ISOLATION AND IDENTIFICATION OF BURKHOLDERIA-PSEUDOMALLEI FROM SOIL USING SELECTIVE CULTURE TECHNIQUES AND THE POLYMERASE CHAIN-REACTION

An environmental soil survey to detect Burkholderia pseudomallei was p erformed during the dry and wet seasons in Darwin, Northern Territory, Australia. Soil was sampled at regular intervals during a 15-month pe riod at different depths from areas which were representative of the l ocal, soil environment. Selective culture techniques using Ashdown's a nd Galimand and Dodin's methods and the polymerase chain reaction (PCR ) using specific 16S rRNA primers were used to detect and identify the organism and determine its distribution within the soil stratum over the change in seasons. Results showed that Ashdown's method gave highe r isolation rates in the dry season, and Galimand and Dodin's method g ave higher isolation rates during the wet season. PCR of the soil enri chment proved to be a more sensitive method than culture and was also a useful confirmatory test in determining the identification of isolat es where biochemical tests gave inconsistent results, The PCR primers were specific and able to detect 10(1) cfu g(-1) soil and 10(4) cfu g( -1) of soil using Ashdown's enrichment broth and Galimand and Dodin's broth, respectively. Overall the isolation of B. pseudomallei was grea test during the dry season and at the higher and lower soil depths, wh ich is contradictory to epidemiological evidence that melioidosis occu rs primarily during the wet season among patients exposed to contamina ted surface soil and water.