Efficacy and Safety of NVX-CoV2373 in Adults in the United States and Mexico.

BACKGROUND: NVX-CoV2373 is an adjuvanted, recombinant spike protein nanoparticle vaccine that was shown to have clinical efficacy for the prevention of coronavirus disease 2019 (Covid-19) in phase 2b-3 trials in the United Kingdom and South Africa, but its efficacy had not yet been tested in North America. METHODS: We conducted a phase 3, randomized, observer-blinded, placebo-controlled trial in the United States and Mexico during the first half of 2021 to evaluate the efficacy and safety of NVX-CoV2373 in adults (≥18 years of age) who had not had severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Participants were randomly assigned in a 2:1 ratio to receive two doses of NVX-CoV2373 or placebo 21 days apart. The primary objective was to determine vaccine efficacy against reverse-transcriptase-polymerase-chain-reaction-confirmed Covid-19 occurring at least 7 days after the second dose. Vaccine efficacy against moderate-to-severe disease and against different variants was also assessed. RESULTS: Of the 29,949 participants who underwent randomization between December 27, 2020, and February 18, 2021, a total of 29,582 (median age, 47 years; 12.6% ≥65 years of age) received at least one dose: 19,714 received vaccine and 9868 placebo. Over a period of 3 months, 77 cases of Covid-19 were noted - 14 among vaccine recipients and 63 among placebo recipients (vaccine efficacy, 90.4%; 95% confidence interval [CI], 82.9 to 94.6; P<0.001). Ten moderate and 4 severe cases occurred, all in placebo recipients, yielding vaccine efficacy against moderate-to-severe disease of 100% (95% CI, 87.0 to 100). Most sequenced viral genomes (48 of 61, 79%) were variants of concern or interest - largely B.1.1.7 (alpha) (31 of the 35 genomes for variants of concern, 89%). Vaccine efficacy against any variant of concern or interest was 92.6% (95% CI, 83.6 to 96.7). Reactogenicity was mostly mild to moderate and transient but was more frequent among NVX-CoV2373 recipients than among placebo recipients and was more frequent after the second dose than after the first dose. CONCLUSIONS: NVX-CoV2373 was safe and effective for the prevention of Covid-19. Most breakthrough cases were caused by contemporary variant strains. (Funded by Novavax and others; PREVENT-19 ClinicalTrials.gov number, NCT04611802.).

Using Adjuvants to Drive T Cell Responses for Next-Generation Infectious Disease Vaccines.

Using adjuvants to drive features of T cell responses to vaccine antigens is an important technological challenge in the design of new and improved vaccines against infections. Properties such as T helper cell function, T cell memory, and CD8+ T cell cytotoxicity may play critical roles in optimal and long-lived immunity through vaccination. Directly manipulating specific immune activation or antigen delivery pathways with adjuvants may selectively augment desired T cell responses in vaccination and may improve the effectiveness and durability of vaccine responses in humans. In this review we outline recently studied adjuvants in their potential for antigen presenting cell and T cell programming during vaccination, with an emphasis on what has been observed in studies in humans as available.

Safety and Immunogenicity of an AS03B-Adjuvanted Inactivated Tetravalent Dengue Virus Vaccine Administered on Varying Schedules to Healthy U.S. Adults: A Phase 1/2 Randomized Study.

