A nucleoside-modified mRNA vaccine forming rabies virus-like particle elicits strong cellular and humoral immune responses against rabies virus infection in mice.

Rabies is a lethal zoonotic disease that threatens human health. As the only viral surface protein, the rabies virus (RABV) glycoprotein (G) induces main neutralizing antibody (Nab) responses; however, Nab titre is closely correlated with the conformation of G. Virus-like particles (VLP) formed by the co-expression of RABV G and matrix protein (M) improve retention and antigen presentation, inducing broad, durable immune responses. RABV nucleoprotein (N) can elicit humoral and cellular immune responses. Hence, we developed a series of nucleoside-modified RABV mRNA vaccines encoding wild-type G, soluble trimeric RABV G formed by an artificial trimer motif (tG-MTQ), membrane-anchored prefusion-stabilized G (preG). Furthermore, we also developed RABV VLP mRNA vaccine co-expressing preG and M to generate VLPs, and VLP/N mRNA vaccine co-expressing preG, M, and N. The RABV mRNA vaccines induced higher humoral and cellular responses than inactivated rabies vaccine, and completely protected mice against intracerebral challenge. Additionally, the IgG and Nab titres in RABV preG, VLP and VLP/N mRNA groups were significantly higher than those in G and tG-MTQ groups. A single administration of VLP or VLP/N mRNA vaccines elicited protective Nab responses, the Nab titres were significantly higher than that in inactivated rabies vaccine group at day 7. Moreover, RABV VLP and VLP/N mRNA vaccines showed superior capacities to elicit potent germinal centre, long-lived plasma cell and memory B cell responses, which linked to high titre and durable Nab responses. In summary, our data demonstrated that RABV VLP and VLP/N mRNA vaccines could be promising candidates against rabies.

Application prospects of the 2BS cell-adapted China fixed rabies virus vaccine strain 2aG4-B40.

BACKGROUND: Rabies is a fatal zoonotic disease whose pathogenesis has not been fully elucidated, and vaccination is the only effective method for protecting against rabies virus infection. Most inactivated vaccines are produced using Vero cells, which are African green monkey kidney cells, to achieve large-scale production. However, there is a potential carcinogenic risk due to nonhuman DNA contamination. Thus, replacing Vero cells with human diploid cells may be a safer strategy. In this study, we developed a novel 2BS cell-adapted rabies virus strain and analysed its sequence, virulence and immunogenicity to determine its application potential as a human diploid cell inactivated vaccine. METHODS AND RESULTS: The 2BS cell-adapted rabies virus strain 2aG4-B40 was established by passage for 40 generations and selection of plaques in 2BS cells. RNA sequence analysis revealed that mutations in 2BS cell-adapted strains were not located at key sites that regulate the production of neutralizing antibodies or virulence in the aG strain (GQ412744.1). The gradual increase in virulence (remaining above 7.0 logLD50/ml from the 40th to 55th generation) and antigen further indicated that these mutations may increase the affinity of the adapted strains for human diploid cells. Identification tests revealed that the 2BS cell-adapted virus strain was neutralized by anti-rabies serum, with a neutralization index of 19,952. PrEP and PEP vaccination and the NIH test further indicated that the vaccine prepared with the 2aG4-B40 strain had high neutralizing antibody levels (2.24 to 46.67 IU/ml), immunogenicity (protection index 270) and potency (average 11.6 IU/ml). CONCLUSIONS: In this study, a 2BS cell-adapted strain of the 2aG4 rabies virus was obtained by passage for 40 generations. The results of sequencing analysis and titre determination of the adapted strain showed that the mutations in the adaptive process are not located at key sequence regions of the virus, and these mutations may enhance the affinity of the adapted strain for human diploid cells. Moreover, vaccines made from the adapted strain 2aG4-B40 had high potency and immunogenicity and could be an ideal candidate rabies virus strain for inactivated vaccine preparation.

