白色念珠菌(Candida albicans)是一種常見的人類病原菌。生物膜的形成被認為是微生物在自然界生存的一種本能和天性，現今的研究也認為，對於致病菌而言，形成生物膜將有助於其感染能力。在白色念珠菌 感染的案例中，生物膜的形成同樣扮演了重要的角色，這樣的情況在院內的感染尤其顯著，因為常用的置入性醫療器材往往成為白色念珠菌貼附形成生物膜的表面，有生物膜貼附的醫療器材不僅僅形成一個感染源，同時因為形成生物膜的白色念珠菌對常用的抗真菌感染藥物產生抗
藥性，更造成在治療上的瓶頸。Mss11 是酵母菌生理上一個重要的轉錄因子，負責調控Flo蛋白質家族的表現，這個蛋白質家族負責細胞和基質以及細胞和細胞間的交互作用，同時也影響了細胞形態的改變和侵入性的細胞移動，最重要的是它們也參與了生物膜的形成。由於白色念珠菌和麵包酵母在演化上相 似，利用序列比對的方式，本研究將白色念珠菌基因庫中的orf19.6309鑑定為CaMSS11. 為了解此基因在白色念珠菌生理上的功能，使用 SAT1flipper cassette將這個基因給刪除，並比較突變株和野生株的差異。結果發現CaMSS11在生理上和ScMSS11有相似之處，其中以生物膜的形成能力最為顯著。另外，突變株貼附於基質的能力並沒有減弱，並且突變株對於細胞壁干擾藥物的抵抗力相對於野生株有明顯改變。綜合我所得到的實驗結果， CaMSS11應該是ScMSS11在白色念珠菌的同源基因，也確
定了它在白色念珠菌生物膜成熟期扮演了重要的角色。

Candida albicans is the most important fungal pathogen of humans. Biofilm is a common phenomenon of natural microbe communities, in which cells attach and grow on a biotic or an anbiotic surface, and has been proved to be highly related to virulence of pathogens. In C. albicans, biofilm formation shows a significant consequence for human health and contributes to implanted medical device-associated infections and drug resistance. The formation of C. albicans biofilm is a complex process that involves different cell phenotypes including cell adhesion, cell-cell interaction, yeast-hyphae morphogenesis and extracellular matrix secretion. Mss11p is a transcription factor and controls flocculation and biofilm formation of the model yeast, Saccharomyces cerevisiae. According to the high similarity between C. albicans and S. cerevisiae, In an effort to search the homolog of ScMSS11 in Candida genome database, orf19.6309 is identified as CaMSS11. To examine the functions of C. albicans MSS11 gene, I have knocked-out the genes by the SAT1 flipper method. In the comparisons between wild type and CaMSS11 mutant strains, the mutant strains cause some phenotypic defects that are similar to that of ScMSS11 deletion strains, including cell morphogenesis, invasion, biofilm formation. The difference in biofilm formation ability is the most significant phenotype. The ability of adhering to surface showed no difference between different strains, while the susceptibility of CaMSS11 deletion strains to cell wall integrity interference reagents is different from wild type (SC5314). Moreover, cell to cell interaction is weaker in the mutant strains and the deletion of CaMSS11 gene does not affect virulence in a mouse model of systemic infection. Together, the results suggest that CaMSS11 is the homolog of ScMSS11 and it plays an important role in Candida biofilm maturation

1. Calderone, R.A., Candida and candidiasis. 2002: ASM press.
2. Odds, F.C., Candida and candidosis. 1979.
3. Kauffman, C.A., Fungal infections. Proc Am Thorac Soc, 2006. 3(1):
p. 35-40.
4. Hung, C.C., et al., Clinical spectrum, morbidity, and mortality of
acquired immunodeficiency syndrome in Taiwan: a 5-year prospective
study. J Acquir Immune Defic Syndr, 2000. 24(4): p. 378-85.
5. Wisplinghoff, H., et al., Nosocomial bloodstream infections in US
hospitals: analysis of 24,179 cases from a prospective nationwide
surveillance study. Clin Infect Dis, 2004. 39(3): p. 309-17.
6. Gudlaugsson, O., et al., Attributable mortality of nosocomial
candidemia, revisited. Clin Infect Dis, 2003. 37(9): p. 1172-7.
7. Wenzel, R.P., Nosocomial candidemia: risk factors and attributable
mortality. Clin Infect Dis, 1995. 20(6): p. 1531-4.
8. Kojic, E.M. and R.O. Darouiche, Candida infections of medical
devices. Clin Microbiol Rev, 2004. 17(2): p. 255-67.
9. Kumamoto, C.A. and M.D. Vinces, Alternative Candida albicans
lifestyles: growth on surfaces. Annu Rev Microbiol, 2005. 59: p. 113-
33.
10. Douglas, L.J., Medical importance of biofilms in Candida infections.
Rev Iberoam Micol, 2002. 19(3): p. 139-43.
11. Ramage, G., et al., Candida biofilms: an update. Eukaryot Cell, 2005.
4(4): p. 633-8.
