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.

Effects of Russell's viper venom fractions on systemic and renal hemodynamics.

Changes in systemic and renal hemodynamics induced by Russell's viper venom are well established. The component of the venom responsible for hemodynamic alteration has not been identified. By Sephadex column chromatography five fractions of Russell's viper (Daboia russellii siamensis) venom were isolated. Each venom fraction consisted of phospholipase A2, proteolytic enzyme, phosphomonoesterase, phosphodiesterase, arginine ester hydrolase and hyaluronidase of varying activities. Hemodynamic effects of each venom fraction including mean arterial pressure, cardiac output, systemic and renal vascular resistance, renal blood flow and glomerular filtration rate were studied in five groups of dogs; each group had four dogs. Minimal hemodynamic changes were observed in dogs receiving venom fraction I. Increased renal vascular resistance with diminution of renal blood flow and glomerular filtration rate was observed in dogs receiving venom fractions II, III, IV and V. A markedly increased renal vascular resistance with maximal decrease in renal blood flow and glomerular filtration rate was caused by fraction III of the venom with highest PLA2 and proteolytic enzyme activities. However, renal hemodynamic changes appeared to correlate better with proteolytic enzyme activity than PLA2 activity. The findings suggested the proteolytic enzyme as an important determinant of hemodynamic alteration. Fractional excretion of Na was increased in dogs injected with venom fraction IV, and is presumed to be due to the inhibition of tubular reabsorption of Na by a natriuretic factor in this venom fraction.

Screening of upregulated genes in suckling mouse central nervous system during the disease stage of rabies virus infection.

The pathogenesis of hydrophobia remains unclear. The aim of this study was to identify the differentially upregulated genes that correlated with disease development in an experimental mouse model to provide better understanding of pathological mechanisms in rabies. The present work employed Clontech mouse array 1.2 II containing 1,176 gene transcripts. Suckling mice were intracerebrally infected with canine rabies virus. The gene expression profiles on day 2, 4 and 6 post inoculation were followed. The results show genes whose expression increased at least twofold above the control, mock-infected brain. The numbers of genes showing altered expression level were 29, 109 and 98 genes on day 2, 4 and 6, respectively. The genes with altered expression were classified into eight major groups, namely immune response, metabolism, receptor and transporter, growth factors, death mediated factors, transcription and translation factors, proteases, and kinases. The numbers of upregulated genes during the disease stage was much higher than during the asymptomatic stage. This suggested that direct interaction between RABV and target cells induced massive destruction of a cellular homeostasis which may lead to functional termination of the CNS.

Genetic analysis of dog rabies viruses circulating in Bangkok.

The genetic diversity of the rabies virus glycoprotein (G) gene isolated from individual rabid dogs (inter-hosts) and within a single infected dog (intra-host) has been analyzed in an effort to better understand selective pressures and population shifts among rabies viruses circulating in Bangkok. Comparison of individual master sequences among inter-hosts revealed that the dog virus isolates circulating in Bangkok were phylogenetically closely related. The ectodomain of the glycoprotein was highly conserved among the virus isolates. Furthermore, the genetic diversity of the G gene within an intra-host was assessed by comparing the cloned sequences in the virus population. The comparisons revealed that rabies virus circulating in an intra-host consisted of closely related heterogenous populations with minor substitutions at nucleotide (0.19%) and amino acid levels.

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.

Potency, sterility and immunogenicity of rabies tissue culture vaccine after reconstitution and refrigerated storage for 1 week.

The economical Thai Red Cross intradermal (TRC-ID) post-exposure rabies treatment schedule is now widely used in Asia. However, directives from WHO and manufacturers mandated that the vaccine be used within 8h after reconstitution of the freeze-dried product. This limits the use of TRC-ID to large animal bite clinics that see several rabies exposed patients daily. This study demonstrated that refrigerated purified chick embryo and Vero cell rabies vaccines can be stored safely for up to 7 days after reconstitution; allowing use of this treatment regimen in clinics that see few rabies exposed subjects. A large project applying this method in a Northern Thai canine rabies endemic province is now in place.

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.