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dc.contributor.advisorSebolai, O. M.
dc.contributor.advisorPohl, C. H.
dc.contributor.authorMochochoko, Bonang Michael
dc.date.accessioned2016-09-01T12:07:11Z
dc.date.available2016-09-01T12:07:11Z
dc.date.issued2016-01
dc.identifier.urihttp://hdl.handle.net/11660/4037
dc.description.abstractEnglish: Cryptococcus neoformans is today recognised as an important human pathogen that has arisen from a non-pathogenic terrestrial fungus. Its interactions (specifically antagonistic interactions with other microbes (such as amoeba) in its natural habitat, soil, has led to cryptococcal cells evolving elegant offensive and defensive strategies to survive in nature. Importantly, these qualities seem to be maintained and activated whenever cryptococcal cells engage in antagonistic interactions with other cell types i.e. when in competition with Pseudomonas cells, which is the subject of chapter two, as well as when engaging in parasitism with macrophages, which is the subject of chapter three. Towards this end, in this dissertation, special attention was given to the role of the secondary metabolite, cryptococcal 3-hydroxy C9:0, in mediating the fate of C. neoformans when in contact with these other cell types. In chapter two, it was sought to explore the interactive outcome between cryptoccocal cells and Pseudomonas cells. Both cell types can, in their individual capacity, lead to the development of pneumonia. While the host body has optimal conditions (temperature, ambient air, etc.) to promote the growth of these two pathogens, their growth may be limited by the available nutrients within the confined space of the host lung. It is therefore reasonable to conclude that these microbes would engage in competition in order to exert territorial dominance over the other. To demonstrate this point in vitro, Pseudomonas cells were co-cultured with cryptococcal cells over a period of time. It was subsequently determined that cryptococcal cells dominated Pseudomonas cells. This domination manifested in a significant reduction (p = 0.05) in the number of Pseudomonas cells over a 24-h period. In order to determine if the observed domination could be as a result of the cryptococcal 3-hydroxy fatty acid, 3-OH-C9:0, negatively affecting the growth of Pseudomonas cells, Pseudomonas cells were (in a separate experiment) directly challenged with this molecule. And here it was shown that the growth of Pseudomonas cells was inhibited in a dose-dependent manner by increasing concentrations of this molecule. Importantly, 3-OH-C9:0 inhibited growth of Pseudomonas cells, possibly leading to their death, via altering their membrane function following the incorporation of this saturated molecule into the bilayer leading to a more rigid membrane. Taken together, these findings suggested cryptococcal 3-OH-C9:0 possesses antimicrobial properties, that when secreted into the extracellular environment, negatively affect the fate of surrounding microbes. In chapter three the role of 3-OH-C9:0 in mediating the function of macrophages was investigated. Macrophages are critical in clearing infecting or invading microbial cells through phagocytosis. However, phagocytosis is a receptor-mediated process that is governed by a balance between pro-signal molecules that promote phagocytosis and anti-signal molecules that inhibit it. Thus, it is not surprising that microbes would counter the action of macrophages by producing anti-signal molecules that would subvert the capability of macrophages to clear infecting cells. Critically, the designed macrophage-challenge study showed that 3-OH-C9:0 induced macrophages to produce the protein alpha-2-HS-glycoprotein. This protein is an anti-inflammatory modulator that is important in maintaining immunological homeostasis. This implies this protein counters the effects of pro-inflammatory modulators, which drive the T-helper 1 response in order to clear microbial infections. Furthermore, it was established that macrophages do not produce 3-hydroxy C9:0, and this highlighted that macrophages would not deliberately produce these anti-signal molecules in order to undermine their responsibility of clearing infections. Based on the findings reported in this dissertation, a picture emerges that clearly points that 3-hydroxy fatty acids are crucial to the survival and pathogenesis of cryptococcal cells. According to the Centers of Disease Control and Prevention, cases of disseminated cryptococcal infections remain a major cause of morbidity and mortality. Thus there are considered efforts to manage cryptococcal infections. The studies presented in the dissertation highlight 3-hydroxy fatty acid biosynthetic route as a potential target for controlling the pathogenesis of this medically important fungus. Towards this end, the usage of animals in modelling disseminated cryptococcal infections should be considered in order to clearly establish the therapeutic benefits of drugs such as aspirin, which has previously been shown to inhibit production of these molecules in a dose-dependent manner.en_ZA
