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Working correlates of protection predict SchuS4-derived-vaccine candidates with improved efficacy against an intracellular bacterium, Francisella tularensis.

ABSTRACT: Francisella tularensis, the causative agent of tularemia, is classified as Tier 1 Select Agent with bioterrorism potential. The efficacy of the only available vaccine, LVS, is uncertain and it is not licensed in the U.S. Previously, by using an approach generally applicable to intracellular pathogens, we identified working correlates that predict successful vaccination in rodents. Here, we applied these correlates to evaluate a panel of SchuS4-derived live attenuated vaccines, namely SchuS4-ΔclpB, ΔclpB-ΔfupA, ΔclpB-ΔcapB, and ΔclpB-ΔwbtC. We combined in vitro co-cultures to quantify rodent T-cell functions and multivariate regression analyses to predict relative vaccine strength. The predictions were tested by rat vaccination and challenge studies, which demonstrated a clear relationship between the hierarchy of in vitro measurements and in vivo vaccine protection. Thus, these studies demonstrated the potential power a panel of correlates to screen and predict the efficacy of Francisella vaccine candidates, and in vivo studies in Fischer 344 rats confirmed that SchuS4-ΔclpB and ΔclpB-ΔcapB may be better vaccine candidates than LVS.

SUBMITTER: De Pascalis R

PROVIDER: S-EPMC9385090 | biostudies-literature | 2022 Aug

REPOSITORIES: biostudies-literature

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Working correlates of protection predict SchuS4-derived-vaccine candidates with improved efficacy against an intracellular bacterium, Francisella tularensis.

De Pascalis Roberto R Frey Blake B Rice Helen M HM Bhargava Varunika V Wu Terry H TH Peterson Ross L RL Conlan J Wayne JW Sjöstedt Anders A Elkins Karen L KL

NPJ vaccines 20220817 1

Francisella tularensis, the causative agent of tularemia, is classified as Tier 1 Select Agent with bioterrorism potential. The efficacy of the only available vaccine, LVS, is uncertain and it is not licensed in the U.S. Previously, by using an approach generally applicable to intracellular pathogens, we identified working correlates that predict successful vaccination in rodents. Here, we applied these correlates to evaluate a panel of SchuS4-derived live attenuated vaccines, namely SchuS4-Δclp ...[more]

PMID: 35977964

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Project description:Francisella tularensis is the etiological agent of the potentially severe infection tularemia. An existing F: tularensis vaccine, the live vaccine strain (LVS), has been used to protect at-risk personnel, but it is not licensed in any country and it has limited efficacy. Therefore, there is a need of a new, efficacious vaccine. The aim of the study was to perform a detailed analysis of the characteristics of the human immune response to F. tularensis, since this will generate crucial knowledge required to develop new vaccine candidates. Nine individuals were administered the LVS vaccine and peripheral blood mononuclear cells (PBMC) were collected before and at four time points up to one year after vaccination. The properties of the PBMC were characterized by flow cytometry analysis of surface markers and intracellular cytokine staining. In addition, the cytokine content of supernatants from F. tularensis-infected PBMC cultures was determined and the protective properties of the supernatants investigated by adding them to cultures with infected monocyte-derived macrophages (MDM). Unlike before vaccination, PBMC collected at all four time points after vaccination demonstrated F. tularensis-specific cell proliferation, cytokine secretion and cytokine-expressing memory cells. A majority of 17 cytokines were secreted at higher levels by PBMC collected at all time points after vaccination than before vaccination. A discriminative analysis based on IFN-γ and IL-13 secretion correctly classified samples obtained before and after vaccination. Increased expression of IFN-γ, IL-2, and MIP-1β were observed at all time points after vaccination vs. before vaccination and the most significant changes occurred among the CD4 transient memory, CD8 effector memory, and CD8 transient memory T-cell populations. Growth restriction of the highly virulent F. tularensis strain SCHU S4 in MDM was conferred by supernatants and protection correlated to levels of IFN-γ, IL-2, TNF, and IL-17. The findings demonstrate that F. tularensis vaccination induces long-term T-cell reactivity, including TEM and TTM cell populations. Individual cytokine levels correlated with the degree of protection conferred by the supernatants. Identification of such memory T cells and effector mechanisms provide an improved understanding of the protective mechanisms against F. tularensis. mechanisms against F. tularensis.

