{"id":{"repo_id":"east-anglia","oai_identifier":"oai:ueaeprints.uea.ac.uk:48103"},"canonical_url":"https://search.dev.ndltd.org/etd/east-anglia/oai:ueaeprints.uea.ac.uk:48103","repository":{"repo_id":"east-anglia","name":"University of East Anglia","base_url":"https://ueaeprints.uea.ac.uk/cgi/oai2"},"display":{"title":"Antimicrobial Carbohydrate Vaccines: Development of Burkholderia pseudomallei immunogens","abstract":"The potential bio-terror threat posed by Burkholderia pseudomallei highlights the need for an effective vaccine. Immunisation and challenge experiments in mice have demonstrated that the capsular polysaccharide (CPS-1) of B. pseudomallei, which is composed of β-1,3-linked 6-deoxy-D-manno-heptopyranose residues, is a promising candidate for vaccine development. This thesis set out to explore routes to potential vaccine candidates for Burkholderia pseudomallei infection based on both bottom-up synthetic monosaccharide constructs and top-down CPS-1 conjugates. Initially this thesis focused on the development of a monosaccharide antigen based on the 6-deoxy-D-manno-heptose building block of CPS-1. The antigens were conjugated to TetHc carrier protein via an 8-(methoxycarbonyl)octyl linker using traditional acyl hydrazide conjugation chemistry. The TetHc monosaccharide conjugates were analysed by SDS-PAGE and MALDI-ToF mass spectrometry before undergoing immunisation trials in sheep. Immunological assessment of the resulting polyclonal antisera was conducted by ELISA, slot-blot and the Octet biosensor system. These studies demonstrated the generation of antibodies that were specific for the cognate monosaccharide. Preliminary investigations showed that the 6-deoxy-D-mannoheptose- derived antibodies reacted positively with the natural CPS-1. The linker was shown to be important in the specificity of the polyclonal sera, leading to cross reactivity with non-parent monosaccharide antigens. A second generation of antigens was developed using a short linker, 3-(methyl mercaptopropionate). The expression and purification of the natural B. pseudomallei CPS-1 was achieved in an avirulent strain of Burkholderia, B. thailandensis E555. Characterisation of the capsular polysaccharide highlighted the co-expression of an α-1,3-mannan polysaccharide. Mild acid hydrolysis of the CPS-1 preparation, combined with capillary electrophoresis was trialled with a view to the generation of oligosaccharide fragments of CPS-1 for conjugation and immunisation studies. While initial carbohydrate conjugates were prepared with a previously described tetanus toxoid Hc fragment, the development and in planta expression of a chemically conjugatable Hepatitis B core virus-like particle system was also achieved.","abstract_html":"The potential bio-terror threat posed by Burkholderia pseudomallei highlights the need for an effective vaccine. Immunisation and challenge experiments in mice have demonstrated that the capsular polysaccharide (CPS-1) of B. pseudomallei, which is composed of β-1,3-linked 6-deoxy-D-manno-heptopyranose residues, is a promising candidate for vaccine development. This thesis set out to explore routes to potential vaccine candidates for Burkholderia pseudomallei infection based on both bottom-up synthetic monosaccharide constructs and top-down CPS-1 conjugates. Initially this thesis focused on the development of a monosaccharide antigen based on the 6-deoxy-D-manno-heptose building block of CPS-1. The antigens were conjugated to TetHc carrier protein via an 8-(methoxycarbonyl)octyl linker using traditional acyl hydrazide conjugation chemistry. The TetHc monosaccharide conjugates were analysed by SDS-PAGE and MALDI-ToF mass spectrometry before undergoing immunisation trials in sheep. Immunological assessment of the resulting polyclonal antisera was conducted by ELISA, slot-blot and the Octet biosensor system. These studies demonstrated the generation of antibodies that were specific for the cognate monosaccharide. Preliminary investigations showed that the 6-deoxy-D-mannoheptose- derived antibodies reacted positively with the natural CPS-1. The linker was shown to be important in the specificity of the polyclonal sera, leading to cross reactivity with non-parent monosaccharide antigens. A second generation of antigens was developed using a short linker, 3-(methyl mercaptopropionate). The expression and purification of the natural B. pseudomallei CPS-1 was achieved in an avirulent strain of Burkholderia, B. thailandensis E555. Characterisation of the capsular polysaccharide highlighted the co-expression of an α-1,3-mannan polysaccharide. Mild acid hydrolysis of the CPS-1 preparation, combined with capillary electrophoresis was trialled with a view to the generation of oligosaccharide fragments of CPS-1 for conjugation and immunisation studies. While initial carbohydrate conjugates were prepared with a previously described tetanus toxoid Hc fragment, the development and in planta expression of a chemically conjugatable Hepatitis B core virus-like particle system was also achieved.","abstract_has_math":false,"creators":["Donaldson, Matthew"],"institution":"University of East Anglia","degree_name":"phd","degree_level":"doctoral","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2013,"date_issued":"2013-09","date_published":"2013-09","updated_at":"2026-07-24T02:11:59Z","subjects":[],"languages":["en"],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":null,"outbound_label":null,"outbound_source":null},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Donaldson, Matthew"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2013-09"]},{"key":"dc:date.issued","label":"Date","values":["2013-09"]},{"key":"dc:publisher.department","label":"Dc Publisher Department","values":["School of Chemistry"]},{"key":"dc:publisher.institution","label":"Dc Publisher Institution","values":["University of East Anglia"]},{"key":"dc:relation.isreferencedby","label":"Dc Relation