12. Douglas, L.J., Candida biofilms and their role in infection. Trends
Microbiol, 2003. 11(1): p. 30-6.
13. Nett, J. and D. Andes, Candida albicans biofilm development,
modeling a host-pathogen interaction. Curr Opin Microbiol, 2006.
9(4): p. 340-5.
14. Blankenship, J.R. and A.P. Mitchell, How to build a biofilm: a fungal
perspective. Curr Opin Microbiol, 2006. 9(6): p. 588-94.
15. Lopez-Ribot, J.L., Candida albicans biofilms: more than
filamentation. Curr Biol, 2005. 15(12): p. R453-5.
16. Richard, M.L., et al., Candida albicans biofilm-defective mutants.
Eukaryot Cell, 2005. 4(8): p. 1493-502.
17. Allison, D.G., The biofilm matrix. Biofouling, 2003. 19(2): p. 139-50.
18. Branda, S.S., et al., Biofilms: the matrix revisited. Trends Microbiol,
2005. 13(1): p. 20-6.
19. Keller, L. and M.G. Surette, Communication in bacteria: an
ecological and evolutionary perspective. Nat Rev Microbiol, 2006.
4(4): p. 249-58.
20. Eberhard, A., Inhibition and activation of bacterial luciferase
synthesis. J Bacteriol, 1972. 109(3): p. 1101-5.
21. Gonzalez, J.E. and N.D. Keshavan, Messing with bacterial quorum
sensing. Microbiol Mol Biol Rev, 2006. 70(4): p. 859-75.
22. Hornby, J.M., et al., Quorum sensing in the dimorphic fungus
Candida albicans is mediated by farnesol. Appl Environ Microbiol,
2001. 67(7): p. 2982-92.
23. Oh, K.B., et al., Purification and characterization of an
autoregulatory substance capable of regulating the morphological
transition in Candida albicans. Proc Natl Acad Sci U S A, 2001.
98(8): p. 4664-8.
24. Chen, H., et al., Tyrosol is a quorum-sensing molecule in Candida
albicans. Proc Natl Acad Sci U S A, 2004. 101(14): p. 5048-52.
25. Ramage, G., et al., Inhibition of Candida albicans biofilm formation
by farnesol, a quorum-sensing molecule. Appl Environ Microbiol,
2002. 68(11): p. 5459-63.
26. Kruppa, M., et al., The two-component signal transduction protein
Chk1p regulates quorum sensing in Candida albicans. Eukaryot Cell,
2004. 3(4): p. 1062-5.
27. Cao, Y.Y., et al., cDNA microarray analysis of differential gene
expression in Candida albicans biofilm exposed to farnesol.
Antimicrob Agents Chemother, 2005. 49(2): p. 584-9.
28. Nobile, C.J. and A.P. Mitchell, Genetics and genomics of Candida
albicans biofilm formation. Cell Microbiol, 2006. 8(9): p. 1382-91.
29. Verstrepen, K.J. and F.M. Klis, Flocculation, adhesion and biofilm
formation in yeasts. Mol Microbiol, 2006. 60(1): p. 5-15.
30. Gagiano, M., et al., Mss11p is a transcription factor regulating
pseudohyphal differentiation, invasive growth and starch metabolism
in Saccharomyces cerevisiae in response to nutrient availability. Mol
Microbiol, 2003. 47(1): p. 119-34.
31. Webber, A.L., M.G. Lambrechts, and I.S. Pretorius, MSS11, a novel
yeast gene involved in the regulation of starch metabolism. Curr
Genet, 1997. 32(4): p. 260-6.
32. Halme, A., et al., Genetic and epigenetic regulation of the FLO gene
family generates cell-surface variation in yeast. Cell, 2004. 116(3): p.
405-15.
33. Bester, M.C., I.S. Pretorius, and F.F. Bauer, The regulation of
Saccharomyces cerevisiae FLO gene expression and Ca2+ -
dependent flocculation by Flo8p and Mss11p. Curr Genet, 2006. 49(6):
p. 375-83.
34. Mortensen, H.D., et al., The Flo11p-deficient Saccharomyces
cerevisiae strain background S288c can adhere to plastic surfaces.
Colloids Surf B Biointerfaces, 2007. 60(1): p. 131-4.
35. Gagiano, M., et al., Msn1p/Mss10p, Mss11p and Muc1p/Flo11p are
part of a signal transduction pathway downstream of Mep2p
regulating invasive growth and pseudohyphal differentiation in
Saccharomyces cerevisiae. Mol Microbiol, 1999. 31(1): p. 103-16.
36. van Dyk, D., I.S. Pretorius, and F.F. Bauer, Mss11p is a central
element of the regulatory network that controls FLO11 expression and
invasive growth in Saccharomyces cerevisiae. Genetics, 2005. 169(1):
p. 91-106.
37. Cao, F., et al., The Flo8 transcription factor is essential for hyphal
development and virulence in Candida albicans. Mol Biol Cell, 2006.
