The yeast Candida is an opportunistic human pathogen often associated with nosocomial infections. Understanding of the fungal virulence factor is limited. Adhesion and biofilm formation on prosthetic devices is likely to be one such factor for Candida infection. Biofilms form a structured community embedded by exopolymeric substances. Catheter associated Candidiasis is problematic since biofilm is hard to eradicate by antifungal therapy alone. / C. parapsilosis is a normal human commensal commonly isolated from human skin. C. parapsilosis is considered to be an emerging fungal pathogen and particularly associated with infections in neonates and catheterized intensive care unit (ICU) patients. Thus investigating the characteristics of biofilm formation helps to understand the biology of C. parapsilosis biofilm and assess the prognosis of the infection. / Also, we still only know a little about the mechanisms of biofilm formation in C. parapsilosis. Genetic studies about C. parapsilosis biofilm, especially in clinical samples are rare. Agglutinnin-like sequence (ALS) genes are commonly studied and well known in C. albicans but not in C. parapsilosis. Recently, some bioinformatics studies had revealed the presence of five ALS genes in C. parapsilosis. Some differentially expressed genes in C. parapsilosis biofilm were also identified in other studies. In this study, our aim is to investigate the characteristics of the clinical isolates of C. parapsilosis and the expressions of the selected genes in the biofilm. / In this study, the demographic characteristics of 61 patients with C. parapsilosis fungemia were studied. Among the patients, 60.6% aged higher than 65 years old. The risk factors of previous antibiotic treatment and venous catheterization were both towered in 72.13% of the patients. No resistant C. parapsilosis against amphotericin B and fluconazole was isolated in the studied samples. A static biofilm formation model was designed to investigate the biofilm formation of the samples using 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H-tetra zolium hydroxide (XTT) assay. Quantitative real-time PCR was carried out to evaluate the temporal expression five ALS gene and one ERG11 gene in the biofilm of the clinical samples. The differential expression of the five ALS genes and ERG11 gene of the high biofilm forming group and low biofilm forming group of the clinical samples were compared. Our result shows that CPAG_05056 and CPAG_00369 were relatively more expressed in the high biofilm forming group and was upregulated at 24 h. CPAG_05314 was relatively more expressed in the high biofilm forming group and was more upregulated at 1.5h. The expression of CPAG_05054 was relatively higher in high biofilm forming group but showed similar expression level at 1.5 h and 24 h for both groups of samples. CPAG_00368 and ERG11 expression was relatively higher in low biofilm forming group and downregulated at 1.5 h and 24 h for both groups of samples. / In conclusion, C. parapsilosis bloodstream infection was prevalence in the elderly. Patients with prolonged hospital stay, being in the ICU, received antibiotic treatments and venous catheter are at a very high risk of infection. The ability to form biofilm in C. parapsilosis showed great variations among the clinical samples tested in our static biofilm formation model. The expression of CPAG_05056, and CPAG_00369 may be involved in the late stage of biofilm formation by C. parapsilosis. Expression of CPAG_05314 is believed to be involved in the early stage of biofilm formation. The expression of CPAG_05054 may be involved in both early and late stage of biofilm formation. CPAG_00368 and ERG11 were downregulated and seemed to show no relation to biofilm formation to both groups in our tested model. The result indicated that these genes may only be partly responsible for biofilm formation. The mechanisms of biofilm formation in C. parapsilosis is far more complicated. / 念珠菌是一種機會性人類病原體，並經常與院內感染有關。現時對於真菌致病因子的了解是有限的。而粘附在人工物質上的和形成生物膜的特性可能是一個念珠菌感染的因素。生物膜形成一個被外在聚合體包圍的結構化的群落。而導管相關性念珠菌引發血流感染是有其存在的問題，因為單靠抗真菌治療是很難根除生物膜。 / 近平滑念珠菌是一種正常的人類定植真菌並分佈於人體皮膚表面。近平滑念珠菌被認為是一個新興的病原真菌，特別是感染嬰兒和有插管的深切治療病房病人。因此，調查生物膜形成的特點能有助於理解近平滑念珠菌生物膜的生物學和有效預防感染。 / 此外，研究人員仍然對近平滑念珠菌生物膜形成的機制了解甚少。而關於近平滑念珠菌生物膜的基因研究，尤其是對其臨床標本研究的是很罕見的。類凝集素基因(ALS)基因的研究目前集中在於白色念珠菌，但不是在近平滑念珠菌。最近，在一些生物信息學研究中揭示了ALS基因亦存在於近平滑念珠菌。而其他在近平滑念珠菌生物膜中差異表達的基因在其他研究中也被確定。本研究的目的是調查近平滑念珠菌的臨床標本的特性和所選擇的基因在臨床標本所形成的生物膜中的表達。 / 在這項研究中，對61個近平滑念珠菌血液感染患者的人口學特徵進行了研究。而其中60.6％的病人年齡高於65歲。原有的抗生素治療，靜脈置管的危險因素則有72.13％的患者伴有。在所研究的臨床樣品中無發現對兩性黴素B和氟康唑有抗性的近平滑念珠菌。利用靜態生物膜形成模型和使用XTT法來檢測臨床標本的生物膜形成。進行了定量實時聚合酶鏈反應來評測五個ALS基因和ERG11基因在臨床樣品的生物膜形成中的時間性表達。五個ALS基因和ERG11基因在高生物膜形成組和低生物膜的形成組的臨床樣本中的差異表達進行了比較。我們的結果表明，CPAG_05056基因和CPAG_00369基因相對更表現在高生物膜成形組，並在二十四小時的時間點上調。CPAG_05314基因相對在高生物膜形成組表達並在一小時半的時間點上調。高生物膜形成組的CPAG_05054基因表達較高，但在一小時半和二十四小時都表現出相似的表達水平。CPAG_00368基因和ERG11基因表達相對在低生物膜形成組較高，而兩組樣品的表達都一小時半和二十四小時下調。 / 本次研究得出的結論是，近平滑念珠菌血流感染在中老年人的患病率較高。長期住院的患者，在深切治療病房，接受抗生素治療和有靜脈插管都會提高感染風險。在測試所用的靜態生物膜形成模型中，近平滑念珠菌臨床樣本之間的生物膜形表現出不同的變化。而的CPAG_05056基因和CPAG_00369基因的表達可能和近平滑念珠菌生物膜形成的後期有關。CPAG_05314基因表達被認為是涉及生物膜形成的早期階段。CPAG_05054基因表達可能涉及生物膜形成的早期和後級。在本次測試模型中，CPAG_00368和ERG11的基因表達在兩個組別下調，可能與生物膜形成沒有直接關係。以上證明所研究的基因可能只涉及生物膜形成過程的一部份而近平滑念珠菌的生物膜形成應更為複雜。 / Man, Wing Chung. / Thesis M.Phil. Chinese University of Hong Kong 2014. / Includes bibliographical references (leaves 91-102). / Abstracts also in Chinese. / Title from PDF title page (viewed on 24, October, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_1291545 |
| Date | January 2014 |
| Contributors | Man, Wing Chung (author.), Hui, Mamie (thesis advisor.), Chinese University of Hong Kong Graduate School. Division of Microbiology. (degree granting institution.) |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography, text |
| Format | electronic resource, electronic resource, remote, 1 online resource (xvii, 116 leaves) : illustrations, computer, online resource |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons "Attribution-NonCommercial-NoDerivatives 4.0 International" License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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