Stray dogs in Bangkok, Thailand: rabies virus infection and rabies antibody prevalence.

In Thailand, the animal most reported rabid is the stray dog. Dog bite related rabies cases in humans account for 70-95% of rabies related deaths. The reported incidence of dog bites is highest in the central part of the country, especially in Bangkok. This epidemiological survey shows that at least five different canine rabies virus types are present in Bangkok. Rabies antigen and antibody prevalence in stray dogs in Bangkok was also investigated. Saliva and serum samples were taken from 3,314 stray dogs, captured between December 2003 and June 2004. One two year-old female was antigen positive by latex agglutination test and the result was confirmed by reverse transcription-polymerase chain reaction (RT-PCR). The overall antibody seroprevalence by enzyme-linked immunosorbent assay was 62% (95% CI: 54, 70%). Antibody seroprevalence was higherfor dogs captured within central Bangkok (86% of 1,208 dogs captured) than in the dogs captured on the outskirts of the greater metropolitan area (49% of 2,106 dogs captured). If our sample of stray dogs is representative, then the seroprevalence achieved from previous vaccination campaigns is insufficient in order to break the rabies transmission cycle among stray and feral dogs.

Prevalence of rabies virus infection and rabies antibody in stray dogs: a survey in Bangkok, Thailand.

To investigate the rabies antigen and antibody prevalences among stray dogs in Bangkok, Thailand, we took both a saliva and serum sample from each of 3314 stray dogs captured once each between December 2003 and June 2004. One 2-year-old female was antigen positive in the latex-agglutination test and confirmed by reverse transcription-polymerase chain reaction. The overall antibody seroprevalence from the enzyme-linked immunosorbent assay that we used was 62% (95% CI: 54, 70%). Antibody seroprevalence was greater for dogs captured within central Bangkok (86% of 1208 dogs captured) than in the dogs captured in the outskirts of the greater metropolitan area (49% of 2106 dogs captured). If our samples of stray dogs are representative, then the seroprevalence achieved from previous vaccination campaigns is too low to protect the dog and human populations.

Immunosuppression in rabies virus infection mediated by lymphocyte apoptosis.

Previously, we demonstrated a depression of cell-mediated immunity in mice by street rabies virus infection. In the present study, we investigated several events during the course of infection and looked for alterations in the host lymphoid cells for evidence of apoptosis. Total cellular RNA was extracted from muscle tissues at the inoculation site of peripherally infected mice at different intervals after infection. Rabies virus mRNA was monitored by reverse transcription-PCR. The length of virus localization at the site of exposure in the muscle was as long as 5 days post-inoculation before the virus entered the central nervous system. At this inoculation site, the virus disappeared transiently between days 7 and 9 after infection but then was restored thereafter until death. Annexin V-fluorescein isothiocyanate staining of splenocytes and thymocytes from mice revealed apoptotic changes in these cells with a marked increase after day 6 of infection. Rabies virus antigen in the brain became detectable 6 days after infection; this occurred parallel to the appearance of apoptosis in the lymphoid cells. There was atrophy of the spleen and thymus, with no evidence of infection. Our results suggest that the interaction between the rabies virus and infected neurons triggers the process of lymphoid cell apoptosis, which reflects the defective operation of the immune system.

Seroprevalence of hepatitis A virus antibody among children and young adults in Bangkok.

OBJECTIVE: To determine the prevalence of hepatitis A virus (HAV) antibodies in various age groups of healthy children and young adults who have not received the hepatitis A vaccine. METHOD: Blood samples were collected from 825 volunteers aged 1-30 years from a well baby clinic and five academic institutions in the Don Mueang area from 1998 to 1999. Serum samples were assayed for specific HAV IgG antibodies using a commercial enzyme-linked immunosorbent assay (ELISA) kit. RESULTS: The seropositivity rate (12.4% overall) in each age group was as follows: 1-3 years, 7.7 per cent; 4-7 years, 6.6 per cent; 8-11 years, 12.4 per cent; 12-15 years, 10.7 per cent and 16-30 years, 25.9 per cent. CONCLUSIONS: In the Don Mueang area of Bangkok, the majority of children (< 16 years) do not have natural immunity against HAV. The use of hepatitis A vaccine for this population should be considered. Pre-vaccination serologic screening for HAV IgG in children may not be worthwhile.

Detection of rabies virus antigen in dog saliva using a latex agglutination test.

Dog bites are responsible for more than 90% of human rabies deaths in Asia. We developed a simple and inexpensive test based on latex agglutination (LA) for rabies virus antigen detection in dog saliva. Rabies virus antigen could be detected by agglutination on a glass slide using latex particles coated with gamma globulin. By evaluation of paired saliva-brain specimens from 238 dogs, the LA test using saliva was 99% specific and 95% sensitive compared to the fluorescent antibody test (FAT) on brain smears. The advantages of the LA test over the standard FAT are that it is comparatively simple and there is no need to kill the animal before examination.

Cell-mediated immunosuppression in mice by street rabies virus not restored by calcium ionophore or PMA.