Dengue disease and its causative agents, the dengue viruses (DENV-1-4), cause high morbidity in tropical and subtropical regions. We evaluated three dosing regimens of the investigational tetravalent AS03B-adjuvanted dengue-purified inactivated vaccine (DPIV+AS03B). In this phase 1/2, observer-blind, placebo-controlled study (NCT02421367), 140 healthy adults were randomized 1:1:2 to receive DPIV+AS03B according to the following regimens: 0-1 month (M), 0-1-6 M, or 0-3 M. Participants received DPIV+AS03B or placebo at M0, M1, M3, and M6 according to their dosing schedule. Primary objectives were 1) to evaluate the safety of DPIV+AS03B for 28 days (D) after each dose; 2) to demonstrate the added value of a booster dose (0-1-6 M versus 0-1 M) based on neutralizing antibody titers to each DENV type (DENV-1-4) at 28 D after the last dose; and, if this objective was met, 3) to demonstrate the benefit of a longer interval between the first and second doses (0-1 M versus 0-3 M). Adverse events (AEs) within 7 D after vaccination tended to be more frequent after DPIV+AS03B doses than placebo; the number of grade 3 AEs was low (≤ 4.5% after DPIV+AS03B; ≤ 2.9% after placebo), with no obvious differences across groups. Within 28 D following each dose, the frequency of unsolicited AEs after DPIV+AS03B appeared higher for three-dose (0-1-6 M) than two-dose (0-1 M and 0-3 M) regimens. No serious AEs were considered related to vaccination, and no potential immune-mediated diseases were reported during the study. All three schedules were well tolerated. Both primary immunogenicity objectives were demonstrated. The 0-3 M and 0-1-6 M regimens were more immunogenic than the 0-1 M regimen.

Molecular Characterization of Hamster-Adapted Yellow Fever Virus.

We previously reported two hamster models for viscerotropic yellow fever virus (YFV) infection: one using a YFV strain (Jiménez), isolated from a fatal human case in Panama in 1974, and the other using the prototype YFV strain (Asibi). Asibi hamster passage 7 (P7) was associated with accumulation of seven amino acid substitutions, including five in the envelope protein. In this study we report the genome sequences of the hamster Jiménez P0 and P10 viruses in which we identified only two amino acid substitutions during passage, one each in the nonstructural proteins NS3 and NS5, indicating a role for the nonstructural proteins in increased YFV viscerotropism in the Jiménez hamster model. Thus, there are multiple molecular mechanisms involved in viscerotropism of YFV in the hamster model. Neither Asibi P7 nor Jiménez P10 viruses were viscerotropic in mice or guinea pigs. Thus, the hamster viscerotropic phenotype did not translate to other laboratory rodent species.

Role of antigen specific T and B cells in systemic and mucosal immune responses in ETEC and Shigella infections, and their potential to serve as correlates of protection in vaccine development.

The generation of robust systemic and mucosal antibody and cell-mediated immune (CMI) responses that are protective, long-lasting, and can quickly be recalled upon subsequent re-exposure to the cognate antigen is the key to the development of effective vaccine candidates. These responses, whether they represent mechanistic or non-mechanistic immunological correlates of protection, usually entail the activation of T cell memory and effector subsets (T-CMI) and induction of long-lasting memory B cells. However, for ETEC and Shigella, the precise role of these key immune cells in primary and secondary (anamnestic) immune responses remains ill-defined. A workshop to address immune correlates for ETEC and Shigella, in general, and to elucidate the mechanistic role of T-cell subsets and B-cells, both systemically and in the mucosal microenvironment, in the development of durable protective immunity against ETEC and Shigella was held at the recent 2nd Vaccines against Shigella and ETEC (VASE) conference in June 2018. This report is a summary of the presentations and the discussion that ensued at the workshop.

Characteristics of regulatory T-cell populations before and after Ty21a typhoid vaccination in children and adults.

Typhoid fever, caused by the pathogen Salmonella enterica serovar Typhi (S. Typhi), is a serious global health concern. Challenge studies with wild type S. Typhi identified associations between gut-homing regulatory T cells (Treg) and development of typhoid disease. Whether oral live-attenuated Ty21a vaccination induces gut-homing Treg remains unclear. Here, we analyze pediatric and adult Treg pre- and post-Ty21a vaccination in an autologous S. Typhi-antigen presentation model to address this knowledge gap. We show that peripheral memory Treg populations change from childhood to adulthood, but not following Ty21a vaccination. Unsupervised dimensionality reduction with t-distributed stochastic neighbor embedding (tSNE) identifies homing, memory, and functional features which evidence age-associated maturation of multifunctional S. Typhi-responsive Treg, which were not impacted by Ty21a vaccination. These findings improve understanding of pediatric regulatory T cells, while identifying age-related differences in S. Typhi-responsive Treg, which may aid in the development of improved pediatric vaccination strategies against S. Typhi.

