Vaccine 31 (2013) 2465–2470 Contents lists available at SciVerse ScienceDirect Vaccine jou rn al hom ep age: www.elsevier.com/locat e/vaccine Pneumococcal Surface Protein A does not affect the immune responses to a combined diphtheria tetanus and pertussis vaccine in mice Fernanda A. Lima a , Eliane N. Miyaji a , Wagner Quintilio b , Isaias Raw a , Paulo L. Ho a,b , Maria Leonor S. Oliveira a,∗ a Centro de Biotecnologia, Instituto Butantan, São Paulo, SP, Brazil b Divisão Bioindustrial, Instituto Butantan, São Paulo, SP, Brazil a r t i c l e i n f o Article history: Received 31 August 2012 Received in revised form 6 February 2013 Accepted 14 March 2013 Available online 26 March 2013 Keywords: Pneumococcal vaccines DTP PspA Adjuvants a b s t r a c t The Pneumococcal Surface Protein A (PspA) is a promising candidate for the composition of a protein vac- cine against Streptococcus pneumoniae. We have previously shown that the whole cell Bordetella pertussis vaccine (wP) is a good adjuvant to PspA, inducing protective responses against pneumococcal infection in mice. In Brazil, wP is administered to children, formulated with diphtheria and tetanus toxoids (DTP w ) and aluminum hydroxide (alum) as adjuvant. A single subcutaneous dose of PspA5–DTP low (a formulation containing PspA from clade 5 and a new generation DTP w , containing low levels of B. pertussis LPS and Alum) induced high levels of systemic anti-PspA5 antibodies in mice and conferred protection against respiratory lethal challenges with two different pneumococcal strains. Here we evaluate the mucosal immune responses against PspA5 as well as the immune responses against the DTP antigens in mice vaccinated with PspA5–DTP low . Subcutaneous immunization of mice with PspA5–DTP low induced high levels of anti-PspA5 IgG in the airways but no IgA. In addition, no differences in the influx of cells to the respiratory mucosa, after the challenge, were observed in vaccinated mice, when compared with control mice. The levels of circulating anti-pertussis, -tetanus and -diphtheria antibodies were equivalent in mice vaccinated with DTP low or PspA5–DTP low . Antibodies induced by DTP low or PspA5–DTP low showed similar ability to neutralize the cytotoxic effects of the diphtheria toxin on Vero cells. Furthermore, combination with PspA5 did not affect protection against B. pertussis and tetanus toxin challenges in mice. Our results support the proposal for a combined PspA-DTP vaccine. © 2013 Elsevier Ltd. All rights reserved. 1. Introduction In spite of the great efficacy of pneumococcal conjugate vaccines against colonization and invasive diseases in children, Streptococ- cus pneumoniae continues to be a threat, mainly in developing countries [1]. One important drawback of conjugate vaccines is the induction of serotype-specific protection that may lead to the substitution of the prevalent pneumococcal serotypes in the vacci- nation area [2]. Protein-based vaccines are being studied as alter- natives. The Pneumococcal Surface Protein A (PspA) is a candidate antigen that has been shown to induce protection against different animal models of pneumococcal infection [3–6]. Protective epi- topes were described in the N-terminal part of the PspA molecule, a region that is exposed to the immune system and display high Abbreviations: PspA, Pneumococcal Surface Protein A; DTP low , diphteria, tetanus and pertussis vaccine formulated with a whole cell pertussis containing low amounts of LPS and alum. ∗ Corresponding author. Tel.: +55 11 3726 7222x2244; fax: +55 11 3726 1505. E-mail address: mloliveira@butantan.gov.br (M.L.S. Oliveira). variability [7,8]. PspA molecules were classified into 6 clades that can be grouped into three families, according to the amino acid sequence of the clade defining region, present at the end of the N- terminal portion [9]. Although this variability may have negative implications for the use of PspA as a vaccine candidate, recombinant N-terminal fragments of PspA from clades 4 (PspA4) and 5 (PspA5) were shown to induce antibodies that display cross-reactivity with PspAs from different clades [10,11]. Effective immune responses were characterized by the induction of high levels of anti-PspA antibodies, with a preferable Th-1 character and the induction of IFN- [12]. Our group has recently proposed that the combina- tion of PspA5 with the Bordetella pertussis whole cell vaccine (wP) could induce such responses [13]. The wP vaccine produced by Instituto Butantan (São Paulo, Brazil) is part of the DTP w vaccine (a formulation containing diphtheria and tetanus toxoids, wP and aluminum hydroxide as adjuvant) that is distributed to Brazilian children by the Ministry of Health, free of charge. This vaccine was introduced in the country more than 20 years ago and has dramat- ically reduced the incidence of pertussis [14]. Despite the safety of the current DTP w vaccine, improvements are being made and a new generation vaccine containing low levels of B. pertussis LPS (DTP low ) 0264-410X/$ – see front matter © 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.vaccine.2013.03.026