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dc.contributor.advisorPohl-Albertyn, C. H.
dc.contributor.advisorAlbertyn, J.
dc.contributor.advisorElls, R.
dc.contributor.authorFourie, Ruan
dc.date.accessioned2016-08-31T09:11:34Z
dc.date.available2016-08-31T09:11:34Z
dc.date.issued2016-01
dc.identifier.urihttp://hdl.handle.net/11660/4027
dc.description.abstractEnglish: Interkingdom interactions between microorganisms facilitate the growth and survival of microbial communities, and the understanding of these interactions can be beneficial to mankind. These interactions may also be detrimental to human health, with the combination of virulence factors promoting the survival of microbial populations during infection. A relevant model for this, is the interaction between Candida albicans and Pseudomonas aeruginosa. These two microorganisms are frequently found together at infection sites. During infection, C. albicans and P. aeruginosa elicit the release of arachidonic acid (AA), utilized by the host to form immunomodulatory compounds termed eicosanoids. Among these is prostaglandins, for example, prostaglandin E2 (PGE2). Prostaglandin E2 can inhibit Th1 and promote Th2 responses. In combination to other eicosanoids, they modulate inflammation in hosts, ultimately affecting the ability of the host to clear infection. Candida albicans and P. aeruginosa are also able to produce immunomodulatory eicosanoids from exogenous AA. This study confirms the production of PGE2 by C. albicans monomicrobial biofilms, together with the production of PGF2α and 15-hydroxyeicosatetraenoic acid (15-HETE) with the use of enzyme-linked immunosorbent assay (ELISA) and LC-MS/MS for PGE2 confirmation. The production of these eicosanoids is also reported here by P. aeruginosa monomicrobial biofilms. This study is the first to identify authentic PGE2 production by P. aeruginosa biofilms. In addition, polymicrobial biofilms were shown to produce significantly more eicosanoids that monomicrobial counterparts, possibly contributing to the increased morbidity during co-infection by these pathogens. Although the pathways and enzymes involved in eicosanoid production by mammalian systems have been well studied, the production of eicosanoids by microorganisms requires much research. This is due to the fact that microorganisms frequently don’t possess homologs to mammalian enzymes responsible for eicosanoid production. Therefore, inhibitors previously identified to inhibit C. albicans PGE2 production, were evaluated in terms of their effect on eicosanoid production by monomicrobial and polymicrobial biofilms of C. albicans and P. aeruginosa. The inhibitors used are acetylsalicylic acid (ASA, a cyclooxygenase inhibitor), ammonium tetrathiomolybdate (ATM, inhibition of copper-dependant enzymes) and nordihydroguaiaretic acid (NDGA, a potent antioxidant inhibiting various enzyme classes). A possible “shift” in eicosanoid production by C. albicans is seen in the presence of ASA as well as ATM. This phenomenon is also seen for P. aeruginosa in the presence of ATM. Interestingly, ASA increased eicosanoid production by P. aeruginosa. The anti-oxidant NDGA decreased eicosanoid production by monomicrobial, as well as polymicrobial biofilms. Different profiles for eicosanoid production obtained between monomicrobial and polymicrobial biofilms in the presence of ASA and ATM were observed suggest the complex interaction of C. albicans and P. aeruginosa in terms of eicosanoid production. In addition, the inhibitors caused dramatic alterations in polymicrobial biofilm morphology. Interestingly, although these inhibitors did not affect C. albicans metabolic activity or biofilm biomass, ASA caused a significant increase in P. aeruginosa metabolic activity. In addition, P. aeruginosa metabolic activity was significantly inhibited by NDGA. The possible clinical relevance of these findings warrant further investigation, as the use of inhibitors, such as the ones used in the present study, could possibly affect virulence and the ability of hosts to clear infection. This study also evaluated the role of a secretable 15-lipoxygenase produced by P. aeruginosa capable of converting AA to 15-HETE, although further research and methodology is needed to elucidate its role. This study is the first to investigate the production of eicosanoids by polymicrobial biofilms of C. albicans and P. aeruginosa. The increased production of these eicosanoids compared to monomicrobial counterparts, suggest that the microbially produced eicosanoids may possibly play a role in pathogen-pathogen interaction, as well as host-pathogen interaction. This may ultimately affect the ability of the host to clear infection. In addition, with the use of inhibitors, the possible involvement of various enzymatic groups can be speculated during polymicrobial eicosanoid production. Further research into this interaction may provide valuable insight into polymicrobial eicosanoid production and may contribute to possible therapeutic intervention strategies during monomicrobial and polymicrobial infection.en_ZA