Diversity of Salmonella Typhi-responsive CD4 and CD8 T cells before and after Ty21a typhoid vaccination in children and adults.

Typhoid fever is a life-threatening disease caused by the human-restricted pathogen Salmonella enterica serovar Typhi (S. Typhi). The oral live attenuated Ty21a typhoid vaccine protects against this severe disease by eliciting robust, multifunctional cell-mediated immunity (CMI), shown to be associated with protection in wild-type S. Typhi challenge studies. Ty21a induces S. Typhi-responsive CD8+ and CD4+ T cells but little is known about the response to this vaccine in children. To address this important gap in knowledge, we have used mass cytometry to analyze pediatric and adult pre- and post-Ty21a vaccination CMI in an autologous S. Typhi antigen presentation model. Here, using conventional supervised analytical tools, we show adult T cells are more multifunctional at baseline than those obtained from children. Moreover, pediatric and adult T cells respond similarly to Ty21a vaccination, but adult responders remain more multifunctional. The use of the unsupervised dimensionality reduction tool tSNE (t-distributed Stochastic Neighbor Embedding) allowed us to confirm these findings, as well as to identify increases and decreases in well-defined specific CD4+ and CD8+ T-cell populations that were not possible to uncover using the conventional gating strategies. These findings evidenced age-associated maturation of multifunctional S. Typhi-responsive T-cell populations, including those which we have previously shown to be associated with protection from, and/or delayed onset of, typhoid disease. These findings are likely to play an important role in improving pediatric vaccination strategies against S. Typhi and other enteric pathogens.

Age-Associated Heterogeneity of Ty21a-Induced T Cell Responses to HLA-E Restricted Salmonella Typhi Antigen Presentation.

Human-restricted Salmonella enterica serovar Typhi (S. Typhi) is the causative agent of typhoid fever-a life-threatening disease of great global health significance, particularly in the developing world. Ty21a is an oral live-attenuated vaccine that protects against the development of typhoid disease in part by inducing robust T cell responses, among which multifunctional CD8+ cytotoxic T lymphocytes (CTL) play an important role. Following Ty21a vaccination, a significant component of adult CTL have shown to be targeted to S. Typhi antigen presented by the conserved major histocompatibility complex (MHC) class Ib molecule, human leukocyte antigen-E (HLA-E). S. Typhi challenge studies have shown that baseline, multifunctional HLA-E responsive T cells are associated with protection from, and delayed onset of, typhoid disease. However, despite the overwhelming burden of typhoid fever in school-aged children, and due to limited availability of pediatric samples, incomplete information is available regarding these important HLA-E-restricted responses in children, even though studies have shown that younger children may be less likely to develop protective cell mediated immune (CMI) responses than adults following vaccination. To address this gap, we have studied this phenomenon in depth by using mass cytometry to analyze pediatric and adult T cell responses to HLA-E-restricted S. Typhi antigen presentation, before and after Ty21a vaccination. Herein, we show variable responses in all age strata following vaccination among T effector memory (TEM) and T effector memory CD45RA+ (TEMRA) cells based on conventional gating analysis. However, by utilizing the dimensionality reduction tool tSNE (t-distributed Stochastic Neighbor Embedding), we are able to identify diverse, highly multifunctional gut-homing- TEM and TEMRA clusters of cells which are more abundant in adult and older pediatric participants than in younger children. These findings highlight a potential age-associated maturation of otherwise conserved HLA-E restricted T cell responses. Such insights, coupled with the marked importance of multifunctional T cell responses to combat infection, may better inform future pediatric vaccination strategies against S. Typhi and other infectious diseases.

Differences Between Pediatric and Adult T Cell Responses to In Vitro Staphylococcal Enterotoxin B Stimulation.

