Rabies; cause, symptom, zoonotic risk and prevention in dogs

05/10/2022

Rabies Vaccines
Vaccines stimulate adaptive immunity, which is antigen-dependent and antigen-specific; therefore, rabies vaccination provides protection specifically from rabies infection. To be licensed in the United States, vaccines must protect at least 88% of vaccinated animals against challenge with virulent virus.5 Multiple vaccines are licensed for use in domestic animals, and inactivated (killed) vaccines are available for use in dogs . Recombinant virus-vectored products are available for cats, and oral modified live vaccines are available for wild animals, but these options are not superior to the inactivated vaccine for dogs.
Rabies Vaccination Recommendations
Rabies vaccine is a core vaccine according to the American Animal Hospital Association (AAHA) and is the only companion animal vaccine required by law in most states.6 In Canada, rabies vaccination of dogs is only required in the province of Ontario.6 Licensed veterinarians are legally required to vaccinate dogs for rabies in most U.S. states. Some states also allow canine rabies vaccines to be administered by licensed or certified veterinary technicians or lay people working under the direct supervision of a licensed veterinarian.3 After vaccination, a rabies vaccination certificate verifying the vaccinate, the health of the vaccinate, the product used, the booster interval, and the administering veterinarian is issued to identify vaccinated dogs.
While there are no clearly defined vaccination site recommendations for dogs as there are in cats, noting which vaccines are administered where is important in case of an adverse reaction. Inactivated rabies virus vaccines can be administered either intramuscularly or subcutaneously. The first vaccination is given per label recommendations at a minimum of 3 months of age due to the potential interference by maternally derived antibodies and a relatively poor immune response in the young.2,5 Regardless of the age of the first vaccination, a booster vaccine is repeated 1 year after the initial vaccine, with subsequent boosters given annually or triennially depending on the labeled duration of immunity of the vaccine used and local public health regulations.2,3,5 A recently published challenge study with virulent rabies virus demonstrated that the duration of immunity conferred by rabies vaccine extends beyond the 3-year label, which may change vaccination schedules in the future.7

Regardless of booster interval, within 28 days after initial vaccination, a peak rabies virus antibody titer is expected, and the animal can be considered immunized.3 The initial vaccine schedule (1 vaccine followed by a booster 1 year later) is inconsistent with other inactivated vaccine protocols, in which 2 sequential doses administered 2 to 4 weeks apart are required to stimulate adequate immunity. Like other inactivated vaccines, the initial dose of a rabies vaccine serves as the “priming” dose. If a dog, after having received only 1 dose of rabies vaccine, is subsequently exposed to virulent rabies virus, exposure to the virulent virus then serves as the second, or immunizing, “dose.” Because the onset of signs of rabies is slow after exposure, there is adequate time for a protective, humoral immune response to develop.6

While no vaccine is 100% effective, rabies infection is rare in vaccinated dogs. In one study, 4.9% of cases of rabid dogs had a history of prior rabies vaccination.8 Vaccination efforts can provide protection for dogs exposed to potentially rabid animals even if they are overdue for a rabies booster vaccine. Results comparing the anamnestic response rate in currently vaccinated animals versus overdue animals indicated that dogs with an out-of-date vaccination status were not inferior in their antibody response following booster rabies vaccination compared with dogs with a current vaccination status.9 The findings of this study led to changes in recommendations for postexposure management of dogs exposed to an animal confirmed or suspected to be rabid, and to advisement that an animal is currently vaccinated and considered immunized immediately after any booster vaccination.3
Rabies Postexposure Management
A dog that has been exposed to a confirmed or suspected rabid animal should immediately receive veterinary medical care for assessment, wound cleansing, and booster vaccination.3 If the exposed dog is current on rabies vaccination, it should be quarantined and observed by the owner for 45 days.3 An exposed dog that is overdue for a rabies vaccine and has documentation of a past vaccine can also be quarantined and observed by the owner for 45 days. If the exposed dog is overdue for a booster vaccine without appropriate documentation, local public health authorities should be consulted to determine the quarantine period and the utility of serologic testing to provide proof of an anamnestic response
If the exposed dog has never been vaccinated for rabies, it should be euthanized immediately; however, if the owner is unwilling to euthanize, strict quarantine for 4 months or longer without direct contact with people may be an option after consultation with local public health authorities.3

