Graphical AbstractHighlights d SARS-CoV-2 uses the SARS-CoV receptor ACE2 for host cell entry d The spike protein of SARS-CoV-2 is primed by TMPRSS2 d Antibodies against SARS-CoV spike may offer some protection against SARS-CoV-2

Swine influenza virus (SIV) and Streptococcus suis are common pathogens of the respiratory tract in pigs, with both being associated with pneumonia. The interactions of both pathogens and their contribution to copathogenesis are only poorly understood. In the present study, we established a porcine precision-cut lung slice (PCLS) coinfection model and analyzed the effects of a primary SIV infection on secondary infection by S. suis at different time points. We found that SIV promoted adherence, colonization, and invasion of S. suis in a two-step process. First, in the initial stages, these effects were dependent on bacterial encapsulation, as shown by selective adherence of encapsulated, but not unencapsulated, S. suis to SIV-infected cells. Second, at a later stage of infection, SIV promoted S. suis adherence and invasion of deeper tissues by damaging ciliated epithelial cells. This effect was seen with a highly virulent SIV subtype H3N2 strain but not with a low-virulence subtype H1N1 strain, and it was independent of the bacterial capsule, since an unencapsulated S. suis mutant behaved in a way similar to that of the encapsulated wildtype strain. In conclusion, the PCLS coinfection model established here revealed novel insights into the dynamic interactions between SIV and S. suis during infection of the respiratory tract. It showed that at least two different mechanisms contribute to the beneficial effects of SIV for S. suis, including capsule-mediated bacterial attachment to SIV-infected cells and capsule-independent effects involving virus-mediated damage of ciliated epithelial cells. R espiratory diseases in swine are responsible for high economic losses in the pig industry worldwide. The upper respiratory tract is a reservoir for a heterogeneous community of (potentially) pathogenic microorganisms and commensals (1). Pneumonia often represents a multifactorial disease complex caused by mixed infections with different pathogens, including viruses and bacteria. Furthermore, unfavorable environmental conditions facilitate the development of the disease (2). Thus, it is termed porcine respiratory disease complex (PRDC) (1). Often primary viral agents, such as porcine reproductive and respiratory syndrome virus, porcine circovirus type 2, and swine influenza virus (SIV), lead to damage of the mucociliary barrier and a decreased immune response, predisposing pigs to secondary infections and pneumonia by opportunistic bacterial pathogens, such as Pasteurella multocida, Mycoplasma hyopneumoniae, and Streptococcus suis (3). However, very little is known about interactions between viral and bacterial pathogens and their role in the copathogenesis of such complex diseases.SIV is an infectious agent causing respiratory disease in pig herds, and it is associated with morbidity of up to 100%. Acute symptoms are high fever, depression, tachypnea, abdominal breathing, and, infrequently, coughing (4). SIV subtypes H1N1, H1N2, and H3N2 are pervasive and cocirculating in the swine population worldwide. Subtype H1N1 viruses...

Streptococci may colonize the epithelium in the airways and other entry sites. While local infection often remains asymptomatic, severe or even fatal diseases occur when streptococci become invasive and spread to different sites in the infected host. We have established porcine respiratory air-liquid interface cultures (ALI) from the porcine lung to analyze the interaction of streptococci with their primary target cells. As representative of the streptococcal family we chose Streptococcus suis (S. suis) that is not only a major swine respiratory pathogen but can also infect humans. Suilysin, a cholesterol-dependent cytolysin (CDC), is an important virulence factor. By comparing a S. suis wt strain with a suilysin-deficient mutant, we demonstrate that suilysin contributes to (i) adherence to airway cells (ii) loss of ciliated cells (iii) apoptosis, and (iv) invasion. Furthermore, we show that cytolytic activity of suilysin is crucial for these effects. A striking result of our analysis was the high efficiency of S. suis-induced apoptosis and invasion upon infection under ALI conditions. These properties have been reported to be less efficient when analyzed with immortalized cells. We hypothesize that soluble effectors such as suilysin are present at higher concentrations in cells kept at ALI conditions and thus more effective. These results should be relevant also for infection of the respiratory tract by other respiratory pathogens.

Coronaviruses and influenza viruses are circulating in humans and animals all over the world. Coinfection with these two viruses may aggravate clinical signs. However, the molecular mechanisms of co-infections by these two viruses are incompletely understood. In this study, we applied airliquid interface (ALI) cultures of well-differentiated porcine tracheal epithelial cells (PTECs) to analyze the co-infection by a swine influenza virus (SIV, H3N2 subtype) and porcine respiratory coronavirus (PRCoV) at different time intervals. Our results revealed that in short-term intervals, prior infection by influenza virus caused complete inhibition of coronavirus infection, while in long-term intervals, some coronavirus replication was detectable. The influenza virus infection resulted in (i) an upregulation of porcine aminopeptidase N, the cellular receptor for PRCoV and (ii) in the induction of an innate immune response which was responsible for the inhibition of PRCoV replication. By contrast, prior infection by coronavirus only caused a slight inhibition of influenza virus replication. Taken together, the timing and the order of virus infection are important determinants in co-infections. This study is the first to show the impact of SIV and PRCoV co-and super-infection on the cellular level. Our results have implications also for human viruses, including potential co-infections by SARS-CoV-2 and seasonal influenza viruses.