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Measuring immune correlates of disease acquisition and protection in the context of a clinical trial is a prerequisite for improved vaccine design. We analysed binding and neutralizing antibody measurements 4 weeks post vaccination as correlates of risk of moderate to severe-critical COVID-19 through 83 d post vaccination in the phase 3, double-blind placebo-controlled phase of ENSEMBLE, an international randomized efficacy trial of a single dose of Ad26.COV2.S. We also evaluated correlates of protection in the trial cohort. Of the three antibody immune markers we measured, we found most support for 50% inhibitory dilution (ID

Anti-spike IgG binding antibody, anti-receptor binding domain IgG antibody, and pseudovirus neutralizing antibody measurements four weeks post-vaccination were assessed as correlates of risk of moderate to severe-critical COVID-19 outcomes through 83 days post-vaccination and as correlates of protection following a single dose of Ad26.COV2.S COVID-19 vaccine in the placebo-controlled phase of ENSEMBLE, an international, randomized efficacy trial. Each marker had evidence as a correlate of risk and of protection, with strongest evidence for 50% inhibitory dilution (ID50) neutralizing antibody titer. The outcome hazard ratio was 0.49 (95% confidence interval 0.29, 0.81; p=0.006) per 10-fold increase in ID50; vaccine efficacy was 60% (43, 72%) at nonquantifiable ID50 (< 2.7 IU50/ml) and rose to 89% (78, 96%) at ID50 = 96.3 IU50/ml. Comparison of the vaccine efficacy by ID50 titer curves for ENSEMBLE-US, the COVE trial of the mRNA-1273 vaccine, and the COV002-UK trial of the AZD1222 vaccine supported consistency of the ID50 titer correlate of protection across trials and vaccine types.

Background Multiple Anthrax vaccines are licensed or in development for post-exposure prophylaxis in individuals 18 to 65 years of age. No information exists on anthrax vaccines in populations over the age of 65. It is critical that we assess the capacity of anthrax vaccines to generate a protective immune response in older individuals. In this study, we compared BioThrax® to a formulation containing a CpG adjuvant (AV7909). Methods We conducted a Phase 2 clinical study to evaluate safety and immunogenicity of three vaccination schedules of the AV7909 vaccine candidate and one vaccination schedule of BioThrax® vaccine in adults over 65 years of age. A total of 305 subjects were enrolled to assess safety and immunogenicity by seroprotection rates, toxin neutralizing antibody titers, and anti-Protective Antigen ELISA titers. Results Compared to BioThrax, AV7909 elicited a more robust immune response in older subjects, especially with three doses of AV7909 at Days 1, 15, and 29, or two doses at Days 1 and 29. These trends were true with both seroprotection rates as defined by the percentage of subjects with 50 percent neutralization factors greater than 0.56, and geometric mean antibody titers. The responses to both AV7909 and BioThax were lower in older subjects compared to those aged 18–50. Conclusion The immunogenicity data suggest that the CpG adjuvant in the AV7909 vaccine helps to elicit a more robust immune response in subjects over the age of 65. Alternative dosing strategies may be considered in this population given the high seroprotection rates with Day 1 and 29, or Day 1, 15, and 29 regimens. Trial Registration: clinicaltrials.gov Identifier: NCT03518125.

The Ebola virus epidemic in West Africa proved to be the largest in the history of filovirus outbreaks, causing the World Health Organization to declare a public health emergency of international concern in August of 2014. In collaboration with domestic and international partners, the Biomedical Advanced Research and Development Authority (BARDA) initiated several vaccine development projects in support of the overall response efforts. The urgency associated with the epidemic triggered the clinical evaluation of lead vaccine candidates starting in late 2014. Here we will discuss development of the lead vaccine candidates for Ebola virus, specifically Zaire ebolavirus.

Vaccinations against the encephalitic alphaviruses (western, eastern, and Venezuelan equine encephalitis virus) are of significant interest to biological defense, public health, and agricultural communities alike. Although vaccines licensed for veterinary applications are used in the Western Hemisphere and attenuated or inactivated viruses have been used under Investigational New Drug status to protect at-risk personnel, there are currently no licensed vaccines for use in humans. Here, we will discuss the need for a trivalent vaccine that can protect humans against all three viruses, recent progress to such a vaccine, and a strategy to continue development to Food and Drug Administration licensure.