dc.description.abstractAfrikaans: Interkoninkryk-interaksies tussen mikro-organismes fasiliteer die groei en oorlewing van mikrobiese populasies en begrip van hierdie interaksies kan vir die mensdom voordelig wees. Hierdie interaksies kan ook nadelig wees vir menslike gesondheid, waar die kombinasie van virulensiefaktore die oorlewing van mikrobiese populasies tydens infeksie bevorder. ‘n Relevante model vir hierdie is die interaksie tussen Candida albicans en Pseudomonas aeruginosa. Hierdie twee mikro-organismes word gereeld saam gevind tydens infeksie. Gedurende infeksie veroorsaak C. albicans en P. aeruginosa die vrystelling van arachidoonsuur (AA), wat gebruik word deur die gasheer vir die produksie van immuunmodulerende verbindings, waarna verwys word as eikosanoïede. Prostaglandiene, insluitend prostaglandien E2 (PGE2) vorm deel van hierdie groep. Prostaglandien E2 kan die Th1-reaksie inhibeer en die Th2-reaksie bevorder. In kombinasie met ander eikosanoïede, kan hierdie verbindings die inflammasie van die gasheer moduleer, wat uiteindelik die vermoë van die gasheer om infeksie te beveg affekteer. Candida albicans en P. aeruginosa kan ook hierdie immuunmodulerende eikosanoïede vanaf eksterne AA produseer. Die huidige studie bevestig die produksie van PGE2 deur C. albicans mono-mikrobiese biofilms, sowel as die produksie van PGF2α en 15-hidroksie-eikosatetraenoësuur (15-HETE) deur die gebruik van ‘n ensiem-gebonde immuunadsorberende toets (ELISA) en LC-MS/MS vir die bevestiging van PGE2. Die produksie van hierdie eikosanoïede deur P. aeruginosa mono-mikrobiese biofilms word ook hier berig. Die huidige studie is die eerste om PGE2-produksie deur P. aeruginosa biofilms the rapporteer. Bykomend word aangetoon dat poli-mikrobiese biofilms aansienlik meer eikosanoïede produseer in vergelyking met die mono-mikrobiese biofilms. Hierdie verhoogde produksie kan moontlik aanleiding gee tot die verhoogde morbiditeit gedurende ko-infeksie deur hierdie patogene. Alhoewel die weë en ensieme wat ‘n rol speel in die produksie van eikosanoïede deur soogdiere goed bestudeer is, kan dieselfde nie gesê word vir die produksie van eikosanoïede deur mikro-organismes nie. Dit is as gevolg van die feit dat mikro-organsismes selde homoloë van soogdier-ensieme vir die produksie van eikosanoïede besit. As gevolg hiervan, was inhibitore, wat voorheen bewys is om PGE2-produksie deur C. albicans te inhibeer, geëvalueer ten opsigte van hul effek op eikosanoïed-produksie deur mono- en poli-mikrobiese biofilms. Die inhibitore wat gebruik is, sluit asetielsalisiensuur (ASA, ‘n sikloöksigenase inhibitor), ammonium-tetratiomolibdaat (ATM, ‘n inhibitor van koper-afhanklike ensieme) en nordihidroguaiariensuur (‘n antioksidant wat ‘n verskeidenheid ensiemklasse inhibeer) in. ‘n Moontlike verskuiwing van ekosanoïed-produksie deur C. albicans in die teenwoordigheid van ASA en ATM is waargeneem. Hierdie verskuiwing is ook gesien in die geval van P. aeruginosa biofilms in die teenwoordigheid van ATM. ‘n Toename in eikosanoïed-produksie deur P. aeruginosa in die teenwoordigheid van ASA is waargeneem. Die anti-oksidant NDGA het eikosanoïed-produksie deur mono- en poli-mikrobiese biofilms inhibeer. Verskillende profiele vir eikosanoïed-produksie deur mono- en poli-mikrobiese biofilms wat met ASA en NDGA behandel is, is verkry, wat ‘n komplekse interaksie tydens ko-inkubasie aandui. Die inhibitore het ook dramatiese verskille in poli-mikrobiese biofilm morfologie veroorsaak. Alhoewel die metaboliese aktiwiteit en biofilm biomassa vand C. albicans nie beïnvloed deur die inhibitore was nie, was daar ‘n verhoging in die metabolise aktiwiteit van P. aeruginosa biofilms in die teenwoordigheid van ASA. Die metabolise aktiwiteit van hierdie bakteriese biofilms was ook aansienlik verlaag deur NDGA. Die kliniese effek van hierdie bevindings sal verder evalueer moet word, aangesien die inhibitore wat gebruik is in hierdie studie moontlik die virulensie van die patogene, sowel as die vermoë van die gasheer om ontslae te raak van infeksie kan beïnvloed. Hierdie studie het ook die rol van die 15-lipoksigenase van P. aeruginosa geëvalueer, maar verdere navorsing is nodig om hierdie faset van interaksie te verstaan. Die huidige studie is die eerste van sy tipe om die produksie van eikosanoïede deur poli-mikrobiese biofilms van C. albicans en P. aeruginosa te evalueer. Die verhoogde produksie hiervan deur poli-mikrobiese biofilms dui moontlik aan dat mikrobiese eikosanoïede ‘n rol kan speel gedurende patogeen-patogeen interaksies, sowel as patogeen-gasheer interaksies. Dit kan moontlik die vermoë van die gasheer om patogene te verwyder beïnvloed. Die studie het ook resultate gegee wat lei tot die moontlike identifikasie van ensieme wat ‘n rol kan speel in die produksie van eikosanoïede deur poli-mikrobiese biofilms. Verdere navorsing kan belangrike inligting wat voordelig kan wees in kliniese behandeling tydens infeksie van C. albicans en P. aeruginosa bied.af
dc.language.isoenen_ZA
dc.publisherUniversity of the Free Stateen_ZA
dc.subjectDissertation ((M.Sc. (Microbial, Biochemical and Food Biotechnology))--University of the Free State, 2016en_ZA
dc.subjectArachidonic aciden_ZA
dc.subjectBiofilmsen_ZA
dc.subjectCandida albicansen_ZA
dc.subjectEicosanoidsen_ZA
dc.subjectPolymicrobialen_ZA
dc.subjectPseudomonas aeruginosaen_ZA
dc.titleIn vitro arachidonic acid metabolism by polymicrobial biofilms of Candida albicans and Pseudomonas aeruginosaen_ZA
dc.typeDissertationen_ZA
dc.rights.holderUniversity of the Free Stateen_ZA


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