Rural Rabies Prevention Project - A ‘One Health’ Experiment in India: An Overview.

Aim: To assess the feasibility of implementing "one health approach" to prevent human rabies and control animal rabies in a rural community. Study Design: Health services research in a rural setting. Place and Duration of Study: A medical college and a veterinary college along with an animal welfare organization delivered a wide array of "integrated services" in three villages' i.e. Kumbalagodu, Thagachikuppe and Gerupalya near Bangalore, India comprising a population of 10,220 persons for a period of two years from December, 2009 to November, 2011. The nearby three villages of Ramohally, Vinayakanagara and Bhimanakuppe with a population of 6,023 persons formed the control group, with no project inputs. Methodology: This consisted of household surveys at the beginning and end of two years ; rabies awareness campaigns; clinical and laboratory surveillance of rabies in dogs; rabies post-exposure prophylaxis (PEP) in humans; pre-exposure rabies prophylaxis (PrEP) by intradermal route in school children, pet dog owners and veterinarians ; mass dog vaccination and deworming; and sero-surveillance in both dogs and humans. Results: The ratio of veterinary and medical manpower in study villages was 1: 11 and dog to human ratio was 1: 23.The information, education and communication materials developed and used were domestic outdoor wall writings (11), domestic indoor annual wall calendar (2000), school book labels (1000); game charts (16); wall posters (65); flip chart (15); rabies educational DVD (1). There were 102 local cable television transmissions on rabies prevention. 69 persons received rabies PEP. PrEP was given to 368 school children, pet dog owners and veterinarians. Sixty one human serum samples were analyzed by rapid fluorescent focus inhibition test for rabies antibody detection. Sixteen veterinarians were trained to use direct rapid immunohistochemical test (dRIT) for rabies diagnosis. Six ruminants were confirmed rabid by dRIT. There were no cases of human rabies. The cost of entire project was US $ 85,958. Conclusion: A blend of medical, veterinary and animal welfare services were successfully delivered through a "one health" approach. Based on this success a "conceptual model" was evolved to propagate its replication in other rural communities across India.

Boosting effect of purified chick embryo cell rabies vaccine using the intradermal route in persons previously immunized by the intramuscular route or vice versa.

BACKGROUND: At present, in the event of re-exposure to rabies, 2 booster doses are recommended for people who have been previously vaccinated with cell culture rabies vaccines by the conventional intramuscular route. As the intradermal route of vaccination is likely to be introduced in the future, we investigated the immune response to a cell culture rabies vaccine after crossing over from the intramuscular to the intradermal route and vice versa. METHODS: Twenty healthy adult volunteers who had received a primary course of rabies vaccination with purified chick embryo cell rabies vaccine by either the intramuscular (n = 10) or intradermal (n = 10) route received booster vaccination with the same vaccine by the alternative route. The regimen used was 0.1 ml of vaccine by the intradermal route at two sites (deltoid area) for the intramuscular group, or 1 ml of vaccine by the intramuscular route (deltoid muscle) to the intradermal group on days 0 and 3. RESULTS: There was a 15-fold rise in the rabies virus neutralizing antibody response both by the intradermal and intramuscular routes of booster vaccination (p < 0.0001). Thus, the change of route of purified chick embryo cell booster vaccination did not alter the anamnestic immune response to the vaccine. No side-effects were observed after vaccination with either of the routes. CONCLUSION: Purified chick embryo cell vaccine was found to be safe and immunologically efficacious following booster vaccination after cross-over from the intradermal to the intramuscular route and vice versa.

An epidemiological study of animal bites in India: results of a WHO sponsored national multi-centric rabies survey.

