Protective action of rifampicin in experimental rabies infection of albino mice

The paper presents the results of the experimental study of the action of rifampicin on the process of rabies infection in albino mice contaminated with 1-10 LD50 of the fixed rabies virus. Exposure to rifampicin in doses of 250 and 500 micrograms/mouse (35-70 mg/kg) resulted in survival of 66.7 and 83.4 per cent of the animals respectively while in the controls it did not exceed 16.6 and 25.0 per cent. The average life-span of the albino mice treated with the antibiotic increased 1.6-2.4-fold in comparison with the controls. The chemotherapeutic index of rifampicin representing the ratio of the maximum tolerance dose to the minimum dose providing the protective action was equal to 20. The protective action was observed either after administration of the antibiotic according to the treatment-and-prophylaxis scheme or after administration of its 2- or 3-fold dose once a day immediately after the contamination.     

Effectiveness of specific treatment of rabies. II. Analysis of cases of human death from rabies following a complete course of treatment with specific agents]

Of 79318 persons treated of rabies in the Uzbek SSR in 1968--1974 there were recorded 16 cases of death of clinically manifest rabies. The course of treatment included injection of rabies gamma-globulin in a dose of 0.5 ml per 1 kg of body weight and the maximal dose of rabies vaccine. The incubation period lasted from 44 to 833 days (229.6 on the average). The authors considered that irrational use of high doses of rabies preparations was the cause of unsuccessful treatment of rabies.     

Oral vaccination of captive arctic foxes with lyophilized SAG2 rabies vaccine

Arctic foxes (Alopex lagopus) were immunized with lyophilized SAG2 oral rabies vaccine. The effectiveness of this vaccine was determined by serologic response and survival to challenge by rabies virus isolated from a red fox from Alaska (USA). No vaccine virus was found in saliva 1-72 hr after ingestion. At 2 wk after vaccination, all foxes had seroconverted, with rabies virus neutralizing antibody levels of 0.2-3.1 IU ml(-1). All vaccinated foxes survived to week 17 after challenge, and hippocampus, pons, and cerebellum were free of rabies virus as determined by direct immunofluorescence testing after death. One of four nonvaccinated foxes survived challenge and was free of rabies virus in neural tissue, and no rabies virus neutralizing antibody was detected in blood. Our results suggest that the lyophilized SAG2 oral rabies vaccine could be effective in arctic and subarctic regions, where freezing air and ground temperatures probably would not reduce its immunogenicity.     

Overexpression of cytochrome C by a recombinant rabies virus attenuates pathogenicity and enhances antiviral immunity

The pathogenicity of individual rabies virus strains appears to correlate inversely with the extent of apoptotic cell death they induce and with the expression of rabies virus glycoprotein, a major inducer of an antiviral immune response. To determine whether the induction of apoptosis by rabies virus contributes to a decreased pathogenicity by stimulating antiviral immunity, we have analyzed these parameters in tissue cultures and in mice infected with a recombinant rabies virus construct that expresses the proapoptotic protein cytochrome c. The extent of apoptosis was strongly increased in primary neuron cultures infected with the recombinant virus carrying the active cytochrome c gene [SPBN-Cyto c(+)], compared with cells infected with the recombinant virus containing the inactive cytochrome c gene [SPBN-Cyto c(-)]. Mortality in mice infected intranasally with SPBN-Cyto c(+) was substantially lower than in SPBN-Cyto c(-)-infected mice. Furthermore, virus-neutralizing antibody (VNA) titers were significantly higher in mice immunized with SPBN-Cyto c(+) at the same dose. The VNA titers induced by these recombinant viruses paralleled their protective activities against a lethal rabies virus challenge infection, with SPBN-Cyto c(+) revealing an effective dose 20 times lower than that of SPBN-Cyto c(-). The strong increase in immunogenicity, coupled with the marked reduction in pathogenicity, identifies the SPBN-Cyto c(+) construct as a candidate for a live rabies virus vaccine.     

Treatment with inactivated cultured antirabies vaccine and antirabies gamma-globulin of persons biten by rabid wolves or those suspected of being rabid]

The authors present the results of using inactivated cultural rabies vaccine from the Vnukovo-32 strain in combination with rabies gamma-globulin for the treatment of 39 persons; of this number 28 were bitten by rabid wolves (the diagnosis was confirmed by laboratory methods), 25 had wounds of dangerous localization, and 3 were children from 7 to 15 years of age. In examining the sera of all the 39 bitten persons it was found that the scheme of combined vaccinations with inactivated cultural ribies vaccine and rabies gamma-globulin, officially acting in the USSR, in case of bites of dangerous localization led to formation of early continuous and lengthy passive and active immunity of adequate intensity.     