Toxic shock syndrome (TSS) is capable of inducing life-threatening fever, rash, and systemic organ failure, though the specific mechanisms behind these symptoms remain poorly understood. Staphylococcal enterotoxin B (SEB) and other superantigens have shown to be important factors in TSS, capable of promoting cross-linking between T cell receptors and major histocompatibility complexes which results in overwhelming T cell activation, proliferation, and cytokine production. The resulting proinflammatory cytokine cascade, often referred to as the "cytokine storm," seems to be critical to the development of disease. Interestingly, clinical studies have shown that children exhibit less severe TSS-associated morbidity than adults, though the mechanism behind this phenomenon has not been addressed. Indeed, despite the fact that most novel antigen exposure occurs early in life, be it from environmentally acquired pathogens or routine vaccination, normal pediatric T cell immune functions remain critically underexplored. This is largely due to difficulty in obtaining enough samples to explore more than a narrow sliver of the cell-mediated immune compartment. To address this limitation, we optimized a T effector (Teff)/circulating T follicular helper (cTFH) cell mass cytometry panel which allowed us to analyze a wide array of T cell populations and effector functions following in vitro SEB stimulation. We show that T cell activation-as measured by CD69 expression-following SEB stimulation is lower in pediatric participants, increasing throughout childhood, and reaching adult levels by around 15 years old. Further, while individual CD4+ effector memory T cell (TEM) effector molecules show limited age-associated differences following SEB stimulation, multifunctional CD4+ TEM are shown to positively correlate with increasing age through adolescence. Individual CD8+ TEM effectors and multifunctional phenotypes also show very strong age-associated increases following SEB stimulation. SEB stimulation has little impact on cTFH activation or functional cellular markers, regardless of age. These results, coupled with the fact that a robust proinflammatory cytokine response seems critical to developing severe TSS, suggest a possible connection between the significantly reduced T cell activation and multifunctional populations following in vitro SEB stimulation in our pediatric participants and clinical observations relating to reduced TSS mortality in children.

Safety, tolerability, and immunogenicity of two Zika virus DNA vaccine candidates in healthy adults: randomised, open-label, phase 1 clinical trials.

BACKGROUND: The Zika virus epidemic and associated congenital infections have prompted rapid vaccine development. We assessed two new DNA vaccines expressing premembrane and envelope Zika virus structural proteins. METHODS: We did two phase 1, randomised, open-label trials involving healthy adult volunteers. The VRC 319 trial, done in three centres, assessed plasmid VRC5288 (Zika virus and Japanese encephalitis virus chimera), and the VRC 320, done in one centre, assessed plasmid VRC5283 (wild-type Zika virus). Eligible participants were aged 18-35 years in VRC19 and 18-50 years in VRC 320. Participants were randomly assigned 1:1 by a computer-generated randomisation schedule prepared by the study statistician. All participants received intramuscular injection of 4 mg vaccine. In VRC 319 participants were assigned to receive vaccinations via needle and syringe at 0 and 8 weeks, 0 and 12 weeks, 0, 4, and 8 weeks, or 0, 4, and 20 weeks. In VRC 320 participants were assigned to receive vaccinations at 0, 4, and 8 weeks via single-dose needle and syringe injection in one deltoid or split-dose needle and syringe or needle-free injection with the Stratis device (Pharmajet, Golden, CO, USA) in each deltoid. Both trials followed up volunteers for 24 months for the primary endpoint of safety, assessed as local and systemic reactogenicity in the 7 days after each vaccination and all adverse events in the 28 days after each vaccination. The secondary endpoint in both trials was immunogenicity 4 weeks after last vaccination. These trials are registered with ClinicalTrials.gov, numbers NCT02840487 and NCT02996461. FINDINGS: VRC 319 enrolled 80 participants (20 in each group), and VRC 320 enrolled 45 participants (15 in each group). One participant in VRC 319 and two in VRC 320 withdrew after one dose of vaccine, but were included in the safety analyses. Both vaccines were safe and well tolerated. All local and systemic symptoms were mild to moderate. In both studies, pain and tenderness at the injection site was the most frequent local symptoms (37 [46%] of 80 participants in VRC 319 and 36 [80%] of 45 in VRC 320) and malaise and headache were the most frequent systemic symptoms (22 [27%] and 18 [22%], respectively, in VRC 319 and 17 [38%] and 15 [33%], respectively, in VRC 320). For VRC5283, 14 of 14 (100%) participants who received split-dose vaccinations by needle-free injection had detectable positive antibody responses, and the geometric mean titre of 304 was the highest across all groups in both trials. INTERPRETATION: VRC5283 was well tolerated and has advanced to phase 2 efficacy testing. FUNDING: Intramural Research Program of the Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health.

