Identification of genes associated with germination of conidia to form mycelia in the fungus Paracoccidioides brasiliensis
Keywords:
Paracoccidioides, spores, fungal, mycelium, germination
Abstract
Introduction. Paracoccidioides brasiliensis is a thermo-dimorphic fungus. At room temperature it grows as a mold that produces conidia, whereas in the vertebrate host it grows as a multiple-budding yeast. The molecular mechanisms involved in the germination from the conidia to the mycelia process remain unknown.Objective. The kinetics of conidia to mycelia germination process were studied in the dimorphic fungus P. brasiliensis. Gene expression during this process was evaluated by construction and analysis of an EST library.
Materials and methods. For the germination kinetics study, P. brasiliensis conidia were isolated as single cell units. Then, they were cultured at 18° C in BHI (brain-heart infusion) broth for 24, 48, 72 and 96 hr. After each perion, they were examined by light microscopy. From conidia harvested at 96 hr, an EST library was constructed; at this stage the gene expression was presumed to be maximal for the germination process.
Results. During the conidia to the mycelia developmental process, the following germination rates were observed: at 24 hr, 11.7±1.2%; at 48 hr, 30±0.6%; at 72 hr, 43±1.3%; and at 96 hr, 66±2.4%. At the 96 hour stage, an EST library was constructed. It consisted of 129 sequences grouped in 4 contigs and 7 singlets for a total of 11 possible genes. Eight of the sequences had not been described previously in other EST libraries of this fungus.
Conclusions. New genes were identified that were expressed during the conidia to the mycelia germination process and may represent genes specific to the germination process.
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References
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28. Bailao AM, Shrank A, Borges CL, Parente JA, Dutra V, Felipe MS, et al. The transcriptional profile of Paracoccidioides brasiliensis yeast cells is influenced by human plasma. FEMS Immunol Med Microbiol. 2007;51:43-57.
29. Ramírez JR. Paracoccidioides brasiliensis: conversion of yeastlike forms into mycelia in submerged culture. J Bacteriol. 1971;105:523-6.
30. Fedorova ND, Khaldi N, Joardar VS, Maiti R, Amedeo P, Anderson MJ, et al. Genomic islands in the pathogenic filamentous fungus Aspergillus fumigatus. PLoS Genet. 2008;4:e1000046.
31. Hand RA, Jia N, Bard M, Craven RJ. Saccharomyces cerevisiae Dap1p, a novel DNA damage response protein related to the mammalian membrane-associated progesterone receptor. Eukaryot Cell. 2003;2:306-17.
32. Gant TM, Wilson KL. Nuclear assembly. Annu Rev Cell Dev Biol. 1997;13:669-95.
33. Tranque P, Hu MC, Edelman GM, Mauro VP. rRNA complementarity within mRNAs: a possible basis for mRNA-ribosome interactions and translational control. Proc Natl Acad Sci USA. 1998;95:12238-43.
2. Restrepo A, Tobon AM. Chapter 266. Paracoccidioides brasiliensis. In: Mandell GL, Bennetts JE, Dollin, R, editors. Principles and practice of infectious diseases. 6th edition. Philadelphia, P.A: Elsevier; 2005. p. 3062-8.
3. Camilo J, Tabares AM, Gómez BL, Aristizábal BE, Cock AM, Restrepo A. The oral route in the pathogenesis of paracoccidioidomycosis: an experimental study in BALB/c mice infected with P. brasiliensis conidia. Mycopathologia. 2001;151:57-62.
4. McEwen JG, Brummer E, Stevens DA, Restrepo A. Effect of murine polymorphonuclear leukocytes on the yeast form of Paracoccidioides brasiliensis. Am J Trop Med Hyg. 1987;36:603-8.
5. Goldman GH, dos Reis E, Duarte DC, de Souza LA, Quiapin AC, Vitorelli PM, et al. Expressed sequence tag analysis of the human pathogen Paracoccidioides brasiliensis yeast phase: identification of putative homologues of Candida albicans virulence and pathogenicity genes. Eukaryot Cell. 2003;2:34-48.
6. Nunes LR, Costa de Oliveira R, Leite DB, da Silva VS, dos Reis E, da Silva ME, et al. Transcriptome analysis of Paracoccidioides brasiliensis cells undergoing mycelium-to-yeast transition. Eukaryot Cell. 2005;4:2115-28.
7. Queiroz-Telles F. Paracoccidioides brasiliensis: Ultra-structural findings. In: Franco LC, Restrepo A, Del Negro G, editors. Paracoccidioidomycosis. Boca Raton, FL: CRC Press; 1994. p. 27-48.
8. Salazar ME, Restrepo A. Morphogenesis of the mycelium-to-yeast transformation in Paracoccidioides brasiliensis. Sabouraudia. 1985;23:7-11.
9. Aristizábal BH, Clemons KV, Stevens DA, Restrepo A. Morphological transition of Paracoccidioides brasiliensis conidia to yeast cells: in vivo inhibition in females. Infect Immun. 1998;66:5587-91.
10. Cock AM, Cano LE, Vélez D, Aristizábal BH, Trujillo J, Restrepo A. Fibrotic sequelae in pulmonary paracocci-dioidomycosis: histopathological aspects in BALB/c mice infected with viable and non-viable Paracoccidioides brasiliensis propagules. Rev Inst Med Trop Sao Paulo. 2000;42:59-66.
