Dr Mary Burtnick
University of Nevada, Reno School of Medicine, USA
Optimization of Serological Assays for the Analysis of Antigen-Specific Antibody Responses in Melioidosis Patients
Melioidosis, caused by Burkholderia pseudomallei, is a severe tropical infectious disease endemic in Southeast Asia and Northern Australia. Delays in diagnosis and treatment of melioidosis can result in high mortality rates and no vaccines currently exist for protection against this important bacterial pathogen. Detection of antibodies against B. pseudomallei antigens may aid in the early diagnosis of melioidosis. This study was aimed at optimizing serological assays for detection of antibody responses against different Burkholderia antigens in melioidosis patients. Using pooled plasma samples obtained from melioidosis patients and healthy donors, we optimized enzyme-linked immunosorbent assay (ELISA) conditions for the analysis of IgG, IgM, IgA and IgG subclasses against five purified Burkholderia antigens. The polysaccharide and protein targets that were selected have previously been identified as serodiagnostic targets and/or potential vaccine candidates and included O-polysaccharide (OPS), capsular polysaccharide (CPS), hemolysin co-regulated protein 1 (Hcp1), alkyl hydroperoxide reductase C (AhpC) and the deubiquitinase TssM. Following optimization of the antigen concentrations and antibody dilutions, the ELISAs were evaluated using plasma samples obtained from culture-confirmed melioidosis patients (n=210) and healthy donors (n=210) from northeast Thailand. Results showed that higher levels of antigen-specific IgG, IgM, IgA, IgG1, IgG2 and IgG3 were present in melioidosis patients compared to healthy donors. In addition, we observed that antibody responses against the proteins were predominantly IgG1 and IgA whereas antibody responses against the polysaccharides were primarily IgG2 and IgA. Collectively, our results indicate that we have developed robust ELISAs that will be useful for characterizing humoral immune responses to Burkholderia antigens in melioidosis patients. We anticipate that these optimized assays will be useful for improving serodiagnosis of melioidosis in clinical settings.
Research in the Burtnick laboratory is focused on identifying the molecular mechanisms used by Burkholderia pseudomallei and Burkholderia mallei to persist within eukaryotic cells. Specifically, we are interested in determining how the virulence-associated secretion systems expressed by these facultative intracellular pathogens facilitate their survival and replication within a variety of human and murine cell types. Our laboratory has over 20 years of experience working with pathogenic Burkholderia species and has significant expertise in areas relating to bacterial genetics, pathogenesis, host-pathogen interactions, vaccine antigen discovery and immune assay development. The main objective of our research is to use the information gained from host-pathogen interaction studies to identify antigens that can be used to develop novel vaccines, diagnostics and immune assays to combat the diseases caused by these important bacterial pathogens.