Human immune responses against Shigella and enterotoxigenic E. coli: Current advances and the path forward.

Robust and well-established immunological assays and firm immune correlates of protection that can predict disease outcome and/or vaccine efficacy are essential to adequately assess human immune responses to infection and vaccination. The availability of reagents and calibrated controls is also critically important to standardize assays and generate comparable results among different laboratories. The workshop "Human Immune Responses against Shigella and ETEC: Current Advances and the Path Forward" held during the VASE meeting provided an opportunity to disseminate and discuss recent advances in the field of Shigella and ETEC immunology, identify research needs, and propose collaborative activities to advance the field. Four presentations featured current knowledge on humoral and cellular immune responses to Shigella and ETEC during infection and vaccination. A discussion followed on immunological methods relevant for clinical studies, immune parameters associated with protection, harmonization of assays among laboratories, and availability of reagents and standards. Specific recommendations proposed to facilitate "the path forward" included supporting communication among scientists, harmonization of assays and sharing of protocols, the creation of a repository of reagents and calibrated controls and distribution of such material to the research community, and expansion of exploratory studies to better understand the interactions between these pathogens and the human immune system and the ensuing responses.

Zika Virus: Recent Advances towards the Development of Vaccines and Therapeutics.

Zika is a rapidly emerging public health threat. Although clinical infection is frequently mild, significant neurological manifestations have been demonstrated in infants born to Zika virus (ZIKV) infected mothers. Due to the substantial ramifications of intrauterine infection, effective counter-measures are urgently needed. In order to develop effective anti-ZIKV vaccines and therapeutics, improved animal models and a better understanding of immunological correlates of protection against ZIKV are required. This review will summarize what is currently known about ZIKV, the clinical manifestations and epidemiology of Zika as well as, the development of animal models to study ZIKV infection, host immune responses against ZIKV, and the current state of development of vaccines and therapeutics against ZIKV.

Importance of Salmonella Typhi-Responsive CD8+ T Cell Immunity in a Human Typhoid Fever Challenge Model.

Typhoid fever, caused by the human-restricted organism Salmonella enterica serovar Typhi (S. Typhi), constitutes a major global health problem. The development of improved attenuated vaccines is pressing, but delayed by the lack of appropriate preclinical models. Herein, we report that high levels of S. Typhi-responsive CD8+ T cells at baseline significantly correlate with an increased risk of disease in humans challenged with a high dose (~104 CFU) wild-type S. Typhi. Typhoid fever development was associated with higher multifunctional S. Typhi-responsive CD8+ T effector memory cells at baseline. Early decreases of these cells in circulation following challenge were observed in both S. Typhi-responsive integrin α4ß7- and integrin α4ß7+ CD8+ T effector memory (TEM) cells, suggesting their potential to home to both mucosal and extra-intestinal sites. Participants with higher baseline levels of S. Typhi-responsive CD8+ T memory cells had a higher risk of acquiring disease, but among those who acquired disease, those with a higher baseline responses took longer to develop disease. In contrast, protection against disease was associated with low or absent S. Typhi-responsive T cells at baseline and no changes in circulation following challenge. These data highlight the importance of pre-existing S. Typhi-responsive immunity in predicting clinical outcome following infection with wild-type S. Typhi and provide novel insights into the complex mechanisms involved in protective immunity to natural infection in a stringent human model with a high challenge dose. They also contribute important information on the immunological responses to be assessed in the appraisal and selection of new generation typhoid vaccines.

Impact of CD4+ T Cell Responses on Clinical Outcome following Oral Administration of Wild-Type Enterotoxigenic Escherichia coli in Humans.