Enhanced activity of rifampicin loaded with polybutyl cyanoacrylate nanoparticles in relation to intracellularly localized bacteria]

Association of rifampicin with polybutylcyanoacrylate nanoparticles provided considerable enhancement of drug antibacterial activity. In vitro nanoparticle-loaded rifampicin was more active against Staphylococcus aureus and Mycobacterium avium, localized in isolated alveolar macrophages. Level of rifampicin in macrophages increased 2-3-fold after incubation with rifampicinloaded nanoparticles comparing to the free drug. High therapeutic efficacy of colloidal rifampicin was demonstrated in vivo. Use of nanoparticles provided 2-fold increase in rifampicin efficacy, comparing with the free drug at treatment of staphylococcus sepsis in mice. Single administration of nanoparticulate rifampicin in the dose 25 mg/kg resulted in 80% survival of mice with salmonellosis, while 50 mg/kg of free rifampicin could provide only 10% survival. It may be considered that high antibacterial efficacy of rifampicin bound to nanoparticles is due to its effective delivery to macrophages.     

The induction of protective immune response in mice vaccinated by recombinant avian adenovirus CELO expressing glycoprotein G of the rabies virus

The recombinant avian adenovirus CELO-gpRb expressing glycoprotein G of rabies virus (strain TS-80, ARRIW&M, Pokrov, Russia) was used for mice vaccination against rabies. Double intramuscular immunization by recombinant CELO-gpRb adenovirus in a dose 10(9) pfu per mouse caused the induction of virus neutralizing antibodies (VNA) synthesis in 78% of mice, while twice repeated intradermal injections of the recombinant adenovirus failed to induce the VNA production. The protection level in groups of vaccinated mice after intracerebral injection of CVS rabies virus in a dose of 100 MLD50 was equal to 45% at single intramuscular immunization and to 91% after twice repeated intramuscular immunization. The recombinant adenoviral vaccine against rabies, based on CELO viral genome, has a good perspective for domestic and wild animal vaccination, not only due to rather high protection level, but also because the production of adenoviral CELO vaccine in chicken embryos is of high technology and inexpensive.     

Experimental inoculation of raccoons (Procyon lotor) with rabies virus of skunk origin.

To determine raccoon (Procyon lotor) susceptibility and serum neutralizing antibody response to a skunk salivary gland rabies virus, raccoons were inoculated with a rabies virus isolated from a naturally-infected striped skunk (Mephitis mephitis). Raccoons were divided into four groups of three animals each. A dilution of the rabies virus suspension, 10(2.4), 10(3.4), or 10(4.8), mouse intracerebral lethal dose50 (MICLD50), was administered into the masseter muscles of each animal. Three negative control animals received only diluent. Saliva and sera were collected on post-inoculation days 35, 63 and 92 for virus isolation and determination of serum neutralizing antibody titer. All animals survived the 92 day observation period and none exhibited the behavioral changes classically associated with clinical rabies virus infections. Rabies virus was not detected in the saliva of any raccoon and two of the three animals receiving the highest inoculum developed serum neutralizing antibodies (SNA). On day 92, a challenge suspension of New York City/Georgia (NYC/GA) strain rabies virus in fox salivary glands (10(3.2) MICLD50) was administered to all 12 raccoons. All animals succumbed to rabies virus except the two animals that had earlier developed SNA. The results of this study provided evidence about the susceptibility of raccoons to a skunk rabies virus and demonstrated that exposed raccoons could survive for at least 92 days following exposure. Furthermore, animals developing SNA under such circumstances were capable of withstanding challenge with rabies virus that was fatal for seronegative raccoons.     

