INDIAN KALA AZAR 



(iii) Alum-carmine and carbol-thionin. Stain in the carmine solution for 24 

 hours. Wash for 30 mins. Stain for 30 min. in carbol-thionin. Differentiate in 

 oil of cloves until the section has a red- violet tint and the protoplasm of the cells 

 a pinkish colour. Pass rapidly through alcohol and clear in xylol.] 



Appearance in cultures. Rogers found that the parasites multiplied in 

 blood from the spleen withdrawn by splenic puncture and also in human 

 blood containing a small amount of a sterile 5 per cent, solution of sodium 

 citrate made slightly acid with citric acid. The most suitable temperature 

 is 22 C. though multiplication also takes place at 27 C. Sub-cultures do 

 not grow. 



In acidulated citrated blood growth is rapid : after incubating for about 

 48 hours a few flagellated forms are visible ; some of these have the shape of 

 a grain of barley while others are much longer. Each parasite has a nucleus 

 and a blepharoplast, the latter giving origin to the flagellum : the anterior 

 flagellated extremity is rounded, the posterior pointed : there is no undulating 

 membrane. 



Leishmania donovani can also be grown on Novy and MacNeal's medium. 



Novy and MacNeal's medium : 



Maceration of beef, ..... 1000 c.c. 



Agar, 



Peptone, 



Salt, 



Normal solution of sodium bicarbonate, 



20 grams. 

 20 

 5 

 10 c.c. 



Dissolve in tubes, sterilize and to two parts of the alkaline agar add 1 part defibrinated 

 rabbit blood. 



Nicolle's medium is easier to prepare and gives better results. 

 Nicolle's medium [N.N.N. medium] : 



Water, - - 900 c.c. 



Salt, 6 grams. 



Agar, 16 



Dissolve, distribute in tubes, sterilize and add to the medium in each tube after 

 liquefying and cooling to 40-50 C. one-third its volume of rabbit blood obtained by 

 cardiac puncture. Slope the tubes for twelve hours, incubate at 37 C. for five days to 

 test the sterility of the medium and then keep them at the ordinary temperature of the 

 laboratory for a few days before sowing them. [The tubes should be sealed to prevent 

 evaporation.] 



[Sub-cultures can be obtained on this medium (Row).] 

 [Laveran and Pettit's medium : 



Peptone (Chapoteaut), - 2 grams. 



Salt, - 6 



Water, - - 900 c.c. 



Dissolve. Distribute in quantities of 15 c.c. in small Roux flasks. Sterilize. 



Add to each flask an equal volume of defibrinated rabbit's blood. The medium 



should form a shallow layer on the bottom of the vessel. 



[This liquid medium is useful for growing rich cultures such as are required 

 for animal inoculation. ] 



Etiology. The life history of Leishmania donovani outside the human 

 body is still imperfectly known. It may be conjectured that kala azar is 

 propagated by a biting insect. Patton from recent experiments is inclined 

 to incriminate the bug, Cimex rotundatus : after feeding these insects on 

 persons suffering from the disease Patton found in them flagellated forms 

 similar to those seen in cultures. 



[Patton has now been able to follow the complete development of Leishmania 

 donovani in both the European and Indian bed bugs (Cimex lectularius and 

 C. rotundatus). The insects were infected by allowing them to feed upon a case 

 of kala azar in whose blood the parasites were present in large numbers. The 



