RESUMO
Histoplasma capsulatum is the causative agent of histoplasmosis, a systemic disease responsible for most reported causes of morbidity and mortality among immunosuppressed individuals. Peptidogalactomannan (pGM) was purified from the yeast cell wall of H. capsulatum isolated from bats, and its structure and involvement in modulating the host immune response were evaluated. Gas chromatography, methylation analysis, and two-dimensional nuclear magnetic resonance (2D-NMR) were used for the structural characterization of pGM. Methylation and 2D-NMR data revealed that pGM comprises a main chain containing α-D-Manp (1 â 6) residues substituted at O-2 by α-D-Manp (1 â 2)-linked side chains, non-reducing end units of α-D-Galf, or ß-D-Galp linked (1â 6) to α-D-Manp side chains. The involvement of H. capsulatum pGM in antigenic reactivity and in interactions with macrophages was demonstrated by ELISA and phagocytosis assay, respectively. The importance of the carbohydrate and protein moieties of pGM in sera reactivity was evaluated. Periodate oxidation abolished much pGM antigenic reactivity, suggesting that the sugar moiety is the most immunogenic part of pGM. Reactivity slightly decreased in pGM treated with proteinase K, suggesting that the peptide moiety plays a minor role in pGM antigenicity. In vitro experiments suggested that pGM is involved in the phagocytosis of H. capsulatum yeast and induction of IL-10 and IFN-γ secretion by peritoneal macrophages from C57BL/6 mice. These findings demonstrated the role of pGM in the H. capsulatum-host interaction.
Assuntos
Glicopeptídeos/química , Glicopeptídeos/farmacologia , Histoplasma/química , Histoplasmose/microbiologia , Macrófagos Peritoneais/efeitos dos fármacos , Mananas/química , Mananas/farmacologia , Animais , Parede Celular/química , Parede Celular/imunologia , Quirópteros/microbiologia , Feminino , Galactose/análogos & derivados , Histoplasma/imunologia , Histoplasma/isolamento & purificação , Histoplasmose/genética , Histoplasmose/imunologia , Humanos , Interferon gama/genética , Interferon gama/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Macrófagos Peritoneais/imunologia , Masculino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos C57BL , Fagocitose/efeitos dos fármacos , CoelhosRESUMO
Histoplasmosis is a worldwide-distributed deep mycosis that affects healthy and immunocompromised hosts. Severe and disseminated disease is especially common in HIV-infected patients. At least 11 phylogenetic species are recognized and the majority of diversity is found in Latin America. The northeastern region of Brazil has one of the highest HIV/AIDS prevalence in Latin America and Ceará State has one of the highest death rates due to histoplasmosis in the world, where the mortality rate varies between 33-42%. The phylogenetic distribution and population genetic structure of 51 clinical isolates from Northeast Brazil was studied. For that morphological characteristics, exoantigens profile, and fungal mating types were evaluated. The genotypes were deduced by a MSLT in order to define local population structure of this fungal pathogen. In addition, the relationships of H. capsulatum genotypes with clinically relevant phenotypes and clinical aspects were investigated. The results suggest two cryptic species, herein named population Northeast BR1 and population Northeast BR2. These populations are recombining, exhibit a high level of haplotype diversity, and contain different ratios of mating types MAT1-1 and MAT1-2. However, differences in phenotypes or clinical aspects were not observed within these new cryptic species. A HIV patient can be co-infected by two or more genotypes from Northeast BR1 and/or Northeast BR2, which may have significant impact on disease progression due to the impaired immune response. We hypothesize that co-infections could be the result of multiple exposure events and may indicate higher risk of disseminated histoplasmosis, especially in HIV infected patients.
