A Comparative Study of the RAPINA and the Virus-Neutralizing Test (RFFIT) for the Estimation of Antirabies-Neutralizing Antibody Levels in Dog Samples.

The mass vaccination of dogs against rabies is a highly rational strategy for interrupting the natural transmission of urban rabies. According to the World Organization for Animal Health (OIE) and the World Health Organization (WHO), the immunization of at least 70% of the total dog population minimizes the risk of endemic rabies. Knowledge of the virus-neutralizing antibody (VNA) level against the rabies virus (RABV) is required to evaluate protective immunity and vaccine coverage of dogs in the field. The rapid focus fluorescent inhibition test (RFFIT) and the fluorescent antibody virus neutralization (FAVN) test are recommended by OIE and WHO to determine the VNA levels in serum. However, these tests are cell culture based and require the use of live viruses and specialized equipment. The rapid neutralizing antibody test (RAPINA) is a novel, immunochromatographic test that uses inactivated virus to estimate the VNA level qualitatively. It is a simple, rapid and inexpensive, although indirect, assay for the detection of VNA levels. The RAPINA has shown good positive and negative predictive values and a high concordance with the RFFIT results. In this study, we compared the performance of the two tests for evaluating the vaccination status of dogs in the Philippines, Thailand and Japan. A total of 1135 dog sera were analysed by the RAPINA and compared to the VNA levels determined by the RFFIT. The overall positive and negative predictive values of the RAPINA were 96.2-99.3% and 84.5-94.8%, respectively, with a concordance (kappa) of 0.946-0.97 among the three countries. The RAPINA results were highly homologous and reproducible among different laboratories. These results suggest that this test is appropriate to survey vaccination coverage in countries with limited resources.

One or three intradermal injections within one week for rabies pre-exposure immunization.

Pre-exposure rabies vaccination is recommended fortravellers to endemic countries and forselected populations in highly endemic regions. It consists of three injections administered over 3-4 weeks. Travellers often ignore or do not have enough time to receive a complete course prior to departure or leave with only one or two injections. In this study, the secondary antibody response in volunteers who received one or three injections within one week is evaluated. Results are compared to the recommended three injection regimen when administered with two booster vaccinations on days 0 and 3 one year later. Long-lasting immune memory in volunteers who received only one time vaccination of 0.1 ml (1 site) intradermally and then received two booster vaccinations of 0.1 ml (1 site) intradermally on days 0 and 3, one or three years later is also reported. All volunteers developed an accelerated and adequate neutralizing antibody response within seven days after they received two boosters. These data suggest that three injections within one week are as immunogenic as three injections within four weeks. One clinic visit with two-site 0.1 ml ID injections or a one-site single 0.1 ml pre-exposure rabies vaccination is enough to prime the host immune memory for at least one to three years.

Three-year experience with 4-site intradermal booster vaccination with rabies vaccine for postexposure prophylaxis.

For booster vaccination of previously immunized persons with potential exposure to rabies, the World Health Organization recommends 2 doses of cell-culture vaccine administered intramuscularly or intradermally on days 0 and 3. We believe that four 0.1-mL intradermal booster doses given on a single day could be used at no risk to the recipient. We studied use of a single booster vaccination on day 0 followed by four 0.1-mL intradermal doses of cell-culture rabies vaccines, and we determined that this is a safe, convenient, and economical regimen for postexposure treatment of previously vaccinated individuals.

Failure of multiple-site intradermal postexposure rabies vaccination in patients with human immunodeficiency virus with low CD4+ T lymphocyte counts.

Human immunodeficiency virus (HIV)-infected patients with low CD4(+) T lymphocyte counts had a poor neutralizing antibody response to pre- and postexposure rabies vaccination. This study of HIV-infected patients with CD4(+) T lymphocyte counts < 200/microL indicated that patients had a poor response after 4-site intradermal vaccinations (4-4-4-0-2-2, doubling the intradermal doses of cell-culture rabies vaccine).

Failure of pre- and postexposure rabies vaccinations in a child infected with HIV.

We report the case of a 6-y-old HIV-infected girl with severe immune deficiency who failed to respond to intramuscular pre-exposure rabies vaccination using human diploid cell rabies vaccine on days 0, 7 and 28. She also failed to respond to an intradermal postexposure rabies regimen using purified verocell rabies vaccine at 4 sites on days 0, 3 and 7 and at 2 sites on days 30 and 90 (double the usual regimen). Sequentially monitored rabies neutralizing antibody titers were below the WHO minimum acceptable level (> 0.15 IU/ml) in all specimens. Rabies prevention in HIV-infected persons with severe immune suppression requires further study.

