Paracoccidioides species complex : ecology, phylogeny, sexual reproduction, and virulence

Paracoccidioidomycosis (PCM) is a deep systemic mycosis caused by human fungal pathogens of the Paracoccidioides genus. The disease is geographically restricted to subtropical areas of Latin America (from south of Mexico to north of Argentina) with a high prevalence in Brazil, Colombia, Venezuela, and Argentina [1]. The annual incidence rate in Brazil is 10–30 infections per million inhabitants, and the mean mortality rate is 1.4 per million inhabitants per year, making this disease the highest cause of mortality among systemic mycoses [2]. PCM is endemic in rural populations and mainly affects individuals engaged in agricultural activities, who inhale aerosols containing fungal material during manipulation of the soil. Molecular evolutionary studies place the genus Paracoccidioides in the thermodimorphic fungal pathogen clade related to the family Ajellomycetaceae (Ascomycetes), which includes the Blastomyces, Histoplasma, and Emmonsia genera, and with which it shares a common ancestor, Lacazia loboi. PCM can be caused by two species Paracoccidioides brasiliensis and P. lutzii [3]. P. brasiliensis has been considered a single species since its discovery, although several studies including molecular and morphological data support the split of P. brasiliensis into two species [3,4]. P. lutzii is composed of a single monophyletic and recombining population so far found in central, southwest, and north Brazil and Ecuador [3–5]. On the other hand, P. brasiliensis contains a complex of at least four different cryptic species (S1, PS2, PS3 and PS4; Figure 1A [6]). P. brasiliensis S1 represents a monophyletic and recombining population widely distributed in South America and has been associated with the majority of cases of PCM detected up until the present time. Strains belonging to P. brasiliensis S1 have previously been recovered from armadillos, soil, and penguin feces [6]. P. brasiliensis PS2 is a paraphyletic and recombining population identified so far only in Brazil and Venezuela [6]. P. brasiliensis PS3 is comprised of a monophyletic and clonal population that has been recovered in humans and armadillos in endemic regions of Colombia [6]. P. brasiliensis PS4 was recently identified and is composed of a monophyletic population of clinical isolates from Venezuela [5,7]. Besides the typical bicorn cocked hat– and barrel-shaped conidia produced by both species, P. lutzii frequently produces elongated rod-shaped conidia, a characteristic feature that may be used for species identification [3]. Because of the difficulties of conidia production in the laboratory and slight morphological differences among species, molecular identification of Paracoccidioides species has become the most common tool of choice. Several molecular markers have already been applied in population studies of the Pararacoccidioides genus, and for multilocus sequencing typing, gp43, arf, b-tub, and hsp70 loci are the best choices for species delineation [4,6]

Ecological study of Paracoccidioides brasiliensis in soil: growth ability, conidia production and molecular detection

<p>Abstract</p> <p>Background</p> <p><it>Paracoccidioides brasiliensis </it>ecology is not completely understood, although several pieces of evidence point to the soil as its most probable habitat. The present study aimed to investigate the fungal growth, conidia production and molecular pathogen detection in different soil conditions.</p> <p>Methods</p> <p>Soils samples of clayey, sandy and medium textures were collected from ground surface and the interior of armadillo burrows in a hyperendemic area of Paracoccidioidomycosis. <it>P</it>. <it>brasiliensis </it>was inoculated in soil with controlled humidity and in culture medium containing soil extracts. The molecular detection was carried out by Nested PCR, using panfungal and species specific primers from the ITS-5.8S rDNA region.</p> <p>Results</p> <p>The soil texture does not affect fungus development and the growth is more abundant on/in soil saturated with water. Some soil samples inhibited the development of <it>P. brasiliensis</it>, especially those that contain high values of Exchangeable Aluminum (H+Al) in their composition. Some isolates produced a large number of conidia, mainly in soil-extract agar medium. The molecular detection was positive only in samples collected from armadillo burrows, both in sandy and clayey soil.</p> <p>Conclusion</p> <p><it>P. brasiliensis </it>may grow and produce the infectious conidia in sandy and clayey soil, containing high water content, mainly in wild animal burrows, but without high values of H+Al.</p

