Melioidosis is an important cause of sepsis in the tropics caused by an Environmental Sciences soil saprophyte -burkholderia pseudomallel (b. Pseudomallel). In many countries, melioidosis is widely under diagnosed due to the lack of rapid tests for the identification of b. Pseudomallei. Besides, two organisms that have given considerable trouble in the identification of b. Pseudomallel in the laboratory are burkholderia cepacia (b. Cepacia) and the newly recognized and closely related, nonpathogenic, burkholderia thailandensis (b. Thailandensis,).the present invention provides a more rapid, sensitive and specific multiplex polymerase chain reaction (pcr) method and use of novel nucleotide fragment pairs as primers to identify and differentiate b. Pseudomallei from b. Thailandensis and b. Cepacia complex organisms and other closely related gram negative bacteria.