Bordetella parapertussis causes the prolonged coughing illness known as pertussis or whooping cough, persisting for weeks within the respiratory tracts of infected hosts but inducing a very poor T cell response relative to that induced by Bordetella pertussis, the more common cause of pertussis. In this study, we examine the contributions of cytokines involved in the clearance of B. parapertussis and immunomodulation that delays effective clearance. The slow elimination of this pathogen from the respiratory tracts of mice coincides with the gradual accumulation of CD4+ T cells in the lungs and B. parapertussis-responsive IFN-γ–producing cells in the spleen. IFN-γ–deficient mice were defective in the accumulation of leukocytes in lungs and in clearance of B. parapertussis from the lungs. In vitro B. parapertussis-stimulated macrophages produced IL-10, which inhibited the generation of the IFN-γ response that is required for protection in vivo. As compared with wild-type mice, IL-10–deficient mice produced significantly higher levels of IFN-γ, had higher numbers of leukocytes accumulated in the lungs, and cleared B. parapertussis more rapidly. Together, these data indicate that B. parapertussis induces the production of IL-10, which facilitates its persistence within infected hosts by limiting a protective IFN-γ response.

Bordetella pertussis , a causative agent of whooping cough, expresses BrkA, which confers serum resistance, but the closely related human pathogen that also causes whooping cough, Bordetella parapertussis , does not. Interestingly, B. parapertussis , but not B. pertussis , produces an O antigen, a factor shown in other models to confer serum resistance. Using a murine model of infection, we determined that O antigen contributes to the ability of B. parapertussis to colonize the respiratory tract during the first week of infection, but not thereafter. Interestingly, an O antigen-deficient strain of B. parapertussis was not defective in colonizing mice lacking the complement cascade. O antigen prevented both complement component C3 deposition on the surface and complement-mediated killing of B. parapertussis . In addition, O antigen was required for B. parapertussis to systemically spread in complement-sufficient mice, but not complement-deficient mice. These data indicate that O antigen enables B. parapertussis to efficiently colonize the lower respiratory tract by protecting against complement-mediated control and clearance.

Bordetella pertussis and Bordetella parapertussis are closely related endemic human pathogens which cause whooping cough, a disease that is reemerging in human populations. Despite how closely related these pathogens are, their coexistence and the limited efficacy of B. pertussis vaccines against B. parapertussis suggest a lack of cross-protective immunity between the two. We sought to address the ability of infection-induced immunity against one of these pathogens to protect against subsequent infection by the other using a mouse model of infection. Immunity induced by B. parapertussis infection protected against subsequent infections by either species. However, immunity induced by B. pertussis infection prevented subsequent B. pertussis infections but did not protect against B. parapertussis infections. The O antigen of B. parapertussis inhibited binding of antibodies to the bacterial surface and was required for B. parapertussis to colonize mice convalescent from B. pertussis infection. Thus, the O antigen of B. parapertussis confers asymmetrical cross-immunity between the causative agents of whooping cough. We propose that these findings warrant investigation of the relative role of B. parapertussis in the resurgence of whooping cough.

A recent resurgence in the number of cases of whooping cough, and other respiratory diseases caused by members of the bordetellae, in vaccinated populations has demonstrated the need for a thorough understanding of vaccine-induced immunity to facilitate more intelligent vaccine design. In this work, we use a murine model of respiratory infection using the highly successful animal pathogen, Bordetella bronchiseptica. Since previously infected animals have been shown to resist re-infection by B. bronchiseptica, we sought to examine the differences between vaccine-induced immunity and infection-induced immunity. Both prior infection and vaccination conferred nearly complete protection in the lungs, however, only prior infection resulted in significant protection in the upper respiratory tract. While immunity induced by prior infection offered significant protection even in the absence of complement or FcgammaRs, vaccination-induced protection required both complement and FcgammaRs. Although vaccination induced higher titers of B. bronchiseptica-specific antibodies, this serum was less effective than infection-induced serum in clearing bacteria from the lower respiratory tract. Together these findings highlight substantial differences between the mechanisms involved in vaccine- and infection-induced protective immunity.