Toward the Development of a Pan-Lyssavirus Vaccine.

In addition to the rabies virus (RABV), 16 more lyssavirus species have been identified worldwide, causing a disease similar to RABV. Non-rabies-related human deaths have been described, but the number of cases is unknown, and the potential of such lyssaviruses causing human disease is unpredictable. The current rabies vaccine does not protect against divergent lyssaviruses such as Mokola virus (MOKV) or Lagos bat virus (LBV). Thus, a more broad pan-lyssavirus vaccine is needed. Here, we evaluate a novel lyssavirus vaccine with an attenuated RABV vector harboring a chimeric RABV glycoprotein (G) in which the antigenic site I of MOKV replaces the authentic site of rabies virus (RABVG-cAS1). The recombinant vaccine was utilized to immunize mice and analyze the immune response compared to homologous vaccines. Our findings indicate that the vaccine RABVG-cAS1 was immunogenic and induced high antibody titers against both RABVG and MOKVG. Challenge studies with different lyssaviruses showed that replacing a single antigenic site of RABV G with the corresponding site of MOKV G provides a significant improvement over the homologous RABV vaccine and protects against RABV, Irkut virus (IRKV), and MOKV. This strategy of epitope chimerization paves the way towards a pan-lyssavirus vaccine to safely combat the diseases caused by these viruses.

Development of mRNA rabies vaccines.

Rabies, primarily transmitted to humans by dogs (accounting for 99% of cases). Once rabies occurs, its mortality rate is approximately 100%. Post-exposure prophylaxis (PEP) is critical for preventing the onset of rabies after exposure to rabid animals, and vaccination is a pivotal element of PEP. However, high costs and complex immunization protocols have led to poor adherence to rabies vaccinations. Consequently, there is an urgent need to develop new rabies vaccines that are safe, highly immunogenic, and cost-effective to improve compliance and effectively prevent rabies. In recent years, mRNA vaccines have made significant progress in the structural modification and optimization of delivery systems. Various mRNA vaccines are currently undergoing clinical trials, positioning them as viable alternatives to the traditional rabies vaccines. In this article, we discuss a novel mRNA rabies vaccine currently undergoing clinical and preclinical testing, and evaluate its potential to replace existing vaccines.

Geographic Distribution of Rabies Virus and Genomic Sequence Alignment of Wild and Vaccine Strains, Kenya.

Rabies, a viral disease that causes lethal encephalitis, kills ≈59,000 persons worldwide annually, despite availability of effective countermeasures. Rabies is endemic in Kenya and is mainly transmitted to humans through bites from rabid domestic dogs. We analyzed 164 brain stems collected from rabid animals in western and eastern Kenya and evaluated the phylogenetic relationships of rabies virus (RABV) from the 2 regions. We also analyzed RABV genomes for potential amino acid changes in the vaccine antigenic sites of nucleoprotein and glycoprotein compared with RABV vaccine strains commonly used in Kenya. We found that RABV genomes from eastern Kenya overwhelmingly clustered with the Africa-1b subclade and RABV from western Kenya clustered with Africa-1a. We noted minimal amino acid variances between the wild and vaccine virus strains. These data confirm minimal viral migration between the 2 regions and that rabies endemicity is the result of limited vaccine coverage rather than limited efficacy.

Immunogenicity of 4-dose Essen intramuscular regimen for rabies post-exposure prophylaxis: A multi-center cross-sectional study in China.

