3.5.16

DIFFERENTIATION OF BURKHOLDERIA CEPACIA ISOLATES BY FATTY ACID PROFILING AND PCR FOR BIOCONTROL OF RALSTONIA SOLANACEARUM


J RICHARDSON1, J ELPHINSTONE2, DE STEAD 2 and RHA COUTTSl

lDepartment of Biology, Imperial College of Science, Technology and Medicine, Prince Consort Road, London SW7 2BB, UK; 2Central Science Laboratory, Sand Hutton, York Y04 1LZ, UK

Background and objectives
Ralstonia solanacearum causes bacterial wilt in tomato and tobacco, and brown rot in potato. It is a plant pathogen of primary economic importance. Although prevalent in tropical and sub-tropical countries, outbreaks have been reported recently in the UK. The soil-borne bacterium Burkholderia cepacia is a known bio-control agent for fungal plant pathogens. It has antagonistic effects towards R. solanacearum. Hence it is a sensible choice as a potential biological control agent against brown rot. Unfortunately B. cepacia causes soft rot in onions and is also an opportunistic pathogen of cystic fibrosis patients [often with fatal consequences]. Recent work has grouped B. cepacia isolates into genomovars [1], with clinical isolates in separate genomovars to environmental isolates. Using genomovar clustering we plan to select an appropriate environmental strain of B. cepacia as a biocontrol agent, which presents no risk of being a potential human pathogen.

Materials and methods
A panel of isolates was compiled, consisting of B. cepacia isolated from the environment, B. cepacia and other Burkholderia spp. from the NCPPB collection. The isolates were then exposed to fatty acid profiling, REP PCR, BOX PCR and ERIC PCR. Each of these techniques produces a distinctive fingerprint for each bacterial strain. The fingerprints were subjected to clustering analysis, producing dendrograms indicating the relatedness of the bacterial strains in the panel.

Results and conclusions
All four techniques successfully grouped the bacteria in the panel according to species. For B. cepacia, each technique produced three clusters within the species grouping. The strains which appeared in each clusters were reasonably well conserved from one dendrogram profile to another. This result confirms fatty acid profiling, REP PCR, BOX PCR and ERIC PCR can be used as an indicator of relatedness between bacterial strains, and to sub-divide species into genomovars. One cluster did not contain any isolates from clinical sources, the environmental isolates from this cluster will be used for further investigations to evaluate their usefulness as a bio-control agents against R. solanacearum.

Reference
1. Vandamme P, Holmes B, Vancanneyt M, Coenye T, Hoste B, Coopman R, Revets H, Lauwers S, Gillis M, Kersters K, Govan JRW, 1997. International Journal of Systematic Bacteriology 47, 1188-1200.