Bordetella parapertussisandBordetella pertussisare closely related species that cause whooping cough, an acute, immunizing disease. Their coexistence in the same host populations at the same time and vaccine studies showing thatB. pertussisvaccines have little effect onB. parapertussisinfection or disease suggest that the protective immunity induced by each does not efficiently cross protect against the other. Although the mechanisms of protective immunity toB. pertussishave been well studied, those ofB. parapertussishave not. The present study explores the mechanism by whichB. parapertussisis cleared from the lower respiratory tract by anamnestic immunity. Serum antibodies are necessary and sufficient for elimination of this bacterium, and CD4+T cells, complement, and neutrophils are required for serum antibody-mediated clearance. Mice lacking immunoglobulin A had no defect in their ability to control or clear infection. Interestingly, serum antibody-mediated clearance ofB. parapertussisdid not require Fc receptors that are required for antibody-mediated clearance ofB. pertussis. Together these data support a model for the mechanism of protective immunity toB. parapertussisthat is similar but distinct from that ofB. pertussis.

Coxiella burnetii is the etiological agent of Q Fever, a disease that is often spread to humans via inhalational exposure to the bacteria from contaminated agricultural sources. Outbreaks have been observed all over the world with larger foci generating interest in vaccination programs, most notably in Australia and the Netherlands. Importantly, exposure rates among military personnel deployed to the Middle East can be relatively high as measured by seroconversion to C. burnetii-specific antibodies. Q Fever has been of interest to the biodefense community over the years due to its low infectious dose and environmental stability. Recent advances in cell-free growth and genetics of C. burnetii also make this organism easier to culture and manipulate. While there is a vaccine that is licensed for use in Australia, the combination of biodefense- and public health-related issues associated with Q Fever warrant the development of a safer and more effective vaccine against this disease.

Background The ever-growing wealth of biological information available through multiple comprehensive database repositories can be leveraged for advanced analysis of data. We have now extensively revised and updated the multi-purpose software tool Biofilter that allows researchers to annotate and/or filter data as well as generate gene-gene interaction models based on existing biological knowledge. Biofilter now has the Library of Knowledge Integration (LOKI), for accessing and integrating existing comprehensive database information, including more flexibility for how ambiguity of gene identifiers are handled. We have also updated the way importance scores for interaction models are generated. In addition, Biofilter 2.0 now works with a range of types and formats of data, including single nucleotide polymorphism (SNP) identifiers, rare variant identifiers, base pair positions, gene symbols, genetic regions, and copy number variant (CNV) location information. Results Biofilter provides a convenient single interface for accessing multiple publicly available human genetic data sources that have been compiled in the supporting database of LOKI. Information within LOKI includes genomic locations of SNPs and genes, as well as known relationships among genes and proteins such as interaction pairs, pathways and ontological categories. Via Biofilter 2.0 researchers can: • Annotate genomic location or region based data, such as results from association studies, or CNV analyses, with relevant biological knowledge for deeper interpretation • Filter genomic location or region based data on biological criteria, such as filtering a series SNPs to retain only SNPs present in specific genes within specific pathways of interest • Generate Predictive Models for gene-gene, SNP-SNP, or CNV-CNV interactions based on biological information, with priority for models to be tested based on biological relevance, thus narrowing the search space and reducing multiple hypothesis-testing. Conclusions Biofilter is a software tool that provides a flexible way to use the ever-expanding expert biological knowledge that exists to direct filtering, annotation, and complex predictive model development for elucidating the etiology of complex phenotypic outcomes.

The Defense Threat Reduction Agency’s Joint Science and Technology Office manages the Chemical and Biological Defense Program’s Science and Technology portfolio. The Joint Science and Technology Office’s mission is to invest in transformational ideas, innovative people and actionable technology development for Chemical and Biological Defense solutions, with the primary goal to deliver Science and Technology products and capabilities to the warfighter and civilian population that outpace the threat. This commentary focuses on one thrust area within this mission: the Vaccine program of the Joint Science and Technology Office’s Translational Medical Division. Here, we will describe candidate vaccines currently in the S&T pipeline, enabling technologies that should facilitate advanced development of these candidates into FDA licensed vaccines, and how the ever-changing biological threat landscape impacts the future of biodefense vaccines.

