Immunogenicity and safety of two-visit, intradermal pre-exposure rabies prophylaxis simultaneously administrated with chimeric live-attenuated Japanese encephalitis vaccine in children living in rabies and Japanese encephalitis endemic country.

BACKGROUND: Reducing the number of doses required for pre-exposure prophylaxis (PrEP) would make it more feasible and cost-effective to implement in children at the highest risk of rabies exposure in Asia. We studied immune response of 2-site intradermal (ID) injection of rabies vaccine on days 0 and 28 for rabies PrEP simultaneously administrated with live-attenuated Japanese encephalitis chimeric virus vaccine (JE-CV) for children living in endemic area. RESEARCH DESIGN AND METHODS: Seronegative children (n = 49) aged 12-16 months were randomized 2:1 into two groups: Group A subjects were vaccinated with 0.1-mL ID injection of purified Vero cell rabies vaccine (PVRV), each at two sites on day (D) 0 and D28; Group B subjects were vaccinated with conventional 0.5-mL intramuscular PVRV on D0, D7 and D28. Both groups received one dose of JE-CV subcutaneously on D0 and D365. Rabies virus neutralizing antibody (RVNA) titers were measured on D0, D42 and D365 after vaccination; Japanese Encephalitis (JE) neutralizing antibody titers were determined on D0, D42, D365 and D379. RESULTS: All children had RVNA ≥ 0.5 IU/mL on D42 (geometric mean titers [GMTs] of RVNA 14.35 IU/mL [Group A] and 14.83 IU/mL [Group B], p > 0.05]). On D365, RVNA GMTs of subjects in group A and B were 1.50 IU/mL and 2.00 IU/mL (p > 0.05), respectively. All children had seroprotection following booster dose of JE-CV. There were no vaccine-related SAEs observed. CONCLUSION: The 2-site ID PrEP with PVRV on days 0 and 28 co-administrated with JE-CV are safe and immunogenic.

A randomized open-label trial of 2-dose or 3-dose pre-exposure rabies prophylaxis among Thai children.

BACKGROUND: World Health Organization changed the recommendation for pre-exposure rabies prophylaxis from 3-dose to 2-dose regimen in 2018. Given limited data of 2-dose regimens in pediatric population, this study aimed to compare the immunogenicity between 2-dose and 3-dose pre-exposure rabies immunization. METHODS: This study was conducted among healthy children aged 2-12 years. They were randomized to 2-dose vaccination (2D) on days 0 and 28 or 3-dose vaccination (3D) on days 0, 7, and 28. Purified Vero cell rabies vaccine (PVRV-Verorab™) was administered intramuscularly. Rabies virus neutralizing antibody (RVNA) titers were measured at 3 time points: 14-day after complete vaccination, 1-year pre-booster vaccination, and 7-day post-booster dose to mimic scenario of rabies exposure. RVNA titers ≥0.5 IU/ml were considered adequate antibody. T cell specific response to rabies vaccine antigen was measured using the interferon-gamma enzyme linked immunospot assay. RESULTS: From September to October 2017, 107 participants (51% males), 78 in 2D group and 29 in 3D group were enrolled. Median age was 5.8 years (IQR 4.4-7.3). All participants had RVNA titers ≥0.5 IU/ml after primary vaccination [GMT 2D: 18.6 (95%CI 15.9-21.8) and 3D: 16.3 (95%CI 13.2-20.1 IU/ml), p = 0.35]. At 1-year prior to receiving the booster, only 80% of the children in 2D group maintained RVNA titers ≥0.5 IU/ml compared to 100% of the children in 3D group (p = 0.01). However, all participants in both groups had RVNA ≥0.5 IU/ml at 7-day post booster vaccination [GMT 2D: 20.9 (95%CI 17.4-25.3) and 3D: 22.2 (95%CI 15.8-31.4) IU/ml (P = 0.75)]. The median number of IFN-γ secreting cells at 7-day post-booster dose was 98 and 128 SFCs per 106 PBMCs in the 2D and 3D groups, respectively (P = 0.30). CONCLUSIONS: Two-dose primary rabies immunization provided adequate antibody at post primary vaccination and post booster. The results support 2-dose regimen of pre-exposure rabies immunization in the pediatric population.

Evaluation of a rapid immunochromatographic test strip for detection of Rabies virus in dog saliva samples.

