Ebola Vaccines

Clinical Description

The initial clinical manifestations of Ebola virus begin 2–21 days after infection (average 8–10 days) with “flu-like” symptoms including fever, fatigue, and myalgia. Within days, abnormalities related to liver function and clotting appear, with associated clinical findings of nausea, abdominal pain, rash, petechiae, diarrhea, and vomiting. As the disease progresses, in addition to the possible development of respiratory disorders, the gastrointestinal tract becomes involved, leading to signs of bleeding from mucosal sites, including melena and hematochezia. In the terminal phases, typically within 1–2 weeks after infection, there is progression to disseminated infection, with abnormal clotting parameters, parenchymal hepatic damage, disseminated bleeding at various sites including but not limited to venipuncture sites and the conjunctiva, and cardiac dysfunction resulting to terminal hypovolemic or septic shock in the absence of bacterial superinfection. The mortality rate ranges from 36% to 90% in those with recognizable infection.

Complications

Clinical complications of Ebola virus arise from infection of targeted cells during the viral lifecycle. In early infection, the virus replicates in reticuloendothelial cells, monocytes, macrophages and dendritic-like cells. Damage to these cell types results in the release of cytokines that likely contribute to early symptoms of infection, including fever. Subsequent damage to the liver and vasculature as the disease progresses results in impaired blood vessel integrity and gives rise to a bleeding diathesis accompanied by hypotension. Ebola virus infection of the adrenal gland (at later stages of disease) may also contribute to patient hypotension. The inability to maintain blood pressure in the setting of fever, blood loss, profuse vomiting, and diarrhea; and shock leads to a high mortality rate, particularly in the absence of effective supportive care. Isolated case reports from the 2014 outbreak suggested that vigorous clinical support improved patient outcomes. Quality of life of EVD survivors has also been shown to be impacted post recovery. There have been multiple reports of Ebola sequelae in survivors with a higher percentage of symptoms such as headache, fatigue, muscle and joint ache as well as memory loss when compared with uninfected subjects. Further, ophthalmic manifestations of Ebola in survivors support persistent inflammation of ocular tissues. A study exploring the association between inflammatory markers and long-term complications experienced by survivors found no correlation and highlights the continued need to understand underlying mechanisms of disease pathogenesis and recovery.

Virology

Ebola virus and the closely related Marburg virus (MARV) are members of the Filoviridae family, named for the filamentous morphology of virions that appear by electron microscopy to be nonuniform in length but with a consistent diameter of approximately 80 nm. There are six species in the genus Ebolavirus: Zaire ebolavirus (Ebola virus/EBOV), Sudan ebolavirus (Sudan virus/SUDV), Taï Forest ebolavirus (previously named Côte d’Ivoire) (Taï Forest virus / TAFV), Reston ebolavirus (Reston virus/RESTV), Bundibugyo ebolavirus (Bundibugyo virus/BDBV) discovered in 2007, and the most recently discovered Bombali ebolavirus (Bombali virus/BOMV). ^, BOMV was discovered in free-tailed bats using a broadly reactive filovirus PCR assay but as of yet, replication competent BOMV has not been isolated from any potential reservoir. Of known filovirus species, RESTV and BOMV are the only members that have not caused any reported human fatalities. ^, Indeed neither RESTV nor BOMV have been associated with any clinical manifestation in humans, including those who seroconverted after exposure.

The 19 kb nonsegmented negative-strand RNA genome of Ebola virus contains a linear array of seven genes, six of which each encode a single protein product. The virus genome consists of seven genes from which at least 10 proteins are known to be encoded. The GP gene encodes for the trimeric membrane anchored glycoprotein (GP) or dimeric secreted glycoprotein (sGP). sGP is the genomic form of GP, a 364 amino acid protein including a stretch of seven uridines that distinguishes it from full length transmembrane anchored GP. Expression of GP results when a nontemplated uridine (U) is inserted by the viral RNA polymerase within the stretch of seven uridines in the antisense genomic RNA forming the 8U variation. Other viral proteins (VP) encoded by the viral genome include VP40/VP24 that form the inner matrix and nucleoprotein (NP), VP30, VP35, and the RNA-dependent RNA polymerase, L that comprise the ribonucleoprotein complex ( Fig. 22.1 ). Nucleocapsid-like filamentous structures can be formed by overexpression of NP, VP24 and VP35 in the absence of other viral proteins. ^, A T7-polymerase-based minireplicon system demonstrated that the minimal replication machinery for Ebola virus requires the cooperative activity of NP, VP30, VP35, and L, while VP40 and GP are sufficient to form virus-like particles. The X-ray structure of the carboxy-terminal domain of Ebola virus VP35, which possesses both its dsRNA binding and inhibitory activity, is now well characterized. In addition to its role in viral RNA replication and in virion assembly, VP35 plays a crucial role in Ebola virus infectivity and inhibition of host innate immune responses including suppression of type I interferon responses, inhibition of RNA interference (RNAi), ^, and impairment of dendritic cell functions. ^, Another member of the replication machinery, VP30, is a transcriptional activator and plays a role in RNA transcription and replication. Recent crystal structure analysis showed VP-30 interaction with the C-terminal domain (CTD) of NP as critical for its functional activities. VP30 also suppresses host cell silencing of viral genes through interaction with the RNAi pathway. Advances in our molecular understanding of Ebola virus proteins have enabled structure–function-guided identification of susceptible targets for therapeutic intervention.