Modeling the Geographic Spread of Rabies in China

In order to investigate how the movement of dogs affects the geographically inter-provincial spread of rabies in Mainland China, we propose a multi-patch model to describe the transmission dynamics of rabies between dogs and humans, in which each province is regarded as a patch. In each patch the submodel consists of susceptible, exposed, infectious, and vaccinated subpopulations of both dogs and humans and describes the spread of rabies among dogs and from infectious dogs to humans. The existence of the disease-free equilibrium is discussed, the basic reproduction number is calculated, and the effect of moving rates of dogs between patches on the basic reproduction number is studied. To investigate the rabies virus clades lineages, the two-patch submodel is used to simulate the human rabies data from Guizhou and Guangxi, Hebei and Fujian, and Sichuan and Shaanxi, respectively. It is found that the basic reproduction number of the two-patch model could be larger than one even if the isolated basic reproduction number of each patch is less than one. This indicates that the immigration of dogs may make the disease endemic even if the disease dies out in each isolated patch when there is no immigration. In order to reduce and prevent geographical spread of rabies in China, our results suggest that the management of dog markets and trades needs to be regulated, and transportation of dogs has to be better monitored and under constant surveillance.     

Vaccination of small Asian mongoose (Herpestes javanicus) against rabies

Oral vaccination of free-ranging wildlife is a promising technique in rabies control. The small Asian mongoose (Herpestes javanicus) is an important reservoir of rabies on several Caribbean islands, but no vaccines have been evaluated for this species. Captive mongooses were used to test the safety and efficacy of the commercially licensed vaccinia-rabies glycoprotein (V-RG) recombinant vaccine and a newly developed genetically engineered oral rabies virus vaccine (SPBNGA-S). In one study using V-RG, no vaccinated animals developed detectable rabies virus-neutralizing antibodies, and all but one died after experimental challenge with rabies virus. In contrast, all animals given SPBNGA-S demonstrated seroconversion within 7 to 14 days after vaccination and survived rabies virus challenge. On the basis of these preliminary results indicating the greater efficacy of SPBNGA-S vs. V-RG vaccine, additional investigations will be necessary to determine the optimal dose and duration of vaccination, as well as incorporation of the SPBNGA-S vaccine into edible bait.     

Validation of the inactivant binary ethylenimine for inactivating rabies virus for veterinary rabies vaccine production.

The rabies vaccine is produced by inactivation of rabies virus propagated on BHK21 cells. In the rabies inactivation process, BEI is added at a final concentration of 1.6 mM to the viral harvest at 37 degrees C, followed by a second dose of BEI at 24 h post-inactivation. Inactivation was confirmed by the mice innocuity test and tissue culture amplification test as per B.P (Vet) 2004. Validation of test procedure is essential as per cGMP requirement. The dose of BEI was validated by using lower and higher concentrations of BEI in inactivation process. The study indicated that BEI at a lower concentration (0.4 mM) was able to inactivate the rabies virus within 30 h and the routine concentration (1.6 mM) of BEI is effective in inactivating rabies virus within 18 h. The amplification test used for confirming the inactivation of the live virus was validated by spiking the sample with different dilutions of pretitrated live rabies virus. The test revealed that the amplification method is sensitive to detect live rabies virus if present in the inactivated sample. The validation of BEI as an inactivant and the amplification test are discussed.     

Biotransformation of rifampicin in pulmonary tuberculosis patients

Biotransformation of rifampicin in 39 tuberculosis patients treated with the drug was studied. The studies showed that biotransformation of rifampicin was most intensive during the first 3--6 hours after its use, which was confirmed by excretion of maximum amounts of rifampicin and its metabolites with the urine. 2--4 weeks after the beginning of the treatment with rifampicin excretion of its metabolism products decreased. When rifampicin was used simultaneously in a single dose with tubazid excretion of rifampicin transformation product and desacetylation of the antibiotic slowed down.     

Sensitivity to Rifampicin: Incidence,Mechanism, and Prevention

Five out of 200 patients taking rifampicin 900 mg twice weekly and three out of 91 patients taking rifampicin who attended an immunology clinic developed intolerance to the drug. Antibodies to rifampicin, which were found in most cases, decreased steadily after the end of treatment but were detectable for up to 16 months. The dose of rifampicin and the blood levels are predominating factors in the occurrence of reactions. Thus the dose should be reduced in patients in whom rifampicin blood levels rise abnormally. When it is important to continue rifampicin treatment despite intolerance antibody titres within 24 hours after administration of the drug must be measured to find when they are lowest, which determines the “unreactive period,” and when a further dose may be safely given.     

