Cross-Reactive and Potent Neutralizing Antibody Responses in Human Survivors of Natural Ebolavirus Infection.

Recent studies have suggested that antibody-mediated protection against the Ebolaviruses may be achievable, but little is known about whether or not antibodies can confer cross-reactive protection against viruses belonging to diverse Ebolavirus species, such as Ebola virus (EBOV), Sudan virus (SUDV), and Bundibugyo virus (BDBV). We isolated a large panel of human monoclonal antibodies (mAbs) against BDBV glycoprotein (GP) using peripheral blood B cells from survivors of the 2007 BDBV outbreak in Uganda. We determined that a large proportion of mAbs with potent neutralizing activity against BDBV bind to the glycan cap and recognize diverse epitopes within this major antigenic site. We identified several glycan cap-specific mAbs that neutralized multiple ebolaviruses, including SUDV, and a cross-reactive mAb that completely protected guinea pigs from the lethal challenge with heterologous EBOV. Our results provide a roadmap to develop a single antibody-based treatment effective against multiple Ebolavirus infections.

Mechanism of human antibody-mediated neutralization of Marburg virus.

The mechanisms by which neutralizing antibodies inhibit Marburg virus (MARV) are not known. We isolated a panel of neutralizing antibodies from a human MARV survivor that bind to MARV glycoprotein (GP) and compete for binding to a single major antigenic site. Remarkably, several of the antibodies also bind to Ebola virus (EBOV) GP. Single-particle EM structures of antibody-GP complexes reveal that all of the neutralizing antibodies bind to MARV GP at or near the predicted region of the receptor-binding site. The presence of the glycan cap or mucin-like domain blocks binding of neutralizing antibodies to EBOV GP, but not to MARV GP. The data suggest that MARV-neutralizing antibodies inhibit virus by binding to infectious virions at the exposed MARV receptor-binding site, revealing a mechanism of filovirus inhibition.

IACUC and Veterinary Considerations for Review of ABSL3 and ABSL4 Research Protocols.

With the recent upswing of infectious disease outbreaks (coronavirus, influenza, Ebola, etc), there is an ever-increasing need for biocontainment animal use protocols to better address the research of emerging diseases and to increase the health of both animals and humans. It is imperative that we as a research community ensure these protocols are conducted with the utmost scrutiny and regulatory compliance for the welfare of the animals as well as the health and safety concerns of the individual conducting these studies. Both the welfare of the animals and the health and safety of the research staff must be balanced with the integrity of the science being studied. Even prior to reviewing biocontainment protocols, the research stakeholders should have professional and collegial interactions across all levels of the proposed project. These stakeholders should include the attending veterinarian, the principal investigator, the sponsor, and any organic institutional health and safety assets (environmental health and safety, occupational health, biosafety personnel, medical personnel, facilities operations and maintenance, etc). At most institutions, these stakeholders are members of the Institutional Animal Care and Use Committee and may not possess the necessary tools to properly assess an Animal Biosafety Level 3 and 4 animal use protocol. It is the goal of this article to review some basic concepts of biocontainment, discuss critical communications and preapprovals, clinical observations, medical interventions and supportive care, scientific and study endpoints, euthanasia criteria, animal manipulations, documentation, training, emergency response and contingency plans, security, and decontamination and provide a scenario-based and informative thought-provoking process Institutional Animal Care and Use Committee members and veterinary staff may consider during Animal Biosafety Level 3 and 4 protocol review. These topics will enhance the ability of all stakeholders to balance the protection of the people with the integrity of the science and ultimately the welfare of the animal.

Physiological effects of the TASER C2 conducted energy weapon.

In previous studies, exposure to conducted energy weapons (CEWs) (such as TASER International's Advanced TASER X26 device) resulted in leg muscle contraction, acidosis, increased blood electrolytes, and other biochemical and physiological changes. In the current study, experiments were performed to examine the effects of exposures to TASER International's "C2" CEW, which is specifically marketed to civilian rather than law-enforcement users. Ten pigs (Sus scrofa) were sedated with an intramuscular injection of Telezol (tiletamine HCl and zolazepam HCl) and intubated. General anesthesia was maintained with an IV propofol infusion. Applications of the C2 device for 30 s resulted in extensive muscle contraction and significant increases in heart rate and hematocrit, and in blood levels of pCO2, lactate, glucose, and potassium, sodium, and calcium ions. Significant decreases were observed in blood oxygen saturation, pO2, and pH. Qualitatively, many of these changes were consistent with previous reports in the literature dealing with studies of muscle stimulation or exercise. The changes in blood pCO2, pO2, electrolytes, lactate, and pH, however, were greater than in a previous study of three repeated 5-s exposures to the X26 CEW commonly used by law-enforcement personnel. On the basis of the results, potential detrimental effects due to use of the "citizen-version" TASER C2 CEW may be more likely than limited intermittent applications of the X26 CEW.

