690 Marine Microbiology 



monium) nitrogen. Growth is not substantially better in any of 

 the more complex media. 



2. Bacteria Requiring Amino Acids. These produce maxi- 

 mum growth in the basal medium supplemented with 0.4 per cent 

 vitamin-free casamino acids ( Medium BAf ) . 



3. Bacteria Requiring Growth Factors. These organisms 

 grow in medium BA if a mixture of vitamins* is added (Medium 

 BAG). 



4. Bacteria Requiring Unidentified Substances in Yeast Ex- 

 tract. These are capable of maximum growth in Medium BY, the 

 basal medium + 0.1 per cent yeast extract, and show little or 

 submaximal growth in previous media. 



5. Bacteria Requiring Unidentified Substances in Both Yeast 

 Extract and Mud Extract. Abundant growth is produced onh' in 

 Medium BYM, the basal medium supplemented with 0.1 per cent 

 yeast extract + 25% v/v mud extract. 



Media were dispensed in 5 ml amounts in 125 x 16 mm 

 tubes which were capped and sterilized at 115 G for ten minutes. 

 Duplicate tubes of each medium were inoculated by loop trans- 

 fer from a five day culture of each organism grown in sea water- 

 mud extract + 0.1 per cent yeast extract. After five days' incuba- 

 tion at 20 C growth responses of the organisms in the test media 

 were compared as optical densities read in a "Spectronic 20" 

 (Bausch and Lonib) photometer set at 600 m/i. Uninoculated 

 tubes served as blanks. Isolates were then assigned to one or 

 other of the five above groups according to which medium sup- 

 ported clear cut maximum growth. Small differences in growth re- 

 sponse were not regarded as significant. 



t Added from a sterile 10 per cent stock solution of Difco "Vitamin-free" 

 Casamino Acids. This solution had been extracted twice with charcoal and 

 reconstituted according to the method of Ford et al. (6). 



* Vitamin mixture /1 00 ml medium: Riboflavin 50 /ig, Ca pantothenate 50 

 H%, biotin 0.2 /ig, thiamin HCl 50 /ug, pyridoxine HCl 50 /ig, pyridoxal HCl 

 50 /ug, nicotinic acid 50 fig, p-aminobenzoic acid 50 jig, choline chloride 2 mg, 

 folic acid 10 /ig, inositol 5 mg, cyanocobalamin 0.2 /ig. 



