Through the use of acetone and ether extraction of brain tissue from newborn mice infected with certain arthropod-borne viruses, it has been possible to demonstrate hemagglutinins for chick erythrocytes associated with the following viruses: dengue Type 1, dengue Type 2, Eastern equine encephalitis, Ilhéus, Japanese B, Ntaya, St. Louis, Sindbis, Uganda S, Venezuelan equine encephalitis, West Nile (Egypt 101 strain), Western equine encephalitis, and yellow fever (viscerotropic and neurotropic strains).

On the basis of the temperature and pH required for reaction, the viruses can be assembled in two groups: A—those that require 37°C. and a pH of about 6.4, comprising Eastern, Venezuelan, and Western equine encephalitis and Sindbis viruses; and B—those that require either 4° or 22°C. and a pH of about 7.0, comprising dengue Types 1 and 2, Ilhéus, Japanese B, Ntaya, St. Louis, Uganda S, West Nile, and yellow fever viruses.

A method of eliminating non-specific inhibitory substances present in sera was developed. The method consists essentially of filtration through Seitz pads.

Extensive serological crossings were found among viruses of each group, while antisera of one group failed consistently to cross-react with antigens of the other.

Antisera deriving from animals immunized with certain viruses for which no hemagglutinins could be developed by the present method, reacted with members of either one or the other group. Thus Semliki Forest virus would appear to belong to Group A, and Russian Far Eastern and louping ill viruses to Group B.

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