This was a WHO sponsored national multi-centric rabies survey and one of its objectives was to find out the incidence of animal bites, anti-rabies treatment practices, Pet dog population and their care. Twenty-one medical colleges chosen with geoscatter representation conducted the survey during February-August, 2003. The survey was conducted in 18 states, covering a population of 52,731 chosen randomly from 8500 households. The annual incidence of animal bites was high, 1.7% and it was more in rural areas (1.8%), children (2.6%) and poor/low income group (75%). The main biting animal was dog (91.5%), mostly stray (63%), followed by cat (4.7%). A high proportion of bite victims did not wash their wounds with soap and water (39.5%), preferred Government hospitals (59.9%) and nerve tissue vaccine (46.9%). The use of rabies immunoglobulin was low (2.1%). A single animal bite episode led to a loss of 2.2 man-days and the cost of medicines including anti-rabies vaccine was Rs.252 (US$6). The recourse to indigenous treatment (45.3%) and local application to wound (36.8%/) was quite prevalent. About 17% of households reported having a pet/domesticated dog and the pet dog: man ratio was 1: 36. Pet dog care/management practices were not satisfactory with a low veterinary consultation (35.5%) and vaccination (32.9%). The situation was slightly better in urban areas. The people also reported the presence (83%) and menace (22.8%) of stray dogs. It is recommended to initiate appropriate community awareness and dog vaccination campaigns and effective stray dog control measures.

An assessment of rabies free status of the Island of Andaman, Nicobar and Lakshadweep: results of the WHO sponsored national multicentric rabies survey.

The Islands of Andaman and Nicobar and Lakshadweep have been reported to be rabies free from time immemorial. Recently, a survey of the islands of Andaman and Nicobar & Lakshadweep was done between July and August 2003 to assess their rabies free status. It was revealed that these islands continue to remain rabies free. However, in Andamans the increasing dog population, poor vigil on import of dogs and lack of laboratory surveillance for rabies posed a threat to this status. The Lakshadweep islands, which are free of dogs, however, faced a threat from the lack of vigil on the entry and presence of cats and poor surveillance for rabies in them.

Development and evaluation of an enzyme immunoassay for rapid diagnosis of rabies in humans and animals.

The presently advocated tests for rapid diagnosis of rabies such as fluorescent antibody test (FAT) is expensive and requires expertise to carry out and interpret the results. In this study we have developed and evaluated a simple enzyme immuno-assay (EIA) to detect rabies antigen in the brain specimens of animals and humans. We have also evaluated the utility of this test in ante mortem diagnosis of human rabies. The brain homogenates of suspected rabid animals (n=250), humans (n=16) and clinical samples like saliva (n=16) and cerebrospinal fluid (CSF, n=16) applied on to ELISA plates coated with rabies antinucleoprotein antibody and the absorbed rabies nucleoprotein antigen was detected using biotinylated anti-nucleoprotein antibody followed by treatment with streptavidin peroxidase conjugate and colour development with OPD. Rabies infected and normal mouse brain homogenates were used as positive and negative controls respectively. The results of this test was evaluated with fluorescent antibody technique (for brain samples) and mice inoculation test (for saliva and CSF samples). A distinct dark brown color was seen in positive control and all positive samples and there was no color development in negative control and samples. The concordance between FAT and EIA was 98.4%. With brain samples, 83.3% with saliva and 91.6% with CSF samples. The specificity of the test was found to be 100%. It can be concluded that the EIA described here is a sensitive, specific and rapid test for post mortem diagnosis of rabies in animals and humans. The utility of this test for ante mortem diagnosis of rabies needs to be further evaluated.

A passive haemagglutination test for rabies antibodies using rabies glycoprotein coupled erythrocytes.

The presently recommended tests for assaying rabies antibodies like mouse neutralization test (MINT) and rapid fluorescent focus inhibition test (RFFIT) are either time consuming or expensive and are generally performed in reference laboratories. There is a need to develop a specific and rapid method for detection of rabies antibodies that can be used to monitor sero-conversion after pre-or post-exposure vaccination. In this study, we have developed a passive haemagglutination (PHA) using purified rabies virus glycoprotein coupled to sheep erythrocytes using chromium chloride (0.04%) as a coupling agent. Two hundred and fifty five serum samples from people vaccinated with different rabies vaccines, 16 paired serum and CSF samples from autopsy confirmed cases of paralytic rabies, and serum samples from 65 normal healthy controls were tested and evaluated in comparison to standard MNT. Among the vaccinees, 250 samples were positive both by MNT and PHA but 5 samples were negative by PHA and positive by MNT. The titres obtained by PHA were lower compared to MNT, but there was significant correlation between the two (r=0.885). The specificity of the test was 99.7% and sensitivity was 100% as compared to MNT. Thus this PHA test promises to be a rapid and specific test for assaying rabies antibodies and may be useful in screening large number of serum samples for sero conversion after vaccination. It may also assist in rapid laboratory confirmation of paralytic rabies cases, based on detection of antibodies in CSF and serum.