BACKGROUND: The 4-dose Essen intramuscular (IM) regimen for rabies post-exposure prophylaxis (PEP) has been recommended by Advisory Committee on Immunization Practices (ACIP) and World Health Organization (WHO), but the large-sample clinical evidence is still limited. METHOD: Rabies virus neutralizing antibodies of 11,752 patients were detected from 409 rabies prevention clinics in 27 provinces in China. Patients with serum collected before or no later than 1 h after injection on the day of the fifth dose (day 28) of 5-dose Essen regimen were included in Group A to observe the immune efficacy of 4-dose Essen IM regimen, and patients with serum collected 14-28 days after injection of the fifth dose were included in Group B to observe the immune efficacy of 5-dose Essen IM regimen. RESULTS: Finally, 2351 cases met the inclusion and exclusion criteria, including 2244 cases in Group A and 107 cases in Group B. The antibody titer of Group A was higher than that of Group B [12.21 (4.15, 32.10) IU/ml vs. 9.41 (3.87, 27.38) IU/ml] (P = 0.002). In Group A, the median antibody titers were 4.01IU/ml, 11.63IU/ml and 29.46IU/ml in patients vaccinated with purified hamster kidney cell vaccine (PHKCV), purified Vero cell vaccine (PVRV), and human diploid cell rabies vaccine (HDCV), respectively, with statistical significance (P < 0.001). CONCLUSIONS: The 4-dose Essen IM regimen could provide satisfactory immune effect, and HDCV induced higher antibody titer than PHKCV or PVRV.

Rabies virus circulation in a highly diverse bat assemblage from a high-risk area for zoonoses outbreaks in the Brazilian Amazon.

Bats are the second most diverse order of mammals and play a central role in ecosystem dynamics. They are also important reservoirs of potentially zoonotic microorganisms, of which rabies virus is the most lethal among the bat-transmitted zoonotic pathogens. Importantly, recent outbreaks of human rabies have been reported from the Brazilian Amazon. Here we present a survey of bat species and rabies virus (RABV) circulation in a bat assemblage in the Marajó region, northern Brazil. Using data from mist-net captures and bioacoustic sampling, 56 bat species were recorded along the Jacundá River basin over a 10-day expedition in November 2022. For the investigation of RABV, we used the direct fluorescent antibody test (DFAT) and the rapid fluorescent focus inhibition test (RFFIT). In total, 159 bat individuals from 22 species were investigated for RABV. Five adults of the common vampire bat, Desmodus rotundus, showed RABV-specific antibodies in serum samples. Additionally, we report on local residents with injuries caused by D. rotundus bites and the occurrence of colonies of non-hematophagous bats from different species roosting inside human residences. This scenario raises concerns about the risks of new cases of human rabies and other zoonotic diseases associated with bats in the region and highlights the need for epidemiological surveillance and mitigation measures to prevent outbreaks of emerging infectious diseases.

Intracellular Pathogens: Infection, Immunity, and Intervention.

Intracellular pathogens comprise a diverse group of pathogens that all share a required location in a host cell to infect, survive, and replicate. Intracellular location allows pathogens to hide from host immune responses, avoid competition with other pathogens, mediate host cellular functions, replicate safely, and cause infection that is difficult to target with therapeutics. All intracellular pathogens have varying routes of infiltration into host cells and different host cell preferences. For example, bacteria Mycobacterium tuberculosis chooses to invade antigen-presenting cells, which allows them to moderate host antigen presentation to memory cells, whereas rabies virus prefers to invade neurons because they have pre-existing innate immunity protection systems. Regardless of the pathway that each intracellular pathogen follows, all share the capacity to cause disease if they succeed in entering host cells. Here, we give an overview of selected intracellular pathogens and infections they cause, immune responses they induce, and intervention strategies used to treat and control them.

Bat Bites and Rabies PEP in the Croatian Reference Centre for Rabies 1995-2020.