Assuntos
Infecções por HIV/genética , Histoplasma/genética , Histoplasmose/genética , Filogenia , Adulto , Brasil/epidemiologia , Feminino , Variação Genética/genética , Genótipo , HIV/genética , HIV/patogenicidade , Infecções por HIV/microbiologia , Infecções por HIV/patologia , Infecções por HIV/virologia , Haplótipos/genética , Histoplasma/patogenicidade , Histoplasmose/microbiologia , Histoplasmose/patologia , Histoplasmose/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
Histoplasmosis is considered one of the most important endemic and systemic mycoses worldwide. Until now few molecular techniques have been developed for its diagnosis. The aim of this study was to develop and evaluate three real time PCR (qPCR) protocols for different protein-coding genes (100-kDa, H and M antigens) using an animal model. Fresh and formalin-fixed and paraffin-embedded (FFPE) lung tissues from BALB/c mice inoculated i.n. with 2.5x106 Histoplasma capsulatum yeast or PBS were obtained at 1, 2, 3, 4, 8, 12 and 16 weeks post-infection. A collection of DNA from cultures representing different clades of H. capsulatum (30 strains) and other medically relevant pathogens (36 strains of related fungi and Mycobacterium tuberculosis) were used to analyze sensitivity and specificity. Analytical sensitivity and specificity were 100% when DNAs from the different strains were tested. The highest fungal burden occurred at first week post-infection and complete fungal clearance was observed after the third week; similar results were obtained when the presence of H. capsulatum yeast cells was demonstrated in histopathological analysis. In the first week post-infection, all fresh and FFPE lung tissues from H. capsulatum-infected animals were positive for the qPCR protocols tested except for the M antigen protocol, which gave variable results when fresh lung tissue samples were analyzed. In the second week, all qPCR protocols showed variable results for both fresh and FFPE tissues. Samples from the infected mice at the remaining times post-infection and uninfected mice (controls) were negative for all protocols. Good agreement was observed between CFUs, histopathological analysis and qPCR results for the 100-kDa and H antigen protocols. We successfully standardized and validated three qPCR assays for detecting H. capsulatum DNA in fresh and FFPE tissues, and conclude that the 100-kDa and H antigen molecular assays are promising tests for diagnosing this mycosis.
Assuntos
Modelos Animais de Doenças , Genes Fúngicos , Histoplasmose/diagnóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Animais , DNA Fúngico/genética , DNA Fúngico/isolamento & purificação , Histoplasma/genética , Histoplasmose/genética , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Sensibilidade e EspecificidadeRESUMO
Three closely related thermally dimorphic pathogens are causal agents of major fungal diseases affecting humans in the Americas: blastomycosis, histoplasmosis and paracoccidioidomycosis. Here we report the genome sequence and analysis of four strains of the etiological agent of blastomycosis, Blastomyces, and two species of the related genus Emmonsia, typically pathogens of small mammals. Compared to related species, Blastomyces genomes are highly expanded, with long, often sharply demarcated tracts of low GC-content sequence. These GC-poor isochore-like regions are enriched for gypsy elements, are variable in total size between isolates, and are least expanded in the avirulent B. dermatitidis strain ER-3 as compared with the virulent B. gilchristii strain SLH14081. The lack of similar regions in related species suggests these isochore-like regions originated recently in the ancestor of the Blastomyces lineage. While gene content is highly conserved between Blastomyces and related fungi, we identified changes in copy number of genes potentially involved in host interaction, including proteases and characterized antigens. In addition, we studied gene expression changes of B. dermatitidis during the interaction of the infectious yeast form with macrophages and in a mouse model. Both experiments highlight a strong antioxidant defense response in Blastomyces, and upregulation of dioxygenases in vivo suggests that dioxide produced by antioxidants may be further utilized for amino acid metabolism. We identify a number of functional categories upregulated exclusively in vivo, such as secreted proteins, zinc acquisition proteins, and cysteine and tryptophan metabolism, which may include critical virulence factors missed before in in vitro studies. Across the dimorphic fungi, loss of certain zinc acquisition genes and differences in amino acid metabolism suggest unique adaptations of Blastomyces to its host environment. These results reveal the dynamics of genome evolution and of factors contributing to virulence in Blastomyces.
