164 THE PRINCIPLES OF IMMUNOLOGY 



and must be determined, in exact work, for the organisms under investi- 

 gation. The more homogeneous the emulsion, the better the phago- 

 cytosis observed. Numerous investigators have shown that under 

 experimental conditions, bacteria killed by chemicals or by heat are 

 phagocyted at precisely the same rate as living organisms. Further- 

 more, the previous staining of the organisms has no deterrent action 

 on phagocytosis. 



Relation of Bacterial Virulence. The relation of bacterial viru- 

 lence to phagocytosis has been the subject of much research since 

 Marchand first showed that virulent streptococci are taken up hardly 

 at all under conditions where avirulent streptococci are phagocyted 

 with avidity. He demonstrated that this difference is not due to the 

 vitality of the bacteria, for when killed by heat at 60 C, 1.8 per cent. 

 HC1, 2.5 per cent. Na 2 CO 3 or 90 per cent, alcohol, the virulent forms 

 show the same resistance to phagocytosis. Wright and also Levaditi 

 showed that the same difference is observable in the case of phago- 

 cytosis, without the intervention of opsonins. Rosenow confirmed 

 Marchand's results by the use of freshly-isolated virulent pneumococci. 

 Reduction of virulence of thirty-six strains by repeated cultivation on 

 media resulted in increased susceptibility to phagocytosis; and a res- 

 toration of virulence by animal passages led again to decreased phago- 

 cytosis. There is, however, no absolute parallelism between virulence 

 and susceptibility to phagocytosis. Markl, von Gruber and Futaki, as 

 well as Lohlein and others, found that anthrax bacilli and plague bacilli 

 when taken from culture material are actively phagocyted in vitro even 

 though highly virulent for animals. If removed from a guinea-pig's 

 peritoneum after having grown there for several hours, they are no 

 longer phagocyted in vitro. In animal experiments they are at first the 

 victims of active phagocytosis in vivo, but after several hours are re- 

 sistant to phagocytosis. Proper staining shows that in the resistant 

 stage the organisms show definite capsule formation. These experi- 

 ments indicate that the resistance is entirely a function of the bacteria, 

 but that there is some interdependence between the bacteria and the 

 opsonin is indicated by the experiments of Ungermann, who worked 

 with pneumococci virulent for mice in doses as small as 0.000,001 c.c., 

 but not injurious for rabbits in doses as large as i.o c.c. He found 

 that mouse serum has no opsonic action and that rabbit serum acts 

 energetically. After repeated cultivation so as to reduce virulence for 

 mice the organisms are opsonized by mouse serum. Von Bockstaele 

 and also Denys and von den Bergh were able to see leucocytes in the 

 presence of a normal serum approach and even break up chains of viru- 

 lent streptococci without engulfing them; if a strong immune serum 

 were added, there resulted active phagocytosis. In summary, these 

 various experiments show that the possession of virulence by an organ- 

 ism confers upon it the power of resisting opsonization, that this power 

 has some relation to the susceptibility of the particular animal whose 

 serum is used for opsonization, that the resistance to opsonization is 

 not lost on the death of the bacteria, and that in certain instances this 