The influence of homologous vs. heterologous challenge virus strains on the serological test results of rabies virus neutralizing assays

The effect that the relatedness of the viral seed strain used to produce rabies vaccines has to the strain of challenge virus used to measure rabies virus neutralizing antibodies after vaccination was evaluated. Serum samples from 173 subjects vaccinated with either purified Vero cell rabies vaccine (PVRV), produced from the Pittman Moore (PM) seed strain of rabies virus, or purified chick embryo cell rabies vaccine (PCECV), produced from the Flury low egg passage (Flury-LEP) seed strain of rabies virus, were tested in parallel assays by RFFIT using a homologous and a heterologous testing system. In the homologous system, CVS-11 was used as the challenge virus in the assay to evaluate the humoral immune response in subjects vaccinated with PVRV and Flury-LEP was used for subjects vaccinated with PCECV. In the heterologous system, CVS-11 was used as the challenge virus in the assay to evaluate subjects vaccinated with PCECV and Flury-LEP was used for subjects vaccinated with PVRV. Although the difference in G protein homology between the CVS-11 and Flury-LEP rabies virus strains has been reported to be only 5.8%, the use of a homologous testing system resulted in approximately 30% higher titers for nearly two-thirds of the samples from both vaccine groups compared to a heterologous testing system. The evaluation of equivalence of the immune response after vaccination with the two different vaccines was dependent upon the type of testing system, homologous or heterologous, used to evaluate the level of rabies virus neutralizing antibodies. Equivalence between the vaccines was achieved when a homologous testing system was used but not when a heterologous testing system was used. The results of this study indicate that the strain of virus used in the biological assays to measure the level of rabies virus neutralizing antibodies after vaccination could profoundly influence the evaluation of rabies vaccines.     

Isolation and low-temperature preservation of liposomes containing rifampicin

The optimal conditions for preparations of rifampicin-containing liposomes were determined with the methods of mechanical shaking, gas dispersion and and reversible phases. It was found that the percentage of rifampicin incorporation into liposomes depended on the molar ratio of the antibiotic to the lipid (the optimal ratio was 1 : 10), the size and structure of liposomes, the amount of cholesterol added and the lipid membrane charge. Incorporation of rifampicin amounted to 16.1 +/- 2.4, 39.2 +/- 3.2 and 60.5 +/- 2.9 per cent with respect to neutral lecithin multilamellar liposes, liposomes prepared with the gas dispersion method and liposomes prepared with the method of reversible phases, respectively. Cholesterol in a molar ratio to lecithin equal to 2 : 5 or higher and dicetyl phosphate imparting the negative charge to the membrane had an inhibitory effect on the drug uptake by liposomes, while stearyl amine having the positive charge had a stimulating effect. The effect of the cryoprotectors glucose, polyvinylpyrrolidone, poly-ethylene glycole-400 and glycerol on low-temperature preservation and storage of rifampicin-containing liposomes was studied. It was shown that 10--15 per cent solutions of sucrose and glucose had the highest cryoprotective effect, when the two-stage method of freezing was used. It provided almost 84 per cent preservation of liposomal rifampicin. Electron microscopy showed that after defrosting liposomes no significant changes in the size and structure of lipid membranes were observed.     

Combined action of penicillin, tetracycline and rifampicin on R. sibirica and R. prowazekii in experiments on chick embryos

The effect of combinations of penicillin, tetracycline and rifampicin on R. prowazekii (the causative agent of typhus) and R. sibirica (the causative agent of tick-borne rickettsiosis of the North Asia) was studied. It was shown that tetracycline and penicillin used in combination had a summation effect on both R. sibirica and R. prowazekii. The dose of each antibiotic was 2 times lower than the doses of the antibiotics used alone. However, R. sibirica was less sensitive to this combination than R. prowazekii: the minimum rickettsiocidic doses of the combination were 0.5 mg of tetracycline + 10000 units of penicillin per embryo with respect to R. sibirica and 0.1 mg of tetracycline + 10000 units of penicillin per embryo with respect to R. prowazekii. The combinations of rifampicin with penicillin or tetracycline in the concentrations used had no rickettsiocidic effect on either R. sibirica or R. prowazekii. However, it should be noted that these combinations had a synergistic action and provided a rickettsiostatic effect on R. prowazekii: the dose of rifampicin in its combination with penicillin was decreased 10 times and in the combination of rifampicin with tetracycline the doses of both rifampicin and tetracycline were decreased 10 times. Still, penicillin even in a dose of 20000 units per embryo had only a rickettsiostatic effect on R. sibirica and R. prowazekii.     