For any of these situations, if at any time during the quarantine period signs suggestive of rabies develop (e.g., paralysis or seizures), the animal should be euthanized and submitted for rabies testing.3

Adverse Rabies Vaccine Reactions
Concerns about adverse vaccine reactions and overvaccination have occasionally caused reluctance to vaccinate dogs. The following are potential vaccine adverse reactions described in dogs and cats:4
Injection-site reactions
Allergic or immune-mediated reactions
Tumorigenesis
Vaccine-induced immunosuppression
Anaphylaxis
Injection-site sarcomas
The most common vaccine-associated adverse events reported in dogs in one study were allergic reactions, local vaccine-site reactions, and nonspecific systemic signs (fever, lethargy, or anorexia).10 In the same study, the reported rate of adverse events within 3 days of vaccination was 38.2 of 10 000 dogs, although it was noted that adverse events are likely underreported.10 If an acute adverse vaccine reaction is suspected, treatment may include antihistamines (e.g., diphenhydramine), anti-inflammatories (steroids or nonsteroidal anti-inflammatory drugs, depending on the reaction), or more targeted therapy (e.g., fluids and epinephrine for anaphylaxis). All adverse vaccine events should be reported to the vaccine manufacturer and the U.S. Department of Agriculture (USDA) Animal and Plant Health Inspection Service Center for Veterinary Biologics

05/10/2022

Clinical Signs of Rabies
Rabies virus infection has classically been divided into 2 main clinical manifestations: the excitatory (“furious”) and the paralytic (“dumb”) forms.2,5 This classification system is arbitrary because rabies can be quite variable in its presentation, and not all animals progress through all 3 clinical stages: prodromal, excitatory, and paralytic.2,5
Initial clinical signs (the prodromal period) are nonspecific and can include minor behavioral changes. These signs, if recognized, usually last 2 to 3 days in dogs.2,5 During the excitatory phase, dogs suddenly become vicious and behave erratically. They may become restless and irritable and have heightened sensitivities to visual and auditory stimuli.5 This stage usually lasts 1 to 7 days; however, some dogs may progress directly from the prodromal stage to the paralytic stage.2 This final stage is characterized initially by weakness and eventually by paralysis. The limb where the wound occurred is initially affected, followed by progression of paralysis to the neck and head. The paralytic phase usually lasts 2 to 4 days and ends in death from respiratory failure.2,5 The course of rabies typically lasts 3 to 8 days in dogs.2
Diagnosis and Treatment of Rabies
There is no premortem test or effective treatment for rabies in dogs. Dogs that have sustained a bite from an unknown or unvaccinated animal should be immediately vaccinated and quarantined or euthanized, as recommended (TABLE 1).3 Elimination of feral animals or wildlife populations that harbor rabies is not economically feasible, nor is it socially or ecologically acceptable. As such, control through immunization is paramount to protecting dogs.

05/10/2022

Rabies is a deadly zoonotic disease of mammals. It is now rarely reported in dogs in the United States owing to widespread vaccination programs effectively eliminating the canine variant. However, education of the public regarding the need to vaccinate dogs and the continued diligent practice of rabies vaccination in dogs will help to prevent re-emergence of this fatal zoonotic infection.
Public Health Significance Of Rabies
Rabies is responsible for tens of thousands of human deaths annually, with infection usually following a bite or scratch from a rabies-infected animal. According to the World Health Organization, rabies-infected dogs serve as the main source of human rabies transmission globally, contributing up to 99% of all rabies cases in humans, while in the Americas, bats are the most common source of human infections.1 In the U.S., the dog rabies variant, once endemic in small animals, has now been nearly eradicated.2 However, wildlife species, which carry their own rabies variants, present a constant danger of reintroduction of rabies to dogs.
Rabies Disease
Rabies is caused by viruses in the genus Lyssavirus. Virtually all mammals are susceptible to rabies infection, including domestic animals (e.g., dogs, cats, cattle, horses) as well as wild mammalian reservoir populations (e.g., raccoons, skunks, foxes, bats) where multiple rabies virus variants are maintained.3,4 Regardless of the species affected, rabies causes an acute, progressive encephalitis.
Pathogenesis of Rabies
Transmission most commonly occurs through a bite or scratch from an infected animal. The incubation period is highly variable and depends on the age of the individual bitten, the degree of innervation of the bite site, the distance from the point of inoculation to the spinal cord or brain, and the variant and amount of virus introduced, as well as other factors.5 In domestic animals, the incubation period is generally 3 to 12 weeks, but can range from several days to months, rarely exceeding 6 months.3 Once the virus is introduced in the body, it enters peripheral nerves and subsequently gains entry into the central nervous system (CNS). After replication within the CNS, the virus moves outward to other body tissues via the peripheral, sensory, and motor nerves

