10 
Frost damage to plants can be decreased by reducing the natural epiphytic 
population of INA + bacteria; significant protection against frost damage 
has been obtained by application of various bacteriocides. The use of 
antagonistic bacterial strains which compete with the natural epiphytic 
flora has also been shown to be effective inder field conditions. 
Antagonists especially suited for this purpose are INA - mutants of the 
naturally occurring INA+ strains. These mutants, being basically adapted 
for epiphytic growth and survival, presumably displace their wild-type 
counterparts by occupying the same physical spaces and utilizing the same 
nutrients. 
The genes responsible for ice nucleation have been identified and cloned. 
The investigators wish to construct INA - mutants of Fh syringae pv. syringae 
and Eh herbicola and evaluate the efficacy of these mutants as biological 
competitors of epiphytic components of IF& + populations. IFPT strains will 
be tagged with antibiotic resistance markers (e.g., rifamycin, nalidixic 
acid). The strains will then be field tested in several locations on plots 
ranging in size from two and a half to twenty-five acres on several types 
of plants. Dr. Brill said he supported the proposal and moved approval. 
Dr. Scandalios seconded the motion. 
Dr. McKinney said the proposal requests a broad approval. He noted that 
the constructs to be released have not yet been characterized , and that 
the host range of the constructs is unknown. Furthermore, the organisms 
may fill other roles in the ecosystem in addition to precipitating frost 
damage; data on these functions are not available. Dr. Brill said the 
investigators would be deleting some host DNA and adding some innocuous 
ENA. He did not predict any changes in the potential pathogenicity or 
survivability of the organisms. 
Dr. Landy asked if Fh_ syringae and Eh_ herbicola are defined as plant patho- 
gens for plants only because of their ice nucleation capabilities; do these 
organisms destroy tissue after frost damage occurs? Dr. Tolin said the 
organisms can be isolated from frost damaged tissue but these bacteria do 
not produce cell wall degrading enzymes. Their activity appears to be 
opportunistic. Dr. Berns asked if the organisms are plant species specific. 
Dr. Tolin said these organisms are species specific in that pear isolates are 
adapted to grow on pears, those from tobacco are adapted for tobacco, etc. 
Dr. King Holmes said rifampicin is a drug used with great circumspection 
in medicine, primarily for treating tuberculosis and leprosy. Resistance 
to rifamycin emerges fairly readily, and even though the drug is very 
useful against a number of pathogens, it hasn't been widely used because 
of concerns about creating resistance. He said rifampicin resistance is 
encoded by a chromosomal gene. He said he would be uneasy if plasmids 
carrying the gene for rifampicin resistance, whether engineered by recom- 
binant DNA technology or not, were disseminated in the environment. He 
noted that the Erwinieae and Pseudomonas exchange genetic information with 
all the Enterobactenaceae . 
flM 
