Identification and characterization of Tc1/mariner-like DNA transposons in genomes of the pathogenic fungi of the Paracoccidioides species complex

BMC Genomics Identification and characterization of Tc1/ mariner-like DNA transposons in genomes of the pathogenic fungi of the Paracoccidioides species complex Marjorie M Marini 0 Tamiris Zanforlin Patrcia C Santos 0 Roberto RM Barros Anne CP Guerra 0 Rosana Puccia Maria SS Felipe Marcelo Brigido Clia MA Soares Jernimo C Ruiz Jos F Silveira Patrcia S Cisalpino 0 0 Departamento de Microbiologia, Instituto de Ciencias Biologicas, Universidade Federal de Minas Gerais , 31270-901 Belo Horizonte, MG , Brazil Background: Paracoccidioides brasiliensis (Eukaryota, Fungi, Ascomycota) is a thermodimorphic fungus, the etiological agent of paracoccidioidomycosis, the most important systemic mycoses in Latin America. Three isolates corresponding to distinct phylogenetic lineages of the Paracoccidioides species complex had their genomes sequenced. In this study the identification and characterization of class II transposable elements in the genomes of these fungi was carried out. Results: A genomic survey for DNA transposons in the sequence assemblies of Paracoccidioides, a genus recently proposed to encompass species P. brasiliensis (harboring phylogenetic lineages S1, PS2, PS3) and P. lutzii (Pb01-like isolates), has been completed. Eight new Tc1/mariner families, referred to as Trem (Transposable element mariner), labeled A through H were identified. Elements from each family have 65-80% sequence similarity with other Tc1/ mariner elements. They are flanked by 2-bp TA target site duplications and different termini. Encoded DDDtransposases, some of which have complete ORFs, indicated that they could be functionally active. The distribution of Trem elements varied between the genomic sequences characterized as belonging to P. brasiliensis (S1 and PS2) and P. lutzii. TremC and H elements would have been present in a hypothetical ancestor common to P. brasiliensis and P. lutzii, while TremA, B and F elements were either acquired by P. brasiliensis or lost by P. lutzii after speciation. Although TremD and TremE share about 70% similarity, they are specific to P. brasiliensis and P. lutzii, respectively. This suggests that these elements could either have been present in a hypothetical common ancestor and have evolved divergently after the split between P. brasiliensis and P. Lutzii, or have been independently acquired by horizontal transfer. Conclusions: New families of Tc1/mariner DNA transposons in the genomic assemblies of the Paracoccidioides species complex are described. Families were distinguished based on significant BLAST identities between transposases and/or TIRs. The expansion of Trem in a putative ancestor common to the species P. brasiliensis and P. lutzii would have given origin to TremC and TremH, while other elements could have been acquired or lost after speciation had occurred. The results may contribute to our understanding of the organization and architecture of genomes in the genus Paracoccidioides. - Background The thermodimorphic fungus Paracoccidioides brasiliensis is the etiological agent of paracoccidioidomycosis, a systemic endemic disease that affects at least 10 million people in Latin America [1]. Infection probably occurs as a result of inhalation of conidia that subsequently transform into yeast forms within the lung, resulting in asymptomatic infection or infection that can progress to acute (sub-acute) and chronic clinical forms of the disease [2]. Paracoccidioidomycosis is the 8th most common cause of death due to chronic/recurrent infections and parasitic diseases in Brazil [3]. The teleomorph (sexual) stage of this fungus is unknown, but analysis of the large rDNA subunit classified it as an Ascomycete, order Onygenales, family Onygenaceae, phylogenetically close to Ajellomyces, a genus that harbors 2 other human dimorphic fungal pathogens, specifically Histoplasma capsulatum and Blastomyces dermatitidis [4]. Molecular analysis by random amplification of polymorphic DNA (RAPD) [5], restriction fragment length polymorphism (RFLP) [6] and electrophoretic karyotyping showed extensive genetic variability among distinct isolates [7,8]. RAPD, a technique popular for its simplicity, has been useful in determining the existence of genetically distinct P. brasiliensis groups, their relationship with geographic distribution, and the similarity among clinical, animal and environmental samples. However, the information has been difficult to compare. Gene polymorphism studies were pioneered by characterization of PbGP43 polymorphism, a gene encoding the fungus immunodominant antigen, in a sample of 17 isolates [9]. By comparing 2 sequenced PCR fragments from the whole gene (exon 1, intron and exon 2) these authors found 21 informative substitution sites, mostly in exon 2, which defined 5-6 genotypes. The maximum-likelihood phylogenetic tree generated with these sequences clearly reflected the presence of specific genetic groups in the species. Subsequently, multilocus genealogy studies demonstrated evolutionary lineages identifying the occurrence of cryptic phylogenetic species that were morphologically indistinguishable by following the criteria of genealogical concordance and non-discordance [10-12] which is used to identify the reduction of gene flow between groups of individuals due to geographic or reproductive isolation barriers. The PbGP43 locus, whose characteristic genotypes have recently been reviewed [13], was the most polymorphic and informative in these studies. Matute et al. [10] analyzed the genetic structure of 65 P. brasiliensis isolates and concluded that they could be grouped into 3 distinct phylogenetic species: S1 (including isolate Pb18), PS2 (including isolate Pb03) and PS3 (composed exclusively of Colombian isolates). In a study of 21 P. brasiliensis isolates, 14 of which had been included in the above study, Carrero et al. [11] came to a similar conclusion for all the isolates, with the exception of isolate Pb01, which they suggested was a new phylogenetic species in the genus Paracoccidioides. Recently, Teixeira et al. [12], analyzing 88 isolates of the fungus, found that 17 of them were genotypically similar, belonging to the Pb01like group. They estimated that the S1/PS2/PS3 species clade and the Pb01-like new species, for which the name Paracoccidioides lutzii was proposed, shared a common ancestor approximately 32 million years ago. These studies suggested that the genus Paracoccidioides consists of at least 4 different, previously unrecognized phylogenetic species: P. brasiliensis S1 (species), PS2 (phylogenetic species 2) and PS3 (phylogenetic species 3), and a 4th quite distinct lineage comprising fungal isolates formerly referred to as Pb01-like [11], now proposed to be a new species, P. lutzii [12]. The multilocus sequence analysis also supported recombination in nature in P. brasiliensis (S1) [10] and P. lutzii [11,12], indicating the presence of sexual reproduction. Thanks to The Broad Institute Fungal Genome Initi (...truncated)