An immunochromatographic test strip for Rabies virus was evaluated with dog saliva samples. The test was initially validated against 237 dogs of known infection status, and then evaluated in the field with 1,290 live dogs. By validation of paired saliva-brain specimens obtained from dogs at necropsy, the saliva strip test was 94.4% specific and 93.0% sensitive when compared to the gold standard fluorescent antibody test (FAT) on brain smears. The sensitivity and specificity of a nested polymerase chain reaction (nPCR) assay using saliva were 100% compared to the FAT results. The performance of strip test with field saliva samples from street dogs had a specificity of 98.7% in comparison to nPCR as the reference method. As the strip test kit can potentially be used outside the laboratory and be applicable as an on-site testing assay, it represents a powerful screening tool for epidemiological surveys and disease control. The test could be useful for the surveillance of rabies in dogs and, in particular, be used to monitor the success of rabies control programs.

Use of latex agglutination test to determine rabies antibodies in production of rabies antisera in horses.

A therapeutic anti-rabies immunoglobulin for human use has been produced mainly in horses. The presently available seroneutralization test, the rapid fluorescent focus inhibition test (RFFIT), is laborious and rather difficult to carry out in horse farms. This study was undertaken to develop a simple latex agglutination test (LAT) for determining rabies antibodies in horse sera. LAT was validated by testing a total of 468 horse serum samples characterized by RFFIT. Of these, 253 of 260 samples with antibody titers of less than 100 IU/ml had agglutination score of 1+, whereas 174 of 208 samples with antibody titers equal to or greater than 100 IU/ml had agglutination scores of 2-4+. Results of LAT correlated with those of RFFIT (r = 0.87, p < 0.0001). LAT has the advantages of being rapid, simple to perform, easy to interpret, and applicable as an on-site testing tool for the estimation of rabies antibodies in horses.

Real-time PCR analysis of dog cerebrospinal fluid and saliva samples for ante-mortem diagnosis of rabies.

The use of a 10-day observation to determine whether a dog is rabid is standard practice. This study was conducted in order to look for evidence of rabies vius in saliva and cerebrospinal fluid (CSF) of suspected live rabid dogs at the time of quarantine by using a SYBR Green real-time RT-PCR based assay for the detection of rabies virus RNA. Saliva and CSF of dogs were collected once on the day of admission for the 10-day quarantine. All test dogs were or became ill and died of rabies within the observation period. Thirteen of 15 dogs (87%) had saliva samples that were positive for rabies RNA. Two dogs with furious rabies had negative saliva samples. Positive CSF samples were found in 4 of 15 dogs (27%) whose saliva samples were positive. The time from sample collection to result was less than 5 hours. Because virus may be absent or present at very low level in both clinical fluids, samples taken for ante-mortem diagnosis cannot definitively rule out rabies.

Genetic typing of feline rabies virus isolated in greater Bangkok, Thailand.

To study the molecular epidemiology of rabies virus that is prevalent among cats in greater Bangkok, Thailand, a total of 17 rabies virus isolates from cats were characterized and compared with 120 rabies virus isolates from dogs. Analyses were performed on the genetic polymorphism in the rabies virus nucleoprotein (N) gene. Rabies virus N gene of isolates was amplified by reverse transcriptionpolymerase chain reaction. The diversity of N gene was revealed by the restriction fragment length polymorphism (RFLP) method. The rabies virus isolates from cats could be classified into 5 types, designated as Dd I-Hf I, Dd II-Hf II, Dd III-Hf I, Dd IV-Hf I, and Dd IV-Hf III. Type Dd I-Hf I was encountered more frequently than the others. It was apparent that no less than five rabies virus types presented in the areas of Bangkok. Moreover, all five RFLP patterns were typical of those which had been observed in dogs. Our findings suggest that there had been viral transmission between the dogs and the cats.

Oral bacterial flora of dogs with and without rabies: a preliminary study in Thailand.

The authors studied the bacterial flora of the dog oral cavity and of bite wounds, Aerobic bacteria were isolated from mouth swabs of 16 normal and 5 rabid dogs as well as from infected dog-bite wounds from 18 patients. A total of 20 different microbial species were recovered from mouth swab cultures. The most frequently isolated organisms were Klebsiella pneumoniae ssp pneumoniae, Escherichia coli, Staphylococcus aureus, Citrobacter freundii, Enterobacter cloacae, Acinetobacter calcoaceticus, and Pasteurella species. There were no differences in the aerobic bacterial flora between rabid and nonrabid dogs. From the cultures of the bite wound swabs, the authors found that almost all of the organisms identified were part of the normal oral flora of the dog. One or more aerobic bacteria were isolated from the infected dog-bite wounds. Two patients had four, 3 had three, 4 had two, and 6 had one of the nine organisms in their wounds. The predominant species of bacteria involved in infection of bite wounds were, as follows: Staphylococcus aureus, Pasteurella multocida, E. coli, Moraxella species, Pasteurella canis, and Enterobacter cloacae. However, three wound cultures had no aerobic bacterial growth. The results of this study show that the infected bite wounds may contain a mixed bacterial flora that colonize human skin and the oral cavity of dogs.