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NAVIGATING PARACOCCIDIOIDES GENOME RESOURCES AT THE BROAD WEBSITE
Author: DESJARDINS, C. A. 1
Institution: 1. Broad Institute; Broad Institute
Abstract:
Genome sequencing provides a wealth of data which can be difficult to manually analyze and explore. The field of genome analysis also lacks many comprehensive graphically driven and intuitive tools for automated analysis of genome data. Here, I will discuss how to acquire, analyze, and interpret genomic data related to Paracoccidioides. The Broad Institute website (www.broadinstitute.org) hosts numerous genomic resources for Paracoccidioides and related dimorphic fungi, such as Histoplasma and Coccidioides, through the Fungal Genome Initiative. The website has data for available for download in a variety of formats, including nucleotide and protein sequences and genetic maps. I will discuss how access and read these different forms of data, with a focus on Paracoccidioides. I will then discuss downstream applications which can be used to process and explore this data, such as command line searching with BLAST and desktop genome visualization and exploration. Examples will be shown throughout. We will explore various analytical tools available on the Broad website, including 1) searching the Paracoccidioides genomes for genes of interest with BLAST and related tools, 2) exploring the genomes with functional queries, in order to identify groups of genes with related functions, and 3) methods to visually compare Paracoccidioides strains, including dot pots and synteny maps. Furthermore, I will discuss many of the analytical tools used in the analyses in the Paracoccidioides genome paper, with a focus on what kinds of questions can be answered using these tools. Some specific examples will include identifying gene families, functional classification of genes, and phylogenetic analysis. Finally, time will be available for discussing bioinformatic solutions to course member questions.
Keywords: genomics, BLAST, gene function, synteny
Lipid mediators of the immune response to fungi
Author: Alexandra Ivo de Medeiros 1
Institution: 1FCFAR-UNESP - Faculdade de Ciências Farmacêuticas (Rodovia Araraquara/Jau Km 1 Araraquara - CEP: 14801-902)
Abstract:
Lipid mediators are a prominent class of molecules derived from arachidonic acid (AA) metabolism involved in the antimicrobial defense of the lung. In general, the lipid mediators leukotriene (LT) B4, cysteinyl-LTs (LTC4, LTD4 and LTE4) and prostaglandin E2 (PGE2) have opposite effects on pulmonary defense mechanisms. We have reported previously, regarding the primary immune response to Histoplasma capsulatum, that the inhibition of endogenous LTs resulted in 100% mortality in sublethally H. capsulatum-infected mice, higher numbers of lung and spleen Colony Forming Unit (CFU), impaired Nitric Oxide (NO) production and inhibited Th1-cytokine expression. Higher susceptibility in 5-LO-/- mice correlates with decreased phagocytic capacity of IgG-opsonized Hc by macrophages when compared to WT. However, exogenous LTB4 and LTC4 restored phagocytosis deficiency in macrophages. Our results demonstrate that LTs are actually required to control chronic fungal infection in both innate and adaptive in primary immune responses in histoplasmosis. Moreover, in subsequent studies we have documented a protective function of LTs in the secondary immune response against H. capsulatum. In the present study, we demonstrate that the protection of WT mice immunized with Cell-Free Antigens (CFAgs) against H. capsulatum infection is associated with the increase of LTB4, NO and IFN-gamma production, and the recruitment of memory T cells. However, the deficiency of LT production in 5-LO KO mice impaired the protection induced by CFAgs. This effect is associated with the inhibition of recruitment and/or activation of effector memory T cells during H. capsulatum infection, resulting in mortality of immunized mice and increased CFU in the lung and spleen. Our studies have contributed to demonstrating the important role for LTs on both sides of host defense against to fungi.
Keywords: Lipid Mediators, Histoplasma capsulatum, Host defense, Cytokines, Leukotrienes
CONVENTIONAL AND MOLECULAR DIAGNOSIS OF ENDEMIC AND OPPORTUNISTIC MYCOSES
Author: GOMEZ, BL 1
Institution: 1. CIB; Corporacion para Investigaciones Biologicas
Abstract:
The definitive diagnosis of invasive fungal infections is usually based upon the isolation and identification of a specific etiologic agent in culture, and/or on the microscopic demonstration of the etiologic agent in histopathologic or other clinical specimens. Immunological tests (antibody and antigen detection) provide rapid results compared with the time required for culture and identification of fungal pathogens, and consequently they are often used as a basis for decisions on management of patients with invasive fungal infections. Tests for the detection of fungal antibodies have long had an important role in the diagnosis of endemic fungal infections, such as histoplasmosis, coccidioidomycosis, and paracoccidioidomycosis, as well as the different forms of aspergillosis that occur in nonimmunocompromised individuals. However, these tests still remain less useful for the opportunistic infections, such as aspergillosis and candidiasis, which occur in immunocompromised individuals. Antigen detection has long been an established method for the diagnosis of cryptococcosis, and the last few years have seen the advent of similar approaches for the detection of histoplasmosis, paracoccidioidomycosis, and penicilliosis. Although the use of monoclonal antibodies has enabled antigen detection tests to be marketed for the diagnosis of aspergillosis and candidiasis, finding circulating antigens that are not transient in nature remains an elusive goal. New molecular tests offer great hope for the rapid detection and identification of difficult-to-culture organisms, and for rapid diagnosis directly from clinical samples. However, few DNA-based tests are currently available for routine clinical use. Numerous “in-house” protocols have been developed for the detection of fungal DNA in clinical specimens, but there has been no standardization of test procedures. In more recent studies, genes specific for the fungus of interest are now being chosen as PCR targets, thus eliminating this specificity problem. Until these molecular procedures are standardized and commercialized, immunological tests will remain essential for rapid diagnosis of invasive fungal infections. This course will review the wide range of methods now available for the diagnosis of endemic and opportunistic invasive fungal infections and will examine the perspectives and future of new methodologies.
Keywords: DIAGNOSIS, CONVENTIONAL DIAGNOSIS, MOLECULAR DIAGNOSIS, ENDEMIC MYCOSES
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