482 G. H. DREW. 



this hypothetical deficiency of Nitrates in warmer seas, it seemed of 

 interest to investigate the distribution and relative activity of denitri- 

 fying bacteria in Tropical waters in comparison to those found in 

 Temperate seas, and it was with this primary object that the present 

 work was undertaken. 



The previous researches most closely related to these investigations 

 are those of Gran (9), who isolated a number of species of denitrifying 

 bacteria from the inshore waters of the Dutch coast. He made use of 

 solutions of Nitrates, Nitrites, or Ammonium salts as the sole source of 

 Nitrogen in his culture media, which contained only a dilute solution 

 of Calcium malate as organic nutrient material for the bacteria. He 

 classifies the bacteria into four groups according to their reactions in 

 pure cultures towards Nitrates or Nitrites : — 



(1) Those which reduce Nitrates and Nitrites to free Nitrogen with- 

 out any Ammonia formation. 



(2) Those which readily reduce Nitrates to Nitrites. The Nitrite 

 disappears slowly without perceptible formation of free Nitrogen, and 

 some Ammonia is formed. 



(3) Those which cannot reduce Nitrates to Nitrites, but which are 

 capable of slowly removing the Nitrate without perceptible formation 

 of free Nitrogen. Though the Nitrites are not reduced, yet they can 

 serve as the sole source of Nitrogen for the growth of the bacteria. 



(4) Those which cannot reduce, and are not capable of assimilating 

 either Nitrates or Nitrites, but will fiourish when Ammonium salts are 

 present. 



In investigations on samples of water taken in the English Channel 

 some ten miles off Plymouth, I was able to recognize species belonging 

 to the second group of Gran's classification, but could not detect the 

 presence of species belonging to any of the other groups, and it would seem 

 probable that these other groups are chiefly composed of littoral forms. 



In fluid culture media inoculated with samples of sea-water and 

 kept at a temperature of 28'' C, Gran found that the formation of 

 Nitrite was detectable in from one to two days, and that eventually all 

 the Nitrate and Nitrite was destroyed in the majority of cases, espe- 

 cially if the cultures were reinoculated at intervals. In my experiments 

 I was able to obtain similar results in cultures kept at 30° C. after 

 eight days ; in cultures kept at 15° C. the first formation of Nitrite 

 was detectable in from five to six days, but denitrification never pro- 

 ceeded beyond this stage. 



Baur (1) showed that the optimum temperature for growth and de- 

 nitrification of the species described by him lay between 20° C. and 25° C, 

 when the bacteria were grown in fluid culture media containing Peptone. 