dc.description.abstractAfrikaans: Cryptococcus neoformans word tans erken as ‘n belnagrike menslike patogeen wat onstaan het vanaf ‘n nie-patogeniese terrestriële fungus. Sy interaksies (veral antagonistiese interaksies) met ander mikrobe (soos amoeba) in sy natuurlike habitat, grond, het daartoe gelei dat Cryptococcus selle elegante aanvals- en verdedigingstrategieë ontwikkel het om in die natuur te oorleef. Dit is van belang dat hierdie eienskaape blyk behou en geaktiveer te word wanneer Cryptococcus selle deelneem aan antagonistiese interaksies met ander tipes selle d.i. wanneer hulle kompeteer met Pseudomonas selle, die onderwerp van hoofstuk twee, asook wanneer hulle deelneem aan parasitisme met makrofage, die onderwerp van hoofstuk drie. Ter nastrewe van hierdie doel, word in hierdie verhandeling veral aandag gegee aan die rol van die sekondêre metaboliet, 3-hidroksie C9:0 (3-OH-C9:0) vanaf Cryptococcus, in bepaling van die lot van C. neoformans wanner dit in kontak is met hierdie ander seltipes. In hoofstuk twee is gepoog om die interaktiewe uitkoms tussen Cryptococcus selle en Pseudomonas selle te ondersoek. Beide seltipes kan individueel lei tot ontstaan van longontsteking. Alhoewel die gasheerliggaam optimale toestande (temperatuur, lug ens.) bevat om die groei van hierdie twee patogene te promoveer, mag hulle groei beperk word deur die beskikbare voedingstowwe binne die beperkte spasie van die gasheerlong. Dit kan dus redelikerwys afgelei word dat hierdie mikrobes sal deelneem aan kompetisie om territorial domanansie oor die ander uit te oefen. Om hierdie punt in vitro te demonstreer is Pseudomonas selle saam met Cryptococcus selle gekweek oor ‘n seker periode. Gevolglik is gevind dat Cryptococcus selle Pseudomonas selle domineer. Hierdie domineering het gemanifesteer as ‘n beduidende afname (p = 0.05) in die getal Pseudomonas selle oor ‘n tydperk van 24-h. Ten einde te bepaal of die waargeneemde dominering kan wees as gevolg van negatiewe invloed van die Cryptococcus 3-hidroksie vetsuur, 3-OH-C9: 0, op die groei van Pseudomonas selle, is die Pseudomonas selle (in 'n aparte eksperiment) direk blootgestel aan hierdie verbinding. Hier is aangetoon dat die groei van Pseudomonas selle in 'n dosis-afhanklike wyse geïnhibeer word deur toenemende konsentrasies van hierdie verbinding. Wat belangrik is, 3-OH-C9:0 het groei van Pseudomonas selle geïnhibeer, wat moontlik kon lei tot hulle dood, via die wysiging van hul membraanfunksie na die invoeging van hierdie versadigde molekule in die dubbellaag, wat lei tot 'n meer rigiede membraan. Hierdie bevindinge dui saam daarop dat Cryptococcus 3OH-C9:0 antimikrobiese eienskappe besit, wat wanneer dit afgeskei word in die ekstrasellulêre omgewing, ʼn negatiewe invloed op die lot van omliggende mikrobes gehad het. In hoofstuk drie word die rol van 3-OH-C9:0 in die bevordering van die funksie van makrofage ondersoek. Makrofage is van kritieke belang in die opklaar van infeksie of indringer mikrobiese selle deur fagositose. Maar fagositose is 'n reseptor bemiddelde proses wat deur 'n balans tussen pro-sein molekules wat fagositose promoveer en anti-sein molekules wat dit inhibeer, gereguleer word. Dus is dit nie verbasend dat mikrobes die werking van makrofage sou teen werk deur die vervaardiging van anti-sein molekules wat die vermoë van die makrofage sou ondermyn om infekterende selle te verwyder nie. Die ontwerpte makrofaag-blootstellingstudie het getoon dat 3-OH-C9:0 makrofage geïnduseer het om die proteïen, alpha-2-HS-glikoproteïen, te produseer. Hierdie proteïen is 'n anti-inflammatoriese modulator wat belangrik is in die handhawing van immunologiese homeostase. Dit impliseer hierdie proteïen die effek van pro-inflammatoriese modulators, wat die T-helper 1 reaksie om mikrobiese infeksies op te klaar dryf, teenwerk. Verder is vasgestel dat makrofage nie 3-hidroksie C9:0 produseer nie, en dit beklemtoon dat makrofage nie doelbewus hierdie anti-sein molekules sou produseer om sodoende hul verantwoordelikheid vir die opklaar van infeksies te vermy nie. Gebaseer op die bevindinge in hierdie verhandeling, verskyn ʼn beeld wat duidelik wys dat die 3-hidroksie vetsure noodsaaklik is vir die oorlewing en patogenese van Cryptococcus selle. Volgens die Centers van Disease Control & Prevention, bly gevalle van verspreide Cryptococcus infeksies 'n groot oorsaak van morbiditeit en mortaliteit. Dus word ernstige pogings aangewernd om Cryptococcus infeksies te bestuur. Die studies voorgedra in hierdie verhandeling, beklemtoon die 3-hidroksie vetsuur biosintetiese weg as 'n potensiële teiken vir die beheer van die patogenese van hierdie medies belangrike fungus. Dit ten doel, moet die gebruik van diere in modellering van verspreide Cryptococcus infeksies oorweeg word om die terapeutiese voordele van middels soos aspirien, wat voorheen reeds aangetoon is om die produksie van hierdie molekules in 'n dosis-afhanklike wyse te inhibeer, duidelik te vestig.af
dc.description.sponsorshipNational Research Foundation (NRF)en_ZA
dc.description.sponsorshipNMDSen_ZA
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.subjectAnti-microbialen_ZA
dc.subjectAnti-inflammatory modulatoren_ZA
dc.subject3-hydroxy C9:0en_ZA
dc.subjectCryptococcusen_ZA
dc.subjectCompetitionen_ZA
dc.subjectImmunityen_ZA
dc.subjectMacrophagesen_ZA
dc.subjectParasitismen_ZA
dc.subjectPseudomonasen_ZA
dc.titleThe role of cryptococcal 3-hydroxy fatty acids in mediating interspecies interactionsen_ZA
dc.typeDissertationen_ZA
dc.rights.holderUniversity of the Free Stateen_ZA


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