Assuntos
Blastomyces/genética , Chrysosporium/genética , Genoma Fúngico , Transcriptoma/genética , Animais , Blastomyces/patogenicidade , Blastomicose/genética , Blastomicose/microbiologia , Chrysosporium/patogenicidade , Histoplasmose/genética , Histoplasmose/microbiologia , Humanos , Macrófagos/microbiologia , Camundongos , Paracoccidioidomicose/genética , Paracoccidioidomicose/microbiologiaRESUMO
The bioactive lipid mediator leukotriene B4 (LTB4) greatly enhances phagocyte antimicrobial functions against a myriad of pathogens. In murine histoplasmosis, inhibition of the LT-generating enzyme 5-lypoxigenase (5-LO) increases the susceptibility of the host to infection. In this study, we investigated whether murine resistance or susceptibility to Histoplasma capsulatum infection is associated with leukotriene production and an enhancement of in vivo and/or in vitro antimicrobial effector function. We show that susceptible C57BL/6 mice exhibit a higher fungal burden in the lung and spleen, increased mortality, lower expression levels of 5-LO and leukotriene B4 receptor 1 (BLT1) and decreased LTB4 production compared to the resistant 129/Sv mice. Moreover, we demonstrate that endogenous and exogenous LTs are required for the optimal phagocytosis of H. capsulatum by macrophages from both murine strains, although C57BL/6 macrophages are more sensitive to the effects of LTB4 than 129/Sv macrophages. Therefore, our results provide novel evidence that LTB4 production and BLT1 signaling are required for a histoplasmosis-resistant phenotype.
Assuntos
Histoplasma/imunologia , Histoplasmose/veterinária , Leucotrieno B4 , Receptores do Leucotrieno B4/imunologia , Doenças dos Roedores/imunologia , Animais , Araquidonato 5-Lipoxigenase/genética , Araquidonato 5-Lipoxigenase/imunologia , Suscetibilidade a Doenças , Inibidores Enzimáticos/farmacologia , Expressão Gênica/imunologia , Histoplasma/patogenicidade , Histoplasmose/genética , Histoplasmose/imunologia , Histoplasmose/metabolismo , Especificidade de Hospedeiro , Interações Hospedeiro-Patógeno , Leucotrieno B4/metabolismo , Leucotrieno B4/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/microbiologia , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/microbiologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Fagocitose/efeitos dos fármacos , Receptores do Leucotrieno B4/genética , Doenças dos Roedores/genética , Doenças dos Roedores/metabolismo , Transdução de Sinais , Baço/efeitos dos fármacos , Baço/imunologia , Baço/microbiologiaRESUMO
BACKGROUND: In the pre-microbiological era, it was widely accepted that diseases, today known to be infectious, were hereditary. With the discovery of microorganisms and their role in the pathogenesis of several diseases, it was suggested that exposure to the pathogen was enough to explain infection. Nowadays, it is clear that infection is the result of a complex interplay between pathogen and host, therefore dependant on the genetic make-up of the two organisms. Dermatology offers several examples of infectious diseases in different stages of understanding of their molecular basis. In this review, we summarize the main advances towards dissecting the genetic component controlling human susceptibility to infectious diseases of interest in dermatology. Widely investigated diseases such as leprosy and leishmaniasis are discussed from the genetic perspective of both host and pathogen. Others, such as rare mycobacterioses, fungal infections and syphilis, are presented as good opportunities for research in the field of genetics of infection.
