EDWARD C. ROSENOW 581 



in a strong light. Lesions such as obtained in these experiments practically never 

 occur in the animals that die from intercurrent causes or that are chloroformed as 

 controls. Serving as a further check is the fact that highly specific localizations were 

 obtained consistently when material from different disease entities was injected by 

 myself and co-workers into animals from a common supply. Streak cultures on blood 

 agar and inoculation into tall tubes of glucose brain broth or agar were made, as a 

 routine, from the blood of injected animals and from lesions and (for controls) from 

 other tissues. Blocks of tissue were fixed in 10 per cent formalin. Paraffin sections 

 were stained with hematoxylin and eosin for cellular changes, and, by a modified 

 gram method, for bacteria. The latter consists, essentially, of staining the sections 

 more deeply than in the gram method and in decolorizing to a light blue intead of to 

 the end-point. In this way, the nuclei of cells appear light blue, the cytoplasm pale 

 blue, and the bacteria deeply purple. Areas of infiltration, where bacteria are most 

 numerous, can thus be readily identified and random search for bacteria obviated. 



Numerous attempts have been made, with only partial success, to find a method 

 of culture that might be relied on to retain elective localizing power and viability for 

 long periods. Preservation of latent life under reduced oxygen tension in tall tubes of 

 meat-mash infusions or in shake cultures containing relatively few colonies in glucose 

 brain agar or ascites glucose agar, in dense suspensions in glycerol (two parts), and 

 saturated sodium chloride solution (one part), have often served to retain viability 

 and elective localizing and other specific properties for a long time. In exceptional 

 instances this was true for as long as eight years when the corresponding strains, 

 grown aerobically, had lost elective localizing power during several daily transfers. 

 Subcultures made rapidly (from four to eight each twenty-four hours) in previously 

 warmed media which afford a gradient of oxygen tension, such as glucose brain broth, 

 often served to maintain specific localizing power and agglutinating properties for 

 many generations. Elective localizing power of streptococci was preserved often for 

 many weeks or months within the pulp chamber or periapical tissues of artificially 

 devitalized teeth in dogs. 



The use of proper cultural methods is not the only requirement for the successful 

 application of the principles involved in the study of elective localization, especially as 

 pertains to localized inflammatory processes in different diseases. Here difficult pro- 

 cedures and judgment of elusive factors are often necessary. We have learned that a 

 mere correct diagnosis may not be sufficient. The condition at the time of study 

 should also be considered. A higher incidence of localization and more marked lesions 

 in animals are obtained if cultures are secured at the time of acute systemic manifesta- 

 tions or during exacerbations of symptoms in chronic conditions than during quies- 

 cent intervals. Great care is exercised in the collection of material for study both as 

 regards the clinical observations and the prevention of contamination by extraneous 

 organisms. Pus from the depths of tonsils is expressed and cultures made from this 

 instead of swabbings from the surface or from single crypts. Material from the na- 

 sopharynx is obtained by swabbings high behind the palate, with cotton swabs on 

 aluminum wire bent to a suitable angle, without touching the tongue. Pus from 

 the depths of pyorrhea pockets or discharging sinuses is aspirated into fine pipettes 

 after the surface has been cleansed with alcohol or some other antiseptic. Pulpless 



