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dc.contributor.advisorUssher, James
dc.contributor.advisorMedlicott, Natalie
dc.contributor.authorRobinson, Andrew Mark
dc.date.available2014-11-18T00:21:28Z
dc.date.copyright2014
dc.identifier.citationRobinson, A. M. (2014). Novel Methods for the Rapid Identification and Susceptibility Testing of Blood Culture Isolates (Thesis, Bachelor of Biomedical Sciences with Honours). University of Otago. Retrieved from http://hdl.handle.net/10523/5229en
dc.identifier.urihttp://hdl.handle.net/10523/5229
dc.description.abstractThe increasing emergence of antimicrobial resistance, such as that mediated by extended-spectrum β-lactamase (ESBL)-producing gram-negative bacteria, make it more likely that patients with sepsis and bloodstream infections (BSI) will receive ineffective empirical treatment. Rapid identification of disease causing agents, coupled with early detection of antimicrobial resistance facilitates the optimisation of essential treatment decisions. Matrix-assisted laser desorption-ionization time of flight (MALDI-ToF) mass spectrometry has recently been applied to the identification of microorganisms directly from blood cultures, reducing the identification process by up to 24-hours. This study sought (i) to determine the optimal method for the rapid identification of isolates directly from blood cultures and (ii) to develop a rapid method to detect β-lactamase-mediated resistance to extended spectrum cephalosporins directly from blood cultures. Two in-house methods for sample preparation were optimized and compared to a commercially available method. Using the conventional scoring criteria, the differential centrifugation protocol correctly identified 86.8% and 67.9% of clinical isolates at the genus- and species- level. This was compared to a quicker method using Sodium dodecyl sulphate (SDS) to mediate blood cell lysis, which correctly identified 83.0% and 62.3% of clinical isolates to the genus- and species- level. Both methods performed similarly to the more expensive commercial method. Results also suggested that the scoring criteria could be altered to increase the number of species-level identification while maintaining accuracy, achieving up to 90.3% species level identifications. To rapidly detect β-lactamase-mediated resistance to extended spectrum cephalosporins, a high-performance liquid chromatography (HPLC) assay was developed and optimized to detect resistance directly from growth-positive blood cultures. With a 1-hour incubation of bacteria with cefotaxime, resistance could be detected with 95.5% sensitivity and 88.9% specificity. This method was better at detecting resistance mediated by group 1 and 9 CTX-M ESBLs, with reduced sensitivity for the detection of resistance mediated by AmpC β-lactamases. Further research is required to investigate additional markers that could improve the detection of other β-lactamases. Both of these methods could be rapidly integrated into the diagnostic microbiology laboratory, thus reducing the time to effective narrow spectrum antimicrobial therapy, and potentially improving patient outcomes and reducing the spread of antimicrobial resistance.
dc.language.isoen
dc.publisherUniversity of Otago
dc.rightsAll items in OUR Archive are provided for private study and research purposes and are protected by copyright with all rights reserved unless otherwise indicated.
dc.subjectMALDI-ToF
dc.subjectESBL
dc.subjectSusceptibility Testing
dc.subjectCefotaxime
dc.subjectBlood Culture
dc.subjectRapid Identification
dc.subjectSepsityper
dc.subjectHPLC
dc.subjectCeftazidime
dc.subjectCefepime
dc.subjectSepsis
dc.subjectClinical Microbiology
dc.titleNovel Methods for the Rapid Identification and Susceptibility Testing of Blood Culture Isolates
dc.typeThesis
dc.date.updated2014-11-17T23:48:09Z
dc.language.rfc3066en
thesis.degree.disciplineMicrobiology and Immunology
thesis.degree.nameBachelor of Biomedical Sciences with Honours
thesis.degree.grantorUniversity of Otago
thesis.degree.levelHonours
otago.interloanyes
otago.openaccessAbstract Only
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