Seroprevalence of lyssaviruses in certain bat species has been proven in the Republic of Croatia, but there have been no confirmed positive bat brain isolates or human fatalities associated with bat injuries/bites. The study included a retrospective analysis of bat injuries/bites, post-exposure prophylaxis (PEP) and geographic distribution of bat injuries in persons examined at the Zagreb Antirabies Clinic, the Croatian Reference Centre for Rabies. In the period 1995-2020, we examined a total of 21,910 patients due to animal injuries, of which 71 cases were bat-related (0.32%). Of the above number of patients, 4574 received rabies PEP (20.87%). However, for bat injuries, the proportion of patients receiving PEP was significantly higher: 66 out of 71 patients (92.95%). Of these, 33 received only the rabies vaccine, while the other 33 patients received the vaccine with human rabies immunoglobulin (HRIG). In five cases, PEP was not administered, as there was no indication for treatment. Thirty-five of the injured patients were biologists or biology students (49.29%). The bat species was confirmed in only one of the exposure cases. This was a serotine bat (Eptesicus serotinus), a known carrier of Lyssavirus hamburg. The results showed that the bat bites were rather sporadic compared to other human injuries caused by animal bites. All bat injuries should be treated as if they were caused by a rabid animal, and according to WHO recommendations. People who come into contact with bats should be strongly advised to be vaccinated against rabies. Entering bat habitats should be done with caution and in accordance with current recommendations, and nationwide surveillance should be carried out by competent institutions and in close collaboration between bat experts, epidemiologists and rabies experts.

A phase 2b, Randomized, double blinded comparison of the safety and efficacy of the monoclonal antibody mixture SYN023 and human rabies immune globulin in patients exposed to rabies.

BACKGROUND: SYN023 is an anti-rabies monoclonal antibody mixture administered as part of post-exposure prophylaxis regimens. The rabies virus neutralizing antibody (RVNA) concentration generally accepted as an adequate immune response to vaccination is ≥ 0.5 IU/mL. METHODS: Within 54 h of potential rabies exposure, 448 patients in two risk substrata of WHO Category III exposure were randomized to receive either 0.3 mg/kg SYN023 or 0.133 mL/kg human rabies immunoglobulin (HRIG) injected in and around the wound site(s) plus a course of rabies vaccination. Patients were followed for safety and absence of rabies for ≥ 365 days. RESULTS: GMT RVNA was higher with SYN023 throughout the 2-week post-treatment period. In the primary analysis group (n = 368), 99.4 % of SYN023 recipients versus 4.5 % of HRIG recipients had protective RVNA levels on Day 4. On Day 8, 98.1 % SYN023 versus 12.2 % HRIG recipients were protected. The SYN023:HRIG ratio of geometric mean titer of RVNA (RVNA GMTs) on Day 8 (19.42) exceeded the 10 % superiority margin (P < 0.0001) indicating higher Day 8 RVNA with SYN023. On Day 99, the SYN023:HRIG RVNA GMT ratio (0.66) was below the non-inferiority margin of 20 % (P = 0.9485) suggesting some moderation of vaccine immune response by SYN023 relative to HRIG. The ratio of percent SYN023:HRIG recipients achieving RVNA ≥ 0.5 IU/mL on Day 99 (0.98) met the non-inferiority margin of 20 % (P = 0.013) indicating anti-rabies immune response with SYN023 was non-inferior to HRIG despite this effect. There were no probable/confirmed rabies cases in any patient. Study regimens were well tolerated. CONCLUSIONS: SYN023 provided higher RVNA than HRIG soon after rabies exposure. By Day 99 post-treatment, GM RVNA with SYN023 was lower than HRIG, however, the percent of SYN023 recipients with a protective response was not inferior at this time point. No rabies cases were reported in the study. The SYN023 safety profile was acceptable. CLINICALTRIALS: gov ID: NCT03961555.

A broadly applicable protein-polymer adjuvant system for antiviral vaccines.