Enterotoxigenic Escherichia coli (ETEC) is a non-invasive enteric pathogen of considerable public health importance, being one of the most common attributable causes of diarrheal illness in infants and young children in developing countries and the most common cause of traveler's diarrhea. To enhance study-to-study consistency of our experimental challenge model of ETEC in volunteers, and to allow concomitant multi-site trials to evaluate anti-ETEC immunoprophylactic products, hundreds of vials, each containing a standardized inoculum of virulent wild-type (wt) ETEC strain H10407 (serotype O78:H11 expressing colonization factor antigen I and heat-labile and heat-stable enterotoxins), were prepared under current Good Manufacturing Practices (cGMP) and frozen. Following thawing, the contents of each vial can be used (diluted as necessary) to prepare consistent challenge inoculum, even at different study sites. A preliminary human experimental challenge study using this cGMP inoculum was conducted on a research isolation ward and the clinical and cell-mediated immune responses evaluated. Of the 6 healthy adult volunteers challenged 83% (5/6) developed diarrhea and 50% developed moderate-to-severe diarrhea (MSD). Moderate and severe diarrhea were defined as passage of ≥ 1 liter or ≥ 3 liters of diarrheal stool respectively. We compared the CD4+ T cell responses of volunteers who developed MSD against those who did not and identified significant differences in ETEC-specific cytokine production and gut homing potential. We furthermore demonstrated that increased expression of the gut-homing molecule integrin α4ß7 by peripheral T follicular helper cells (pTfh) correlated with decreased stool volume and increased ETEC-specific IgA B memory cell (BM) development. Collectively, despite small numbers of volunteers, our results indicate a potential role for CD4+ T cells, in particular pTfh, in modulating disease outcome following exposure to wt ETEC in a volunteer experimental challenge model.

Characterization of Yellow Fever Virus Infection of Human and Non-human Primate Antigen Presenting Cells and Their Interaction with CD4+ T Cells.

Humans infected with yellow fever virus (YFV), a mosquito-borne flavivirus, can develop illness ranging from a mild febrile disease to hemorrhagic fever and death. The 17D vaccine strain of YFV was developed in the 1930s, has been used continuously since development and has proven very effective. Genetic differences between vaccine and wild-type viruses are few, yet viral or host mechanisms associated with protection or disease are not fully understood. Over the past 20 years, a number of cases of vaccine-associated disease have been identified following vaccination with 17D; these cases have been correlated with reduced immune status at the time of vaccination. Recently, several studies have evaluated T cell responses to vaccination in both humans and non-human primates, but none have evaluated the response to wild-type virus infection. In the studies described here, monocyte-derived macrophages (MDM) and dendritic cells (MoDC) from both humans and rhesus macaques were evaluated for their ability to support infection with either wild-type Asibi virus or the 17D vaccine strain and the host cytokine and chemokine response characterized. Human MoDC and MDM were also evaluated for their ability to stimulate CD4+ T cells. It was found that MoDC and MDM supported viral replication and that there were differential cytokine responses to infection with either wild-type or vaccine viruses. Additionally, MoDCs infected with live 17D virus were able to stimulate IFN-γ and IL-2 production in CD4+ T cells, while cells infected with Asibi virus were not. These data demonstrate that wild-type and vaccine YFV stimulate different responses in target antigen presenting cells and that wild-type YFV can inhibit MoDC activation of CD4+ T cells, a critical component in development of protective immunity. These data provide initial, but critical insight into regulatory capabilities of wild-type YFV in development of disease.

Salmonella Typhi-specific multifunctional CD8+ T cells play a dominant role in protection from typhoid fever in humans.

