[11]

MEASURING RESPIRATORY ACTIVITY OF PSEUDOMONAS AERUGINOSA FOR STS-107 EXPERIMENT BACTER.  S.A. Barton, S.C. Broadaway, and B. H. Pyle.  Dept. of Microbiology, Montana State University, Bozeman.

   An Exotoxin A (ETA) producing P. aeruginosa ATCC 29260 will be flown on STS-107 to study their growth, physiology, and virulence.  Washed bacteria suspended in water will be inoculated into defined medium (MSDM2) and incubated at 37єC for 24 hours.   As defined by the flight timeline, the reagents must be stored for 9 days at 5єC, 1 day at 37єC, and incubated for 7 days at 5єC before analysis.  This study was aimed to determine if stored INT solutions would remain active during storage and be taken up by the cells during incubation.  INT was dissolved in water at 4.9 mg/ml and either used immediately or stored for 7 days at 5єC, followed by 24 hour storage at 37°C.  P. aeruginosa was grown at 37єC in defined medium to ca.10⁹ CFU/ml, followed by the addition of INT with subsequent incubation at 5єC for 7 days or at room temperature for 2 hours.  INT formazan crystals could be seen as dark spots inside the SYBR Green stained cells during epifluorescent microscopy, with numbers comparable to enumeration by R2A agar plate counts.  Incubation at room temperature for 2 hours produced a higher percent of INT positive cells compared to incubation at 5°C.  INT dissolved in water can be stored at 5^oC without loss of activity.  The flight experiment procedure gives reliable results, which permit detection of changes in the proportion of respiring cells that may be caused by microgravity.

 (Supported by NASA and ESA)