Although protein subunit vaccines generally have acceptable safety profiles with precise antigenic content, limited immunogenicity can lead to unsatisfactory humoral and cellular immunity and the need for vaccine adjuvants and delivery system. Herein, we assess a vaccine adjuvant system comprising Quillaja Saponaria-21(QS-21) and cobalt porphyrin polymeric micelles that enabling the display of His-tagged antigen on its surface. The nanoscale micelles promote antigen uptake and dendritic cell activation to induce robust cytotoxic T lymphocyte response and germinal center formation. Using the recombinant protein antigens from influenza A and rabies virus, the micelle adjuvant system elicited robust antiviral responses and protected mice from lethal challenge. In addition, this system could be combined with other antigens to induce high titers of neutralizing antibodies in models of three highly pathogenic viral pathogens: Ebola virus, Marburg virus, and Nipah virus. Collectively, our results demonstrate this polymeric micelle adjuvant system can be used as a potent nanoplatform for developing antiviral vaccine countermeasures that promote humoral and cellular immunity.

Isolation and characterization of rabies monoclonal antibody charge variants.

Current postexposure prophylaxis of rabies includes vaccines, human rabies immunoglobulin (RIG), equine RIG, and recombinant monoclonal antibodies (mAb). In the manufacturing of rabies recombinant mAb, charge variants are the most common source of heterogeneity. Charge variants of rabies mAb were isolated by salt gradient cation exchange chromatography (CEX) to separate acidic and basic and main charge variants. Separated variants were further extensively characterized using orthogonal analytical techniques, which include secondary and tertiary structure determination by far and near ultraviolet circular dichroism spectroscopy. Charge and size heterogeneity were evaluated using CEX, isoelectric focusing (IEF), capillary-IEF, size exclusion chromatography, sodium dodecyl sulfate polyacrylamide gel electrophoresis, and western blotting. Antigen binding affinity was assessed by enzyme linked immuno-sorbent assay and rapid florescence foci inhibition test. Results from structural and physicochemical characterizations concluded that charge variants are formed due to posttranslational modification demonstrating that the charge heterogeneity, these charge variants did neither show any considerable physicochemical change nor affect its biological function. This study shows that charge variants are effective components of mAb and there is no need of deliberate removal, until biological functions of rabies mAb will get affected.

Three in one: An effective and universal vaccine expressing heterologous tandem RBD trimer by rabies virus vector protects mice against SARS-CoV-2.

The rapid emergence of Severe Acute Respiratory Syndrome Coronavirus type 2 (SARS-CoV-2) variants, coupled with severe immune evasion and imprinting, has jeopardized the vaccine efficacy, necessitating urgent development of broad protective vaccines. Here, we propose a strategy employing recombinant rabies viruses (RABV) to create a universal SARS-CoV-2 vaccine expressing heterologous tandem receptor-binding domain (RBD) trimer from the SARS-CoV-2 Prototype, Delta, and Omicron strains (SRV-PDO). The results of mouse immunization indicated that SRV-PDO effectively induced cellular and humoral immune responses, and demonstrated higher immunogenicity and broader SARS-CoV-2 neutralization compared to the recombinant RABVs that only expressed RBD monomers. Moreover, SRV-PDO exhibited full protection against SARS-CoV-2 in the challenge assay. This study demonstrates that recombinant RABV expressing tandem RBD-heterotrimer as a multivalent immunogen could elicit a broad-spectrum immune response and potent protection against SARS-CoV-2, making it a promising candidate for future human or veterinary vaccines and offering a novel perspective in other vaccine design.

Dog ecology and rabies control including canine vaccination coverage: Impacts from a survey in Madagascar.

