Pathogenesis of Staphylococcus aureus is increasingly recognized to be driven by powerful toxins. Staphylococcus aureus employs up to six pore-forming toxins to subvert the human host defense and to promote bacterial invasion: alpha-hemolysin that disrupts epithelial and endothelial barriers and five leukocidins that lyse phagocytes involved in bacterial clearance. Previously, we described two human monoclonal antibodies (mAbs), ASN-1 that neutralizes alpha-hemolysin and four leukocidins (LukSF-PV, LukED, HlgAB, HlgCB), and ASN-2 that inactivates the 5th leukocidin, LukGH. In this study we tested the individual and combined effects of ASN-1 and ASN-2 in multiple in vitro models employing relevant human target cells. We found that diverse S. aureus isolates with different genetic backgrounds (based on MLST- and spa-typing) and antibiotic sensitivity (both MRSA and MSSA) displayed greatly different cytotoxin expression patterns influenced by the type of growth medium used. Both mAbs were required to fully prevent the lysis of human neutrophils exposed to the mixture of recombinant cytotoxins or native toxins present in the culture supernatants of S. aureus isolates. Flow cytometry confirmed the protective effects of ASN-1 + ASN-2 (known as ASN100) on granulocytes, monocytes, NK-cells and T-lymphocytes. ASN-1 alone preserved the integrity of a 3D-primary culture of human tracheal/bronchial mucociliary epithelial tissue infected with S. aureus. We conclude that simultaneous inhibition of alpha-hemolysin and five leukocidins by ASN100 blocks cytolytic activity of S. aureus towards human target cells in vitro.

The majority of antibodies induced by influenza neuraminidase (NA), like those against hemagglutinin (HA), are relatively specific to viruses isolated within a limited time window, as seen in serological studies and the analysis of many murine monoclonal antibodies (MAbs). We report three broadly reactive human MAbs targeting N1 NA. Two were isolated from a young adult vaccinated with trivalent influenza vaccine (TIV), which inhibited N1 NA from viruses isolated from humans over a period of a hundred years. The third antibody, isolated from a child with acute mild H7N9 infection, inhibited both group 1 N1 and group 2 N9 NAs. In addition, the antibodies cross-inhibited the N1 NAs of highly pathogenic avian H5N1 influenza viruses. These antibodies are protective in prophylaxis against seasonal H1N1 viruses in mice. This study demonstrates that human antibodies to N1 NA with exceptional cross-reactivity can be recalled by vaccination and highlights the importance of standardizing the NA antigen in seasonal vaccines to offer optimal protection. IMPORTANCE Antibodies to the influenza virus NA can provide protection against influenza disease. Analysis of human antibodies to NA lags behind that of antibodies to HA. We show that human monoclonal antibodies against NA induced by vaccination and infection can be very broadly reactive, with the ability to inhibit a wide spectrum of N1 NAs on viruses isolated between 1918 and 2018. This suggests that antibodies to NA may be a useful therapy and that the efficacy of influenza vaccines could be enhanced by ensuring the appropriate content of NA antigen.

Inflammatory signals enhance the susceptibility of human PMNs to lysis by LukGH rendering this toxin dominant among the S. aureus leukocidins in vitro.