It is known that rabies virus can suppress the host immune system. In this study we demonstrate a depression of cell-mediated immunity in mice, peripherally infected with Thai street rabies virus. The cell-mediated cytolysis of spleen cells from mice increased transiently on day 5 after infection and declined rapidly thereafter until death. The proliferation of spleen cells stimulated with a T-cell mitogen such as phytohemagglutinin or concanavalin A, was significantly suppressed during the course of infection. There was also a marked suppression of IL-2 secretion in parallel with a decrease of the T-cell proliferative response to mitogen. The suppression of T-cell proliferation was not restored by treatment with a calcium ionophore (A 23187) or phorbol 12-myristate-13 acetate (PMA).

Comparison of separated-lymphocyte and whole-blood lymphocyte proliferation assays for evaluating rabies-induced cellular immunity.

A whole-blood lymphocyte proliferation assay was compared to a standard method requiring the isolation of peripheral blood mononuclear cells (PBMC). Both methods were used to measure the cell-mediated immune responses to rabies in rabies-vaccine recipients. Whole-blood cells gave moderately higher lymphoproliferative responses in terms of stimulation indices than did separated-PBMC. The results obtained from these two methods can be considered equivalent for the purpose of quantitating cellular reactivity to rabies. The use of whole blood has advantages over the standard isolated-PBMC method.

Maternal antibodies against rabies in Thai puppies. A preliminary study.

Serum antibodies to rabies virus were measured in 32 Thai puppies before primary vaccination. Only five showed evidence of rabies antibody by ELISA testing and they had no rabies virus neutralizing activity detectable by RFFIT. Immunologic ignorance of these young dogs leaves a large part of the canine population susceptible to rabies. This could be one reason for the failure of canine rabies control efforts in Thailand.

Radioactive and non-radioactive lymphocyte proliferation assays for measuring rabies-specific cellular immunity.

We compared the radioactive lymphocyte proliferation assay for measuring rabies specific cell-mediated immunity in a group of 42 rabies vaccine recipients with a new technique using rabies-stimulated lymphocytes in a colorimetric test kit (CellTiter 96, Promega Corporation, USA). Results of both tests were good agreement (Kappa = 0.68), however, they did not show good correlation in degree (magnitude) of positivity. In addition, the conventional assay showed a higher degree of sensitivity.

Human immune response to rabies nucleocapsid and glycoprotein antigens.

Antibodies to two components of rabies virus, nucleocapsid (N) and glycoprotein (G), were compared in 11 rabies patients with those in nine recipients of Vero cell rabies vaccine. All rabies vaccinees had antibodies to N and G components by day 10 after the first vaccine injection. A similar but not identical response was observed in three out of 11 rabies patients. Serum antibodies appeared in rabies patients as early as 3 days after onset of the first symptoms of the disease. In these antibody-positive rabies patients, levels of both antibodies, but particularly of anti-N antibody, were lower than in the vaccinated group. Our results suggest that the process of immune recognition and of antibody development in human rabies is more likely to occur early in the pre-clinical phase, and that reactivity to N protein may be crucial for elicitation of neutralizing antibody.

Molecular epidemiology of rotaviruses associated with pediatric diarrhea in Bangkok, Thailand.

Rotavirus diarrhea in 453 pediatric patients (29.8% of 1,518) was studied in greater Bangkok during 1985 to 1987. The disease persisted all year, increasing in incidence from August to January (30 to 50%). Polyacrylamide gel electrophoresis of rotavirus RNA from these patients and from an additional 46 patients of a 1982 to 1983 epidemic revealed 26 electropherotypes, 4 with short (S) and 22 with long (L) RNA profiles. Of the analyzed specimens, 85.5% were L forms. Only one or a few electropherotypes predominated in each epidemic, whereas others appeared sporadically at low frequencies. Shifts in the predominant electropherotypes were observed in every epidemic. Of these, 126 strains were tested for subgroup and serotype by monoclonal antibody enzyme immunoassay. Serotype 4 prevailed from 1982 to 1983, while serotype 1 was encountered more frequently than serotypes 2 and 4 from 1985 to 1987. A complete correlation was found between the electrophoretic migration of segments 10 and 11 and the serologically defined subgroup specificity. Distinct electropherotypes occurred within the same serotype, and strains with the identical electropherotype always showed the same serotype specificity. No specific electropherotype or serotype correlated with patient age. In this study, atypical rotaviruses and mixed infections with different rotaviruses were identified.

Proximal motor neuropathy, IgA paraproteinaemia and anti-myelin-associated glycoprotein reactivity.

We report a case with proximal motor neuropathy associated with benign IgA lambda paraproteinaemia. Immunoblot demonstrated reactivity to myelin-associated glycoprotein and not to P2 protein of peripheral nerve. Dramatic improvement of the polyneuropathy was observed with steroid treatment alone within 4 weeks.

Polyacrylamide gel electrophoresis and silver staining for detection of rotavirus in stools from diarrheic patients in Thailand.

Detection of rotavirus by polyacrylamide gel electrophoresis in combination with silver staining and by enzyme-linked immunosorbent assay showed 96.7% identical results in tests with 1,304 stool specimens from diarrheic patients. The polyacrylamide gel electrophoresis method can be modified to reduce cost and working time. Phenol extraction of stools, however, is essential in maintaining the sensitivity of the method.