BACKGROUND: Rabies virus (RABV; species Lyssavirus rabies) is causing one of the oldest zoonotic diseases known to mankind, leading to fatal encephalomyelitis in animals and humans. Despite the existence of safe and effective vaccines to prevent the disease, an estimated 99% of human rabies deaths worldwide are caused by dog-mediated rabies with children at the highest risk of infection. Rabies has been endemic in Madagascar for over a century, yet there has been little research evaluating local knowledge and practices impacting on the rabies control and prevention. Thus, this study was undertaken to better understand the dog ecology including canine vaccine coverage and to assess knowledge and practices of dog owners and veterinarians. METHODOLOGY: A cross-sectional study was conducted among 123 dog-owning households in thirteen fokontanys in Mahajanga from July 4 to September 13, 2016. Single and multi-member dog-owning households in the study area on the day of the interview were eligible for inclusion and purposively selected with the support of a local guide. The survey included a household questionnaire capturing information on the dog's demographics, husbandry practices, knowledge and practices towards rabies and its control measures; the dog ecology questionnaire collected dog characteristics, vaccination status and husbandry practices. All households that reported a dog bite incident, were invited to participate in a dog bite questionnaire. In addition, direct observations of roaming dogs were conducted to assess dog population demographics and to document behavioural characteristics. Two veterinarians were purposively selected and took part in an interview during the survey period, providing information on rabies control activities, including dog-care practices in the area. Descriptive and inferential data analyses were performed using Epi Info version 7.1.5.0 (CDC Atlanta, USA). RESULTS: We recorded a total of 400 dogs, of which 338 (84.5%) were owned amongst 123 households. More than half (67.8%) of owned dogs were between 1 to 5 years old and 95.6% were kept for guarding purposes. 45% of the surveyed dogs had free access to roam outside the premises. The majority (85.4%) of dog owners were knowledgeable that a dog bite could potentially transmit RABV to humans. 19 dog bites were reported and of these 73.6% were caused by the owner's or a neighbour's dog. In 6 of the 19 cases, children between 7 and 15 years of age were the victims. Dog vaccination coverage against rabies was 34% among owned dogs. Of the participants aware of a veterinarian, the majority (55/82) indicated that they accessed veterinarian services at irregular intervals. The main obstacles to vaccinations cited by dog owners were limited financial resources and difficulty accessing veterinary care. CONCLUSION: This study contributes to enhanced understanding of the dog ecology including canine vaccine coverage as well as knowledge and practices of dog owners in Madagascar. Most dogs in the study area were accessible for preventive vaccination through their owners, however only one third of the investigated canine population was vaccinated against rabies. Concerted national efforts towards rabies prevention and control should aim to address financial challenges and access to veterinary services.

Construction of Vero cell-adapted rabies vaccine strain by five amino acid substitutions in HEP-Flury strain.

Rabies virus (RABV) causes fatal neurological disease. Pre-exposure prophylaxis (PrEP) and post-exposure prophylaxis (PEP) using inactivated-virus vaccines are the most effective measures to prevent rabies. In Japan, HEP-Flury, the viral strain, used as a human rabies vaccine, has historically been propagated in primary fibroblast cells derived from chicken embryos. In the present study, to reduce the cost and labor of vaccine production, we sought to adapt the original HEP-Flury (HEP) to Vero cells. HEP was repeatedly passaged in Vero cells to generate ten- (HEP-10V) and thirty-passaged (HEP-30V) strains. Both HEP-10V and HEP-30V grew significantly better than HEP in Vero cells, with virulence and antigenicity similar to HEP. Comparison of the complete genomes with HEP revealed three non-synonymous mutations in HEP-10V and four additional non-synonymous mutations in HEP-30V. Comparison among 18 recombinant HEP strains constructed by reverse genetics and vesicular stomatitis viruses pseudotyped with RABV glycoproteins indicated that the substitution P(L115H) in the phosphoprotein and G(S15R) in the glycoprotein improved viral propagation in HEP-10V, while in HEP-30V, G(V164E), G(L183P), and G(A286V) in the glycoprotein enhanced entry into Vero cells. The obtained recombinant RABV strain, rHEP-PG4 strain, with these five substitutions, is a strong candidate for production of human rabies vaccine.

Protective Activity of Inactivated Rabies Vaccine Using Flagellin-Based Adjuvant.