The major transcript of the GP gene is a secreted dimer, while the trimeric surface glycoprotein (GP) is synthesized only after addition of a nontemplated adenosine by the viral polymerase to yield the only surface virion protein. Ebola virus particles can infect almost every cell type in the body except lymphocytes. ^, Particle attachment is facilitated by several cellular surface proteins such as phosphatidylserine binding proteins, C-type lectins, DC-SIGN, TIM-1, and Ax1; and an unknown mechanism induces uptake of virions by macropinocytosis into intracellular compartments, a process that allows for large virus particles to travel to late endosomes and lysosomes where they are subjected to low-pH, and viral GP is proteolytically processed. Cathepsin cleavage of GP within these acidified internal compartments reveals an occluded receptor binding site, allowing binding of GP to the endosomal/lysosomal cholesterol transporter Niemann-Pick C1 (NPC1), the cellular receptor for filoviruses. ^{, , ,} A 3.4 Å X-ray structure of GP trimer lacking several regions was solved after cocrystallization with neutralizing antibody KZ52 that was isolated from an Ebola virus survivor. Several subsequent structures and mapping of the cathepsin cleavage site permitted speculation of a model for the cathepsin-cleaved GP protein and greater molecular understanding of GP function, and the mechanism of neutralization by antibodies targeting the cleaved form of GP. ^, Following this, the crystal structure of unliganded and uncleaved GP was solved at 2.2 Å resolution. Availability of this structure enabled studies on conformational changes that occur in GP after antibody or ligand binding.

Pathogenesis as It Relates to Prevention

Ebola virus enters the body through abrasions in the skin or exposure to mucosal surfaces, either by contact with bodily fluids, infected blood or close personal contact ( Fig. 22.2 ). It is thought that the virus traffics rapidly from these sites, possibly by transport on dendritic or other cells bearing DC-SIGN or related lectins, which bind the virus through their recognition of glycans on GP. Cells of the reticuloendothelial system in the liver, lung, and spleen are the initial targets of infection, with subsequent spread to the endothelium. In an effective immune-mediated prevention strategy, viral recognition would most likely have to occur early after infection by cells of the innate and adaptive immune system, as both CD8 ⁺ T-cell and humoral responses are detected in Ebola survivors. The mechanism of viral clearance is not fully understood but seems to require cellular immune responses. ^,

Diagnosis and Prevention

A diagnosis of Ebola virus disease (EVD) is suggested when a patient presents with symptoms of fever, flu-like illness, petechial rash, and disseminated bleeding. Definitive diagnosis is best made using a reverse transcription polymerase chain reaction (RT-PCR) detection assay. Additional documentation can be made through the measurement of IgM antibodies to viral proteins and various more specialized research assays. EVD diagnosis in the beginning of the 2014 West African outbreak heavily relied on RT-PCR which entailed a multistep process and required complex laboratory machinery, usually scarce in regions where Ebola outbreaks occur. This outbreak underscored the need for a quick, field-deployable methods to test samples from Ebola suspected individuals and their contacts. In response, a rapid genetic test called GeneXpert, previously used to diagnose pathogens including tuberculosis, HIV, malaria, was adapted for Ebola and enabled results to be obtained within hours. The Xpert Ebola assay, granted EUA by the FDA in 2015, involves a manual sample inactivation step after which the assay is fully automated and integrates all the steps required to detect Ebola virus in a given sample. ^, More recently, FDA has approved marketing of a rapid antigen test—OraQuick Ebola Rapid Antigen test for use in suspected EVD cases. OraQuick reveals the presence VP40 from any variant of Zaire Ebolavirus in blood within 15–25 minutes, although results should be followed-up with the RT-PCR detection assay that has greater sensitivity and specificity. Due to the emergence in humans of new species of Ebolavirus like the Bundibugyo ebolavirus and sequence diversity between Ebolavirus species, which rapid diagnostic tests could be limited in detecting, more than one diagnostic assay should be used in order to prevent false negatives.