Comparative evaluation of a simple indirect immunofluorescence test and mouse neutralization test for assaying rabies antibodies.

In this study, we have developed and evaluated a simple indirect immunofluorescence test (IIFT) to detect rabies antibodies in a two-step immunofluorescence assay. One hundred and eighty five serum samples from people who had taken different rabies vaccines and 8 pairs of serum and CSF samples from confirmed paralytic rabies cases were tested by IIFT and results evaluated in comparison to standard mouse neutralization test (MNT). Though the titres of rabies antibodies obtained with IIFT were 2-4 times lesser in comparison to MNT, a significant correlation was seen between the two tests (R = 0.883). The specificity of this IIFT was found to be 97.9% and the sensitivity was 97.2%. These results indicate that this simple and rapid IIFT can be used to screen large number of serum samples to monitor sero-conversion after pre or post exposure vaccination and may also assist in rapid ante-mortem diagnosis of atypical human rabies.

Evaluation of two intradermal vaccination regimens using purified chick embryo cell vaccine for post-exposure prophylaxis of rabies.

BACKGROUND: Post-exposure prophylaxis for rabies with cell culture vaccines by the conventional intramuscular regimen is very expensive. The World Health Organization has advocated two cost-effective intradermal regimens with cell culture vaccines for use in developing countries. We evaluated these two regimens--the 2-site and the 8-site regimens--in terms of immunogenicity, safety and tolerance in people with category I exposure to rabies. METHOD: Eighty-two subjects who had mild category I exposure to rabies were immunized using a purified chick embryo cell vaccine. The first regimen given to 43 subjects, consisted of intradermal administration of 0.2 ml of vaccine at 2 sites on days 0, 3 and 7 and at one site on days 28 and 90. The second regimen, given to 39 subjects, consisted of intradermal administration of 0.1 ml of vaccine at 8 sites on day 0, at 4 sites on day 7 and at one site on days 28 and 90. The mouse neutralization test was used to estimate titres of rabies neutralizing antibody in these subjects on different days after vaccination. The subjects were followed up for 1 year. RESULTS: Both regimens produced adequate neutralizing antibody titres from day 14 onwards, though the second regimen produced a more rapid antibody response and significantly higher titres (p < 0.001) on all days tested. There were minimal side-effects and both regimens were well tolerated. CONCLUSION: Both the 2-site and 8-site intradermal regimens with purified chick embryo cell vaccine produce adequate levels of neutralizing antibodies but the 8-site regimen appears to be more immunogenic. The feasibility of using these cost-effective regimens in routine practice needs to be further evaluated under the field conditions prevalent in India.

Post-exposure prophylaxis with purified vero cell rabies vaccine during pregnancy--safety and immunogenicity.

This study was conducted with the main objective of determining the safety and immunogenicity of purified vero cell rabies vaccine (PVRV) during pregnancy. Twenty nine pregnant women exposed to rabies were vaccinated with PVRV as per the Essen regimen advocated by World Health Organization. None of the women experienced any adverse side effects to the vaccine. The intrauterine growth and development monitored by ultrasound examination was found to be normal and the outcome of pregnancy was satisfactory. There were no congenital anomalies in any of the infants born and they were healthy and had normal growth and development during the one year follow-up period. The rabies neutralizing antibody titers from day 14 to day 365 following vaccination in these women was adequate and well above the minimum protective level of 0.5 iu/ml of serum. Protective levels of antibodies were also present in serum of some of the babies tested, for up to 3 months of age. The mothers and infants followed for one year period were doing well at the end of the study period. Consequently, PVRV was found safe and immunogenically efficacious during pregnancy.

Post exposure rabies prophylaxis with Purified Verocell Rabies Vaccine: a study of immunoresponse in pregnant women and their matched controls.

