Dogs that develop rabies post-vaccination usually manifest the paralytic subtype.

Rabies infection can manifest as either encephalitic (furious) or paralytic (dumb) types, with a ratio of approximately 2:1 in dogs. The clinical type of rabies that develops post-vaccination has only been reported in studies from one country, all with similar findings. We report a study of 36 rabid dogs with obtainable vaccination history, presenting to The Queen Saovabha Memorial Institute, Bangkok, Thailand during 2002-2008. Dogs were classified into encephalitic or paralytic types. Of 22 non-vaccinated dogs, 16 (73%) had the encephalitic type. In contrast, of the 14 vaccinated dogs, 10 (71%) had the paralytic type, a difference that was significant (p=0.016). Recent studies on canine brains have shown that lymphocyte response is more pronounced in paralytic rabies at the brainstem level, whereas viral burden is greater in the encephalitic form. We postulate partial immune response in the vaccinated dogs might influence rabies to manifest as the paralytic type. These results can serve as a natural experiment that can help explain the basis for the differences between the paralytic and encephalitic forms of canine rabies.

Intracellular Spread of Rabies Virus Is Reduced in the Paralytic Form of Canine Rabies Compared to the Furious Form.

Studies of the furious and paralytic forms of canine rabies at the early stage of disease have shown a more rapid viral colonization of the cerebral hemispheres in the furious form, as measured by viral antigen within neuronal cell bodies and viral RNA levels. Measurement of cellular processes separate from neuronal cell body provides a visual record of the spread of rabies virus which occurs across synapses. In this study, the amount of rabies viral antigen within cell processes was quantitatively assessed by image analysis in a cohort of naturally rabies infected non-vaccinated dogs (5 furious and 5 paralytic) that were sacrificed shortly after developing illness. Measurements were taken at different levels of the spinal cord, brain stem, and cerebrum. Results were compared to the amount of rabies viral antigen in neuronal cell bodies. Generally, the amount of rabies viral antigen in cell processes decreased in a rostral direction, following the pattern for the amount of rabies viral antigen in neuronal cell bodies and the percentage of involved cell bodies. However, there was a delay in cell process involvement following cell body involvement, consistent with replication occurring in the cell body region and subsequent transport out to cell processes. Greater amounts of antigen were seen in cell processes in dogs with the furious compared to paralytic form, at all anatomic levels examined. This difference was even evident when comparing (1) neurons with similar amounts of antigen, (2) similar percentages of involved neurons, and (3) anatomic levels that showed 100% positive neurons. These findings suggest that intracellular transport of the virus may be slower in the paralytic form, resulting in slower viral propagation. Possible mechanisms might involve host-specific differences in intracellular virus transport. The latter could be cytokine-mediated, since previous studies have documented greater inflammation in the paralytic form.

Reduced viral burden in paralytic compared to furious canine rabies is associated with prominent inflammation at the brainstem level.

BACKGROUND: The mechanisms that differentiate rabies infections into furious and paralytic forms remain undetermined. There are no neuropathological features in human brains that distinguish furious and paralytic rabies. This could be due to methodology and/or examination of specimens late in the disease course.In this study, postmortem examination of brain (5 furious and 5 paralytic) and spinal cord (3 furious and 3 paralytic) specimens was performed in 10 rabies-infected dogs, sacrificed shortly after developing the illness. Rabies virus (RABV) antigen (percentage of positive neurons, average antigen area in positive neurons and average antigen area per neuron) and RNA were quantified at 15 different central nervous system (CNS) regions. The distribution and degree of inflammation were also studied. RESULTS: More RABV antigen was detected in furious rabies than paralytic in many of the CNS regions studied. Caudal-rostral polarity of viral antigen distribution was found in both clinical forms in order from greatest to least: spinal cord, brainstem, cerebellum, midline structures (caudate, thalamus), hippocampus, and cerebrum. In contrast, RABV RNA was most abundant in the cerebral midline structures. Viral RNA was found at significantly higher levels in the cerebral cortex, thalamus, midbrain and medulla of dogs with the furious subtype. The RNA levels in the spinal cord were comparable in both clinical forms. A striking inflammatory response was found in paralytic rabies in the brainstem. CONCLUSIONS: These observations provide preliminary evidence that RABV antigen and RNA levels are higher in the cerebrum in furious rabies compared to the paralytic form. In addition, brainstem inflammation, more pronounced in paralytic rabies, may impede viral propagation towards the cerebral hemispheres.

