In studies of the mechanism of immunity to Bordetella pertussis in a murine respiratory infection model, we have previously demonstrated that natural infection of immunization with a whole cell vaccine induces a potent protective immune response, which is mediated by T-helper type-l (Thl) cells. In contrast an acellular vaccine generates Th2 cells and is associated with delayed bacterial clearance following respiratory challenge. In the present study we have investigated the apparent Thl/Th2 cell dichotomy in acquired immunity and have examined the factors that affect their induction or detection. The cytokine profiles of B. pertussis-specific T cells in immune animals were determined using antigen-stimulated ex vivo spleen cells or CD4+ T-cell lines and clones established in the presence of interleukin-2 (IL-2) or IL-4. Antigen-specific T cells derived from mice immunized with the acellular vaccine were almost exclusively of the Th2 cell type. In contrast, T-cell lines and clones established following respiratory infection or immunization with the whole cell vaccine were predominantly of the Thl type. However, a proportion of T cells from convalescent mice, especially when cultured in the presence of IL-4, secreted IL-4 and IL-5 with or without detectable IL-2 and interferon-y (IFN-y), suggesting that ThO or Th2 cells were also primed during natural infection in vivo. Furthermore, when mice were assessed 6 months after infection, spleen cells produced significant levels of IL-4 and IL-5, which were not evident at 6 weeks. The route of immunization and the genetic background of the mice were also found to influence the preferential priming of Thl cells, and this was directly related to the level of protection against respiratory or intracerebral (i.c.) challenge. Our findings underline the critical role ofCD4+ Thl cells in immunity to B. pertussis, but also demonstrate that a number of factors in the in vivo priming and in vitro restimulation can skew the apparent dominance of one Th cell type over another.