05/10/2022

Vaccination Requirements
All dogs vaccinated against rabies for the first time must be vaccinated at least 4 weeks (28 days) before traveling.
Puppies must NOT be vaccinated against rabies before they are 3-months (12 weeks or 84 days) old. The rabies certificate must include the puppy’s age or date of birth.
Adult dogs (15 months or older) must show a history of previous rabies vaccination with at least one vaccine given after they were 3-months old and one current booster rabies vaccination. With this record, adult dogs don’t need to wait 4 weeks before traveling.

16/08/2022

Rabies is a viral disease that is transmitted through the saliva or nervous system tissues of an infected mammal to another mammal. The rabies virus infects the central nervous system and causes severely distressing neurological symptoms, disease in the brain, and, ultimately, death.
Rabies is a zoonotic disease, which means that it can pass from other animals to humans. Rabies is the deadliest disease on earth with a 99.9% fatality rate.
How is Rabies transmitted?
Infection usually occurs following a bite or scratch from an infected animal, and the rabies virus is transmitted through the saliva of the host animal. Most often, the virus is passed to human populations through dogs (95% of worldwide cases), but the other species have been identified as important reservoirs of the rabies virus, including bats, raccoons, skunks, foxes, and coyotes.
While not as prevalent, transmission can also occur when saliva comes into direct contact with mucous membranes (i.e., eyes, nose, mouth), and very rarely through inhalation of aerosolized saliva, and through corneal and internal organ transplantation.
There have been cases where butchering raw meat from rabid animals has transmitted the infection, presumably through infectious neural tissue coming into contact with open wounds in the skin.
Where does Rabies occur?
Rabies is found on every continent except Antarctica. In Africa, the Middle East, and Asia, canine rabies is a wide-spread problem and contributes to over 90% of rabies cases world-wide. In developed nations and island nations, rabies is either well-controlled amongst domesticated animals, or it is non-existent.
Today, over 90% of rabies deaths are in Africa, Asia and the Middle East where canine rabies is widespread.
How big is the problem in the world?
Rabies kills people, domestic animals (such as dogs and cattle) and causes financial hardship when people have to pay for vaccination after bite wounds. It is estimated that more than 5.5 billion people live at daily risk of rabies.
How many people die each year from Rabies?
Estimates suggest that over 5.5 billion people live with the daily risk of rabies, with 59,000 deaths every year. Over 95% of these deaths are in Africa and Asia, with the majority occurring from rabid dog bites. Around half of the people who die are children.
In western nations, deaths are rare (1-3 deaths per year in the United States), with cases of clinical rabies occurring typically in people who did not realize that they had been exposed.
Is Rabies always fatal?
Yes, there is no effective treatment once clinical symptoms appear. Rabies has the highest case-fatality rate of any infectious disease known to man, because there is no proven cure or treatment available once there are signs of an infection.
However, if proper medical treatment (post-exposure prophylaxis, PEP) is received immediately after exposure to the bite or scratch of a rabid animal, rabies infection can be halted before symptoms of the disease are present, and the disease can be prevented.

