Cellular Reactions in the Dermal Connective Tissue of the Hamster to Leishmania brasiliensis.

tained through the courtesy of Dr. Q. Geiman of the Department of Comparative Pathology and Tropical Medicine of Harvard University, and was originally isolated by him from a human case of cutaneous leishmaniasis in Peru. As a routine, the strain was grown in N.N.N. (Nicolle, Novy and MacNeal) medium incubated at 22 C. After 8 to 9 days, the young parasites in the water of condensation were washed down the surface of the agar several times with a capillary pipette and a concentration of approximately 50,000 parasites per cmm. was obtained. The counts were made with a hemocytometer after killing a sample of the parasites with a weak solution of formalin and homogenizing as far as possible. The standard dose consisted of approximately 3,000,000 parasites (i.e., 0.06 cc. of the above amount). Adult Syrian hamsters (Cricetus auratus), obtained through the courtesy of the Ralston Purina Company, were used and weighed app oximately 90 gm. The inoculations were made in the superficial subcutaneous connective tissue of the anterior abdominal wall which had previously been shaved, wiped with alcohol, and rinsed with salt solution. In 5 animals (those killed at 8 and 24 hours and 4, 7 and 171 days after inoculation), a small amount of India ink was used with the inoculum to mark the place of inoculation. In the other 15 animals, the inoculation was made in one place and 2 injections of India ink were made on either side of the site of the injection of parasites, at a distance of 0.5 cm., to mark the injected area. Complete necropsies of every animal were made immediately after death and sections of their skin, liver, spleen, bone marrow, lymph node, thymus, heart, intestine, kidney and lung were cut. Wet (Zenker-formol hematoxylineosin-azure II) and dry smears (Giemsa or MayGrtinwald-Giemsa) of liver, spleen and bone marrow were made of the animals killed after 8, 24 and 48 hours, and after 7, 14, 32, 36 and 171 days. Cultures were made in N.N.N. medium of the spleen, liver and bone marrow from the animals killed 4, 7, 32 and 171 days after infection and of the spleen alone from the animals killed 1 and 14 days after infection. Neutral Zenkerformalin was used as a fixative. The tissues were embedded in nitrocellulose in accordance with Maximow's technic.' The majority of the sections were stained by the hematoxylin-eosin-azure II process, but some were stained with iron hematoxylin. Sections of 8 or 10M were cut. The first lot of 5 hamsters was inoculated with material from culture 1A44 (8 days old). A second lot of 15 hamsters was inoculated with culture 7A 44 (9 days old). In the first experiment 4 controls were used. Of these, one was inoculated with water of condensation, another with the same material mixed with a little India ink and Received for publication, August 5, 1942. Rockefeller Foundation Fellow. The author is indebted to Professors W. H Taliaferro and W. Bloom for their suggestions in planning these experiments and to Miss E. Bohlman for her help with the drawings. * The classification of Leishmania brasiliensis as a species apart from Leishmania tropica is a subject which requires more experimentation. Discussions of this subject may be found in Falci (Gior. ital. di dermat. e sif. 74: 1109, 1933) and Strong (Stitt's Diagnosis, Prevention and Treatment of Tropical Diseases, Philadelphia, Blakiston, 1942). 1. Buchsbaum, R. and Loosli, C. G.: Methods of Tissue Culture In Vitro and Outlines of Histological Methods, University of Chicago Press, Chicago, 1936.