Proteomic-based identification of novel EV-derived protein antibodies biomarkers for melioidosis diagnosis

Melioidosis, caused by Burkholderia pseudomallei ( Bp ), is a life-threatening disease characterized by diverse clinical manifestations and limited diagnostic capabilities. Extracellular vesicles (EVs) have emerged as critical carriers of novel antibody targets for serodiagnosis. In this study, we established a Bp -infected BEAS-2B cell model ( Bp /BEAS-2B) and isolated EV from both Bp and Bp /BEAS-2B cells to generate EV proteome, identifying potential antigenic biomarkers for melioidosis diagnosis. Bioinformatics analysis identified PPEP and POMCR proteins as candidate antigens, with BLF1 and omp A serving as positive controls. Using a self-developed IgM-ELISA, serum samples from 43 melioidosis patients and 47 healthy volunteers were analyzed to detect antibodies against these antigens. Anti-POMCR IgM demonstrated exceptional diagnostic performance, with an AUC of 0.9872 (95% CI: 0.9713-1.003), sensitivity of 93.02% and specificity of 97.92% at a cutoff value of OD 450 = 0.118. Similarly, IgM against PPEP, BLF1, and omp A also showed high diagnostic accuracy, with AUC values of 0.969, 0.9621, and 0.976, respectively. The accuracy of anti-POMCR and anti-PPEP were 96.43% and 95.54%, respectively, equivalent to anti-omp A (93.75%) and anti-BLF1 (91.96%). Antibodies to EV-derived proteins effectively differentiated melioidosis patients from other bacterial infections and healthy volunteers, highlighting their clinical potential as diagnostic tools for melioidosis.