In an outbreak situation, contact tracing is essential to halt spread of the disease through the population and has proven successful in the control of several outbreaks when vaccines and therapeutics were not available. ^, When an index case is identified, strict public health measures should be implemented, including use of adequate personal protective equipment (PPE), cleaning and decontamination of all equipment, isolation and quarantine to reduce transmission, and identification and testing of all contacts of the index case followed by immediate quarantine of any contacts that also develop signs of illness. Collaboration between health care workers and local community representatives is crucial to improve communication with the population and ensure dissemination and understanding of public health directives. In order to provide efficient basic health care during an outbreak, the number of treatment centers needs to be in accordance with the size of the susceptible population. Centers for Disease Control (CDC) and WHO established guidelines declare a country to be considered free of Ebola virus if during a 42-day window (twice the length of the maximum incubation period) no patients are diagnosed to be Ebola positive.

Incidence and Prevalence

Until 2014, outbreaks of Ebola virus infection occurred sporadically and remained localized, primarily in the countries of sub-Saharan Africa, including the Democratic Republic of Congo (DRC), the Republic of Congo (Brazzaville), Côte d’Ivoire, Sudan, and Uganda ( Fig. 22.3 ). Depending on the proximity to larger population centers and the virulence of the Ebolavirus species, outbreaks ranged in size from dozens to hundreds of fatalities, resulting in cumulative mortality numbers greater than 10,000 over the past four and a half decades. Outbreaks have mainly centered in the rain forest regions of northern Congo and the DRC, particularly but not exclusively at the beginning of the rainy season in October or November.

In more recent years, two protracted epidemics totaling tens of thousands of cases have occurred. The first large epidemic was declared in early 2014, lasting until June 2016, and affected three countries in West Africa—Guinea, Liberia, and Sierra Leone, causing 28,646 cases and 11,323 deaths according to official reports. Case numbers exceeded those of all previous outbreaks combined. While retrospective investigation placed the first case in December 2013 in rural Guinea at a triple-border area among the three countries, health authorities first recognized the outbreak in March 2014. The delay in the initial detection coupled with an initially modest response that was insufficient to track and contain case contacts allowed for a rapid spread of the virus across borders among the three West African countries. Due to international travel from the area of outbreak, Ebola viruspositive patients were identified for the first time in more distant countries, such as Nigeria, Mali, Spain, and the United States. Sequencing of clinical viral isolates from the outbreak revealed that the circulating strain, named Makona, was a new EBOV variant.

The second-largest EVD epidemic occurred in May 2018 in Northeast DRC, the 10th outbreak in this country since the virus was first reported in 1976. ^, Transmission continued through April 2020 totaling 3481 reported cases and 2229 deaths. Community distrust in the response and in being admitted to Ebola treatment facilities (with some patients traveling long distances to seek care and seeding new transmission chains in cities throughout the area) and insecurity due to an ongoing armed conflict in the region were the main difficulties faced in controlling this epidemic. This epidemic was DRC’s longest EVD outbreak and the second largest in number of reported cases in the world after the 2014–2016 West Africa epidemic.

These two epidemics were significant for the development and implementation of Ebola vaccines. The 2014–2016 West Africa epidemic spurred the advanced clinical development of several vaccine candidates, and an efficacy trial was completed at the final part of the outbreak in Guinea (first semester of 2015). ^,

On the other hand, the 2018–2020 DRC epidemic saw the first extensive use of Ebola vaccines in an outbreak and over 300,000 people were vaccinated.

Subsequent outbreaks were reported in DRC from June to November 2020 (Equateur Province, 130 cases) and in January–February 2021 (North Kivu Province, 12 cases) and in Guinea from January to June 2021 (Nzérékoré Region, 23 cases). ^{, ,} These outbreaks demonstrated a pattern of more frequent Ebolavirus reemergence in at least some areas of West Africa and DRC.