There is a need to advance our ability to conduct credible human risk assessments for inhalational anthrax associated with exposure to a low number of bacteria. Combining animal data with computational models of disease will be central in the low-dose and cross-species extrapolations required in achieving this goal. The objective of the current work was to apply and advance the competing risks (CR) computational model of inhalational anthrax where data was collected from NZW rabbits exposed to aerosols of Ames strain Bacillus anthracis. An initial aim was to parameterize the CR model using high-dose rabbit data and then conduct a low-dose extrapolation. The CR low-dose attack rate was then compared against known low-dose rabbit data as well as the low-dose curve obtained when the entire rabbit dose–response data set was fitted to an exponential dose–response (EDR) model. The CR model predictions demonstrated excellent agreement with actual low-dose rabbit data. We next used a modified CR model (MCR) to examine disease incubation period (the time to reach a fever >40 °C). The MCR model predicted a germination period of 14.5 h following exposure to a low spore dose, which was confirmed by monitoring spore germination in the rabbit lung using PCR, and predicted a low-dose disease incubation period in the rabbit between 14.7 and 16.8 days. Overall, the CR and MCR model appeared to describe rabbit inhalational anthrax well. These results are discussed in the context of conducting laboratory studies in other relevant animal models, combining the CR/MCR model with other computation models of inhalational anthrax, and using the resulting information towards extrapolating a low-dose response prediction for man.

Repeated low-level exposures to biological agents could occur before or after the remediation of an environmental release. This is especially true for persistent agents such as B. anthracis spores, the causative agent of anthrax. Studies were conducted to examine aerosol methods needed for consistent daily low aerosol concentrations to deliver a low-dose (less than 10(6) colony forming units (CFU) of B. anthracis spores) and included a pilot feasibility characterization study, acute exposure study, and a multiple 15 day exposure study. This manuscript focuses on the state-of-the-science aerosol methodologies used to generate and aerosolize consistent daily low aerosol concentrations and resultant low inhalation doses to rabbits. The pilot feasibility characterization study determined that the aerosol system was consistent and capable of producing very low aerosol concentrations. In the acute, single day exposure experiment, targeted inhaled doses of 1 × 10(2), 1 × 10(3), 1 × 10(4), and 1 × 10(5) CFU were used. In the multiple daily exposure experiment, rabbits were exposed multiple days to targeted inhaled doses of 1 × 10(2), 1 × 10(3), and 1 × 10(4) CFU. In all studies, targeted inhaled doses remained consistent from rabbit-to-rabbit and day-to-day. The aerosol system produced aerosolized spores within the optimal mass median aerodynamic diameter particle size range to reach deep lung alveoli. Consistency of the inhaled dose was aided by monitoring and recording respiratory parameters during the exposure with real-time plethysmography. Overall, the presented results show that the animal aerosol system was stable and highly reproducible between different studies and over multiple exposure days.

SUMMARYIn a recent experiment, we found that mice previously infected withBordetella pertussiswere not protected against a later infection withBordetella parapertussis, while primary infection withB. parapertussisconferred cross-protection. This challenges the common assumption made in most mathematical models for pathogenic strain dynamics that cross-immunity between strains is symmetric. Here we investigate the potential consequences of this pattern on the circulation of the two pathogens in human populations. To match the empirical dominance ofB. pertussis, we made the additional assumption thatB. parapertussispays a cost in terms of reduced fitness. We begin by exploring the range of parameter values that allow the coexistence of the two pathogens, with or without vaccination. We then track the dynamics of the system following the introduction of anti-pertussis vaccination. Our results suggest that (1) in order forB. pertussisto be more prevalent thanB. parapertussis, the former must have a strong competitive advantage, possibly in the form of higher infectivity, and (2) because of asymmetric cross-immunity, the introduction of anti-pertussis vaccination should have little effect on the absolute prevalence ofB. parapertussis. We discuss the evidence supporting these predictions, and the potential relevance of this model for other pathogens.

