Generation and characterization of a polyvalent vaccine against Salmonella typhi and evaluation of porin-specific immune responses in humans

Abstract

Salmonella (S.) enterica infections are an important global health problem affecting mainly low-income countries with scarce sanitation conditions. Despite the presence of two vaccines against typhoid fever, more than 20 million individuals suffer from enteric fever annually, resulting in more than 200,000 lethal cases per year. The vaccines available in the market exhibit short-term protection, are not suitable for younger infants and require special temperature conditions for transport. Therefore, there is a significant need for identification and study of new molecular targets from Salmonella that contribute to develop safe and cost-effective vaccines against Typhoid fever that induce long-term protection. Outer membrane proteins (Omps) from S. typhi represent sui generis molecular targets that induce long-term protective immune responses in animal models and humans. In the present work we focused on the development and characterization of a vaccine formulation based on S. typhi OmpC/OmpF porins. As a second aim, we studied the specific immune responses directed against OmpC/OmpF porins from the vaccine strain Ty21a after its administration to healthy volunteers. The microencapsulation of S. typhi porins into poly(lactic-co-glycolic acid) (PLGA) protected the multimeric structure of the vaccine antigen from acidic degradation and enhanced its immunogenicity following oral administration to mice. Moreover, the vaccine induced strong S. typhi-specific B cell responses in Peyer’s patches and mesenteric lymph nodes. Hence, this PLGA-microencapsulated porin-based formulation substantially enhanced the efficacy of oral vaccination against S. typhi and may represent a suitable formulation to be tested in clinical trials. In the second part of this work, the administration of Ty21a to healthy volunteers allowed us to study the evolution of the immune responses elicited against S. typhi porins. B cell responses, namely antibody and cellular measurements, as well as characterization of T cell responses was assessed before vaccination and at day 7, 21 and 56 after the last vaccine dose administration. The majority of the participants (80-85%) exhibited specific anti-porin IgM/IgG antibodies in serum, with a significant increase at all time points tested that peaked at day 7 after vaccination for IgM and at day 21 for IgG. Likewise, Ty21a administration induced a significant increase in circulating Antibody Secreting Cells (ASCs) at days 21 and 56. Moreover, vaccinated subjects displayed high frequencies of CD4+ cells expressing the activation marker CD40L that could be detected at days 21 and 56 after vaccination. Additionally, measurement of the specific IgA levels in stool samples showed an increase in antibody levels at day 56 after vaccination. Altogether, these results showed that porinspecific immune responses presented in a concerted manner upon priming of immune cells with the outer membrane proteins contained in the vaccine strain Ty21a. In sum, this work describes the generation of a vaccine formulation containing the well characterized S. typhi porins OmpC/OmpF, and demonstrates that even within the pool of antigenic determinants contained in the vaccine strain Ty21a; porins are capable of inducing detectable immune responses by conventional laboratory assays, reinforcing the notion that these proteins represent promising targets for vaccination.