Assuntos
Predisposição Genética para Doença/genética , Interações Hospedeiro-Patógeno/genética , Dermatopatias Infecciosas/genética , Histoplasmose/genética , Humanos , Leishmania/genética , Leishmaniose/genética , Hanseníase/genética , Mycobacterium leprae/genética , Paracoccidioidomicose/genética , Fatores de Risco , Sífilis Cutânea/genética , Tuberculose Cutânea/genéticaRESUMO
BACKGROUND: In the pre-microbiological era, it was widely accepted that diseases, today known to be infectious, were hereditary. With the discovery of microorganisms and their role in the pathogenesis of several diseases, it was suggested that exposure to the pathogen was enough to explain infection. Nowadays, it is clear that infection is the result of a complex interplay between pathogen and host, therefore dependant on the genetic make-up of the two organisms. Dermatology offers several examples of infectious diseases in different stages of understanding of their molecular basis. In this review, we summarize the main advances towards dissecting the genetic component controlling human susceptibility to infectious diseases of interest in dermatology. Widely investigated diseases such as leprosy and leishmaniasis are discussed from the genetic perspective of both host and pathogen. Others, such as rare mycobacterioses, fungal infections and syphilis, are presented as good opportunities for research in the field of genetics of infection.
INTRODUÇÃO: Durante a era pré-microbiológica, era comum a visão de que doenças, hoje sabidamente infecciosas, eram hereditárias. Com a descoberta dos microorganismos e seu papel na patogênese de diversas patologias, chegou-se a propor que a exposição ao patógeno era condição suficiente para explicar infecção. Hoje, está claro que infecção é o resultado de uma complexa interação entre patógeno e hospedeiro, dependendo portanto, em última análise, do make-up genético de ambos os organismos. A dermatologia oferece diversos exemplos de doenças infecciosas em diferentes graus de entendimento de suas bases moleculares. Nesta revisão, resumimos os principais avanços na direção da dissecção do componente genético controlando suscetibilidade do ser humano a doenças infecciosas de importância na dermatologia. Doenças amplamente estudadas, como a hanseníase e a leishmaniose, são discutidas sob o ponto de vista da genética tanto do hospedeiro quanto do patógeno. Outras, como micobacterioses raras, micoses e sífilis, são apresentadas como boas oportunidades para pesquisa na área de genética de infecção.
Assuntos
Humanos , Predisposição Genética para Doença/genética , Interações Hospedeiro-Patógeno/genética , Dermatopatias Infecciosas/genética , Histoplasmose/genética , Leishmania/genética , Leishmaniose/genética , Hanseníase/genética , Mycobacterium leprae/genética , Paracoccidioidomicose/genética , Fatores de Risco , Sífilis Cutânea/genética , Tuberculose Cutânea/genéticaRESUMO
BACKGROUND: Histoplasmosis and paracoccidioidomycosis (PCM) have increased in Spain in recent years, due firstly to the migration from endemic regions and secondly to travelers returning from these regions. In non-endemic areas, diagnosis of both diseases is hampered by the lack of experience, long silent periods, and the resemblance to other diseases such as tuberculosis and sarcoidosis. METHODS: A total of 39 cases of imported histoplasmosis and 6 cases of PCM diagnosed in the Spanish Mycology Reference Laboratory since 2006 were analyzed. Microbiological diagnosis was performed using classical methods and also a specific real-time polymerase chain reaction (RT-PCR) assay for each microorganism. RESULTS: We had 9 cases of probable histoplasmosis in travelers and 30 cases in immigrants, 29 of whom were defined as proven. Paracoccidioidomycosis (PCM) cases were either immigrants or people who had lived for a long period of time in endemic regions, all of whom were classified as proven cases. Cultures showed a good sensitivity in detecting Histoplasma capsulatum in immigrants with proven histoplasmosis (73%); however, growth was very slow. The fungus was never recovered in traveler patients. Paracoccidioides brasiliensis was isolated in a culture only in one case of the proven PCM. Serological methods were not very reliable in immunocompromised patients with histoplasmosis (40%). A PCR-based technique for histoplasmosis detected 55.5% of the cases in travelers (probable cases) and 89% of the cases in immigrants (proven). The PCR method for PCM detected 100% of the cases. CONCLUSIONS: These kinds of mycoses are increasingly frequent in non-endemic areas, and newer and faster techniques should be used to reach an early diagnosis. The RT-PCR techniques developed appear to be sensitive, specific, and fast and could be helpful to detect those mycoses. However, it is also essential that physicians perform differential diagnosis in individuals coming from endemic areas.