03/06/2022

13/05/2022

11/08/2021

Pathogenesis
Rabies virus is inoculated into muscle and subcutaneous tissues in the saliva of a biting animal (Figure 1). There is a delay in movement of the virus at the site of inoculation during the incubation period that lasts for weeks to months. Rabies virus binds to nicotinic acetylcholine receptors at the neuromuscular junction and travels toward the spinal cord within axons of peripheral nerves by retrograde fast axonal transport at a rate of approximately 50–100 mm per day. The virus disseminates within axons in the CNS along neuroanatomical pathways. Rabies virus replicates in neurons and causes neuronal dysfunction by uncertain mechanisms, which is likely responsible for the clinical features and fatal outcome of the disease. Behavioral changes occur in rabies, which usually leads to transmission by biting in infected animals. Subsequently, there is centrifugal (away from the CNS) spread along nerves to multiple organs, including the salivary glands in animals that transmit the virus. Rabies virus is secreted in the saliva in vectors and transmission occurs to other hosts by biting. Bats, raccoons, skunks, and foxes are important rabies vectors in North America, and dogs are the most important vector worldwide. Bat bites may not be recognized and there may even be no known contact with bats.

11/08/2021

Lyssaviruses
Rabies virus (RABV) is the prototype virus of the genus Lyssavirus (from the Greek lyssa meaning “rage”) in the family Rhabdoviridae (from the Greek rhabdos meaning “rod”) of the order Mononegavirales (MNV). RABV, the causative agent of classic rabies in animals and humans, is a highly neurotropic virus in the mammalian host invariably causing a fatal encephalomyelitis once the infection is established and has reached the brain. RABV is distributed worldwide among specific mammalian reservoir hosts comprising various carnivore and bat species. Fifteen other lyssaviruses, which share certain morphological and structural characteristics with RABV, have been identified. Only 6 of the 16 currently recognized lyssaviruses within the Lyssavirus genus have caused a rabies-like encephalomyelitis in humans. Of note, of the 16 lyssaviruses recognized only RABV has multiple host reservoirs, while the other lyssaviruses are exclusively associated with bat reservoirs (Marston et al., 2018). The 16 lyssaviruses segregate into 2 phylogroups based on phylogenetic analyses. Phylogroup I includes the classic (prototype) RABV, Duvenhage virus (DUVV), European bat lyssavirus, type 1 (EBLV-1), and type 2 (EBLV-2), Australian bat lyssavirus (ABLV), Aravan virus (ARAV), Khujand virus (KHUV), Irkut virus (IRKV), Bokeloh bat lyssavirus (BBLV), Ikoma lyssavirus (IKOV), Lleida bat lyssavirus (LLEBV), and Gannoruwa bat lyssavirus (GBLV), the newest species to be characterized and considered an independent species with phylogroup I (Gunawardena et al., 2016; Hanlon et al., 2005; Kuzmin, Niezgoda, et al., 2008). All phylogroup I lyssaviruses are transmitted by bats; only RABV is adapted to and spread by carnivores as their reservoir host. The evolutionary relationship between bat transmitted and carnivore transmitted lyssaviruses is not well understood (Rupprecht, Kuzmin, & Meslin, 2017).

Phylogroup II includes LBV, Mokola virus (MOKV), and Shimoni bat virus (SHIBV), the newest species to be characterized and considered an independent species within phylogroup II (Kuzmin et al., 2010). West Caucasian bat virus (WCBV), which does not crossreact serologically with any members of the two phylogroups, could tentatively belong to a third phylogroup (Freuling et al., 2011) (See Chapters 4 and 5). Phylogenetic analyses suggest that all lyssaviruses have originated from a precursor bat virus (Rupprecht et al., 2017).

