Effective protection against secondary pneumococcal pneumonia by oral vaccination with attenuated Salmonella delivering PspA antigen in mice☆
Highlights
► Secondary pneumococcal infection exacerbates influenza-induced pneumonia. ► Oral vaccination with RASV protect mice against secondary pneumococcal pneumonia. ► Oral RASV induce long-term protection. ► Antiviral treatment synergistically enhances protective efficacy of oral RASV.
Introduction
Influenza is a common cause of acute respiratory inflammation in humans. While influenza virus can be lethal, considerable numbers of deaths are due to secondary bacterial infections. For instance, the majority of deaths in the “1918 Spanish flu” pandemic resulted from secondary infection by bacterial pneumonia, mostly with Streptococcus pneumoniae [1], [2]. A recent study of the 2009 H1N1 virus pandemic showed that numerous fatal cases had bacterial coinfection with S. pneumoniae [3].
Vaccination has been one of the most effective tools to prevent influenza infection; however, the vaccines against secondary pneumococcal pneumonia are not yet commercially available. In infants, pneumococcal conjugate vaccine prevents approximately 30% of pneumonias associated with respiratory viruses [4]; however, polysaccharide-based vaccines are not ideal for secondary bacterial infection prevention because some non-vaccine serotypes have poor immunological responses [5]. Therefore, there is a need to develop an improved and effective vaccine based on conserved antigens across all capsular serotypes to induce more effective and durable immune responses that could potentially protect against secondary bacterial infections.
Previous studies have identified several S. pneumoniae protein antigens that may be useful vaccine candidates [6], [7], [8]. Among these, pneumococcal surface protein A (PspA) is a choline-binding surface protein that inhibits complement-mediated phagocytosis and binding to lactoferrin [9], [10]. Previous studies indicated that PspA antigen is a promising vaccine target for protection against secondary bacterial pneumonia [11], [12]. Whereas S. pneumoniae mutants that disrupt the gene for neuraminidase A or hyaluronidase did not display different colonization rates, the mutant for PspA decreased colonization in the lungs of mice previously infected with influenza virus [12]. In addition, intranasal vaccination with PspA plus poly(I:C) as mucosal adjuvant resulted in protection against secondary pneumococcal pneumonia [11].
Live recombinant attenuated Salmonella enteric serovar Typhimurium vaccine (RASV) strain has successfully been used as carrier for exogenous protein antigens [13]. We previously reported that oral administration of RASV delivering PspA antigen resulted in long-lasting IgG and IgA antibodies in both systemic and mucosal compartments and efficient protection against pulmonary S. pneumoniae infection in mice [14]. In this study, we determined whether oral vaccination with RASV delivering PspA antigen can induce protective immunity against secondary pneumococcal pneumonia in the murine model. Of note, we found that oral vaccination of RASV delivering PspA antigen effectively protected mice from secondary pneumococcal infection and showed synergistic effects with the antiviral drug oseltamivir.
Section snippets
Virus and bacteria
A/Puerto Rico/8/34 (PR8; H1N1) strains were grown in the allantoic fluid of 11-day-old embryonated chicken eggs and incubated for 3 days at 37 °C. Allantoic fluid was harvested and filtered with a 0.25-μm-pore-size syringe filter (Pall, Port Washington, NY). S. pneumoniae virulent capsular type 3 strain WU2 was grown in 100 ml of THY medium until late log phase, adjusted with 3% glycerol, and frozen in 1-ml aliquots containing 109 colony-forming units (cfu)/ml. RASV synthesizing rPspA-RX1 was
Secondary pneumococcal infection exacerbates influenza-induced pneumonia
In order to establish a murine secondary pneumococcal pneumonia model, WT B6 mice were intratracheally challenged with S. pneumoniae (WU2; 1 × 104 cfu/mouse) 1 week after intratracheal infection with influenza (PR8; 1 × 102 pfu/mouse) (Fig. 1A). To determine the optimal conditions, we tried several different doses of PR8 and WU2 strains and then selected a dose combination on the basis of morbidity and mortality results (data not shown). Each infection of PR8 or WU2 strain alone resulted in a slow
Discussion
As pneumococcal infection frequently has severe clinical outcomes [17], recommendations call for infants, children, and elderly adults to be vaccinated with pneumococcal vaccine [18], [19]. However, a significant proportion of children are colonized with S. pneumoniae [20], [21] and thus are at risk of secondary pneumococcal infection during the influenza season [22]. Our results suggest that pneumococcal vaccination during the influenza season can prevent lethal secondary pneumococcal
Conflict of interest
The authors declare that no competing interests exist.
Acknowledgement
We thank Dr. David E. Briles at the University of Alabama, Birmingham, for providing strain WU2.
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Funding statement: This work was supported by a National Research Foundation of Korea (NRF) grant funded by the Korean government (MEST) (Nos. 2010-0029133 and 2012-0000805) and by Bill and Melinda Gates Foundation Grant #37863.