By working with numerous strains of various species of Leishmania Ross, 1903, and with one species of Leptomonas Kent, 1880, it was possible for us to verify differences in the agglutination between the two above named genera, by the method proposed by Noguchi. An immune serum against Leishmania brasiliensis Vianna, 1911, did not agglutinate a strain of Leptomonas ctenocephali (Fantham, 1912). This verification was made but once and was not checked out; but it agrees with the experiments of Noguchi (9, 10) Kligler (43), and Wagner and Koch (44).
With this method differences were always noticed in the behaviour of different species of Leishmania, in the presence of anti-Leishmania brasiliensis serum. Immune sera against Leishmania brasiliensis have never agglutinated Leishmania tropica, Leishmania donovani, Leishmania infantum and Leishmania canis, a finding that agrees with Noguchi's verification.
The behaviour of different strains of Leishmania brasiliensis cultivated in Noguchi's medium with anti-brasiliensis sera demonstrated an intimate correlation among the various strains, as agglutination frequently takes place. Results were not so clear as in Noguchi's findings and we can see from table 1 and 2 that homologous sera not always agglutinated the same strain or other strains of Leishmania brasiliensis. Positive reactions were, in addition to this, frequently weak and after a superficial examination one would conclude that immunological differences exist between various strains of Leishmania brasiliensis. Microscopical examination of the cultures show that the lack of agglutination is due to the dying of the flagellates in the culture media containing homologous sera, a fact that will be discussed more in detail in a future paper. Agglutination occurs probably only in tubes in which flagellates were inoculated in an amount sufficient to be visible, or in cases in which the sera were not sufficiently potent to kill the flagellates immediately after inoculation, thus allowing the mass of flaggelates to increase.
Lysis of the flagellates is also a cause of failure of Noguchi's method, so one will be led to conclude that immunization of the rabbits must not be too prolonged, as lysins can appear in the blood in amounts that will mask the agglutinating power of the serum. To avoid this, it is sufficient to heat the immune serum and the hemoglobin before preparing the culture media for this reaction.