30 General Bacteriology. 



EXERCISE XXIV. STUDY OF CELL GROUPING. 



HANGING DROP PREPARATIONS. 



a. Make hanging-drop preparations from bouillon cultures prepared above and also 

 from those supplied. 



b. Examine with oil-immersion objective and assign organisms to their proper place, 

 as determined by cell grouping, in the following scheme : 



Name. Sketch. 



Isolated 



( Bacilli 

 Filaments 



(. Cocci 



Plane surface, Tetrads 



C Regular 



Masses -j Irregular 



I 



I Zoogloea 



IMPRESSION PREPARATIONS. The exact relation of cell to cell as they develop in the 

 colony can frequently be determined with greater accuracy by studying a "contact prep- 

 aration" which is prepared as follows: 



a. Melt a gelatin tube and slope it, when solid make a streak culture of B. mycoides 

 FLUEGG.E and when growth has taken place dip the tube in hot water to loosen gelatin 

 which is then slipped out of the tube. 



b. Lower gently a clean cover-glass over the surface. Apply a slight pressure by 

 tapping glass. Raise coyer-glass by one edge taking care that natural arrangement of 

 adherent bacteria is not disturbed. 



c. Thoroughly air dry the same, then fix and stain in the ordinary manner. 



d. Examine the thinner layers noticing the arrangement of cells with reference to 

 each other and draw a sufficient number to illustrate this relationship. 



AGAR HANGING-DROP CULTURES. 



a. Melt a tube of agar and cool to 43 C. 



b. Sterilize a cover- glass by passing it two or three times through the flame quickly. 



c. With the needle make a streak on the cover-glass about 3 mm. long of B. subtilis. 



d. With the loop place a drop of liquid agar so as to cover up streak. 



e. Seal cover-glass to hollow ground slide. Incubate and later examine and sketch. 



EXERCISE XXV. STUDY OP INVOLUTION FORMS. 



a. Grow Bacillus subtilis (EHRENB.) MIG. in bouillon and also in a solution con- 

 taining 0.1% asparagin, 10% sugar, and by means of stained cover- glass preparations 

 compare the individual organisms in each casein regard to their form and size. The de- 

 generated or involution forms are more apparent by staining. Draw several cells illus- 

 trating a variety of involution forms. 



b. Examine a culture of Bacterium diphtheriae (LOEFFLER) MIG. on Loeffler's blood 

 serum. Read M. & R. 5. 



