Methods: Here, we describe an enzyme-linked immunosorbent assay (ELISA) using the synthetic recombinant LipL32 (rLipL32) protein expressed in Escherichia coli for the detection of Leptospira-specific antibodies in human serum samples. The rLipL32-based ELISA was compared with a microscopic agglutination test (MAT), which is currently used as the gold standard for the diagnosis of leptospirosis.

Results: Our results showed that all the MAT-positive serum samples were positive for Leptospira-specific IgG in an ELISA, while 65% (n = 13) of these samples were also positive for Leptospira-specific IgM. In the MAT-negative serum samples, 80% and 55% of the samples were detected as negative by an ELISA for Leptospira-specific IgM and IgG, respectively.

METHODOLOGY/PRINCIPAL FINDINGS: With the availability of the B. pseudomallei whole genome sequence, we undertook to identify genes encoding the known immunogenic outer membrane protein A (OmpA). Twelve OmpA domains were identified and ORFs containing these domains were fully annotated. Of the 12 ORFs, two of these OmpAs, Omp3 and Omp7, were successfully cloned, expressed as soluble protein and purified. Both proteins were recognised by antibodies in melioidosis patients' sera by Western blot analysis. Purified soluble fractions of Omp3 and Omp7 were assessed for their ability to protect BALB/c mice against B. pseudomallei infection. Mice were immunised with either Omp3 or Omp7, subsequently challenged with 1x10(6) colony forming units (cfu) of B. pseudomallei via the intraperitoneal route, and examined daily for 21 days post-challenge. This pilot study has demonstrated that whilst all control unimmunised mice died by day 9 post-challenge, two mice (out of 4) from both immunised groups survived beyond 21 days post-infection.