The rapid and accurate diagnosis of bacterial pathogens causing diseases and infections has been a goal of clinicians and microbiologists since the discovery of microbes as disease-causing agents. Conventional methods of diagnosis rely upon physiological identification of bacteria through a series of chemical, biological and immunological tests. The use of these tests however requires culturing and sub-culturing of bacterial samples to obtain usable numbers of bacterial cells. The waiting period that occurs between sample collection from a patient and the conclusion of a diagnosis can often exceed a week, a time period that is too long for patients to receive adequate treatment. Recently, nucleic acid amplification based detection methods such as PCR (polymerase chain reaction) have been developed for pathogen diagnosis. Although such methods have eliminated the need for culturing, the sample labeling and amplification are still time-consuming and prone to contamination which would lead to false positive result. In addition, the range of pathogenic species that can be detected is limited. Moreover, virulence factors present in pathogens are also extremely important for prescription of appropriate antibiotics. Consequently, there is a need for development of rapid and high throughput tests for detecting both pathogen species as well as their virulence potential. Surface plasmon resonance (SPR) is a label-free technology to study biomolecular interactions in real-time. It has been employed increasingly to study antibody-antigene interactions, protein-protein interactions and DNA protein interactions. SPR used in the imaging mode (SPRi) provides additional benefits, such as higher throughput, multiplexing, and reduced sample consumptions. Our hypothesis is that SPRi technology can be used to simultaneously identify bacterial species or strains as well as virulence factors present in the bacteria. Through this grant proposal, support is requested to design and validate species-specific or virulence factor- specific DNA probes, and to demonstrate proof-of-concept with 20 pathogenic bacterial species and 10 virulence factor genes (including resistant genes and drug efflux pump genes). The proposed experiments will also explore chemistry for probe immobilization (chip fabrication) and compare the SPRi results to data obtained with microarray methods used during probe validation process.
Emergence and dissemination of multi-drug resistant pathogens have become a serious global health problem. There is an unmet clinical need for rapid, comprehensive and accurate tests to detect presence of pathogens as well as their virulence potential. We propose a strategy to detect multiple bacterial pathogens and virulence factors simultaneously using imaging surface plasmon resonance.
| Golanbar, Galarah D; Lam, Christopher K; Chu, Yi-Ming et al. (2011) Phenotypic and molecular characterization of Acinetobacter clinical isolates obtained from inmates of California correctional facilities. J Clin Microbiol 49:2121-31 |