狂犬病毒CTN—1株在Vero细胞上的适应传代研究

本文报导了用我国狂犬病毒固定毒人二倍体细胞适应株（ＣＴＮ－１）进行Ｖｅｒｏ细胞适应传代研究。通过连续传代培养,滴度可达８．０１ｏｇＬＤ５０／ｍｌ,达到了ＷＨＯ规定的不需浓缩的标准。病毒用０．０１ＭＯＩ感染细胞其产量与１Ｍｏｌ感染量相仿。病毒增殖高峰在４－５天,维持达１５天无明显下降,且可连续收获４－５次。因此,该毒种符合ＷＨＯ提出的疫苗生产毒种要求,可用于狂犬病疫苗生产。     

Preliminary evaluation of Raboral V-RG® oral rabies vaccine in Arctic foxes (Vulpes lagopus)

We tested the Raboral V-RG? recombinant oral rabies vaccine for its response in Arctic foxes (Vulpes lagopus), the reservoir of rabies virus in the circumpolar North. The vaccine, which is currently the only licensed oral rabies vaccine in the United States, induced a strong antibody response and protected foxes against a challenge of 500,000 mouse intracerebral lethal dose 50% of an Arctic rabies virus variant. However, one unvaccinated control fox survived challenge with rabies virus, either indicating a high resistance of Arctic foxes to rabies infection or a previous exposure that induced immunity. This preliminary study suggested that Raboral V-RG vaccine may be efficacious in Arctic foxes.     

高效价冻干人用狂犬病疫苗暴露后免疫程序的研究

初步确定高效价冻干人用狂犬病疫苗（6．0IU／剂）暴露后免疫程序。制备高效价的冻干人用狂犬病疫苗（6．0IU／剂）,以狂犬病街毒CNX8601和BD06分别攻击小鼠和比格犬的咬肌,接种不同效价的狂犬病疫苗,以RFFIT法检测中和抗体,根据动物死亡情况,计算暴露后疫苗保护率,对不同效价的疫苗进行中和抗体测定和保护率统计分析。在以小鼠为实验动物的疫苗保护率研究中,冻干人用狂犬病疫苗（3．1IU／剂）0／3／7／14／28免疫程序的保护率为40．6％,高效价的冻干人用狂犬病疫苗（6．0IU／剂）0／3／14免疫程序的保护率为56．2％,中和抗体比较,P〈0．05,2组间有显著性差异;在以比格犬为实验动物的保护效果研究中,冻干人用狂犬病疫苗的保护率（3．1IU／剂）为70％;高效价的冻干人用狂犬病疫苗（6．0IU／剂）的保护率为80％,中和抗体的比较,P〉0．05,没有显著性差异。高效价冻干人用狂犬病疫苗暴露后免疫程序可初步确定为0、3、14d免疫。     

Characterization of saturable binding sites for rabies virus.

A specific, saturable receptor for rabies virus was analyzed on cultured cells of neural or non-neural origin. Viral attachment kinetics were enhanced by DEAE-dextran, an effect which in turn enhanced the apparent infectivity of the virus inoculum. Under optimized conditions, the attachment of metabolically labeled ERA strain rabies virus obeyed the laws of mass action, whereby the amount of virus bound to cells varied proportionally with the concentration of cells or virus. Attachment was sensitive to changes of temperature and pH, did not require divalent cations such as Mg2+ or Ca2+, and occurred despite prior treatment of cells with proteolytic or sialic acid-specific enzymes. Saturation of the cell surface with rabies virus could be accomplished with 3 X 10(3) to 15 X 10(3) attached virions per cell. Competition for the rabies receptor occurred with rabies nonpathogenic variant virus, RV194 -2, and vesicular stomatitis virus. Reovirus type 3, another neurotropic virus, failed to inhibit rabies virus binding, and West Nile virus only slightly inhibited rabies virus binding, suggesting independent cellular receptors were recognized by these viruses. Isolated rabies virus glycoprotein failed to compete in an equivalent manner. However, solubilization of BHK-21 cells with octylglucoside yielded a chloroform-methanol-soluble extract which blocked rabies virus attachment. The binding inhibition activity of this extract was resistant to proteases but could be destroyed by phospholipases and neuraminidase, suggesting a phospholipid or glycolipid component at the receptor site. These data provide evidence for a rhabdovirus-common mechanism for cellular attachment to cells in culture.