The application of infrared imaging employing focal plane array Fourier transform infrared (FPA-FTIR) instrumentation for the identification of bacteria was investigated. FPA-FTIR spectroscopy was shown to provide new opportunities for bacteria identification with unprecedented reliability and throughput by allowing 102--103 FTIR spectra to be acquired simultaneously from surface areas of 90 x 90 to 200 x 200 mum with a spatial resolution of ∼6 mum. The combination of data redundancy and spatial resolution afforded by infrared imaging made it possible to acquire highly reproducible spectra from bacterial films. A protocol for enhancing the reliability of bacteria identification by transmission-mode FPA-FTIR spectroscopy was developed by optimizing spectral acquisition parameters, spectral processing and data analysis; using the differentiation of two Campylobacter species as a test case. The results for this test case were compared with those obtained from three alternate FTIR spectral acquisition modes. The optimized protocol was employed for the generation of a spectral database of foodborne bacteria, containing over 1,000,000 spectra acquired by infrared imaging of 36 species from 19 genera. The development of a modular hierarchical clustering (MHC) model, in combination with the use of a region selection algorithm, allowed all species in the database to be differentiated from each other down to the species level based on differences in their infrared absorption profiles. A validation study involving the identification of well-characterized isolates by comparison of their spectra to those in the database demonstrated the robustness of the MHC model. In a further study employing 44 strains of Clostridium botulinum, the discriminatory power of FPA-FTIR spectroscopy was compared with that of pulsed-field gel electrophoresis, and the region selection algorithm was applied to identify growth medium-independent spectral regions that allowed for the differentiation of Group I and Group II C. botulinum strains in two blind validation studies. The research carried out also demonstrated the high-throughput potential of bacteria identification by infrared imaging when combined with the use of a microarray system for sample deposition. Overall, the novel FPA-FTIR spectroscopy-based bacteria identification protocol developed in this work provides a rapid-response and reagent-free technique suitable for routine use in both food and clinical microbiology laboratories.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.111855 |
| Date | January 2007 |
| Creators | Prévost Kirkwood, Jonah. |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Food Science and Agricultural Chemistry.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 002611006, proquestno: AAINR32318, Theses scanned by UMI/ProQuest. |
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