BACKGROUND: Typhoid fever, caused by the human-restricted organism Salmonella Typhi (S. Typhi), is a major public health problem worldwide. Development of novel vaccines remains imperative, but is hampered by an incomplete understanding of the immune responses that correlate with protection. METHODS: Recently, a controlled human infection model was re-established in which volunteers received ~10(3) cfu wild-type S. Typhi (Quailes strain) orally. Twenty-one volunteers were evaluated for their cell-mediated immune (CMI) responses. Ex vivo PBMC isolated before and up to 1 year after challenge were exposed to three S. Typhi-infected targets, i.e., autologous B lymphoblastoid cell-lines (B-LCL), autologous blasts and HLA-E restricted AEH B-LCL cells. CMI responses were evaluated using 14-color multiparametric flow cytometry to detect simultaneously five intracellular cytokines/chemokines (i.e., IL-17A, IL-2, IFN-g, TNF-a and MIP-1b) and a marker of degranulation/cytotoxic activity (CD107a). RESULTS: Herein we provide the first evidence that S. Typhi-specific CD8+ responses correlate with clinical outcome in humans challenged with wild-type S. Typhi. Higher multifunctional S. Typhi-specific CD8+ baseline responses were associated with protection against typhoid and delayed disease onset. Moreover, following challenge, development of typhoid fever was accompanied by decreases in circulating S. Typhi-specific CD8+ T effector/memory (TEM) with gut homing potential, suggesting migration to the site(s) of infection. In contrast, protection against disease was associated with low or no changes in circulating S. Typhi-specific TEM. CONCLUSIONS: These studies provide novel insights into the protective immune responses against typhoid disease that will aid in selection and development of new vaccine candidates.

Activation of Salmonella Typhi-specific regulatory T cells in typhoid disease in a wild-type S. Typhi challenge model.

Salmonella Typhi (S. Typhi), the causative agent of typhoid fever, causes significant morbidity and mortality worldwide. Currently available vaccines are moderately efficacious, and identification of immunological responses associated with protection or disease will facilitate the development of improved vaccines. We investigated S. Typhi-specific modulation of activation and homing potential of circulating regulatory T cells (Treg) by flow and mass cytometry using specimens obtained from a human challenge study. Peripheral blood mononuclear cells were obtained from volunteers pre- and at multiple time-points post-challenge with wild-type S. Typhi. We identified differing patterns of S. Typhi-specific modulation of the homing potential of circulating Treg between volunteers diagnosed with typhoid (TD) and those who were not (No TD). TD volunteers demonstrated up-regulation of the gut homing molecule integrin α4ß7 pre-challenge, followed by a significant down-regulation post-challenge consistent with Treg homing to the gut. Additionally, S. Typhi-specific Treg from TD volunteers exhibited up-regulation of activation molecules post-challenge (e.g., HLA-DR, LFA-1). We further demonstrate that depletion of Treg results in increased S. Typhi-specific cytokine production by CD8+ TEM in vitro. These results suggest that the tissue distribution of activated Treg, their characteristics and activation status may play a pivotal role in typhoid fever, possibly through suppression of S. Typhi-specific effector T cell responses. These studies provide important novel insights into the regulation of immune responses that are likely to be critical in protection against typhoid and other enteric infectious diseases.

Complex adaptive immunity to enteric fevers in humans: lessons learned and the path forward.

Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever, and S. Paratyphi A and B, causative agents of paratyphoid fever, are major public health threats throughout the world. Although two licensed typhoid vaccines are currently available, they are only moderately protective and immunogenic necessitating the development of novel vaccines. A major obstacle in the development of improved typhoid, as well as paratyphoid vaccines is the lack of known immunological correlates of protection in humans. Considerable progress has been made in recent years in understanding the complex adaptive host responses against S. Typhi. Although the induction of S. Typhi-specific antibodies (including their functional properties) and memory B cells, as well as their cross-reactivity with S. Paratyphi A and S. Paratyphi B has been shown, the role of humoral immunity in protection remains undefined. Cell mediated immunity (CMI) is likely to play a dominant role in protection against enteric fever pathogens. Detailed measurements of CMI performed in volunteers immunized with attenuated strains of S. Typhi have shown, among others, the induction of lymphoproliferation, multifunctional type 1 cytokine production, and CD8(+) cytotoxic T-cell responses. In addition to systemic responses, the local microenvironment of the gut is likely to be of paramount importance in protection from these infections. In this review, we will critically assess current knowledge regarding the role of CMI and humoral immunity following natural S. Typhi and S. Paratyphi infections, experimental challenge, and immunization in humans. We will also address recent advances regarding cross-talk between the host's gut microbiota and immunization with attenuated S. Typhi, mechanisms of systemic immune responses, and the homing potential of S. Typhi-specific B- and T-cells to the gut and other tissues.