11. McEwen JG, Bedoya V, Patino MM, Salazar ME, Restrepo A. Experimental murine paracoccidiodomycosis induced by the inhalation of conidia. J Med Vet Mycol. 1987;25:165-75.
12. Cano LE, Brummer E, Stevens DA, Restrepo A. An evaluation of the enzyme-linked immunoabsorbent assay (ELISA) for quantitation of antibodies to Paracoccidioides brasiliensis. J Med Vet Mycol. 1986;24:467-75.
13. Cano LE, Gómez B, Brummer E, Restrepo A, Stevens DA. Inhibitory effect of deferoxamine or macrophage activation on transformation of Paracoccidioides brasiliensis conidia ingested by macrophages: reversal by holotransferrin. Infect Immun. 1994;62:1494-6.
14. González A, Aristizábal BH, Gómez EC, Restrepo A, Cano LE. Short report: Inhibition by tumor necrosis factor-alpha-activated macrophages of the transition of Paracoccidioides brasiliensis conidia to yeast cells through a mechanism independent of nitric oxide. Am J Trop Med Hyg. 2004;71:828-30.
15. González A, de Gregori W, Vélez D, Restrepo A, Cano LE. Nitric oxide participation in the fungicidal mechanism of gamma interferon-activated murine macrophages against Paracoccidioides brasiliensis conidia. Infect Immun. 2000;68:2546-52.
16. Restrepo BI, McEwen JG, Salazar ME, Restrepo A. Morphological development of the conidia produced by Paracoccidioides brasiliensis mycelial form. J Med Vet Mycol. 1986;24:337-9.
17. Felipe MS, Andrade RV, Petrofeza SS, Maranhao AQ, Torres FA, Albuquerque P, et al. Transcriptome characterization of the dimorphic and pathogenic fungus Paracoccidioides brasiliensis by EST analysis. Yeast. 2003;20:263-71.
18. Felipe MS, Torres FA, Maranhao AQ, Silva-Pereira I, Pocas-Fonseca MJ, Campos EG, et al. Functional genome of the human pathogenic fungus Paracoccidioides brasiliensis. FEMS Immunol Med Microbiol. 2005;45:369-81.
19. Bastos KP, Bailao AM, Borges CL, Faria FP, Felipe MS, Silva MG, et al. The transcriptome analysis of early morphogenesis in Paracoccidioides brasiliensis mycelium reveals novel and induced genes potentially associated to the dimorphic process. BMC Microbiol. 007;7:29.
20. Restrepo A, Salazar ME, Cano LE, Patiño MM. A technique to collect and dislodge conidia produced by Paracoccidioides brasiliensis mycelial form. J Med Vet Mycol.1986;24:247-50.
21. Restrepo A, Jiménez BE. Growth of Paracoccidioides brasiliensis yeast phase in a chemically defined culture medium. J Clin Microbiol.1980;12:279-81.
22. Calich VL, Purchio A, Paula CR. A new fluorescent viability test for fungi cells. Mycopathologia.1979;66:175-7.
23. Marra MA, Kucaba TA, Hillier LW, Waterston RH. High-throughput plasmid DNA purification for 3 cents per sample. Nucleic Acids Res.1999;27:e37.
24. Ewing B, Green P. Base-calling of automated sequencer traces using phred. II. Error probabilities. Genome Res.1998;8:186-94.
25. Huang X, Madan A. CAP3: A DNA sequence assembly program. Genome Res.1999;9:868-77.
26. White O, Kerlavage AR. TDB: new databases for biological discovery. Methods Enzymol.1996;266:27-40.
27. Tavares AH, Silva SS, Dantas A, Campos EG, Andrade RV, Maranhao AQ, et al. Early transcriptional response of Paracoccidioides brasiliensis upon internalization by murine macrophages. Microbes Infect.2007;9:583-90.
28. Bailao AM, Shrank A, Borges CL, Parente JA, Dutra V, Felipe MS, et al. The transcriptional profile of Paracoccidioides brasiliensis yeast cells is influenced by human plasma. FEMS Immunol Med Microbiol. 2007;51:43-57.
29. Ramírez JR. Paracoccidioides brasiliensis: conversion of yeastlike forms into mycelia in submerged culture. J Bacteriol. 1971;105:523-6.
30. Fedorova ND, Khaldi N, Joardar VS, Maiti R, Amedeo P, Anderson MJ, et al. Genomic islands in the pathogenic filamentous fungus Aspergillus fumigatus. PLoS Genet. 2008;4:e1000046.
31. Hand RA, Jia N, Bard M, Craven RJ. Saccharomyces cerevisiae Dap1p, a novel DNA damage response protein related to the mammalian membrane-associated progesterone receptor. Eukaryot Cell. 2003;2:306-17.
32. Gant TM, Wilson KL. Nuclear assembly. Annu Rev Cell Dev Biol. 1997;13:669-95.
33. Tranque P, Hu MC, Edelman GM, Mauro VP. rRNA complementarity within mRNAs: a possible basis for mRNA-ribosome interactions and translational control. Proc Natl Acad Sci USA. 1998;95:12238-43.
How to Cite
1.
García AM, Hernández O, Aristizábal BH, Cano LE, McEwen JG, Restrepo Ángela. Identification of genes associated with germination of conidia to form mycelia in the fungus Paracoccidioides brasiliensis. biomedica [Internet]. 2009 Sep. 1 [cited 2024 May 17];29(3):403-12. Available from: https://revistabiomedica.org/index.php/biomedica/article/view/12
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