Tab A - Page 17 
used In plant disease control will not be used. The ONA sequences or segments 
flagging the deletions provide a means for positive Identification of strains 
carrying the deletions by dot-blot hybridization with the respective ONA probe. 
This makes possible the large-scale monitoring of bacterial populations on a 
routine basis to determine survival and lateral dissemination of strains. (I) 
Prior to field application, the mutant strains were tested In a growth 
chamber and/or greenhouse, to verify that they did not Induce frost or other 
Injury to plants and were not capable of colonizing their leaves to a greater 
extent than the original strain. These preliminary contained experiments 
provided data on the ability of the mutant INA" bacteria to displace wild 
h 
strains from leaf surfaces. In the growth chamber and greenhouse the plants 
were sprayed with suspensions of INA" deletion mutants. The plants were then 
challenge-sprayed with INA* parental or heterologous strains or allowed to be 
+ c 
naturally colonized with wild INA strains. Frost injury was assessed for 
/ 
plants sprayed with either or both types of bacteria as well as for nonsprayed 
✓ 
controls after growth chamber exposure to subzero temperatures . (Z) * 
Survival, lateral dissemination, and population dynamics of INA” mutants 
and INA* populations In the field and the specificity if any, of INA’ and INA* 
bacterial Interactions on leaf surfaces will be studied In parallel. To 
facilitate these studies, the strains will be marked with antibiotic resistance 
mutations not used in clinical medicine. so that they can be- selectively 
reisolated. Measurements of the applied bacteria will be made at 10m, 100m and 
lkm in all directions from the site of application. Measurement of bacteria 
carrying the appropriate genetic marker will be made before treatment and 
weekly for the first month after treatment. Thereafter, treatment will be at 
14 
[ 211 ] 
