Molecular Investigation Confirms Myotis Genus Bats as Common Hosts of Polychromophilus in Brazil

Plasmodium spp. and some other blood parasites belonging to the order Haemosporida are the focus of many epidemiological studies worldwide. However, haemosporidian parasites from wild animals are largely neglected in scientific research. For example, Polychromophilus parasites, which are exclusive to bats, are described in Europe, Asia, Africa, and Oceania, but little is known about their presence and genetic diversity in the New World. In this study, 224 samples of bats from remaining fragments of the Atlantic Forest and Pantanal biomes, as well as urbanized areas in southern and southeastern Brazil, were analyzed for the presence of haemosporidian parasites by PCR of the mitochondrial gene that encodes cytochrome b (cytb). The PCR fragments of the positive samples were sequenced and analyzed by the Bayesian inference method to reconstruct the phylogenetic relationships between Polychromophilus parasites from bats in Brazil and other countries. Sequences from Brazilian lineages of Polychromophilus were recovered in a clade with sequences from Polychromophilus murinus and close to the one Polychromophilus sequence obtained in Panama, the only available sequence for the American continent. This clade was restricted to bats of the family Vespertilionidae and distinct from Polychromophilus melanipherus, a parasite species mainly found in bats of the family Miniopteridae. The detection of Polychromophilus and the genetic proximity to P. murinus were further confirmed with the amplification of two other genes (clpc and asl). We also found a Haemosporida parasite sequence in a sample of Noctilio albiventris collected in the Pantanal biome, which presents phylogenetic proximity with avian Haemoproteus sequences. Morphological and molecular studies are still needed to conclude and describe the Polychromophilus species in Brazilian Myotis bats in more detail and to confirm Haemoproteus parasites in bats. Nevertheless, these molecular results in Brazilian bats confirm the importance of studying these neglected genera.Fil: da Silva Mathias, Bruno. Universidade de Sao Paulo; BrasilFil: Minozzo, Guilherme Augusto. No especifíca;Fil: Biondo, Alexander Welker. Universidade Federal do Paraná; BrasilFil: de Oliveira Jorge Costa, Jaciara. Universidade de Sao Paulo; BrasilFil: Sousa Soares, Herbert. Universidade de Santo Amaro; BrasilFil: Marcili, Arlei. Universidade de Sao Paulo; BrasilFil: de Oliveira Guimarães, Lilian. Instituto Pasteur; BrasilFil: a Clares dos Anjos, Carolina. Universidade de Sao Paulo; BrasilFil: Pires Dos Santos, Andrea. Purdue University; Estados UnidosFil: Riediger, Irina Nastassja. No especifíca;Fil: Fecchio, Alan. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Patagonia Norte. Centro de Investigación Esquel de Montaña y Estepa Patagónica. Universidad Nacional de la Patagonia "San Juan Bosco". Centro de Investigación Esquel de Montaña y Estepa Patagónica; ArgentinaFil: Bueno, Marina Galvão. Fundación Oswaldo Cruz; BrasilFil: Pinho, João Batista. Universidade Federal do Mato Grosso do Sul; BrasilFil: Kirchgatter, Karin. Universidade de Sao Paulo; Brasi

