organs tested and the survival of ∆SPI-13 within chicken macrophages remained unaltered.ConclusionsThe results of this study show that SPI-13 contributes to the pathogenesis of S. Enteritidis in streptomycin pre-treated mice but not in day-old chickens and raises the possibility that SPI-13 may play a role in pathogenesis and the host adaptation/restriction of Salmonella serovars.

Critically ill COVID-19 patients infected with SARS-CoV-2 display signs of generalized hyperinflammation. Macrophages trigger inflammation to eliminate pathogens and repair tissue, but this process can also lead to hyperinflammation and resulting exaggerated disease. The role of macrophages in dysregulated inflammation during SARS-CoV-2 infection is poorly understood. We used SARS-CoV-2 infected and glycosylated soluble SARS-CoV-2 Spike S1 subunit (S1) treated THP-1 human-derived macrophage-like cell line to clarify the role of macrophages in pro-inflammatory responses. Soluble S1 upregulated TNF-α and CXCL10 mRNAs, and induced secretion of TNF-α from THP-1 macrophages. While THP-1 macrophages did not support productive SARS-CoV-2 replication, virus infection resulted in upregulation of both TNF-α and CXCL10 genes. Our study shows that S1 is a key viral component inducing inflammatory response in macrophages, independently of virus replication. Thus, virus-infected or soluble S1-activated macrophages may become sources of pro-inflammatory mediators contributing to hyperinflammation in COVID-19 patients.

bDimethyl adenosine transferase (KsgA) performs diverse roles in bacteria, including ribosomal maturation and DNA mismatch repair, and synthesis of KsgA is responsive to antibiotics and cold temperature. We previously showed that a ksgA mutation in Salmonella enterica serovar Enteritidis results in impaired invasiveness in human and avian epithelial cells. In this study, we tested the virulence of a ksgA mutant (the ksgA::Tn5 mutant) of S. Enteritidis in orally challenged 1-day-old chickens. The ksgA:: Tn5 mutant showed significantly reduced intestinal colonization and organ invasiveness in chickens compared to those of the wild-type (WT) parent. Phenotype microarray (PM) was employed to compare the ksgA::Tn5 mutant and its isogenic wild-type strain for 920 phenotypes at 28°C, 37°C, and 42°C. At chicken body temperature (42°C), the ksgA::Tn5 mutant showed significantly reduced respiratory activity with respect to a number of carbon, nitrogen, phosphate, sulfur, and peptide nitrogen nutrients. The greatest differences were observed in the osmolyte panel at concentrations of >6% NaCl at 37°C and 42°C. In contrast, no major differences were observed at 28°C. In independent growth assays, the ksgA::Tn5 mutant displayed a severe growth defect in high-osmolarity (6.5% NaCl) conditions in nutrient-rich (LB) and nutrient-limiting (M9 minimum salts) media at 42°C. Moreover, the ksgA::Tn5 mutant showed significantly reduced tolerance to oxidative stress, but its survival within macrophages was not impaired. Unlike Escherichia coli, the ksgA::Tn5 mutant did not display a cold-sensitivity phenotype; however, it showed resistance to kasugamycin and increased susceptibility to chloramphenicol. To the best of our knowledge, this is the first report showing the role of ksgA in S. Enteritidis virulence in chickens, tolerance to high osmolarity, and altered susceptibility to kasugamycin and chloramphenicol.

Understanding host-virus interactions is essential for the development of effective interventions against respiratory syncytial virus (RSV), a paramyxovirus that is a leading cause of viral pneumonia in infants. RSV induces autophagy following infection, although the viral factors involved in this mechanism are unknown.