Alphavirus replication in cultured cells and infected animals is inhibited by antiproteinase agents.

The influence of different antiproteinase agents on alphavirus replication was examined. Sindbis virus multicycle replication in cultured cells was suppressed by N-tosyl-phenylalanine chloromethyl ketone (TPCK), an inhibitor of chymotrypsin-like proteinases, and by aprotinin, an inhibitor of a wide spectrum of proteinases. Antiviral activity of TPCK was also demonstrated in Sindbis virus-infected animals. Parenteral injections of TPCK in infected mice reduced virus titers in brain and blood. The possible mechanism(s) of antiviral action of the antiproteinase agents are discussed.

Radial haemolysis in gel used in serological studies on Venezuelan equine encephalomyelitis virus.

Radial haemolysis in gel was successfully used for the detection of antibody in convalescents after laboratory infection with Venezuelan equine encephalomyelitis (VEE) virus at remote intervals after infection (up to 23 years). The antibodies reacted only with VEE antigen but not with Sindbis and Chikungunya alphavirus antigens. Radial haemolysis in gel was sensitive as the haemagglutination inhibition test but the antibody could be titrated by the diameter of the haemolysis zone omitting the serum dilutions. Slight deviations in the virus dose used for sensitization of erythrocytes or in pH values were not critical. For the test, the sera were heated at 56 degrees C for 30 min. Both sera stored for long periods and those collected on paper disks could be used.

Passive hemolysis-in-gel with Togaviridae arboviruses.

Radial hemolysis-in-agarose gel (HIG) with members of the genera Alphavirus and Flavivirus is described. The test can be reproduced with goose and sheep erythrocytes sensitized at the optimal pH for each virus. Human (O group), rooster, guinea pig, mouse and hamster erythrocytes were found to be unsuitable. The titer of immune ascitic fluids in HIG coincides with the titer in hemagglutination inhibition. Investigation of convalescent sera after laboratory infection suggests the possiblity of HIG application for serologic diagnosis of arbovirus infection.

Radial haemolysis in gel: a comparative study with Chikungunya, Japanese encephalitis and rubella viruses, representatives of three genera of Togaviridae family.

Optimal conditions of radial haemolysis in gel (HIG) with Chikungunya (Chik) and Japanese encephalitis (JE) viruses were developed and compared with those of rubella virus. Sheep erythrocytes were found preferable for HIG with Chik and JE viruses, unlike rubella virus, where goose erythrocytes gave best results. Rooster erythrocytes were completely negative in the Chik and JE HIG, again differing from rubella virus, which was practically as good with rooster as with goose erythrocytes. Sheep erythrocytes with rubella virus gave incomplete reactions in HIG. Sensitization of erythrocytes with JE and Chik viruses is better, when borate-phosphate, and not phosphate buffer, is used. Optimum pH values for both viruses should be regarded. Sucrose-acetone, freon and borate-saline antigens from infected mice brain may be used for HIG with Chik and JE viruses.

Simple method for preparation of immune sera to envelope and nucleocapsid antigens of Semliki Forest virus.

Immune sera to the envelope and nucleocapsid components of Zaisan 260 and R 16225 strains of Semliki Forest virus (SFV) were prepared with the use of disrupted, partially purified virus.

Composition of the Zaysan virion.

The sedimentation coefficient of Zaysan virion ribonucleic acid (RNA), estimated by sucrose density gradient centrifugation, was 42 S. Pancreatic ribonuclease digested the viral RNA to acid-soluble fragments. Two viral structural proteins with apparent molecular weight of 50,000 and 30,000 daltons were identified by polyacrylamide gel electrophoresis. A dense, RNA-rich particle, containing the smaller polypeptide, was isolated after mild detergent treatment of Zaysan virus.