Neutralizing antibodies to rabies following injection of rabies immune globulin into gluteal fat or deltoid muscle.

BACKGROUND: This experiment was carried out to determine whether there is any difference in circulating rabies antibody when rabies immune globulin is administered into fat or muscle tissue. METHODS: Blood samples were taken at 24 and 48 hours after administering 40 IU/kg body weight of purified equine rabies immune globulin (ERIG) into deltoid muscle, or fatty gluteal tissue of grossly obese subjects. RESULTS: Both groups revealed barely detectable antibody levels. CONCLUSIONS: No conclusion was possible concerning the absorption kinetics of immune globulin from fat or muscle. However, it was evident that circulating antibody levels, using the recommended 40 IU/kg dose of ERIG, were extremely low and probably less than the required protective level at the possible bite site. This study supports current recommendations for local infiltration of virus inoculation sites with human or equine rabies immune globulin.

Immunogenicity of rabies postexposure booster injections in subjects who had previously received intradermal preexposure vaccination.

BACKGROUND: Preexposure rabies vaccination is recommended using the full dose intramuscular or less expensive reduced dose intradermal method. The reliability of the reduced dose intradermal preexposure regimen is still controversial. The objective of this study was to determine whether it will mount a predictable accelerated immune response after a simulated rabies exposure. METHOD: One hundred and thirty-eight veterinary students received intradermal or intramuscular preexposure vaccination using a potent batch of purified chick embryo rabies vaccine. They then received booster injections one year later. RESULTS: Subjects who received intradermal rabies preexposure vaccination, using 0.1 mL of a potent tissue culture vaccine on days 0, 7, and 28, had a lower postexposure booster antibody response 1 year later than subjects given the preexposure series intramuscularly. A significant number showed an unsatisfactory early anamnestic response. Residual neutralizing antibodies, 1 year after the intramuscular preexposure series, were also significantly higher in the intramuscular than in the 0.1 mL dose intradermal group. However, all study subjects had antibody titers above the minimum recommended level of 0.5 IU/mL by day 14. CONCLUSIONS: We conclude that not all subjects who received an intradermal preexposure vaccine series may be fully protected during the first 5 days after an exposure. Rabies immune globulin, injected into bite wounds and followed by a complete postexposure vaccine series, may be indicated if such a patient experiences a severe rabies exposure.

Economic issues in postexposure rabies treatment.

Rabies remains a worldwide public health problem even though means to control this disease are known. Logistic problems and cultural barriers for effective dog control in many countries and the high cost of human postexposure treatment, account for much of the remaining worldwide human toll. Efforts to make vaccines, with effective and safe tissue or avian culture products and immune globulin, more affordable have been only marginally successful. Second generation rabies vaccines (purified Vero-, chick and duck embryo cell products) are effective, safe and less expensive than human diploid cell rabies vaccine. Reduced dose intradermal postexposure vaccination works, is affordable, and has helped abolish the use of dangerous and poorly immunogenic brain tissue-derived vaccines in Thailand, the Philippines, and Sri Lanka. The use of purified equine rabies immune globulin has been found to be safe and cost-effective. It is, unfortunately, in short supply worldwide. Preexposure rabies vaccination for travelers to endemic regions is recommended and should be administered by the intramuscular route whenever possible.

Antibody response after a four-site intradermal booster vaccination with cell-culture rabies vaccine.

The current World Health Organization recommendation for booster vaccination of previously immunized individuals with potential exposure to rabies is two doses of vaccine intramuscularly or intradermally on days 0 and 3. We report responses to two types of postexposure treatment of healthy individuals who had received preexposure rabies vaccination 1 year previously. Group A individuals received four intradermal doses (one-fifth of the diluent volume of vaccine per dose) on day 0, and group B individuals received two intramuscular doses on days 0 and 3. Immunogenicity of the two booster regimens was assessed by titrating the amount of neutralizing antibody (Nab). We found that the booster doses of vaccine produced remarkable responses in all subjects. Nab titers of > or = 0.5 IU/mL (acceptable antibody level for protection against rabies) were detected in all subjects on day 14, and they were shown to be consistently high 1 year after the booster vaccination. We also found that the Nab titers for group A were significantly higher (two- to eightfold) than those for group B on days 5, 14, 150, and 360 after the initial booster vaccination (P < .05). Our study shows that the four-site intradermal booster regimen with use of one-fifth of the diluent volume of cell-culture rabies vaccine on day 0 is associated with a significantly higher antibody response than is the conventional booster regimen for subsequent postexposure rabies treatment of individuals who have received preexposure rabies vaccination with cell-culture rabies vaccine 1 year previously.

