Physiology: Characteristics in Semisynthetic and Chemically Defined Liquid Media 



worker is safe from aerosols when removing cells with the spinning wire brad, and the procedure 

 does not release agar into the cell suspension (4). The suspensions were drawn off and diluted to 

 absorbance readings of 1.0 or less at 660 nm. 



Anaerobic, aerobic, aerobic without COt , or aerobic + increased CO, cultural environments 

 were produced in test tubes with absorbent cotton plugs and combinations of saturated pyro- 

 gallol, 207f KOH, 10% NajCOj, and 1 M KH, PO4 ; rubber stoppers were then inserted in the 

 tops of the tubes (9). Virtually all of the broth cultures were incubated with Na^ CO, -KHj PO4 

 (CO2 ) seals. In general, air with increased CO2 tension was created by adding 0.3 ml each of 10% 

 Na2C03 and 1 M KHj PO4 to the cotton plug. The test tubes were placed on a rotary shaker at 

 70 rpm at 37°C; A's were read at chosen intei^vals. Other commonly used bacteriological, serolog- 

 ical, or chemical methods have been described (6). 



RESULTS 



A. Detemiination of basal growth conditions and development of a basal medium. 



In the initial "probing" experiments, we examined culture conditions and media fonnula- 

 tions which would support consistent growth of small inocula and provide maximal cell density. 

 The experience of others and our own experimental findings showed that the major factors which 

 affect growth of the LDB are size and condition of the inoculum and the presence of air. carbon 

 dioxide, pyruvate, alpha-ketoglutaric acid, and cysteine. Certain of these results are shown below. 



Figure 1. Relationship of dry weiglit to absorbance (660 nm) of the inoculum of Legionnaires" disease bacterium 

 strain Philadelphia 1. grown on the F-G medium. Cells used were obtained from CYE agar and were singles or 

 doublets of small, cigar-shaped bacilli. 



31 



