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Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy. / Natalia, Kapel

Swansea University Author: Natalia, Kapel

Abstract

Infectious diseases remain a serious threat to human life and health as well as having important economical factor. One way of successful combating diseases is designing the most appropriate treatment plan following the correct diagnosis. Therefore, there is a need for a method combining reproducibi...

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Published: 2013
URI: https://cronfa.swan.ac.uk/Record/cronfa42219
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spelling 2018-08-29T14:57:36.8112225 v2 42219 2018-08-02 Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy. 78abde3be61f76dc797a455bc7bb6014 Natalia Kapel Natalia Kapel true false 2018-08-02 Infectious diseases remain a serious threat to human life and health as well as having important economical factor. One way of successful combating diseases is designing the most appropriate treatment plan following the correct diagnosis. Therefore, there is a need for a method combining reproducibility, precision and speed. The aim of this work was to evaluate the potential of micro-Raman spectroscopy for identifying bacteria at different taxonomic levels, strains revealing different antibiotic resistance profiles, and for phylogenetic investigation. The project was based on a selection of bacteria: Staphylococcus aureus (6571, Cowanl), Staphylococcus epidermidis (1457, 9142), Escherichia coli including wild- types (strain B, K12, Top 10), transformants expressing ampicillin and kanamycin resistance (Top10Amp, Top10Kan) and clinical isolates expressing extended-spectrum beta- lactamases (ESBL). Following a precise and detailed protocol, Raman spectra were recorded from bacterial colonies grown overnight on a Colombia Blood Agar. In order to remove background fluorescence, rolling-circle filter procedure was applied. The most critical peaks for differentiation between organisms as well as for characterising each microorganism were determined. The spectral data were analyzed using principal component and cluster analysis techniques. As expected, the degree of separation decreased in the order genus&amp;rarr;species&amp;rarr;strain. It was determined that DNA/RNA, proteins and amino-acids are responsible for the differentiation between strains on a lower level of similarity with more influence of the constituents of the bacterial envelope between more closely related organisms. Raman spectroscopy was capable of differentiating between susceptible and resistant strains as well as monitoring whether the organism has been grown under antibiotic pressure. Based on triplex PCR, clinical isolates of ESBL strains were assigned to one of the phylogenetic group characterising Esherichia genus and it was revealed that within CTX- M TEM-1 there were two distinct clusters of D and B2 groups. Overall we have demonstrated that the combination of micro-Raman spectroscopy, microbiology and bioinformatics has the potential for the successful discrimination of bacteria species and strains, for the determination of antibiotic resistance profiles and investigating phylogenetic grouping in a clinical environment. EThesis Medicine.;Analytical chemistry. 31 12 2013 2013-12-31 COLLEGE NANME Swansea University Medical School COLLEGE CODE Swansea University 2018-08-29T14:57:36.8112225 2018-08-02T16:24:28.4641899 Swansea University Medical School Swansea University Medical School Natalia Kapel 1 0042219-02082018162437.pdf 10797921.pdf 2018-08-02T16:24:37.4330000 Output 21029868 application/pdf E-Thesis true 2018-08-02T16:24:37.4330000 false
title Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
spellingShingle Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
Natalia, Kapel
title_short Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
title_full Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
title_fullStr Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
title_full_unstemmed Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
title_sort Rapid species identification and antimicrobial susceptibility testing using Raman spectroscopy.
author_id_str_mv 78abde3be61f76dc797a455bc7bb6014
author_id_fullname_str_mv 78abde3be61f76dc797a455bc7bb6014_***_Natalia, Kapel
author Natalia, Kapel
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publishDate 2013
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hierarchy_top_title Swansea University Medical School
hierarchy_parent_id swanseauniversitymedicalschool
hierarchy_parent_title Swansea University Medical School
department_str Swansea University Medical School{{{_:::_}}}Swansea University Medical School{{{_:::_}}}Swansea University Medical School
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description Infectious diseases remain a serious threat to human life and health as well as having important economical factor. One way of successful combating diseases is designing the most appropriate treatment plan following the correct diagnosis. Therefore, there is a need for a method combining reproducibility, precision and speed. The aim of this work was to evaluate the potential of micro-Raman spectroscopy for identifying bacteria at different taxonomic levels, strains revealing different antibiotic resistance profiles, and for phylogenetic investigation. The project was based on a selection of bacteria: Staphylococcus aureus (6571, Cowanl), Staphylococcus epidermidis (1457, 9142), Escherichia coli including wild- types (strain B, K12, Top 10), transformants expressing ampicillin and kanamycin resistance (Top10Amp, Top10Kan) and clinical isolates expressing extended-spectrum beta- lactamases (ESBL). Following a precise and detailed protocol, Raman spectra were recorded from bacterial colonies grown overnight on a Colombia Blood Agar. In order to remove background fluorescence, rolling-circle filter procedure was applied. The most critical peaks for differentiation between organisms as well as for characterising each microorganism were determined. The spectral data were analyzed using principal component and cluster analysis techniques. As expected, the degree of separation decreased in the order genus&amp;rarr;species&amp;rarr;strain. It was determined that DNA/RNA, proteins and amino-acids are responsible for the differentiation between strains on a lower level of similarity with more influence of the constituents of the bacterial envelope between more closely related organisms. Raman spectroscopy was capable of differentiating between susceptible and resistant strains as well as monitoring whether the organism has been grown under antibiotic pressure. Based on triplex PCR, clinical isolates of ESBL strains were assigned to one of the phylogenetic group characterising Esherichia genus and it was revealed that within CTX- M TEM-1 there were two distinct clusters of D and B2 groups. Overall we have demonstrated that the combination of micro-Raman spectroscopy, microbiology and bioinformatics has the potential for the successful discrimination of bacteria species and strains, for the determination of antibiotic resistance profiles and investigating phylogenetic grouping in a clinical environment.
published_date 2013-12-31T03:55:25Z
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