The genus Bordetella includes a group of closely related mammalian pathogens that cause a variety of respiratory diseases in a long list of animals ( B. bronchiseptica ) and whooping cough in humans ( B. pertussis and B. parapertussis ). While past research has examined how these pathogens are eliminated from the lower respiratory tract, the host factors that control and/or clear the bordetellae from the upper respiratory tract remain unclear. We hypothesized that immunoglobulin A (IgA), the predominant mucosal antibody isotype, would have a protective role against these mucosal pathogens. IgA −/− mice were indistinguishable from wild-type mice in their control and clearance of B. pertussis or B. parapertussis , suggesting that IgA is not crucial to immunity to these organisms. However, naïve and convalescent IgA −/− mice were defective in reducing the numbers of B. bronchiseptica in the upper respiratory tract compared to wild-type controls. Passively transferred serum from convalescent IgA −/− mice was not as effective as serum from convalescent wild-type mice in clearing this pathogen from the tracheae of naive recipient mice. IgA induced by B. bronchiseptica infection predominantly recognized lipopolysaccharide-containing O-antigen, and antibodies against O-antigen were important to bacterial clearance from the trachea. Since an IgA response contributes to the control of B. bronchiseptica infection of the upper respiratory tract, immunization strategies aimed at inducing B. bronchiseptica -specific IgA may be beneficial to preventing the spread of this bacterium among domestic animal populations.

Bordetella pertussis causes whooping cough, an endemic respiratory disease that is increasing in prevalence despite vaccination efforts. Although host immunity is modulated by virulence factors of this pathogen, it is unclear what host factors are required to overcome their effects. Here, we investigate an apparent relationship between the effects of pertussis toxin and tumor necrosis factor (TNF)- alpha . B. pertussis grew efficiently and caused moderate pathology in wild-type mice, whereas TNF- alpha (-/-) mice had higher numbers of bacteria and leukocytes in lungs, experienced more airway resistance, and died of the infection. Interestingly, an isogenic B. pertussis strain lacking pertussis toxin did not induce these effects in TNF- alpha (-/-) mice and behaved similarly in wild-type and TNF- alpha -deficient hosts. Together, these results indicate that TNF- alpha is essential for the host to overcome the effects of pertussis toxin, allowing both control of B. pertussis numbers and regulation of the inflammation induced by infection.

Bordetella pertussis,B. parapertussis, andB. bronchisepticaare closely related species associated with respiratory disease in humans and other mammals. WhileB. bronchisepticahas a wide host range,B. pertussisandB. parapertussisevolved separately from aB. bronchiseptica-like progenitor to naturally infect only humans. Despite very different doubling times in vitro, all three establish similar levels of infection in the mouse lung within 72 h. Recent work has revealed separate roles for Toll-like receptor 4 (TLR4) in immunity toB. pertussisandB. bronchiseptica, while no role for TLR4 duringB. parapertussisinfection has been described. Here we compared the requirement for TLR4 in innate host defense to these organisms using the same mouse infection model. WhileB. bronchisepticacauses lethal disease in TLR4-deficient mice,B. pertussisandB. parapertussisdo not. Correspondingly, TLR4 is critical in limitingB. bronchisepticabut notB. pertussisorB. parapertussisbacterial numbers during the first 72 h. Interestingly,B. bronchisepticainduces a TLR4-dependent cytokine response that is considerably larger than that induced byB. pertussisorB. parapertussis. Analysis of their endotoxins using RAW cells suggests thatB. bronchisepticalipopolysaccharide (LPS) is 10- and 100-fold more stimulatory thanB. pertussisorB. parapertussisLPS, respectively. The difference in LPS stimulus is more pronounced when using HEK293 cells expressing human TLR4. Thus, it appears that in adapting to infect humans,B. pertussisandB. parapertussisindependently modified their LPS to reduce TLR4-mediated responses, which may compensate for slower growth rates and facilitate host colonization.

