Characterization of Protective Mucosal and Systemic Immune Responses Elicited by Pneumococcal Surface Protein PspA and PspC Nasal Vaccines against a Respiratory Pneumococcal Challenge in Mice

Pneumococcal surface protein A (PspA) and PspC are virulence factors that are involved in the adhesion of Streptococcus pneumoniae to epithelial cells and/or evasion from the immune system. Here, the immune responses induced by mucosal vaccines composed of both antigens as recombinant proteins or delivered by Lactobacillus casei were evaluated. None of the PspC vaccines protected mice against an invasive challenge with pneumococcal strain ATCC 6303. On the other hand, protection was observed for immunization with vaccines composed of PspA from clade 5 (PspA5 or L. casei expressing PspA5) through the intranasal route. The protective response was distinguished by a Th1 profile with high levels of immunoglobulin G2a production, efficient complement deposition, release of proinflammatory cytokines, and infiltration of neutrophils. Intranasal immunization with PspA5 elicited the highest level of protection, characterized by increased levels of secretion of interleukin-17 and gamma interferon by lung and spleen cells, respectively, and low levels of tumor necrosis factor alpha in the respiratory tract.Pneumococcal diseases kill more than 1 million children worldwide every year. The situation is worse in developing countries, where 90% of deaths occur. In Latin America, there are at least 1.6 million cases of pneumococcal disease every year, killing 18,000 children (53). While appropriate treatment, including the use of antibiotics and good nutrition, lowers the incidence of pneumococcal diseases, vaccines are the most efficacious way of preventing them. The existing pneumococcal conjugate vaccine dramatically reduces diseases, disabilities, and deaths, but elevated cost and protection restricted to included serotypes have prevented its implementation in large-scale immunization programs in developing countries. For these reasons, there is considerable interest in using conserved pneumococcal protein antigens as vaccines to provide cost-effective broad protection in all age groups. A number of leading candidates have been shown to elicit protection in mice (10, 51); among these antigens, two of the most promising candidates are pneumococcal surface protein A (PspA) and PspC.An additional concern in the development of cost-effective vaccines against pneumococcal disease is the route of immunization. Human vaccines are traditionally administered intramuscularly by needle inoculation, which brings the risk of transmitting blood-borne pathogens such as human immunodeficiency virus and hepatitis viruses (20). Furthermore, the cost of equipment and well-trained personnel for delivering vaccines by parenteral routes is several times higher than the cost of the vaccines themselves. This aspect is extremely important for vaccine implementation in large-scale immunization programs for developing countries. Mucosal delivery of pediatric vaccines has become an explicit goal of the WHO (20). Immunization via mucosal surfaces would greatly increase the ease of vaccination and would be more readily acceptable than parenteral immunization in many populations. Therefore, the move from injection to mucosal application would be very positive from economical, logistical, and safety standpoints. Mucosal immune responses are also important for the prevention of many infectious diseases because they represent the first barrier from the hosts that pathogens must evade.Research into the host immune response to pneumococcal diseases has focused primarily on the role of innate and adaptative humoral immune responses. However, in the last few years, attention has been drawn to cellular immune responses against Streptococcus pneumoniae, with interesting results. The majority of these studies analyzed cellular aspects of innate immunity and proposed that lymphocytes, neutrophils, and macrophages orchestrate effective immune responses without the presence of specific antibodies. In this context, proinflammatory cytokines promote an adequate milieu for pneumococcal clearance (22, 24, 25, 31, 34, 50, 54). A Th1-biased immune response has also been shown to be engaged in the resolution of pneumococcal infection in humans (21). Nevertheless, inflammatory cell influx into the lung and mucosal responses must be regulated to avoid exacerbated tissue injury. This is evidenced in recent studies of the role of γδ T cells and/or anti-inflammatory cytokines, such as interleukin-10 (IL-10), in pneumococcal infection (26, 42, 55).Protective immune responses against invasive pneumococcal disease and colonization were shown using pneumococcal whole-cell vaccines (28, 46) or recombinant proteins as mucosal vaccines (2, 6, 7, 9, 40). In recent approaches, lactic acid bacteria (3, 11, 18, 37), which are able to activate and modulate the innate immune system (35, 42), were used for pneumococcal antigen presentation, with promising results.Very few works compared pulmonary and systemic immune responses induced by pneumococcal antigens using parenteral and mucosal immunizations (13). The present study aims at investigating local and systemic cellular and humoral immune responses required for protection against invasive intranasal (i.n.) challenge with S. pneumoniae strain ATCC 6303 using PspA and PspC antigens administered by both routes, without the use of adjuvants, or presented by Lactobacillus casei through the nasal route.