Novel vaccine candidates of Bordetella pertussis identified by reverse vaccinology.

Whooping cough is a disease caused by Bordetella pertussis, whose morbidity has increased, motivating the improvement of current vaccines. Reverse vaccinology is a strategy that helps identify proteins with good characteristics fast and with fewer resources. In this work, we applied reverse vaccinology to study the B. pertussis proteome and pangenome with several in-silico tools. We analyzed the B. pertussis Tohama I proteome with NERVE software and compared 234 proteins with B. parapertussis, B. bronchiseptica, and B. holmessi. VaxiJen was used to calculate an antigenicity value; our threshold was 0.6, selecting 84 proteins. The candidates were depurated and grouped in eight family proteins to select representative candidates, according to bibliographic information and their immunological response predicted with ABCpred, Bcepred, IgPred, and C-ImmSim. Additionally, a pangenome study was conducted with 603 B. pertussis strains and PanRV software, identifying 3421 core proteins that were analyzed to select the best candidates. Finally, we selected 15 proteins from the proteome study and seven proteins from the pangenome analysis as good vaccine candidates.

Design of a Multi-Epitope Vaccine against Tuberculosis from Mycobacterium tuberculosis PE_PGRS49 and PE_PGRS56 Proteins by Reverse Vaccinology.

Tuberculosis is a disease caused by Mycobacterium tuberculosis, representing the second leading cause of death by an infectious agent worldwide. The available vaccine against this disease has insufficient coverage and variable efficacy, accounting for a high number of cases worldwide. In fact, an estimated third of the world's population has a latent infection. Therefore, developing new vaccines is crucial to preventing it. In this study, the highly antigenic PE_PGRS49 and PE_PGRS56 proteins were analyzed. These proteins were used for predicting T- and B-cell epitopes and for human leukocyte antigen (HLA) protein binding efficiency. Epitopes GGAGGNGSLSS, FAGAGGQGGLGG, GIGGGTQSATGLG (PE_PGRS49), and GTGWNGGKGDTG (PE_PGRS56) were selected based on their best physicochemical, antigenic, non-allergenic, and non-toxic properties and coupled to HLA I and HLA II structures for in silico assays. A construct with an adjuvant (RS09) plus each epitope joined by GPGPG linkers was designed, and the stability of the HLA-coupled construct was further evaluated by molecular dynamics simulations. Although experimental and in vivo studies are still necessary to ensure its protective effect against the disease, this study shows that the vaccine construct is dynamically stable and potentially effective against tuberculosis.

Characterization and use in neutralization assays of avian influenza codon-optimized H5 and H7 retroviral pseudotypes.

Influenza is a relevant problem for public and animal health, with a significant economic impact. In recent years, outbreaks of avian influenza virus have resulted in devastating losses in the poultry industry worldwide, and although its transmission to humans is very rare, there is always a potential risk for an even more severe outbreak. Currently, vaccination is considered the most effective tool for the control and prevention of influenza infections in both humans and animals. The maintenance of animal welfare and the successful implementation of animal health programs depend on the timely administration of vaccines, which must comply with quality specifications indicated by health authorities; for example, the capability to ensure a minimum antibody titer. The production of viral antigens used in these tests can pose a biosafety risk, and some viral strains can be difficult to grow. Therefore, new biotechnological alternatives are required to overcome these disadvantages. In this study, we produced pseudotypes carrying H5 and H7 hemagglutinins from lowly and highly pathogenic avian influenza viruses. These pseudotypes were used in neutralization assays to detect neutralizing antibodies in avian sera, which were confirmed positive by inhibition of the hemagglutination test. Our results showed that the pseudotype neutralization assay is a viable alternative for the detection of neutralizing antibodies, by demonstrating subtype specificity and requiring reduced biosafety requirements. Therefore, it represents a versatile platform that can facilitate technology transfer protocols between laboratories, and an immediate application in serological tools for quality control of veterinary vaccines against avian influenza.

[Reverse vaccinology: strategy against emerging pathogens]. / Vacunología reversa: estrategia contra patógenos emergentes.

New technologies in vaccinology are capable of achieving fast development, as well as large-scale production of effective and safe vaccines. Reverse vaccinology is an in silico methodology, which studies different characteristics of infectious agents, in order to identify antigens that are good vaccine candidates, without the need of traditional culture. This strategy is based on bioinformatics tools, that in a simple, safety and inexpensive way, reduces time and effort significantly in the new vaccine design, against traditional vaccinology. In recent years, the rapid spread of infections by emerging pathogens requires prompt development of new vaccines. Bioinformatic strategies joined with the latest next-generation vaccines allow the selection of vaccine candidates in a short time, which is relevant in the development of new vaccines against pathogens with pandemic potential.

Las nuevas tecnologías en vacunología son capaces de lograr un desarrollo rápido, así como una producción a gran escala de vacunas seguras y eficaces. La vacunología reversa es una metodología in silico que estudia diferentes características de los agentes infecciosos, con el objetivo de identificar antígenos que sean buenos candidatos vacunales, sin la necesidad del cultivo tradicional. Esta estrategia se basa en el uso de herramientas bioinformáticas, por lo que es una metodología sencilla, segura, económica y que reduce de forma significativa el tiempo de diseño de una vacuna, en comparación con la vacunología tradicional. En los últimos años, la rápida diseminación de infecciones por patógenos emergentes ha requerido del desarrollo oportuno de nuevas vacunas. Las estrategias bioinformáticas aunadas a los más recientes diseños de vacunas de nueva generación permiten la selección de candidatos vacunales en corto tiempo, lo cual es muy importante en el desarrollo de nuevas vacunas contra patógenos con potencial pandémico.

Stability of retroviral pseudotypes carrying the hemagglutinin of avian influenza viruses under various storage conditions.

Retroviral pseudotypes are broadly used as safe instruments to mimic the structure and surface of highly pathogenic viruses. They have been employed for the discovery of new drugs, as diagnostic tools in vaccine studies, and part of serological assays. Because of their widespread use in research and their potential as tools for quality control, it is important to know their shelf life, stability, and best storage conditions. In this study, we produced pseudotypes carrying the lacZ reporter gene and the hemagglutinin (HA) of avian influenza virus subtypes H5 and H7 to investigate their stability under various storage conditions. We produced pseudotypes with titers of approximately 106 RLU/mL, which decreased to 105-106 RLU/mL after short-term storage at 4 °C (up to 4 weeks). Stability was maintained after long-term storage at -20 °C (up to 12 months), even under storage variations such as freeze-thaw cycles. We conclude that, although the titers decreased by 1 log10 under the different storage conditions, the remaining titers can be readily applicable in many techniques, such as neutralization assays. These findings show that large quantities of retroviral pseudotypes can be safely stored for short- or long-term use, allowing standardization and reduced variation in assays involving retroviral pseudotypes.