502 



SCIENCE 



[Vol. liVI, No. 1453 



The first tests were carried out at Pago Pago 

 in American Samoa, but tliey were later sup- 

 plemented by several tests at Berkeley, Cali- 

 fornia, witli material collected in Samoa. In 

 the tests just mentioned, the ordinary Omelian- 

 sky solutions for nitrite and nitrate formation 

 were employed, and they were inoculated in 

 two series, one with sea water and the other 

 with calcareous sand taken from beneath that 

 same sea water. Twenty-five ee. portions of 

 sea water and about one gram of the calcareous 

 sand were used as inocula. After three weeks, 

 there was no trace of nitrite or nitrate in the 

 sea water inoculation, but very good nitrifica- 

 tion in the calcareous sand inoculations. The 

 first result is, therefore, in agreement with 

 that of Berkeley, though reached in ignorance 

 of Berkeley's experiments. It will be noted, 

 however, that Berkeley used sea water media 

 containing 2 per cent, of (N"H4)2S04, which is 

 very different from the weak salt media of the 

 Omeliansky solution which contains only .1 per 

 cent. (NH4)2S04. In fact, there seems to be 

 no real reason for the use of such high concen- 

 trations of ammonium sulfate, and especially 

 in the presence of a concentrated salt solution 

 like sea water. Nevertheless, the agreement in 

 the results of the two tests is noteworthy. Be- 

 fore discussing further the significance of my 

 second result, namely, with the calcareous sand 

 inoculations, it is best to describe some sub- 

 sequent experiments. Thinking that the period 

 of incubation may have been too short in the 

 sea water cultures above descril>ed, since the 

 nitrifying bacteria could not in any ease be 

 expected to be present in sea water in great 

 numbers, I repeated the experiments on my 

 return to California, from Samoa, and allowed 

 the culture to run for eight months in one case. 

 The results were, however, just the same as in 

 the first series. The inoculations with sea 

 water gave no tests for nitrite or nitrate, and 

 the inoculations with calcareous sand taken 

 directly below that sea water gave marked 

 nitrification. 



Even these tests did not entirely satisfy me, 

 however, because I still thought that the num- 

 ber of nitrifying bacteria in sea water might 

 be so small as to render possible their total 

 absence from a 25 cc. sea water inoculum. An 



opportunity to make further tests came, how- 

 ever, during my continued studies on marine 

 bacteria and the lime precipitation problem 

 during the past summer (1922), this time at 

 the Tortugas Laboratory of the Carnegie In- 

 stitution of Washington. On June 9, two 150 

 ee. portions of Omeliansky's solution were 

 placed in one liter Erlenmeyer fiasks and ster- 

 ilized. When the culture solutions were cool, 

 they were inoculated as follows : one with 

 about ten grams of calcareous sand obtained 

 from the sea boittom near the Loggerhead Key 

 shore; the other with 150 oc. of sea water ob- 

 tained immediately above the calcareous sand. 

 On June 16, tests were made with Tromms- 

 dorff's reagent of the cultures, which were 

 incubated at room temperature (about 27° to 

 31° C. constantly). No test for nitrite was 

 obtained in either culture. On June 22,, the 

 tests were made again. This time, the cal- 

 careous sand inoculation showed marked 

 nitrite production, whereas the other culture 

 showed nothing. A further test made on July 

 21 gave the same results as that on June 22. 



All of these itests, taken in conjunction with 

 those of Berkeley and possibly those of Isaat- 

 ehenko, compel us to the conclusion that either 

 the nitrifying bacteria are absent from the sea 

 water, or ithey can not function in such concen- 

 trated salt solutions. The former alternative is 

 probably the correct one, since it will be re- 

 membered that my firsit tests were with much 

 weaker salt solutions, and as Miss Meek and 

 I have shown in a paper soon to appear in the 

 Journal of General Physiology, the nitrifying 

 bacteria can withstand very high salt concen- 

 trations. It may, therefore, be concluded with 

 reasonable safety that the nitrifying bacteria 

 are absent from open sea water and that, there- 

 fore, no nitrification occure in such sea water. 

 What may be the state of affairs in small areas 

 of the sea close to land and harboring much 

 organic matter is not directly relevant to this 

 particular inquiry. I hope ito secure some 

 information on that point soon. 



Not the least interesting feature of my tests 

 on this question, however, is ithe result ob- 

 tained with the calcareous sand inoculations. 

 It is remarkable that calcareous sand, which is 

 in constant contact with sea water, should har- 