Genus Paracoccidioides: Species Recognition and Biogeographic Aspects

Background: Paracoccidioidomycosis is a systemic mycosis caused by Paracoccidioides brasiliensis (species S1, PS2, PS3), and Paracoccidioides lutzii. This work aimed to differentiate species within the genus Paracoccidioides, without applying multilocus sequencing, as well as to obtain knowledge of the possible speciation processes. Methodology/Principal Findings: Single nucleotide polymorphism analysis on GP43, ARF and PRP8 intein genes successfully distinguished isolates into four different species. Morphological evaluation indicated that elongated conidia were observed exclusively in P. lutzii isolates, while all other species (S1, PS2 and PS3) were indistinguishable. To evaluate the biogeographic events that led to the current geographic distribution of Paracoccidioides species and their sister species, Nested Clade and Likelihood Analysis of Geographic Range Evolution (LAGRANGE) analyses were applied. The radiation of Paracoccidioides started in northwest South America, around 11–32 million years ago, as calculated on the basis of ARF substitution rate, in the BEAST program. Vicariance was responsible for the divergence among S1, PS2 and P. lutzii and a recent dispersal generated the PS3 species, restricted to Colombia. Taking into account the ancestral areas revealed by the LAGRANGE analysis and the major geographic distribution of L. loboi in the Amazon basin, a region strongly affected by the Andes uplift and marine incursions in the Cenozoic era, we also speculate about the effect of these geological events on the vicariance between Paracoccidioides and L. loboi. Conclusions/Significance: The use of at least 3 SNPs, but not morphological criteria, as markers allows us to distinguish among the four cryptic species of the genus Paracoccidioides. The work also presents a biogeographic study speculating on how these species might have diverged in South America, thus contributing to elucidating evolutionary aspects of the genus Paracoccidioides

Caracterização da transição micélio-levedura em Paracoccidioides brasiliensis e sua relação com a expressão do gene do choque térmico 70(HSP70)

O fungo termo dimórfico, Paracoccidioides brasiliensis é o agente etiológico da Paracoccidioidomicose (PCM), a micose sistêmica mais prevalente da América Latina. Este fungo vem sendo frequentemente isolado de amostras clínicas, tecidos de tatu (Dasypus novemcinctus) e recentemente foi também isolado de cão. Este trabalho avaliou a transição de micélio para levedura (M-L), a termo tolerância e o perfil de virulência em nove isolados de P. brasiliensis (quatro de pacientes humanos, quatro de tatus e um de cão), bem como a sua relação com a seqüência parcial e expressão do gene hsp70 (Heat Shock Protein 70) através de Real Time RT-PCR. Tanto os dados morfológicos como moleculares se mostraram variáveis dentre os diferentes isolados. Alguns destes dados, como sequenciamento e morfologia leveduriforme corroboram com a divisão de nossos isolados nas duas espécies crípticas simpátricas previamente propostas por Matute et al (2006). Nossos resultados confirmam que a HSP70 pode ser um importante fator de virulência por estar associado à termo tolerância, mas sua expressão parece não ser diretamente associada a altos padrões de virulência.The thermo dimorphic fungus Paracoccidioides brasiliensis is the etiological agent of Paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in Latin America. The previous phylogenetic species recognition proved the existence of, at least, three cryptic species in this pathogen. In this work we evaluated the mycelia to yeast (M-Y) transition, thermo tolerance and virulence profiles of nine isolates of P. brasiliensis, (including members of two of the three species) as well as its relation to the partial sequence and expression of hsp70 gene. It was observed a large phenotypic variability concerning the M-Y transition. The isolates Bt84 and T10 took more time to convert to the yeast form. These same isolates presented stretched yeast cells at 36°C, instead of the typical round cells. It was also observed arthroconidia production during the M-Y transition for some of the nine isolates studied. The hsp70 expression showed to be variable among our isolates. The partial sequencing of hsp70 gene resulted in a Neighbour Joining tree that divided our isolates in two main groups. Our data confirm that hsp70 gene might be an important virulence factor, associated with the thermo tolerance, but its expression does not seem to be directly related to high virulence profiles. We also presented some preliminary results about mycological characters that could be important candidates for morphologic markers for species recognition, as well as the partial sequencing of one member of the hsp70 gene family that allowed the separation of our isolates in two clusters, that correspond to the two sympatric cryptic species that occur in our PCM hyper endemic area (Botucatu, SP, Brazil).Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Inteins in pathogenic fungi: a phylogenetic tool and perspectives for therapeutic applications