Rabies in Asia: the classical zoonosis.

Rabies remains a constant threat to humans throughout much of Asia. The dog is the main reservoir and vector with wildlife playing a very minor role. No Asian country or region has been declared rabies free by WHO in over two decades and there is evidence of canine rabies spread to new regions during the past 10 years. We now have the knowledge and technology to control canine rabies. The main barrier in managing this costly endemic is lack of motivation by authorities to address this issue along with regional inability of public health and livestock (agriculture) officials to tackle this issue in cooperation and coordination. Rabies is one of the first recognized zoonoses and a model for a true "One Health" management goal where human; veterinary, and government officials must work together in harmony to defeat this disease.

Detection of rabies viral RNA by TaqMan real-time RT-PCR using non-neural specimens from dogs infected with rabies virus.

To determine the burden of rabies in developing countries, a reliable and accurate diagnostic test for the examination of the brains of animals is needed. Recently, the number of samples and carcasses submitted to rabies diagnostic units has been declining. Methods for obtaining tissues from different regions of the brain are even more difficult, and direct florescent antibody examination may fail if the samples decomposed. The spread of rabies virus to peripheral non-nervous tissues starts early during the pre-clinical phase. It has been shown that saliva and skin biopsies taken at the neck and containing hair follicles can be used in the ante-mortem diagnosis of rabies in humans. Obtaining oral swab samples, whisker or hair follicles from the heads of canines is easy and practical and can be performed without special equipment. The objective of this study was to determine whether these non-neural specimens can be used for the detection of rabies viral RNA. The RNAs extracted from these specimens were tested using a real-time reverse transcriptase polymerase chain reaction (RT-PCR). The sensitivity of the TaqMan real-time RT-PCR analysis using samples from dogs confirmed to be infected with rabies virus was 84.6% (55/65), 81.8% (54/66) and 66.7% (44/66) when using oral swab samples, extracted whisker follicles and extracted hair follicles; the specificity of all specimen types was 100%. The negative predictive values were 77.8%, 74.4% and 61.4%, respectively. Although the rate of positivity when combining the three non-neural specimen types was increased to 86.4%, this level of sensitivity was not sufficient to help physicians whether to administer post exposure prophylaxis. However, these oral swab and whisker specimens may serve to enhance epidemiological surveillance; such data will contribute in the planning of rabies control programs.

Comprehensive proteome analysis of hippocampus, brainstem, and spinal cord from paralytic and furious dogs naturally infected with rabies.

Paralytic and furious forms are unique clinical entities of rabies in humans and dogs. However, molecular mechanisms underlying these disorders remained unclear. We investigated changes in proteomes of the hippocampus, brainstem and spinal cord of paralytic and furious dogs naturally infected with rabies compared to noninfected controls. Proteins were extracted from these tissues and analyzed by two-dimensional gel electrophoresis (2-DE) (n = 6 gels/region in each group, a total of 54 gels were analyzed). From >1000 protein spots visualized in each gel, spot matching, quantitative intensity analysis, and ANOVA with Tukey's posthoc multiple comparisons revealed 32, 49, and 67 protein spots that were differentially expressed among the three clinical groups in the hippocampus, brainstem and spinal cord, respectively. These proteins were then identified by quadrupole time-of-flight mass spectrometry and tandem mass spectrometry (Q-TOF MS and MS/MS), including antioxidants, apoptosis-related proteins, cytoskeletal proteins, heat shock proteins/chaperones, immune regulatory proteins, metabolic enzymes, neuron-specific proteins, transcription/translation regulators, ubiquitination/proteasome-related proteins, vesicular transport proteins, and hypothetical proteins. Among these, 13, 17, and 41 proteins in the hippocampus, brainstem and spinal cord, respectively, significantly differed between paralytic and furious forms and thus may potentially be biomarkers to differentiate these two distinct forms of rabies. In summary, we report herein for the first time a large data set of changes in proteomes of the hippocampus, brainstem and spinal cord in dogs naturally infected with rabies. These data will be useful for better understanding of molecular mechanisms of rabies and for differentiation of its paralytic and furious forms.