Assuntos
Emigração e Imigração/estatística & dados numéricos , Histoplasma/isolamento & purificação , Histoplasmose/diagnóstico , Paracoccidioides/isolamento & purificação , Paracoccidioidomicose/diagnóstico , Viagem/estatística & dados numéricos , Adulto , África , Antifúngicos/administração & dosagem , América Central , Doenças Endêmicas , Feminino , Histoplasmose/tratamento farmacológico , Histoplasmose/epidemiologia , Histoplasmose/genética , Humanos , Masculino , Pessoa de Meia-Idade , Paracoccidioidomicose/tratamento farmacológico , Paracoccidioidomicose/epidemiologia , Paracoccidioidomicose/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , América do Sul , Espanha/epidemiologia , Adulto JovemRESUMO
Introducción: La histoplasmosis es una micosis causada por Histoplama capsulatum: La enfermedad tiene una alta prevalencia en países del continente americano donde los climas tropical y subtropical favorecen el crecimiento del hongo. En la susceptibilidad a la infección se ha descrito la participación de factores inmunológicos, ocupacionales y genéticos. En cuanto a los factores genéticos, los genes del Sistema Principal de Histocompatibilidad (SPH) se han asociado con la susceptibilidad a la histoplasmosis y otras enfermedades. En el humano estos genes se localizan en el cromosoma seis, son muy polimórficos, y sus productos son antígenos HLA, impotantes en la regulación de la respuesta inmunológica contra agentes extraños. Factores genéticos asociados a la histoplasmosis: Estudios realizados en la población mexicana han mostrado que los alelos HLA-B17 y HLA-B22, son frecuentes en pacientes con histoplasmosis pulmonar; el último contenido en el haplotipo extendido [HLA-A2, B22, Cw5]. Por la frecuencia en estos pacientes lo alelos referidos podrían servir como marcadores para la enfermedad. Por lo anterior, se sugiere un efecto predisponentes para la infección y el desarrollo de histoplasmosis en individuos que presentan estos alelos
Assuntos
Antígenos HLA/genética , Antígenos HLA , Histoplasmose/genética , Histoplasmose/imunologia , Imunogenética , Complexo Principal de Histocompatibilidade/genética , Marcadores Genéticos/genética , Técnicas de Tipagem MicológicaRESUMO
Immunologic and occupational aspects of the susceptible population exposed to Histoplasma capsulatum, the causative agent of histoplasmosis were analyzed in the Mexican State of Guerrero. Three areas were studied, Juxtlahuaca, Olinala, and Coyuca; in the first two, their populations refer contact with bat guano and/or avian excreta, which contain nutrients for fungal growth, while the Coyuca population referred no contact with the above mentioned excreta. Previous infection with H. capsulatum was determined by histoplasmin-skin test, and the response was higher in men than in women (93.87, 85.71, and 6.6% for men, and 78.94, 66.6, and 0% for women) in Juxtlahuaca, Olinala, and Coyuca, respectively. Labor activities related to a persistent contact with the fungus were considered as an occupational risk factor, histoplasmin-skin test reached 88.57 and 36.36% of positive response in individuals with high and low risk activities. A high percentage of histoplasmin responses was observed in subjects with constant contact with H. capsulatum, such as, cave-tourist guides, peasants, and game-cock handlers, and generally they developed the largest diameter of skin reactions. Genetic risk factor was determined by studying the gene frequency of the Major Histocompatibility Complex antigens in a sample of individuals and their degree relatives in Juxtlahuaca, Olinala, and Coyuca. Significant differences were found for HLA-B22 and B17 antigens in Juxtlahuaca, and for HLA-B22 in Olinala, in comparison to the usual gene frequency observed in the normal Mexican population. HLA results were important, considering that HLA-B22 was previously found to be possibly related to pulmonary histoplasmosis in Guerrero.