XI International Meeting on Paracoccidioidomycosis

XI International Meeting on Paracoccidioidomycosis

Resume:136-1

Poster (Painel)

136-1	CHROMOSOMAL POLYMORPHISM, GENE MAPPING, AND SYNTENIC RELATIONSHIPS IN THE PATHOGENIC FUNGI Sporothrix schenckii AND Sporothrix brasiliensis.
Authors:	Alexandre Augusto Sasaki (UNIFESP - Universidade Federal de São Paulo) ; Geisa Ferreira Fernandes (UNIFESP - Universidade Federal de São Paulo) ; Luciano dos Santos Feitosa (UNICASTELO - Universidade Camilo Castelo Branco) ; Zoilo Pires Camargo (UNIFESP - Universidade Federal de São Paulo)

Abstract

Sporotrichosis is a chronic mycosis caused by the dimorphic fungus Sporothrix schenckii. Recent studies have shown that this specie is a complex composed by cryptic species and among them Sporothrix brasiliensis, new proposed specie. In Brazil, a continental country, S. schenckii and S. brasiliensis are the most common species. Six clinical and one environmental S. schenckii isolates and three clinical S. brasiliensis isolates, originated from different Brazilian geographic areas, were cultured on BHI broth medium at 37•C for 10 days under agitation. Yeast cells (2x10℘8 cells/ml) were washed 3 times with wash buffer and mixed with low melting point agarose (1%), and then, the suspension was allowed to solidify inside casting mold. Inserts were unmolded and zymolyase solution was added. After overnight incubation at 37•C, zymolyase solution was replaced with cell lyses solution and incubated overnight at 50•C. The solution was replaced with stock solution and stored at 4•C until use. PFGE was performed in a Gene Navigator apparatus, with homogeneous and interpolated pulses for 7 days. Then, the gel was stained with ethidium bromide and photographed. Chromosomal molecules were transferred to a nylon membrane and hybridized with 9 probes (β-tubulin, calmodulin, catalase, chitin synthase I, Internal Transcribed Spacer, Pho85 cyclin-dependent kinase, Protein Kinase C, G Protein α subunit, topoisomerase II), genomic DNA and 8 Mb chromosomal band from S. schenckii isolate. DNA hybridization was used to establish and to compare the electrophoretic karyotypes of isolates from different Brazilian geographic areas. Chromosomal polymorphism was observed among the isolates; 4 isolates presented the same electrophoretic profile (3 S. schenckii and 1 S. brasiliensis), while other isolates presented distinct profiles (4 S. schenckii and 2 S. brasiliensis). Gene mapping allowed the identification of synteny groups and the use of isolated whole chromosomal band to probe chromoblots indicated the existence of repetitive sequences, contributing to a better understanding of the structure and organization of the fungus genome. This study represents the first comparative mapping among different Sporothrix spp. isolates.਀吀栀椀猀眀漀爀欀眀愀猀猀甀瀀瀀漀爀琀攀搀戀礀䌀䄀倀䔀匀Ⰰ 䌀一倀焀愀渀搀䘀䄀倀䔀匀倀⸀

Keyword: Sporothrix schenckii, Sporothrix brasiliensis, Chromosomes, Karyotype, PFGE

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