Infection by the Gram-negative pathogen Burkholderia pseudomallei results in the disease melioidosis, acquired from the environment in parts of southeast Asia and northern Australia. Clinical symptoms of melioidosis range from acute (fever, pneumonia, septicemia, and localized infection) to chronic (abscesses in various organs and tissues, most commonly occurring in the lungs, liver, spleen, kidney, prostate and skeletal muscle), and persistent infections in humans are difficult to cure. Understanding the basic biology and genomics of B. pseudomallei is imperative for the development of new vaccines and therapeutic interventions. This formidable task is becoming more tractable due to the increasing number of B. pseudomallei genomes that are being sequenced and compared. Here, we compared three B. pseudomallei genomes, from strains MSHR668, K96243 and 1106a, to identify features that might explain why MSHR668 is more virulent than K96243 and 1106a in a mouse model of B. pseudomallei infection. Our analyses focused on metabolic, virulence and regulatory genes that were present in MSHR668 but absent from both K96243 and 1106a. We also noted features present in K96243 and 1106a but absent from MSHR668, and identified genomic differences that may contribute to variations in virulence noted among the three B. pseudomallei isolates. While this work contributes to our understanding of B. pseudomallei genomics, more detailed experiments are necessary to characterize the relevance of specific genomic features to B. pseudomallei metabolism and virulence. Functional analyses of metabolic networks, virulence and regulation shows promise for examining the effects of B. pseudomallei on host cell metabolism and will lay a foundation for future prediction of the virulence of emerging strains. Continued emphasis in this area will be critical for protection against melioidosis, as a better understanding of what constitutes a fully virulent Burkholderia isolate may provide for better diagnostic and medical countermeasure strategies.

Background With the abundance of information and analysis results being collected for genetic loci, user-friendly and flexible data visualization approaches can inform and improve the analysis and dissemination of these data. A chromosomal ideogram is an idealized graphic representation of chromosomes. Ideograms can be combined with overlaid points, lines, and/or shapes, to provide summary information from studies of various kinds, such as genome-wide association studies or phenome-wide association studies, coupled with genomic location information. To facilitate visualizing varied data in multiple ways using ideograms, we have developed a flexible software tool called PhenoGram which exists as a web-based tool and also a command-line program. Results With PhenoGram researchers can create chomosomal ideograms annotated with lines in color at specific base-pair locations, or colored base-pair to base-pair regions, with or without other annotation. PhenoGram allows for annotation of chromosomal locations and/or regions with shapes in different colors, gene identifiers, or other text. PhenoGram also allows for creation of plots showing expanded chromosomal locations, providing a way to show results for specific chromosomal regions in greater detail. We have now used PhenoGram to produce a variety of different plots, and provide these as examples herein. These plots include visualization of the genomic coverage of SNPs from a genotyping array, highlighting the chromosomal coverage of imputed SNPs, copy-number variation region coverage, as well as plots similar to the NHGRI GWA Catalog of genome-wide association results. Conclusions PhenoGram is a versatile, user-friendly software tool fostering the exploration and sharing of genomic information. Through visualization of data, researchers can both explore and share complex results, facilitating a greater understanding of these data.

The recognition of bacterial lipopolysaccharide (LPS) by host Toll-like receptor (TLR)4 is a crucial step in developing protective immunity against several gram negative bacterial pathogens. Bordetella bronchiseptica and B. pertussis stimulate robust TLR4 responses that are required to control the infection, but a close relative, B. parapertussis, poorly stimulates this receptor, and TLR4 deficiency does not affect its course of infection. This led us to hypothesize that inefficient TLR4 stimulation enables B. parapertussis to evade host immunity. In a mouse model of infection, B. parapertussis grew rapidly in the lungs, but no measurable increase in TLR4-mediated cytokine, chemokine, or leukocyte responses were observed over the first few days of infection. Delivery of a TLR4 stimulant in the inoculum resulted in a robust inflammatory response and a 10- to 100-fold reduction of B. parapertussis numbers. As we have previously shown, B. parapertussis grows efficiently during the first week of infection even in animals passively immunized with antibodies. We show that this evasion of antibody-mediated clearance is dependent on the lack of TLR4 stimulation by B. parapertussis as co-inoculation with a TLR4 agonist resulted in 10,000-fold lower B. parapertussis numbers on day 3 in antibody-treated wild type, but not TLR4-deficient, mice. Together, these results indicate that inefficient TLR4 stimulation by B. parapertussis enables it to avoid host immunity and grow to high numbers in the respiratory tract of naïve and immunized hosts.