The present controlled clinical trial evaluates the immunoresponse to Purified Verocell Rabies Vaccnine (PVRV) by Essen schedule of vaccination during Pregnancy. Seventeen Pregnant women with history of animal bites who received PVRV as per Essen regimen were matched for the confounding variables of age, socio-economic status and doses of PVRV received with seventeen "Non-pregnant women". The mean age was about 24 years, majority (70.6%) belonging to middle socio-economic group and received 3 doses of PVRV. Contrary to the expectations the rabies neutralizing antibody titres were slightly higher in pregnant women (except day 180) but the difference was not significant (P > 0.2). Both the groups of women had antibody titres above protective level (0.5 IU/ml) from day 14 till day 365 thus indicating immunogenic efficacy of PVRV by Essen regimen during Pregnancy.

Response to purified chick embryo cell rabies vaccine administered intradermally for post-exposure prophylaxis.

BACKGROUND: Post-exposure immunization of rabies with presently available cell culture vaccines in the regular five-dose intramuscular schedule is expensive for developing countries. In view of the latest World Health Organization recommendation on the use of the abbreviated intradermal regimen (now routinely followed in Thailand), we evaluated this regimen for the purified chick embryo cell vaccine. METHODS: Twenty-five subjects who had nursed or casually handled a rabies patient were administered purified chick embryo cell vaccine. RESULTS: There was 100% seroconversion and all the subjects developed neutralizing antibody levels higher than the adequate level of 0.5 i.u./ml. Only minor side-effects were observed in some subjects. CONCLUSION: The feasibility of using this regimen in routine practice needs to be evaluated further.

Human rabies: epidemiological and laboratory studies in 80 cases.

Epidemiological and laboratory studies were conducted on 80 clinically diagnosed human rabies cases. Incidence of rabies was more in adult males (55%) and boys (37.5%) than adult females (5%) and girls (2.5%) and more in rural areas (76.2%) than urban areas (23.8%). Dogs constituted the main vector of transmission (92.5%). Incomplete course of antirabic vaccine had been received by 31% of cases and 2.5% of cases had complete course of antirabic vaccine. Among the laboratory tests conducted 3.8% of corneal smears and 2.5% of saliva smears were positive by fluorescent antibody technique. Rabies virus was isolated from 3.8% of saliva and 3% of cerebrospinal fluid samples. Rabies neutralising antibody titre ranged from 1:64 to 1:128 in the serum of patients who were fully vaccinated.

Reduction in the dosage schedule of BPL-inactivated neural tissue vaccine for rabies prophylaxis in man.

There exist two different immunization schedules for neural tissue rabies vaccine (NTV) for human use in India, the amount of vaccine given by the schedule recommended by Central Research Institute (CRI), Kasauli, being higher than that recommended by Pasteur Institute of India (PII), Coonoor. A study was therefore undertaken to assess the feasibility of reducing the CRI dosage schedule for rabies prophylaxis. The antirabies antibody response in laboratory animals and human subjects following 7, 10 and 14 daily doses of NTV with or without administering rabies immune globulin (RIG) was much higher than the minimum protective level of 0.5 IU/ml of serum. Based on these results, the CRI schedule could be reduced from 14 x 5 ml of NTV to 10 x 5 ml in class II and class III rabies exposure cases.

Oral infectivity of street and fixed rabies virus strains in laboratory animals.

Oral transmission of rabies could be produced in laboratory animals like mice, guinea pigs and hamsters using challenge virus strain (CVS) and 2 strains of street virus. Study of virus pathway following ingestion suggested predominant neural spread to brain and centrifugal spread to non neural organs like heart and kidneys. However it was found that virus dose required for oral infection was relatively very high. The role of such a transmission in nature needs to be further evaluated, keeping in view the high dose of virus required for oral infectivity and the frequency of consumption of brain by carnivorous animals.

Evaluation of the new rapid rabies immunodiagnosis technique.

The rapid rabies enzyme immuno-diagnosis (RREID) kit was evaluated for its sensitivity and specificity in comparison to fluorescent antibody technique (FAT) and biological test (BT) for the diagnosis of rabies. While 93 per cent correlation was observed in case of fresh brain samples, 71 per cent correlation was noted with glycerol preserved samples, where RREID was more sensitive than FAT. RREID test can be employed for rapid diagnosis of rabies, when facilities for FAT are not available. However, to test, at one time, small number of specimens, the RREID kit needs to be modified.