Lyssavirus species share many of the biologic and physicochemical features that are associated with other viruses of the Rhabdoviridae family. These include the bullet-shaped virus morphology, helical nucleocapsid (NC) or ribonucleoprotein (RNP) core, and general organization of the viral RNA (vRNA) genome and structural proteins. In contrast to all other rhabdoviruses, however, lyssaviruses are not transmitted by insect vectors and have adapted to direct transmission. The five structural proteins of the lyssavirus particle (virion) include a nucleocapsid protein (N), phosphoprotein (P), matrix protein (M), glycoprotein (G), and RNA-dependent RNA polymerase or large protein (L). These lyssavirus proteins generally share many of the biologic functions that the same viral proteins have in other rhabdoviruses. Some of the structural proteins of lyssaviruses, on the other hand, can differ dramatically in their antigenic properties and in their post-translational modifications to convey different, often specific properties that distinguish lyssaviruses from other rhabdoviruses. Lyssaviruses, like other rhabdoviruses, also use similar mechanisms to enter susceptible cells (albeit, they may use different receptors) express and replicate their genome RNA, assemble and release mature progeny virions from the plasma membrane or internal membranes of infected cells.

11/08/2021

Rabies virus is particularly useful for the study of neuronal circuits because of its ability to spread transsynaptically in the retrograde direction.63–65 Although rabies viruses are restricted to nonpromoter-based targeting because of their negative-strand RNA nature, infected cells remain viable for weeks66 and the virus can amplify from even a single viral particle.67 In 2007, Wickersham and his collaborators developed a rabies virus variant that spread only to the cells monosynaptically connected to the infected neurons, allowing tests of the functional role of neurons one synapse away from the initially infected cell

11/08/2021

Innate immune response in the periphery
RABV is inoculated in the skin, subcutaneous tissues, or in muscle by bites or scratches. The entry of the virus by the host is rapidly detected by host defense mechanisms in the periphery. The IFN response triggered at the site of entry has an antiviral effect. Evidence has been indirectly obtained by comparing the viral load in the thigh muscle of two groups of mice, parental mice and mice lacking the type I IFN receptor (IFNAR) after injection of the hindlimb with CVS. The viral load measured by the accumulation of viral RNA was increased in mice lacking IFNAR compared to parental strain of mice (Chopy, Detje, Lafage, Kalinke, & Lafon, 2011). This observation suggests that some viral particles might be readily eliminated at this early step of infection.

The cells-producing type I IFN at the site of the injection are likely sentinel cells (pDCs or macrophages). Infection of these cells is dispensable to mount an innate immune response, as shown in vitro when macrophages could be activated by addition of inactivated RABV to the culture (Nakamichi, Inoue, Takasaki, Morimoto, & Kurane, 2004). Nevertheless, there is in vitro experimental evidence that RABV can infect bone-marrow-derived conventional DCs (cDCs) and macrophages in vitro. Despite nonproductive infection, RABV triggers the production of IFNs, cytokines, and chemokines in these cells (Faul et al., 2010; Nakamichi et al., 2004). In cell culture, maturation of cDCs in the presence of RABV is controlled by IFN, which production might rely on the recognition of intracytoplasmic RABV RNAs through RIG-I and mda-5 receptors and not TLR7 (Faul et al., 2010), a characteristic of cDCs (Eisenacher, Steinberg, Reindl, & Krug, 2007).

However, it seems that virulent RABV strains trigger weaker DCs activation versus attenuated strains. Experiments were performed with highly attenuated recombinant RABV (rRABV) genetically modified to allow the expression of chemokines or multiple copies of G protein in a search for more effective rabies vaccines. These RABVs trigger a stronger activation of cDCs in the periphery than the parental rRABV strains (Li, McGettigan, Faber, Schnell, & Dietzschold, 2008; Pulmanausahakul et al., 2001; Wen et al., 2011). Experiments comparing the capacity of dog RABV strains (DRV-NG11 from a dog in Nigeria, or DRV from a dog in Mexico RABV) and of highly attenuated recombinant RABV strains (CVS-B2C or TrisGAS strain) to activate DCs, showed that in contrast to the attenuated RABV strain, the dog RABV strains poorly stimulate the DCs in vitro and as a consequence trigger a lower antibody immune response than the attenuated RABV strains (Gnanadurai et al., 2015; Yang et al., 2015). The poor activation of DCs by the dog RABV strain from Mexico results from the low binding of this virus to DCs (Yang et al., 2015). These data suggest that DC activation inversely correlates with the pathogenicity of the RABV strains.