Project description:There are no defined correlates of protection for any intracellular pathogen, including the bacterium Francisella tularensis, which causes tularemia. Evaluating vaccine efficacy against sporadic diseases like tularemia using field trials is problematic, and therefore alternative strategies to test vaccine candidates like the Francisella Live Vaccine Strain (LVS), such as testing in animals and applying correlate measurements, are needed. Recently, we described a promising correlate strategy that predicted the degree of vaccine-induced protection in mice given parenteral challenges, primarily when using an attenuated Francisella strain. Here, we demonstrate that using peripheral blood lymphocytes (PBLs) in this approach predicts LVS-mediated protection against respiratory challenge of Fischer 344 rats with fully virulent F. tularensis, with exceptional sensitivity and specificity. Rats were vaccinated with a panel of LVS-derived vaccines and subsequently given lethal respiratory challenges with Type A F. tularensis. In parallel, PBLs from vaccinated rats were evaluated for their functional ability to control intramacrophage Francisella growth in in vitro co-culture assays. PBLs recovered from co-cultures were also evaluated for relative gene expression using a large panel of genes identified in murine studies. In vitro control of LVS intramacrophage replication reflected the hierarchy of protection. Further, despite variability between individuals, 22 genes were significantly more up-regulated in PBLs from rats vaccinated with LVS compared to those from rats vaccinated with the variant LVS-R or heat-killed LVS, which were poorly protective. These genes included IFN-γ, IL-21, NOS2, LTA, T-bet, IL-12rβ2, and CCL5. Most importantly, combining quantifications of intramacrophage growth control with 5-7 gene expression levels using multivariate analyses discriminated protected from non-protected individuals with greater than 95% sensitivity and specificity. The results therefore support translation of this approach to non-human primates and people to evaluate new vaccines against Francisella and other intracellular pathogens.

Project description:Francisella tularensis is a highly infectious bacterial pathogen that invades and replicates within numerous host cell types. After uptake, F. tularensis bacteria escape the phagosome, replicate within the cytosol, and suppress cytokine responses. However, the mechanisms employed by F. tularensis to thrive within host cells are mostly unknown. Potential F. tularensis mutants involved in host-pathogen interactions are typically discovered by negative selection screens for intracellular replication or virulence. Mutants that fulfill these criteria fall into two categories: mutants with intrinsic intracellular growth defects and mutants that fail to modify detrimental host cell processes. It is often difficult and time consuming to discriminate between these two possibilities. We devised a method to functionally trans-complement and thus identify mutants that fail to modify the host response. In this assay, host cells are consistently and reproducibly infected with two different F. tularensis strains by physically tethering the bacteria to antibody-coated beads. To examine the efficacy of this protocol, we tested phagosomal escape, cytokine suppression, and intracellular replication for F. tularensis ΔripA and ΔpdpC. ΔripA has an intracellular growth defect that is likely due to an intrinsic defect and fails to suppress IL-1β secretion. In the co-infection model, ΔripA was unable to replicate in the host cell when wild-type bacteria infected the same cell, but cytokine suppression was rescued. Therefore, ΔripA intracellular growth is due to an intrinsic bacterial defect while cytokine secretion results from a failed host-pathogen interaction. Likewise, ΔpdpC is deficient for phagosomal escape, intracellular survival and suppression of IL-1β secretion. Wild-type bacteria that entered through the same phagosome as ΔpdpC rescued all of these phenotypes, indicating that ΔpdpC failed to properly manipulate the host. In summary, functional trans-complementation using bead-bound bacteria co-infections is a method to rapidly identify mutants that fail to modify a host response. Francisella tularensis is a facultative intracellular bacterial pathogen and is the causative agent of the disease tularemia. F. tularensis enters host cells through phagocytosis, escapes the phagosome, and replicates in the host cell cytosol while suppressing cytokine secretion [1]-[4]. Although substantial progress has been made in understanding the intracellular life cycle of F. tularensis, the F. tularensis proteins responsible for manipulating many host cell pathways are unknown. Identifying novel host-pathogen effector proteins is difficult because there is no rapid method to reliably distinguish between bacterial proteins that modify host processes and proteins that are involved in bacterial processes that are required for the bacteria to survive or replicate in the intracellular environment. The ability to identify mutants that are deficient for host-pathogen interactions is important because it can aid in prioritizing the investigation of genes of interest and in downstream experimental design. Moreover, certain mutant phenotypes, such as decreased phagosomal escape, hinder investigation of other potential phenotypes. A method to specifically complement these phenotypes would allow for further characterizations of certain F. tularensis mutants. Thus we sought to develop a method to easily identify and functionally complement mutants that are deficient for interactions with the host.

Dataset Information

Working correlates of protection predict SchuS4-derived-vaccine candidates with improved efficacy against an intracellular bacterium, Francisella tularensis.

Publications

Working correlates of protection predict SchuS4-derived-vaccine candidates with improved efficacy against an intracellular bacterium, Francisella tularensis.

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