A simple and rapid immunochromatographic test kit for rabies diagnosis.

In rabies endemic countries, funds and infrastructure are often insufficient to employ the approved gold standard for the definitive diagnosis of rabies: the direct fluorescent test. In the present study, two types (type 1 and 2) of an ICT kit were evaluated for detection of rabies. These were developed using monoclonal antibodies which recognize epitope II and III of the nucleoprotein of rabies virus. Both kits specifically detected all rabies virus strains and there was no cross reactivity with Lyssaviruses (Lagos, Mokola and Duvenhage), Rhabdovirus (VSV and Oita 296/1972) and other common canine-pathogenic viruses. In type 1, a single type of monoclonal antibody was used. It was capable of detecting recombinant nucleoprotein and showed sensitivity of 95.5% (42/44) and specificity of 88.9% (32/36) using brain samples from rabid dogs. In contrast, type 2 which was made of two different monoclonal antibodies had a lower sensitivity of 93.2% (41/44) and higher specificity of 100% (36/36). These ICT kits provide a simple and rapid method for rabies detection. They need neither cold chain for transportation nor complicated training for personnel. This diagnostic test is suitable for rabies screening, particularly in areas with a high prevalence of rabies and where the fluorescent antibody test is not available.

Problems in human rabies post-exposure prophylaxis management.

This is a prospective study of problems encountered with rabies-exposed patients in an animal bite clinic in a rabies endemic region. Five hundred successive patients were analyzed. The study focused mainly on issues that are not well covered in current WHO and US-CDC rabies manuals. Those dealing with immunoglobulin administration, managing severe and unusual injuries, logistic and financial issues headed the list. Animal bite patients often present challenging problems, which are not or inadequately addressed in official publications. Experience and good clinical judgment are essential in preventing human rabies.

Intratesticular injection of a balanced zinc solution for permanent sterilization of dogs.

Five male mature dogs with normal sex organs and normal pre-study semen evaluation received intratesticular injections of a sterile solution of balanced zinc gluconate. Semen without sperm could be collected on days 26, 30, 35 and 51 from 4 dogs. The dogs achieved penile erection but no ejaculation before these days. There were no significant adverse effects and no change in the dog behavior during the time that they were observed. Histopathological findings documented virtually complete fibrosis of seminiferous tubules and Leydig cells on days 60 and 75. This preliminary study concludes that balanced zinc gluconate may be used to sterilize dogs.

Six criteria for rabies diagnosis in living dogs.

OBJECTIVE: The authors studied the predictive value of six criteria for clinical diagnosis of rabies in living dogs. DESIGN: Identify and test the criteria in a retrospective and prospective study. MATERIAL AND METHOD: Both studies were conducted at the Rabies Diagnostic Unit, Queen Saovabha Memorial Institute, Thai Red Cross Society, Bangkok. The authors reviewed 1,170 dogs that were kept under observation for 10 days after they exhibited abnormal behavior. To test the predictive value of the six criteria, a prospective study involving 450 rabies suspected dogs was also performed. RESULTS AND CONCLUSION: The six criteria demonstrated 90.2% sensitivity, 96.2% specificity and 94.6% accuracy for the clinical diagnosis of rabies. They can be used for a presumptive diagnosis and may help in prioritizing post-exposure treatments and institute urgent rabies control measures.

Rabies control in South and Southeast Asia.