Inteins or internal proteins are coding sequences that are transcribed and translated with flanking sequences (exteins). After translation, the inteins are excised by an autocatalytic process and the host protein assumes its normal conformation and develops its expected function. These parasitic genetic elements have been found in important, conserved proteins in all three domains of life. Most of the eukaryotic inteins are present in the fungi kingdom and the PRP8 intein is one of the most widespread inteins, occurring in important pathogens such as Cryptococcus neoformans (varieties grubii and neoformans), Cryptococcus gattii, Histoplasma capsulatum and Paracoccidioides brasiliensis. The knowledge of conserved and non-conserved domains in inteins have opened up new opportunities for the study of population variability in pathogenic fungi, including their phylogenetic relationships and recognition or diagnoses of species. Furthermore, inteins in pathogenic fungi should also be considered a promising therapeutic drug target, since once the autocatalytic splicing is inhibited, the host protein, which is typically vital, will not be able to perform its normal function and the fungal cell will not survive or reproduce

Inteins in pathogenic fungi: a phylogenetic tool and perspectives for therapeutic applications

Inteins or "internal proteins" are coding sequences that are transcribed and translated with flanking sequences (exteins). After translation, the inteins are excised by an autocatalytic process and the host protein assumes its normal conformation and develops its expected function. These parasitic genetic elements have been found in important, conserved proteins in all three domains of life. Most of the eukaryotic inteins are present in the fungi kingdom and the PRP8 intein is one of the most widespread inteins, occurring in important pathogens such as Cryptococcus neoformans (varieties grubii and neoformans), Cryptococcus gattii , Histoplasma capsulatum and Paracoccidioides brasiliensis . The knowledge of conserved and non-conserved domains in inteins have opened up new opportunities for the study of population variability in pathogenic fungi, including their phylogenetic relationships and recognition or diagnoses of species. Furthermore, inteins in pathogenic fungi should also be considered a promising therapeutic drug target, since once the autocatalytic splicing is inhibited, the host protein, which is typically vital, will not be able to perform its normal function and the fungal cell will not survive or reproduce

Paracoccidioides brasiliensis AND Paracoccidioides lutzii, A SECRET LOVE AFFAIR

Para comemorar o centenário de aniversário do Prof. Dr. Carlos da Silva Lacaz, os autores fazem um breve relato dos estudos sobre a biologia, ecologia e filogenia molecular que culminaram na revelação da espécie Paracoccidioides lutzii, que havia permanecido escondida por mais de um século ao lado de Paracoccidioides brasiliensis. O professor Lacaz exerceu papel central no desenvolvimento desta área do conhecimento, pois manteve interesse permanente nas pesquisas deste fungo e da paracoccidioidomicose, visando principalmente proporcionar benefícios aos pacientes acometidos por esta micose.To commemorate Prof. Carlos da Silva Lacaz's centennial anniversary, the authors have written a brief account of a few, out of hundreds, biological, ecological, molecular and phylogenetic studies that led to the arrival of Paracoccidioides lutzii, hidden for more than a century within Paracoccidioides brasiliensis. Lacaz's permanent interest in this fungus, and particularly his conviction on the benefits that research on paracoccidioidomycosis would bring to patients, were pivotal in the development of the field