Rickettsia prowazekii has been tested for biological warfare due to the high mortality that it produces after aerosol transmission of very low numbers of rickettsiae. Epidemic typhus, the infection caused by these obligately intracellular bacteria, continues to be a threat because it is difficult to diagnose due to initial non-specific symptoms and the lack of commercial diagnostic tests that are sensitive and specific during the initial clinical presentation. A vaccine to prevent epidemic typhus would constitute an effective deterrent to the weaponization of R. prowazekii; however, an effective and safe vaccine is not currently available. Due to the cytoplasmic niche of Rickettsia, CD8+ T-cells are critical effectors of immunity; however, the identification of antigens recognized by these cells has not been systematically addressed. To help close this gap, we designed an antigen discovery strategy that uses cell-based vaccination with antigen presenting cells expressing microbe's proteins targeted to the MHC class I presentation pathway. We report the use of this method to discover a protective T-cell rickettsial antigen, RP884, among a test subset of rickettsial proteins.

Summary It has been suggested that tumour cell lysis by gamma‐radiation induces a tumoral antigen release eliciting an immune response. It is not clear how a specific immune response in cervical cancer patients is developed after radiotherapy. This study is an attempt to investigate the role of the human papillomavirus type 16 (HPV‐16) E7‐specific T helper response before and after radiotherapy. Lymphocytes were isolated from 32 cervical cancer patients before and after radiotherapy and from 16 healthy women. They were stimulated for 12 hr with autologous HPV‐16 E7‐pulsed monocyte‐derived dendritic cells or directly with HPV‐16 E7 synthetic peptides: E751–70, E765–84 and E779–98. The cells were stained for CD4, CD69, intracellular interferon‐γ (IFN‐γ) and interleukin‐4 (IL‐4) cytokines and analysed by flow cytometry. A specific CD4+ CD69+ IFN‐γ+ immune response against HPV‐16 E779–98 peptide was observed in 10 of 14 patients (71·4%) after treatment, compared with 4 of 14 (28·5%) before radiotherapy (P = 0·039); however, this response was not associated with a successful clinical response. Before treatment, 5 of 31 patients showed a HPV‐16 E779–98‐specific T helper type 2 (Th2) response. Interestingly, this response was significantly associated with a decrease in disease‐free survival (P = 0·027). These results suggest that a Th2‐type cellular response could be useful as a predictor of recurrence and poor prognosis. An increase of the HPV‐specific immune response was observed after radiotherapy; however, it is not enough to control completely the disease after treatment. Our results support that the E7‐specific T‐cell IFN‐γ response in cervical cancer patients, rather than reflecting the host’s capability of controlling tumour growth, might be an indicator for disease severity.

The obligately intracellular bacteria Rickettsia infect endothelial cells and cause systemic febrile diseases that are potentially lethal. No vaccines are currently available and current knowledge of the effective immune response is limited. Natural and experimental rickettsial infections provide strong and cross-protective cellular immunity if the infected individual survives the acute infection. Although resistance to rickettsial infections is attributed to the induction of antigen-specific T cells, particularly CD8+ T cells, the identification and validation of correlates of protective cellular immunity against rickettsial infections, an important step towards vaccine validation, remains a gap in this field. Here, we show that after a primary challenge with Rickettsia typhi in the C3H mouse model, the peak of anti-Rickettsia CD8+ T cell-mediated responses occurs 7 days post-infection (dpi), which coincides with the beginning of rickettsial clearance. At this time point, both effector-type and memory-type CD8+ T cells are present, suggesting that 7 dpi is a valid time point for the assessment of CD8+ T cell responses of mice previously immunized with protective antigens. Based on our results, we suggest four correlates of cellular protection for the assessment of protective rickettsial antigens: 1) production of IFN-γ by antigen experienced CD3+CD8+CD44high cells, 2) production of Granzyme B by CD27lowCD43low antigen-experienced CD8+ T cells, 3) generation of memory-type CD8+ T cells [Memory Precursor Effector Cells (MPECs), as well as CD127highCD43low, and CD27highCD43low CD8+ T cells], and 4) generation of effector-like memory CD8+ T cells (CD27lowCD43low). We propose that these correlates could be useful for the general assessment of the quality of the CD8+ T cell immune response induced by novel antigens with potential use in a vaccine against Rickettsia.

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Natural and experimental rickettsial infections provide strong and cross-protective immunity if the individual survives the acute infection. Although resistance to rickettsial infections is attributed to the induction of antigen-specific T cells, particularly CD8+ T cells, the characterization of the cellular immune response has not been systematically addressed. Here, we report the systematic, kinetic and ex vivo analysis of CD8+ T cells from spleen, liver, lungs and lymph nodes by flow cytometry with specific panels to address the activation, effector, and memory phases of the anti-rickettsial response in C3H/HeN mice infected with R. typhi. Our findings suggest that the instauration of the effector phase occurs around 7dpi which coincides with the rickettsial clearance. This phase was characterized by an increased frequency of CD8+ T cells with a phenotype compatible with effector cells:CD44+CD43+CD27-/+ and CD44+CD43-/+CD127-; along with an increased production of IFN-γ and GZM B. A more robust production of IFN-γ and GZM-B was detected in the target organs, highlighting the compartmentalization of the immune response. Furthermore, at 7dpi, differences in the production of IFN-γ and GZM-B between controls and mice immunized with a vaccine candidate were also observed. These findings are relevant for vaccine development, providing insights about meaningful time points and specific correlates of protection that can be used as a paradigm for anti-rickettsial vaccine testing.