Isreferencedby","values":["https://ueaeprints.uea.ac.uk/id/eprint/48103/"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]},{"key":"dc:type.qualificationlevel","label":"Dc Type Qualificationlevel","values":["doctoral"]},{"key":"dc:type.qualificationname","label":"Dc Type Qualificationname","values":["phd"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://ueaeprints.uea.ac.uk/id/eprint/48103/1/M_Donaldson_2013.pdf"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["The potential bio-terror threat posed by Burkholderia pseudomallei highlights the need for an effective vaccine. Immunisation and challenge experiments in mice have demonstrated that the capsular polysaccharide (CPS-1) of B. pseudomallei, which is composed of β-1,3-linked 6-deoxy-D-manno-heptopyranose residues, is a promising candidate for vaccine development. This thesis set out to explore routes to potential vaccine candidates for Burkholderia pseudomallei infection based on both bottom-up synthetic monosaccharide constructs and top-down CPS-1 conjugates. Initially this thesis focused on the development of a monosaccharide antigen based on the 6-deoxy-D-manno-heptose building block of CPS-1. The antigens were conjugated to TetHc carrier protein via an 8-(methoxycarbonyl)octyl linker using traditional acyl hydrazide conjugation chemistry. The TetHc monosaccharide conjugates were analysed by SDS-PAGE and MALDI-ToF mass spectrometry before undergoing immunisation trials in sheep. Immunological assessment of the resulting polyclonal antisera was conducted by ELISA, slot-blot and the Octet biosensor system. These studies demonstrated the generation of antibodies that were specific for the cognate monosaccharide. Preliminary investigations showed that the 6-deoxy-D-mannoheptose- derived antibodies reacted positively with the natural CPS-1. The linker was shown to be important in the specificity of the polyclonal sera, leading to cross reactivity with non-parent monosaccharide antigens. A second generation of antigens was developed using a short linker, 3-(methyl mercaptopropionate). The expression and purification of the natural B. pseudomallei CPS-1 was achieved in an avirulent strain of Burkholderia, B. thailandensis E555. Characterisation of the capsular polysaccharide highlighted the co-expression of an α-1,3-mannan polysaccharide. Mild acid hydrolysis of the CPS-1 preparation, combined with capillary electrophoresis was trialled with a view to the generation of oligosaccharide fragments of CPS-1 for conjugation and immunisation studies. While initial carbohydrate conjugates were prepared with a previously described tetanus toxoid Hc fragment, the development and in planta expression of a chemically conjugatable Hepatitis B core virus-like particle system was also achieved."]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["Antimicrobial Carbohydrate Vaccines: Development of Burkholderia pseudomallei immunogens"]}]}],"canonical_facts":{"dc:creator":["Donaldson, Matthew"],"dc:date":["2013-09"],"dc:date.issued":["2013-09"],"dc:description.abstract":["The potential bio-terror threat posed by Burkholderia pseudomallei highlights the need for an effective vaccine. Immunisation and challenge experiments in mice have demonstrated that the capsular polysaccharide (CPS-1) of B. pseudomallei, which is composed of β-1,3-linked 6-deoxy-D-manno-heptopyranose residues, is a promising candidate for vaccine development. This thesis set out to explore routes to potential vaccine candidates for Burkholderia pseudomallei infection based on both bottom-up synthetic monosaccharide constructs and top-down CPS-1 conjugates. Initially this thesis focused on the development of a monosaccharide antigen based on the 6-deoxy-D-manno-heptose building block of CPS-1. The antigens were conjugated to TetHc carrier protein via an 8-(methoxycarbonyl)octyl linker using traditional acyl hydrazide conjugation chemistry. The TetHc monosaccharide conjugates were analysed by SDS-PAGE and MALDI-ToF mass spectrometry before undergoing immunisation trials in sheep. Immunological assessment of the resulting polyclonal antisera was conducted by ELISA, slot-blot and the Octet biosensor system. These studies demonstrated the generation of antibodies that were specific for the cognate monosaccharide. Preliminary investigations showed that the 6-deoxy-D-mannoheptose- derived antibodies reacted positively with the natural CPS-1. The linker was shown to be important in the specificity of the polyclonal sera, leading to cross reactivity with non-parent monosaccharide antigens. A second generation of antigens was developed using a short linker, 3-(methyl mercaptopropionate). The expression and purification of the natural B. pseudomallei CPS-1 was achieved in an avirulent strain of Burkholderia, B. thailandensis E555. Characterisation of the capsular polysaccharide highlighted the co-expression of an α-1,3-mannan polysaccharide. Mild acid hydrolysis of the CPS-1 preparation, combined with capillary electrophoresis was trialled with a view to the generation of oligosaccharide fragments of CPS-1 for conjugation and immunisation studies. While initial carbohydrate conjugates were prepared with a previously described tetanus toxoid Hc fragment, the development and in planta expression of a chemically conjugatable Hepatitis B core virus-like particle system was also achieved."],"dc:format":["application/pdf"],"dc:identifier.uri":["https://ueaeprints.uea.ac.uk/id/eprint/48103/1/M_Donaldson_2013.pdf"],"dc:language":["en"],"dc:publisher.department":["School of Chemistry"],"dc:publisher.institution":["University of East Anglia"],"dc:relation.isreferencedby":["https://ueaeprints.uea.ac.uk/id/eprint/48103/"],"dc:title":["Antimicrobial Carbohydrate Vaccines: Development of Burkholderia pseudomallei immunogens"],"dc:type":["Thesis"],"dc:type.qualificationlevel":["doctoral"],"dc:type.qualificationname":["phd"]},"updated_at":"2026-07-24T02:11:59Z"}