STUDIES UPON THE ETIOLOGY OF DENGUE FEVER : II. CULTIVATION AND NATURE OF THE VIRUS.

We have described the cultivation of a minute organism, upon the special media devised by Noguchi for growing spirochetal organisms, that gives rise to pyrexial and leucocytic reactions in the guinea pig. The reaction in the animal following the injection of culture filtrate is identical with that induced by the inoculation of human dengue blood. While it is possible, it is hardly probable that the infection of animals with cultures of this microorganism several generations removed from the original is due to a mechanically transferred virus with which the visible microorganism cultivated became accidentally associated. Furthermore, it is unlikely that such a virus would remain viable and be carried over to subcultures in sufficient numbers to infect the animal. The microorganisms appear in culture as globoid bodies measuring from 0.1 to 0.3micro in diameter and are arranged singly, in pairs, and in short chains. They readily pass through the Berkefeld filter (N and V) and the filtrate gives rise to a characteristic reaction in the inoculated guinea pig. The filtrate yields in subplants the same globoid bodies of the original culture. Initial cultures have been obtained directly from the blood of the human case as well as from the blood of guinea pigs reacting to the human material. However, only early generations retain the degree of virulence necessary to cause the experimental reaction, the culture of remote generation failing to infect. As far as known this minute organism has characters in common with the globoid bodies of Flexner and Noguchi obtained from cases of poliomyelitis. In plasma semisolid medium the striking feature indicative of growth is colonization, which in itself serves to differentiate the opalescence of the medium occasioned by disintegration of the contained tissue. As regards the presence of spirochetal organisms in the blood of dengue fever, we may state that examination was made of the material from human cases and from. a large number of inoculated animals. Despite the most careful search with the dark-field microscope and repeated examinations of blood specimens stained by the best methods, we have been unable to find any demonstrable spiral organism in the blood of dengue patients or in the experimentally inoculated animal. The minute organism herein described is frequently present in cultures of the blood of human dengue and of animals inoculated with dengue material. The inoculation of the culture into guinea pigs produces a response comparable to that occurring in the human case, and that induced by the injection of human dengue blood in guinea pigs. While we are of the opinion that the anaerobic globular bodies described may bear etiological relationship to dengue, we realize that further proof and confirmation of our work are required to establish the connection.