At the site of injection, muscle cells can be infected as observed in experimental rabies in skunks with a wild-type RABV strain (Charlton & Casey, 1979, 1981). In vitro, muscle infection could be reproduced in the mouse muscle myoblast G-8 cells using the Japanese Nishigahara RABV strain and its derivative, the Ni-CE strain causing lethal and subclinical infections, respectively, after intramuscular injection in mice. Compared to NI-CE, only the Nishigahara RABV strain triggers stable viral replication in muscle cells (Yamaoka et al., 2013, 2017). It has been demonstrated that the muscle cell infection by the Nishigahara RABV strain and entry into the peripheral nerve are promoted through the INF antagonist of the phosphoprotein of the Nishigahara RABV strain, mainly by the N-terminally truncated isoforms of the phosphoprotein (Okada et al., 2016; Yamaoka et al., 2013, 2017). These observations suggest that the capacity of a RABV strain to infect muscle cells correlates with the pathogenicity of the strain.

Therefore, it seems that virulent RABV strains use selected mechanisms allowing them to escape, at least partially, the local host innate immune response early after they have entered the body. These mechanisms could contribute to limit early elimination of viral particles and promote virus entry into the NS.

11/08/2021

Pathogenesis
Rabies virus is usually transmitted in the saliva of a biting animal, although transmission has rarely been documented from aerosolized virus in caves and laboratories and by corneal transplantation. After inoculation of rabies virus into muscles or subcutaneous tissues, there may be amplification of the virus in muscle at the site of exposure, accounting for the long incubation period. Rabies virus binds to the nicotinic acetylcholine receptor and spreads within axons of peripheral nerves by retrograde fast axonal transport at a rate of about 50 to 100 mm per day and reaches the spinal cord and brain and disseminates throughout the CNS along neuroanatomic pathways. Rabies virus replicates in neurons and causes neuronal dysfunction by uncertain mechanisms, which is likely responsible for the clinical features and fatal outcome of the disease. Behavioral changes occur in rabies, which leads to transmission by biting in infected animals. There is centrifugal spread of the virus from the brain to the salivary glands, which is important for transmission of virus in rabies vectors. Rabies virus may be present in an animal's saliva before the onset of symptoms or signs of rabies.

11/08/2021

Macroevolutionary dynamics of rabies virus. Maximum likelihood phylogenetic tree of the nucleoprotein gene showing relationships among major RABV lineages. Lineage names follow Troupin et al. (2016), with additional annotations following Streicker et al. (2010) and Velasco-Villa et al. (2017). Branch colors indicate the suspected reservoir association. Host genera are abbreviated as follows: A = Artibeus; An = Antrozous; C = Canis; Ca = Callithrix; Ce = Cerdocyon; D = Desmodus; H = Histiotus; Hr = Herpestes; L = Lasiurus; La = Lasionycteris; M = Myotis; Me = Mephitis; Mel = Melogale; Mol = Molossus; Ny = Nyctinomops; N = Nycticeius; P = Perimyotis; Pa = Parastrellus; Pt = Plecotus; S = Spilogale; T = Tadarida; V = Vulpes.

11/08/2021

Rabies virus (RABV) thrives in a realm of extremes and paradoxes that challenge our view of how viruses evolve and sustain transmission in natural and novel hosts. RABV is universally lethal yet able to maintain itself without driving populations of its generally long-lived and slow-reproducing bat and carnivore hosts extinct. RABV has the biological capacity to infect any mammalian species and cross-species transmission events are readily observed in nature, yet diverse viral variants rely on single host species for their independent, long-term perpetuation. Finally, despite apparent host specificity over ecological timescales, the evolutionary history of RABV is dominated by host shifts both within and between bats and carnivores (Fig. 3.1). This potential to establish transmission cycles in novel host species remains largely unpredictable. RABV achieves each of these seemingly improbable feats with a nonsegmented, single-stranded genome of approximately 12,000 nucleotides encoding only five genes (N, P, M, G, and L). Moreover, like many other RNA viruses, the molecular evolution of RABV is inherently constrained. The absence of a proofreading mechanism in the virus-encoded RNA polymerase leads to high rates of spontaneous mutation (Drake, 1993). While these mutations potentially provide a diverse population of genetic variants upon which selection might act (sometimes referred to as a “quasispecies” or “mutant spectrum”), most mutations will be deleterious and pleiotropic. Thus, despite constant generation of novel genetic diversity, pathways for adaptive evolution are likely to be highly constrained in RABV (Holmes, Woelk, Kassis, & Bourhy, 2002). How has a virus with such an extreme life history, small genome, and constrained evolution become a global threat to human and animal health?

