探討剔除白色念珠菌 CaORF19.3360、CaORF19.4965 和 CaORF19.6268 對白色念珠菌黏附、型態變化和生長情況之影響

Abstract

白色念珠菌（Candida albicans）是一種伺機性病原真菌，共生於人體當中。當宿主免疫功能低落時，白色念珠菌就能造成口腔或生殖道的感染。目前已知在細菌和真菌中，FLO 基因可以做出細胞壁的蛋白，並且該基因包含了串聯重複的序列（tandem repeat sequence），而此基因與免疫原性的改變、黏附能力（adhesion）和致病力（pathogenesis）有關聯。因此想知道若白色念珠菌基因序列中帶有重複序列的現象， 是否會與致病力有關。因此選擇了 CaORF19.3360、CaORF19.4965 和 CaORF19.6268 這三個也包含編碼重複序列的基因進行本研究。利用包含一個藥物篩選標記的 SAT1 flipper cassette ，分別建構出此三個基因的雙套剔除株，並進行型態變化、生長曲線和生物膜形成的測驗。結果顯示，與型態相關的芽管生成、菌絲生成和侵犯力的實驗中，三個基因的雙套剔除株和野生株沒有明顯差異；在生長速率的測驗中，三個基因雙套剔除株和野生株也無明顯差異；而在生物膜的測驗中，CaORF19.3360 單套剔除株的生物膜穩定性較野生型和雙套剔除株的差，CaORF19.4965 雙套剔除株的生物膜穩定性較野生型和單套剔除株的差，CaORF19.6268雙套剔除株的生物膜穩定性明顯的比野生株和單套剔除株差。

Candida albicans is an opportunistic fungal pathogens. It is a commensal microorganism in human. When the host is immunocompromised, it could cause oral and vaginal mucosal infection. In bacteria and yeasts, FLO encodes cell wall proteins containing repeat sequence. The FLO gene is associated with alterations in immunogenicity, adhesion, and pathogenesis. Hence, it was suggested that repeat sequences may be associated with pathogenesis. CaORF19.3360, CaORF19.4965 and CaORF19.6268 genes of Candida albicans also contains repeat sequences in the coding region. Hence, I am interested to know whether those genes may affect pathogenesis. I used the SAT1 flipper cassette, containing a drug resistance marker, to disrupt the coding sequences of CaORF19.3360, CaORF19.4965 and CaORF19.6268 genes, separately. Then, I determined the effect on their morphogenesis, growth rates and biofilm formation. In conclusion, there is no significant difference between the homozygous knock-out strains and the wild-type strain in their morphogenesis and growth rates under the tested conditions. In biofilm formation test, CaORF19.3360 heterozygous knock-out strain is worse than the homozygous strain and the wild-type, CaORF19.4965 homozygous knock-out strain is worse than the heterozygous knock-out strain and the wild-type and CaORF19.6268 homozygous knock-out strain is evidently worse than the heterozygous knock-out strain and the wild-type.