Problems with rabies postexposure management: a survey of 499 public hospitals in Thailand.

Rabies is still a major public health problem in Asia. The incidence of known annual human cases in India alone has recently been revised from 20,000 to 30,000, and over 500,000 patients are given some form of postexposure rabies treatment. Only China, Peninsular Malaysia, Singapore, and Thailand are reporting a significant decrease in the prevalence of this disease in humans. Over 150,000 courses of postexposure treatment (PET) are given in Thailand every year. To determine remaining barriers to further reduction of the number of human rabies deaths, we carried out a questionnaire study of government hospitals throughout the Kingdom.

An immunogenicity and efficacy study of purified chick embryo cell culture rabies vaccine manufactured in Japan.

Purified chick embryo cell rabies vaccine manufactured by the Chemo-Sero-Therapeutic Institute(Kaketsuken) at Kumamoto, Japan (Kaketsuken) was submitted to an immunogenicity and efficacy study. 52 severely rabies exposed patients were treated with the conventional five doses intramuscular WHO approved ('Essen') postexposure schedule. This included the administration of 40 IU kg-1 of equine rabies immune globulin on Day 0. A control group of equally severely exposed subjects were treated with human diploid cell rabies vaccine manufactured by the Swiss Serum and Vaccine Institute as well as human rabies immune globulin. There were no deaths in either group in the more than 2 years follow-up period. Subjects treated with the chick embryo vaccine showed greater suppression of the neutralizing antibody response by the equine rabies immune globulin than those given the human diploid cell vaccine and human rabies immune globulin. A group of 20 less severely rabies exposed patients who received only the chick embryo vaccine without immune globulin all had antibody titers greater than the WHO minimal acceptable level on Day 14, 30, 90 and 180. Fourteen subjects among the severely exposed vaccine and immune globulin study group were given vaccine boosters on Day 180 because of low antibody titers. It is concluded that chick embryo rabies vaccine manufactured by Kaketsuken is an immunogenic and effective rabies vaccine, but that the potency of future batches must be increased to provide a greater safety margin.

Immune response to simulated postexposure rabies booster vaccinations in volunteers who received preexposure vaccinations.

Several studies of the efficacy of intradermal postexposure rabies vaccination have shown that this procedure is safe, effective, and cost saving. Less is known about the reliability of the present World Health Organization (WHO)-approved intradermal preexposure regimen, which consists of three 0.1-mL doses that are generally given on days 0, 7, and 28. Previous studies have shown that neutralizing antibody responses are lower and of shorter duration in subjects given the reduced-dose intradermal regimen. Thus, it is still uncertain whether the WHO-recommended single intramuscular or intradermal booster injections given on days 0 and 3 would prevent death in all cases. In this preliminary study, we evaluated titers of neutralizing antibody in Thai student volunteers given two simulated postexposure boosters, as recommended by WHO, and we compared these volunteers to a group given vaccine intramuscularly. We observed a lower, although adequate, accelerated immune response in those given the preexposure series and postexposure boosters intradermally.

Immune response to tissue culture rabies vaccine in subjects who had previous postexposure treatment with Semple or suckling mouse brain vaccine.

Nerve tissue derived Semple and suckling mouse brain rabies vaccines are still widely used. Patients who experience a new rabies exposure and who were given such vaccines decades earlier are not uncommon in rabies endemic countries. The World Health Organization recommends that persons who have had a previous course of a potent tissue or avian culture rabies vaccine and are reexposed, be given booster infections on days 0 and 3 without rabies immune globulin. Persons who have had previous pre- or postexposure vaccination with a vaccine of unproven potency, should receive a full course of tissue or avian cell vaccine and immune globulin in the event of a new exposure. This study evaluated the immune response in 98 Thai patients who gave a history of rabies postexposure treatment with Semple or suckling mouse brain vaccines 10-50 years previously. The majority (81) had an anamnestic response and developed neutralizing antibodies that were above the recommended minimal acceptable level (0.5 IU ml-1) on day 7. This suggests that they still had immunological memory. A minority of 18% had antibody titers below this level on day 7. However, they all developed titers above 0.5 IU ml-1 on days 14 and 30. Failure to have an accelerated response to revaccination by day 7 did not appear to be related to age or time elapsed since previous nerve tissue derived vaccine administration. It was not possible to predict which subject will or will not have an acceptable level of antibody before day 14. Rabies exposed patients who give a prior history of vaccination with an unknown or nerve tissue derived vaccine should therefore be treated as if they had never been vaccinated.