We have the knowledge and tools to eliminate the threat of canine rabies but this disease, nevertheless, remains a public health threat in many parts of the world. Lack of motivation by governments, cultural issues and inadequate funding remain barriers. This is amazing since the number of human rabies deaths worldwide is greater than that from polio, meningococcal meningitis, Japanese encephalitis, yellow fever, SARS, bird flue and other scourges that attract more attention. Safe and effective vaccines are now widely available. Reduced dose effective and less expensive post-exposure vaccination regimens have helped eliminate nerve tissue vaccines in Thailand, Philippines and Sri Lanka. India and Pakistan, the major users of dangerous nerve tissue derived Semple type vaccine, are now considering following suite. Immediate wound care and prompt use of a potent vaccine will save a majority of infected persons. Rabies immunoglobulin, injected into and around bite wounds, provides added safety for the severely exposed. The high cost of rabies immunoglobulin and tissue culture vaccines are remaining barriers, but new manufacturers and the use of intradermal vaccination schedules can reduce costs. Ultimately, it is the need to control rabies in dogs that must occupy most of our attention. The tools are available, but attitudes must change before they can be applied. There have been many new developments since publication of the last WHO rabies expert committee report in 1992 (new version in print)] and we will address those that have practical applicability.

Survival of naturally infected rabid dogs and cats.

A total of 1820 dogs and 332 cats that appeared ill or had bitten humans or animals were observed for >or=10 days. Of these, 957 dogs and 94 cats that were confirmed to be rabid survived <10 days after admission to our institution. This study supports current recommendations that dogs and cats that are suspected of being rabid should be euthanized and examined or, if this is inappropriate, confined and observed for 10 days.

Dog rabies in Bangkok.

Canine rabies remains a serious public health problem in Thailand. The Queen Saovabha Memorial Institute (QSMI) of the Thai Red Cross Society is the principal rabies diagnostic center in central Thailand. The retrospective study of canine rabies cases collected between 1994-1999 revealed that: (1) The prevalence of rabid dogs has decreased, and was associated with an overall decrease in the number of animals submitted for examination. However, the percentages of dogs with positive Fluorescent antibody test (FA) remained the same at approximately 40 per cent. (2) About 62 per cent of rabid dogs were less than one year old. Dogs at this age are more active and most have not been adequately vaccinated. Approximately 80 per cent of rabid dogs had never been vaccinated against rabies.

Risk of rabies after mammal bites in Thai children.

The data were collected by questionnaire at the Outpatient Department of Chulalongkorn University Hospital, Bangkok Thailand. This study revealed that 52 per cent of children had pet mammals in their home of which 67 per cent were dogs. 23.6 per cent of these children gave at least one history of a mammal bite inside (53.4%) or outside (46.6%) their house. Mammal bites of children could be found at all ages. However, most were in the age group of 10-14 years (42.3%), and 5-9 years (39.7%). The most common site of injury was on the legs (56.6%) and hands (30.7%). 31.7 per cent and 68.3 per cent of the bitten children incurred WHO category II and III potential rabies exposures (moderate and severe). 61.9 per cent had performed wound cleansing on each bite injury site and 34 per cent did not. 72 per cent of the children who had mammal bites received no post-exposure rabies treatment and 85.7 per cent did not make any effort to capture or observe the animal who had bitten them. Only 10.6 per cent observed the animals for 10 days or more. It was concluded that children are at considerable risk of mammal bites and that they are not receiving optimal care in this canine rabies endemic region and that 50 per cent of human rabies cases in Thailand were in children under 15 years of age.

Post-exposure rabies treatment in pigs.

A rabid dog invaded a Thai pig farm and severely mauled 11 adult pigs. This offered an opportunity to study efficacy of a human type post-exposure vaccine regimen with and without rabies immunoglobulin. A commercial veterinary tissue culture rabies vaccine and equine rabies immune globulin were used. All pigs survived for 1 year following the exposure. All animals developed detectable rabies neutralizing antibodies on day 7 and levels over 0.5IU/ml on day 14. This small study suggests that post-exposure rabies treatment using a proven human regimen, applied to valuable farm animals, can be safe and effective.