424 ON THE ARTIFICIAL CULTURE OF MARINE PLANKTON ORGANISMS. 



or else putrefaction will set in and the cultures will be irrevocably lost." 

 As a matter of fact, we have found that such organic infusions are 

 unnecessary, when dealing with plankton diatoms, and it has not been 

 our practice to employ them (cf., however, p. 445). 



Miquel obtained cultures of single species of diatoms either by 

 picking out individual diatoms under the microscope and introducing 

 them into the prepared water, or by adding a small quantity of water 

 containing a mixture of diatoms and other organisms to some prepared 

 water, and subdividing this into a number of tubes. If the subdivision 

 has been carried out sufficiently some of the tubes may contain one 

 kind of diatom only, from which fresh cultures can be made. In this 

 way, by repeated subdivision, cultures can be obtained which, by 

 inoculating fresh quantities of prepared water from time to time, 

 may, with care, be maintained indefinitely. Such cultures, however, 

 must practically always contain bacteria, and Miquel distinguishes 

 them from bacteria-free cultures, which he terms " Cultures des Dia- 

 tomees a I'etat de purete absolue." The latter he found very difficult 

 to obtain, but, through repeated washing in sterile water, followed by 

 fractional subdivision, he succeeded in getting some in which he could 

 find no trace of bacteria by ordinary bacteriological methods (cf. Miquel 

 11, p. 155 ; cf. also Kichter, 16-18). 



We propose to call any diatom culture, which can be carried on 

 practically indefinitely by inoculating fresh supplies of prepared 

 water, a '^persistent" culture, the terra '"imre" culture being reserved for 

 cultures which can be proved to contain not more than one organism. 

 We are not satisfied that we have yet succeeded in obtaining cultures 

 of the latter kind. For the most part our persistent cultures contain 

 one species of diatom only, and are free from all organisms larger than 

 small flagellates. 



In our earlier experiments with plankton diatoms, we obtained per- 

 sistent cultures, containing a single species of diatom, by both of the 

 methods recommended by Miquel. We, however, have rarely succeeded 

 by picking out single diatoms or chains of diatoms, for although we 

 have passed the selected diatom through several changes of sterilized 

 sea-water, the resulting cultures, even when the diatoms have multiplied 

 to some extent, have generally shown evidence of contamination by 

 harmful organisms, and have soon died down. Only in one of the 

 earliest experiments, and in one more recent, has complete success 

 resulted. In the first case a small chain of six or eight frustules 

 of Skeletonema costatum, picked out in April, 1905, gave rise to a 

 culture which still persists (Nov., 1909). Subcultures can still be 

 obtained even from tlie original flask inoculated in April, 1905. In the 