What is an acceptable delay in rabies immune globulin administration when vaccine alone had been given previously?

Rabies immune globulins (RIG) are not always available. Rabies-exposed patients often present to medical centers, particularly in canine rabies infested regions, after a vaccine series has been started without immune globulin administration. It is known that rabies immune globulin can result in suppression of the neutralizing antibody response which usually yields detectable antibodies by day 7. We have shown that it can be administered with a delay of up to 5 days after the start of vaccine treatment without significant antibody suppression within the first month. This study utilized the WHO approved multisite Thai Red Cross intradermal postexposure regimen. Effective use of rabies immune globulin in severe and multiple wounds, particularly in small children, may require dilution of the RIG in normal saline to provide a volume adequate for infiltration of all wounds.

Efficacy study of a new albumin-free human diploid cell rabies vaccine (Lyssavac-HDC, Berna) in 100 severely rabies-exposed Thai patients.

A newly developed human diploid cell rabies vaccine (Lyssavac-HDC), produced without added serum albumin and with an effort to remove the virus-inactivating beta-propriolactone prior to addition of the gelatin, L-cysteine and potassium phosphate stabilizer, was tested for safety immunogenicity, adverse reactions and efficacy in 100 severely rabies-exposed Thais. All patients also received human rabies immune globulin and vaccine was administered using the conventional 5-dose intramuscular schedule of one dose on days 0, 3, 7, 14 and 28. One hundred percent of a subgroup of 40 subjects, where blood had been collected, had neutralizing antibodies greater than 0.5 IU ml-1 on days 28 and 90 and all had detectable titers on days 7, 14, 28, 90, 180 and 360. All patients could be followed for at least 1 year and remained well. No significant side-effects from this vaccine were noted.

Immunogenicity of purified duck embryo rabies vaccine (Lyssavac-N) with use of the WHO-approved intradermal postexposure regimen.

The World Health Organization (WHO) has recommended the following regimen for administration of intradermal postexposure rabies vaccines (tissue or avian cultures): 0.1 mL of the vaccine given intradermally at two sites on days 0, 3, and 7 and at one site on days 28 and 90. WHO did not specify which types of vaccines should be used when following this regimen. We evaluated the efficacy of purified duck embryo rabies vaccine (Lyssavac-N) under the above regimen conditions; although purified duck embryo rabies vaccine is highly immunogenic when given intramuscularly, it differs significantly from the other rabies vaccination products in that it is not a solution, but rather, a suspension with an added preservative. Lyssavac-N was found to produce lower mean neutralizing antibody titers with the WHO-recommended intradermal regimen. However, when the volume of each dose was increased from 0.1 mL to 0.2 mL and the same schedule of administration was followed, satisfactory titers were obtained. We concluded that the WHO intradermal postexposure rabies vaccine regimen should only be used with vaccines that have been subjected to immunogenicity studies with satisfactory results.

Immune response to rabies vaccine in Alaskan dogs: failure to achieve a consistently protective antibody response.

Previous studies in Thailand and Tunisia have shown that one injection of dog pre-exposure rabies vaccine does not produce a lasting antibody titre in a significant group of animals. We therefore duplicated the Thai study in a small North American community using healthy, owned dogs. A tissue culture vaccine of known high antigenicity was given intramuscularly as one primary injection and antibody titres were determined by the rapid fluorescent focus inhibition test on days 14, 30, 60, 180 and 360. Titres were less than 0.5 i.u./mL in 27% of dogs bled at 2 months, 24% at 6 months, and in 33% one year after the primary vaccination. In rabies endemic regions, it may be hazardous to rely on the previous vaccine history of a biting dog when making post-exposure management decisions. A retrospective study of antibody levels in previously vaccinated dogs in North America also indicated that a single injection of vaccine often failed to result in adequate titres.