Rabies is a zoonotic disease with high lethality. Most human deaths are associated with the bites received from dogs and cats. Vaccination is the most effective method of preventing rabies disease in both animals and humans. In this study, the ability of an adjuvant based on recombinant Salmonella typhimurium flagellin to increase protective activity of the inactivated rabies vaccine in mice was evaluated. A series of inactivated dry culture vaccine for dogs and cats "Rabikan" (strain Shchelkovo-51) with addition of an adjuvant at various dilutions were used. The control preparation was a similar series of inactivated dry culture vaccine without an adjuvant. Protective activity of the vaccine preparations was evaluated by the NIH potency test, which is the most widely used and internationally recommended method for testing effectiveness of the inactivated rabies vaccines. The value of specific activity of the tested rabies vaccine when co-administered with the adjuvant was significantly higher (48.69 IU/ml) than that of the vaccine without the adjuvant (3.75 IU/ml). Thus, recombinant flagellin could be considered as an effective adjuvant in the composition of future vaccine preparations against rabies virus.

Bayesian modeling of post-vaccination serological data suggests that yearly vaccination of dog aged <2 years old is efficient to stop rabies circulation in Cambodia.

Rabies control remains challenging in low and middle-income countries, mostly due to lack of financial resources, rapid turnover of dog populations and poor accessibility to dogs. Rabies is endemic in Cambodia, where no national rabies vaccination program is implemented. The objective of this study was to assess the short and long-term vaccination-induced immunity in Cambodian dogs under field conditions, and to propose optimized vaccination strategies. A cohort of 351 dogs was followed at regular time points following primary vaccination only (PV) or PV plus single booster (BV). Fluorescent antibody virus neutralization test (FAVNT) was implemented to determine the neutralizing antibody titer against rabies and an individual titer ≥0·5 IU/mL indicated protection. Bayesian modeling was used to evaluate the individual duration of protection against rabies and the efficacy of two different vaccination strategies. Overall, 61% of dogs had a protective immunity one year after PV. In dogs receiving a BV, this protective immunity remained for up to one year after the BV in 95% of dogs. According to the best Bayesian model, a PV conferred a protective immunity in 82% of dogs (95% CI: 75-91%) for a mean duration of 4.7 years, and BV induced a lifelong protective immunity. Annual PV of dogs less than one year old and systematic BV solely of dogs vaccinated the year before would allow to achieve the 70% World Health Organization recommended threshold to control rabies circulation in a dog population in three to five years of implementation depending on dog population dynamics. This vaccination strategy would save up to about a third of vaccine doses, reducing cost and time efforts of mass dog vaccination campaigns. These results can contribute to optimize rabies control measures in Cambodia moving towards the global goal of ending human death from dog-mediated rabies by 2030.

Comparative analysis of rabies pathogenic and vaccine strains detection by RIG-I-like receptors.

Rabies virus (RABV) is a lethal neurotropic virus that causes 60,000 human deaths every year globally. RABV infection is characterized by the suppression of the interferon (IFN)-mediated antiviral response. However, molecular mechanisms leading to RABV sensing by RIG-I-like receptors (RLR) that initiates IFN signaling currently remain elusive. Here, we showed that RABV RNAs are primarily recognized by the RIG-I RLR, resulting in an IFN response in the infected cells, but this response varied according to the type of RABV used. Pathogenic RABV strain RNAs, Tha, were poorly detected in the cytosol by RIG-I and therefore caused a weak antiviral response. However, we revealed a strong IFN activity triggered by the attenuated RABV vaccine strain RNAs, SAD, mediated by RIG-I. We characterized two major 5' copy-back defective interfering (5'cb DI) genomes generated during SAD replication. Furthermore, we identified an interaction between 5'cb DI genomes, and RIG-I correlated with a high stimulation of the type I IFN signaling. This study indicates that wild-type RABV RNAs poorly activate the RIG-I pathway, while the presence of 5'cb DIs in the live-attenuated vaccine strain serves as an intrinsic adjuvant that strengthens its efficiency by enhancing RIG-I detection thus strongly stimulates the IFN response.

