Abstract
Staphylococcus aureus (SA) infections cause high mortality and morbidity in humans. Being central to its pathogenesis, S. aureus thwarts the host defense by secreting a myriad of virulence factors, including bicomponent, pore-forming leukotoxins. While all vaccine development efforts that aimed at achieving opsonophagocytic killing have failed, targeting virulence by toxoid vaccines represents a novel approach to preventing mortality and morbidity that are caused by SA. The recently discovered leukotoxin LukAB kills human phagocytes and monocytes and it is present in all known S. aureus clinical isolates. While using a structure-guided approach, we generated a library of mutations that targeted functional domains within the LukAB heterodimer to identify attenuated toxoids as potential vaccine candidates. The mutants were evaluated based on expression, solubility, yield, biophysical properties, cytotoxicity, and immunogenicity, and several fully attenuated LukAB toxoids that were capable of eliciting high neutralizing antibody titers were identified. Rabbit polyclonal antibodies against the lead toxoid candidate provided potent neutralization of LukAB. While the neutralization of LukAB alone was not sufficient to fully suppress leukotoxicity in supernatants of S. aureus USA300 isolates, a combination of antibodies against LukAB, α-toxin, and Panton-Valentine leukocidin completely neutralized the cytotoxicity of these strains. These data strongly support the inclusion of LukAB toxoids in a multivalent toxoid vaccine for the prevention of S. aureus disease.
Highlights
Staphylococcus aureus, which is a pervasive human pathogen, is a leading cause of life-threatening community and hospital-acquired infections world-wide
With pET24a (+) dual system, as LukA and LukB are co-expressed within the same cell by two different plasmids, it is a better system for introduction of mutations while screening a mutation library
We found that the toxicity and physicochemical properties of LukABWT from the two systems are comparable (Table S1)
Summary
Staphylococcus aureus, which is a pervasive human pathogen, is a leading cause of life-threatening community and hospital-acquired infections world-wide. This Gram-positive bacterium is often associated with a range of diseases, from mild skin and soft tissue infections to invasive bacteremia, septic arthritis, endocarditis, and osteomyelitis [1]. Methicillin-resistant S. aureus (MRSA) causes ~80,000 invasive infections and 11,000 deaths per year in the United States alone [3]. S. aureus expresses a myriad of virulence factors, including cell surface attachment factors, capsular polysaccharides, enzymes, immune modulatory molecules, pore-forming toxins, and superantigens that aimed at establishing the infection or colonization as well as immune evasion [5]. Among the pore-forming toxins, S. aureus produces single component alpha hemolysin (Hla or α-toxin) and bicomponent pore-forming toxins (BCPFTs), Panton-Valentine leukocidin (PVL; composed of LukS-PV and LukF-PV), Leukocidin AB (LukAB), Leukocidin ED (LukED), and γ-hemolysins
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