A Replication-Defective Human Type 5 Adenovirus-Based Trivalent Vaccine Confers Complete Protection against Plague in Mice and Nonhuman Primates.

Currently, no plague vaccine exists in the United States for human use. The capsular antigen (Caf1 or F1) and two type 3 secretion system (T3SS) components, the low-calcium-response V antigen (LcrV) and the needle protein YscF, represent protective antigens of Yersinia pestis We used a replication-defective human type 5 adenovirus (Ad5) vector and constructed recombinant monovalent and trivalent vaccines (rAd5-LcrV and rAd5-YFV) that expressed either the codon-optimized lcrV or the fusion gene designated YFV (consisting of ycsF, caf1, and lcrV). Immunization of mice with the trivalent rAd5-YFV vaccine by either the intramuscular (i.m.) or the intranasal (i.n.) route provided protection superior to that with the monovalent rAd5-LcrV vaccine against bubonic and pneumonic plague when animals were challenged with Y. pestis CO92. Preexisting adenoviral immunity did not diminish the protective response, and the protection was always higher when mice were administered one i.n. dose of the trivalent vaccine (priming) followed by a single i.m. booster dose of the purified YFV antigen. Immunization of cynomolgus macaques with the trivalent rAd5-YFV vaccine by the prime-boost strategy provided 100% protection against a stringent aerosol challenge dose of CO92 to animals that had preexisting adenoviral immunity. The vaccinated and challenged macaques had no signs of disease, and the invading pathogen rapidly cleared with no histopathological lesions. This is the first report showing the efficacy of an adenovirus-vectored trivalent vaccine against pneumonic plague in mouse and nonhuman primate (NHP) models.

Aerosolized Ebola vaccine protects primates and elicits lung-resident T cell responses.

Direct delivery of aerosolized vaccines to the respiratory mucosa elicits both systemic and mucosal responses. This vaccine strategy has not been tested for Ebola virus (EBOV) or other hemorrhagic fever viruses. Here, we examined the immunogenicity and protective efficacy of an aerosolized human parainfluenza virus type 3-vectored vaccine that expresses the glycoprotein (GP) of EBOV (HPIV3/EboGP) delivered to the respiratory tract. Rhesus macaques were vaccinated with aerosolized HPIV3/EboGP, liquid HPIV3/EboGP, or an unrelated, intramuscular, Venezuelan equine encephalitis replicon vaccine expressing EBOV GP. Serum and mucosal samples from aerosolized HPIV3/EboGP recipients exhibited high EBOV-specific IgG, IgA, and neutralizing antibody titers, which exceeded or equaled titers observed in liquid recipients. The HPIV3/EboGP vaccine induced an EBOV-specific cellular response that was greatest in the lungs and yielded polyfunctional CD8+ T cells, including a subset that expressed CD103 (αE integrin), and CD4+ T helper cells that were predominately type 1. The magnitude of the CD4+ T cell response was greater in aerosol vaccinees. The HPIV3/EboGP vaccine produced a more robust cell-mediated and humoral immune response than the systemic replicon vaccine. Moreover, 1 aerosol HPIV3/EboGP dose conferred 100% protection to macaques exposed to EBOV. Aerosol vaccination represents a useful and feasible vaccination mode that can be implemented with ease in a filovirus disease outbreak situation.

Repeated automated plasmapheresis in goats (Capra hircus): a clinically safe and long-term refinement to current antibody recovery techniques.

Automated plasmapheresis is an optimal method of plasma collection because the donor is a part of a closed loop where whole blood is withdrawn and separated and packed cells are returned in a serial fashion until the desired amount of plasma is obtained. The typical approach to collection of antibody-rich plasma involves withdrawal of whole blood from vaccinated animals, yielding approximately 1000 ml plasma from each animal, which is euthanized after this process. In the present study, 32 goats (Capra hircus) were vaccinated and conditioned for restraint in a modified Panepinto sling. Each animal was monitored clinically, including complete and differential blood counts and serum chemistries 24 h before and 24 to 48 h after each procedure. A jugular vein was surgically prepped, a 16-gauge needle catheter was placed, and the animal was attached to an automated plasmapheresis machine. After plasma removal, return of the resuspended packed blood cells, and infusion of 500 ml 0.9% NaCl, the animal was disconnected from the machine, the catheter removed, and the animal returned to the barn. There were no clinically significant changes in either the complete blood counts or the clinical chemistries during the course of this study. These 32 animals produced 240,000 ml of immunoglobulin-rich plasma over the course of this project and more than 949,000 ml of plasma to date. This study identifies a refinement in current antibody-recovery techniques and potentially reduces the number of animals necessary to produce bioreagents on a long-term and continual basis.