138 REPORTS ON INVESTIGATIONS AND PROJECTS. 



Cultures were made on various solid media, and pure cultures were iso- 

 lated. The chief media employed were: 



1. Potassium malate (C2H=OH<gQQg->) i.o gram. 



Sodium phosphate (NasHPOi, 12H2O) 0.25 gram. 



Potassium nitrate (KNO3) O-S gram. 



Sea-water 1,000.0 c.c. 



Agar-agar 12.0 grams. 



The medium was only filtered through glass wool, so that a very slight 

 floccular precipitate of calcium phosphate was retained. 



2. Petone 2.0 grams. 



Potassium nitrate (KNO3) 0.5 gram. 



Sea-water 1,000.0 c.c. 



Agar-agar 12.0 grams. 



By plating samples of surface-water from various positions round Tortu- 

 gas as far as possible removed from influence of the land, and collected on 

 sunny days, an average of 14 colonies per i c.c. sea-water was obtained. 

 These colonies appeared to be of two kinds when grown on peptone agar, 

 one much more plentiful than the other. Subcultures made from these colo- 

 nies in Gran's medium showed that the bacteria forming the most common 

 type of colony produced an active denitrification, while the others grew very 

 slowly in this medium and produced no denitrification. 



The characteristics of the denitrifying form are as follows: 



On the potassium malate or peptone agar media colonies are visible as 

 minute white specks after 6 to 8 hours when the room temperature averages 

 29.5° C. After about 18 hours the colonies are well developed. They are 

 white in color ; circular, but with finely irregular outline, and have a granu- 

 lar appearance. Superficial colonies are much elevated at first, but, as growth 

 proceeds, spread rapidly over the surface of the agar. Deep colonies remain 

 small, circular, and discrete. 



Growth is somewhat more rapid on peptone agar than on the potassium 

 malate agar, and the older colonies develop a brownish tinge in the center 

 when growing on the former medium. On gelatin peptone (0.5 per cent 

 peptone in sea-water and kept at between 20° and 25° C. to insure the 

 medium remaining solid) growth was very slow. In stab cultures growth 

 proceeded slowly from the surface downwards, leaving a funnel-shaped 

 depression of liquefied gelatin. 



Acid formation occurs in dextrose, levulose, mannite, and cane sugar, but 

 not in lactose media. 



Growth is inhibited at a temperature of 10° C, but takes place slowly at 



Growth is much retarded by exposure to bright sunlight, but the bacteria 

 are not killed by a 10 hours' exposure. 



The bacteria are facultative anaerobes, but growth under anaerobic condi- 

 tions is very slow. 



In Gran's medium growth is rapid, but no growth occurs if the potassium 

 nitrate be omitted, or if the calcium malate be replaced by calcium carbonate. 

 Growth in a pure solution of peptone in sea-water is very slight, but becomes 

 abundant if potassium nitrate be added, when denitrification quickly ensues. 

 The most rapid growth was produced in sea-water containing 0.2 per cent 

 peptone, o.i per cent potassium malate, and 0.5 per cent potassium nitrate, 

 and in this clear medium a slight floccular precipitate, presumably of calcium 

 salts derived from the sea-water, was soon formed. Growth was also rapid. 