17(1): p. 295-307.
38. Liu, H., J. Kohler, and G.R. Fink, Suppression of hyphal formation in
Candida albicans by mutation of a STE12 homolog. Science, 1994.
266(5191): p. 1723-6.
39. Gimeno, C.J., et al., Unipolar cell divisions in the yeast S. cerevisiae
lead to filamentous growth: regulation by starvation and RAS. Cell,
1992. 68(6): p. 1077-90.
40. Lee, K.L., H.R. Buckley, and C.C. Campbell, An amino acid liquid
synthetic medium for the development of mycelial and yeast forms of
Candida Albicans. Sabouraudia, 1975. 13(2): p. 148-53.
41. Bedell, G.W. and D.R. Soll, Effects of low concentrations of zinc on
the growth and dimorphism of Candida albicans: evidence for zinc-
resistant and -sensitive pathways for mycelium formation. Infect
Immun, 1979. 26(1): p. 348-54.
42. Reuss, O., et al., The SAT1 flipper, an optimized tool for gene
disruption in Candida albicans. Gene, 2004. 341: p. 119-27.
43. Ramage, G., et al., Characteristics of biofilm formation by Candida
albicans. Rev Iberoam Micol, 2001. 18(4): p. 163-70.
44. Kuhn, D.M., et al., Uses and limitations of the XTT assay in studies of
Candida growth and metabolism. J Clin Microbiol, 2003. 41(1): p.
506-8.
45. Antachopoulos, C., et al., Rapid susceptibility testing of medically
important zygomycetes by XTT assay. J Clin Microbiol, 2006. 44(2): p.
553-60.
46. Meletiadis, J., et al., Colorimetric assay for antifungal susceptibility
testing of Aspergillus species. J Clin Microbiol, 2001. 39(9): p. 3402-8.
47. Thein, Z.M., Y.H. Samaranayake, and L.P. Samaranayake, In vitro
biofilm formation of Candida albicans and non-albicans Candida
species under dynamic and anaerobic conditions. Arch Oral Biol,
2007. 52(8): p. 761-7.
48. He, M., et al., In vitro activity of eugenol against Candida albicans
biofilms. Mycopathologia, 2007. 163(3): p. 137-43.
49. Zupan, J. and P. Raspor, Quantitative agar-invasion assay. J
Microbiol Methods, 2008. 73(2): p. 100-4.
50. Chen, J., et al., Crk1, a novel Cdc2-related protein kinase, is required
for hyphal development and virulence in Candida albicans. Mol Cell
Biol, 2000. 20(23): p. 8696-708.
51. Emes, R.D. and C.P. Ponting, A new sequence motif linking
lissencephaly, Treacher Collins and oral-facial-digital type 1
syndromes, microtubule dynamics and cell migration. Hum Mol
Genet, 2001. 10(24): p. 2813-20.
52. Biswas, S., P. Van Dijck, and A. Datta, Environmental sensing and
signal transduction pathways regulating morphopathogenic
determinants of Candida albicans. Microbiol Mol Biol Rev, 2007.
71(2): p. 348-76.
53. Chaffin, W.L., et al., Cell wall and secreted proteins of Candida
albicans: identification, function, and expression. Microbiol Mol Biol
Rev, 1998. 62(1): p. 130-80.
54. Dean, N., Yeast glycosylation mutants are sensitive to
aminoglycosides. Proc Natl Acad Sci U S A, 1995. 92(5): p. 1287-91.
55. Sutherland, I.W., The biofilm matrix--an immobilized but dynamic
microbial environment. Trends Microbiol, 2001. 9(5): p. 222-7.
56. Wingender J, J.K.-E., Flemming H-C, Interaction between
extracellular polysaccharides and enzymes., in Microbial
Extracellular Polymeric Substances., N.T.R. Wingender J, Flemming
H-C, Editor. 1999, Springer: Berlin. p. 231-251.
57. Zogaj, X., et al., The multicellular morphotypes of Salmonella
typhimurium and Escherichia coli produce cellulose as the second
component of the extracellular matrix. Mol Microbiol, 2001. 39(6): p.
1452-63.
58. Maira-Litran, T., et al., Immunochemical properties of the
staphylococcal poly-N-acetylglucosamine surface polysaccharide.
Infect Immun, 2002. 70(8): p. 4433-40.
59. Eisman, B., et al., The Cek1 and Hog1 mitogen-activated protein
kinases play complementary roles in cell wall biogenesis and
chlamydospore formation in the fungal pathogen Candida albicans.
Eukaryot Cell, 2006. 5(2): p. 347-58.
60. Cullen, P.J., et al., Defects in protein glycosylation cause SHO1-
dependent activation of a STE12 signaling pathway in yeast. Genetics,
2000. 155(3): p. 1005-18.
61. Lee, B.N. and E.A. Elion, The MAPKKK Ste11 regulates vegetative
growth through a kinase cascade of shared signaling components.
Proc Natl Acad Sci U S A, 1999. 96(22): p. 12679-84.