Immunogenicity and Safety of a Purified Vero Rabies Vaccine-Serum Free, Compared With 2 Licensed Vaccines, in a Simulated Rabies Post-Exposure Regimen in Healthy Adults in France: A Randomized, Controlled, Phase 3 Trial.

BACKGROUND: A next-generation Vero cell rabies vaccine (PVRV-NG2) was developed using the same Pitman-Moore strain as in the licensed purified Vero cell vaccine (PVRV; Verorab) and the human diploid cell vaccine (HDCV; Imovax Rabies®). METHODS: This dual-center, modified, double-blind, phase 3 study evaluated the immunogenic non-inferiority and safety of PVRV-NG2 with and without concomitant intramuscular human rabies immunoglobulin (HRIG) versus PVRV + HRIG and HDCV + HRIG in a simulated post-exposure prophylaxis (PEP) regimen. Healthy adults ≥18 years old (N = 640) were randomized 3:1:1:1 to PVRV-NG2 + HRIG, PVRV + HRIG, HDCV + HRIG, or PVRV-NG2 alone (administered as single vaccine injections on days [D] 0, D3, D7, D14, and 28, with HRIG on D0 in applicable groups). Rabies virus neutralizing antibodies (RVNA) titers were assessed pre- (D0) and post-vaccination (D14, D28, and D42) using the rapid fluorescent focus inhibition test. Non-inferiority, based on the proportion of participants achieving RVNA titers ≥0.5 IU/mL (primary objective), was demonstrated if the lower limit of the 95% CI of the difference in proportions between PVRV-NG2 + HRIG and PVRV + HRIG/HDCV + HRIG was >-5% at D28. Safety was assessed up to 6 months after the last injection. RESULTS: Non-inferiority of PVRV-NG2 + HRIG compared with PVRV + HRIG and HDCV + HRIG was demonstrated. Nearly all participants (99.6%, PVRV-NG2 + HRIG; 100%, PVRV + HRIG; 98.7%, HDCV + HRIG; 100%, PVRV-NG2 alone) achieved RVNA titers ≥0.5 IU/mL at D28. Geometric mean titers were similar between groups with concomitant HRIG administration at all time points. Safety profiles were similar between PVRV-NG2 and comparator vaccines. CONCLUSIONS: In a simulated PEP setting, PVRV-NG2 + HRIG showed comparable immunogenicity and safety to current standard-of-care vaccines. CLINICAL TRIALS REGISTRATION: NCT03965962.

What would be the impact on the rabies risk of reducing the waiting period before dogs are imported? A modelling study based on the European Union legislation.

AIMS: Lyssavirus rabies (RABV) is responsible for a major zoonotic infection that is almost always lethal once clinical signs appear. Rabies can be (re)introduced into rabies-free areas through transboundary dog movements, thus compromising animal and human health. A number of measures have been implemented to prevent this happening, one of which is the waiting period (WP) after anti-rabies vaccination and serological testing. This WP ensures that antibodies assessed through the serological test are due to the vaccine, not to infection. Indeed, if antibodies are due to RABV infection, the dog should display clinical signs within this WP and would not therefore be imported. METHODS AND RESULTS: Within a framework of quantitative risk assessment, we used modelling approaches to evaluate the impact of this WP and its duration on the risk of introducing rabies via the importation of dogs into the European Union. Two types of models were used, a classical stochastic scenario tree model and an individual-based model, both parameterised using scientific literature or data specifically applicable to the EU. Results showed that, assuming perfect compliance, the current 3-month waiting period was associated with a median annual number of 0.04 infected dogs imported into the EU. When the WP was reduced, the risk increased. For example, for a 1-month WP, the median annual number of infected dogs imported was 0.17 or 0.15 depending on the model, which corresponds to a four-fold increase. CONCLUSION: This in silico study, particularly suitable for evaluating rare events such as rabies infections in rabies-free areas, provided results that can directly inform policymakers in order to adapt regulations linked to rabies and animal movements.