11/08/2021

After targeted binding of virus to its receptor(s) on host cells, virus is internalized by endocytosis. RABV, like VSV, may also enter the cell through coated pits and uncoated vesicles (viropexis or pinocytosis), which often incorporate several (two to five) virions per vesicle (Tsiang, Derer, & Taxi, 1983). As part of the internalization process, whether by receptor-mediated endocytosis (via the endocytic pathway) or through coated pits, fusion between the viral and endosomal membranes is activated in the acidic environment (pH 6.3–6.5) within the endosomal compartment. At the threshold pH for fusion activation, a series of specific and discrete conformational changes in RABV G takes place whereby it assumes at least three structurally distinct ‘conformational’ states; for review see (Albertini et al., 2012).

Prior to binding to the cellular receptor, the G on the virion surface is in its ‘native’ state. After the virus attaches to the receptor and is internalized, the G is ‘activated’ to a hydrophobic state, which enables it to interact with the hydrophobic endosomal membrane. Upon entering the endosomal compartment and low pH environment of the cellular compartment, the fusion capacity of the G is activated via a major structural change in the G that exposes the fusion domain, which interacts with the target cell membrane. By further rearrangements in the G, the viral and cell membrane are brought into close vicinity such that (via hemifusion) a fusion pore may develop (Albertini et al., 2012).

The low pH-induced exposition of the fusion domain, which is thought to lie between amino acids 102 and 179 (Durrer, Gaudin, Ruigrok, Graf, & Brunner, 1995), is not to be confused with the proposed fusogenic domain (amino acids 360–386) on the RABV G (Morimoto, Ni, & Kawai, 1992). The functional state of the RABV G in which it acquires fusogenic activity is correlated with at least one specific conformational epitope. This epitope, which appears to be formed by combining two separate regions, the neurotoxin-like region (residues 189–214) of RABV G and the conformational antigenic site III (residues 330–340), is abrogated when the G is exposed to acidic conditions (Kankanamge, Irie, Mannen, Tochikura, & Kawai, 2003; Sakai et al., 2004). The transition from virus interaction to generating the fusion pore is a high-cost energy step and depends on the integrity and correct folding of the G trimers directly involved in the fusion process. It has been shown that more than one trimer of G is required to build a competent fusion site.

After low-pH fusion, the G assumes a reversible ‘fusion-inactive’ conformation, which makes the G monomer appear longer than the ‘native’ conformation and assume selective antigenic distinctions (Kankanamge et al., 2003). The fusion-inactivated G, which is no longer relevant to the fusion process is highly sensitive to cellular proteases and appears to be in a dynamic equilibrium with the ‘native’ G that is regulated by lowering and raising the pH (Gaudin, Tuffereau, Segretain, Knossow, & Flamand, 1991; Gaudin, Raux, Flamand, & Ruigrok, 1996). Interestingly, the fusion-inactive conformation serves the G in another capacity. During nascent viral protein synthesis, the G assumes an ‘inactive state-like’ conformation, protecting the G posttranslationally from fusing with the acid nature of Golgi vesicles while it is transported through the Golgi stacks to the cell surface. At the cell surface, the G acquires its ‘native’ conformation and structure (Gaudin et al., 1999). The MAbs that recognize specific low pH-sensitive conformational epitopes of the G can identify certain acid-induced conformational changes, as well as detect the various stages of nascent G monomer folding and its association with molecular chaperones like BiP and calnexin (Gaudin, 1997; Kankanamge et al., 2003; Maillard & Gaudin, 2002).