MORPHOLOGY 637 



cells are seen in the films but very few of them contain gonococci. Towards the 

 third day, the number of gonococci increases and a large proportion of the leucocytes 

 contain the organism. A little later still the epithelial cells disappear, and the 

 majority of the gonococci are intra- cellular and are so numerous that about 15-20 

 per cent, of the leucocytes are invaded. At a later stage of the disease secondary 

 infections take a part in the inflammatory process and as the acute symptoms pass 

 off the epithelial cells again become numerous ; but it is only when the disease 

 enters upon the chronic stage that gonococci are again found within them and the 

 pus cells diminish in number. 



Staining reactions. The gonococcus stains easily with the basic aniline 

 dyes and is gram-negative : it is upon the latter fact that its identification is 

 based (G. Roux). 



A. Stain first with a single stain such as dilute carbol-fuchsin or carbol- 

 thionin. All the organisms are stained. 



B. Stain a film with carbol-violet, examine under the microscope and then 

 treat by Gram's method. The gonococci will be decolourized and the only 

 organisms which will retain the violet are organisms of secondary infection 

 such as staphylococci, the diplococcus described by Legrain, Bumm, etc. 



C. A film should also be double stained by one of the methods based upon 

 the principle that if a film be stained by Gram's method the gram-positive 

 organisms will be the only organisms stained, and if at this stage some other 

 dye in contrast to violet be applied the gram-negative organisms become 

 stained by it. 



The following is a list of the special methods suitable for staining films 

 for the detection of the gonococcus : 



Steinschneider's method. 1. Stain with Ehrlich's violet, then with Gram's 

 iodine solution, decolourize in absolute alcohol and wash in water. 



2. Stain with an aqueous solution of vesuvin, wash, dry, and mount. 



The gonococci and the ground work are stained brown, and the gram- 

 positive organisms violet. 



Nicolle's method. Recommended. 1. Stain with carbol-violet, then with 

 Gram's iodine. Decolourize in acetone-alcohol (p. 143), wash in water. 



2. Stain with a drop of a diluted alcoholic solution of fuchsin for a few 

 seconds. 



Saturated alcoholic (95 per cent.) solution of fuchsin, - 5 c.c. 



Distilled water, - 100 



Wash, dry, and mount. 



The gonococci are stained by the fuchsin, the other organisms by the violet. 



Plato's method. This method depends upon the use of neutral red, a 

 reagent which stains the living gonococci in fresh pus cells, leaving the leuco- 

 cytes, the extra-cellular gonococci and the secondary micro-organisms, 

 whether intra-cellular or extra-cellular, unstained. 



The method is useful for detecting gonococci but cannot be used for 

 permanent preparations. 



The staining solution should be made up immediately before being used : 

 Saturated aqueous solution of neutral-red, - 1 c.c. 



Normal saline solution, - - 100 



Mix a drop of pus with a loopful of the stain on a slide, cover with a cover- 

 glass and examine under the microscope. 



Wahl's method. Wahl recommends the following stain for staining gono- 

 cocci in sections. The solution keeps well : 



Saturated alcoholic solution of auramine, - 2 c.c. 



95 per cent, alcohol, - - 1*5 ,, 



Saturated alcoholic solution of thionin, - 2 



Saturated aqueous solution of methyl green, - 3 ,, 



Distilled water, 6 



