Burkholderia pseudomallei is a causative agent of melioidosis, a fatal community acquired septicemia in Southeast Asia and Northern Australia. A protease has been proposed to be one of the major pathogenic factors to play a significant role in melioidosis. We have used phage display technology to identify peptides binding to B. pseudomallei protease. By screening a constrained cyclic heptapeptide library, five independent clones with affinity to this protease were isolated and the amino acid sequences were determined. The cyclic heptapeptides from two of the phage clones (Cys-Phe-Phe-Met-Pro-His-Thr-Phe-Cys) were identical and showed the strongest phage-protease interaction as detected by ELISA. Four of the five selected phages at the amount of 10(13) phages could inhibit B. pseudomallei protease activity by approximately 50%.