Produção de prostaglandinas e leucotrienos por Paracoccidioides brasiliensis

Paracoccidioides brasiliensis, é o agente da paracoccidioidomicose, a micose profunda endêmica na América Latina. Produção de eicosanóides, como prostaglandinas e leucotrienos, durante as infecções fúngicas tem mostrado desempenhar um papel crítico na sobrevivência e/ou crescimento fúngico, bem como na modulação da resposta imune do hospedeiro. As células hospedeiras são uma fonte desses mediadores, porém outra fonte em potencial pode ser o próprio fungo. O objetivo do nosso estudo foi avaliar se cepas do P. brasiliensis com diferentes graus de virulência (Pb18, Pb265, BT79, Pb192) produzem, prostaglandina E2 (PGE2) e leucotrieno B4 (LTB4). Além disso, verificamos se P. brasiliensis pode usar fontes exógenas de ácido araquidônico (AA), bem como as vias metabólicas dependentes das enzimas cicloxigenase (COX) e lipoxigenase (5-LO), para a produção de PGE2 e LTB4, respectivamente. Finalmente, uma possível associação entre esses eicosanóides e a viabilidade do fungo também foi avaliada. Demonstramos, usando ensaios de Elisa, que todas as cepas de P. brasiliensis, independente do seu grau de virulência, produzem altos níveis de PGE2 e LTB4 após 4h cultura que foram reduzidos após 8h. No entanto, em ambos os tempos da cultura, foram detectados níveis mais elevados de eicosanóides, quando as culturas foram complementadas com uma fonte exógena de AA. Diferentemente, o tratamento com indometacina, um inibidor da COX, ou MK886, um inibidor da 5-LO, induz a uma redução nos níveis de PGE2 e LTB4, respectivamente, bem como a viabilidade do fungo. Os dados fornecem evidências de que P. brasiliensis tem a capacidade de metabolizar, tanto AA endógeno como exógeno por vias dependentes da COX e 5-LO enzimas que participam da produção de PGE2 e LTB4, respectivamente, que são indispensáveis para a sua viabilidade.Paracoccidioides brasiliensis, is the agent of paracoccidioidomycosis, the most prevalent deep mycosis in Latin America. Production of eicosanoids, such as prostaglandins and leukotrienes, during fungal infections is theorized to play a critical role on fungal survival and/or growth as well as on host immune response modulation. Host cells are one source of these mediators; however another potential source may be the fungal itself. The purpose of our study was to assess whether P. brasiliensis strains with different degree of virulence (Pb18, Pb265, PbBT79, Pb192) produce both, prostaglandin E2 (PGE2) and leukotriene B4 (LTB4). Moreover, we asked if P. brasiliensis can use exogenous sources of arachidonic acid (AA), as well as metabolic pathways dependent on cyclooxygenase ( COX) and lipooxygenase (5-LO ) enzymes, for PGE2 and LTB4 production , respectively. Finally, a possible association between these eicosanoides and fungus viability was assessed. We have demonstrated, using Elisa assays, that all P. brasiliensis strains, independently of their virulence degree, produce high PGE2 and LTB4 levels on 4h culture that were reduced after 8 h. However, in both culture times, higher eicosanoides levels were detected when culture medium was supplemented with exogenous AA. Differently, treatment with indomethacin, a COX inhibitor, or MK886, a 5-LO inhibitor, induces a reduction on PGE2 and LTB4 levels, respectively, as well as in fungus viability. The data provide evidence that P. brasiliensis has the capacity to metabolize either endogenous or exogenous AA by pathways dependent on COX and 5-LO enzymes for producing PGE2 and LTB4, respectively, that are critical for its viability.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Identificação e purificação de prostaglandinas e leucotrienos produzidos por Paracoccidioides brasiliensis

A paracoccidioidomicose (PCM) é uma micose sistêmica causada pelo fungo Paracoccidioides brasiliensis. O estabelecimento da paracoccidioidomicose-infecção ou paracoccidioidomicose-doença depende das interações parasita-hospedeiro que podem resultar em uma resposta imune mais ou menos eficaz por parte deste último. Embora vários dos mecanismos decorrentes dessa interação já estejam descritos, estudos sobre a participação de outros fatores potenciais podem ampliar sobremaneira o nosso entendimento sobre a imunopatogênese da PCM. Nesse sentido, os eicosanóides como as prostaglandinas (PGs) e leucotrienos (LTs) devem ser avaliados. Em trabalhos anteriores detectamos que o fungo induz a produção de PGs e LTs por monócitos humanos que, de uma forma autócrina, suprimem ou aumentam respectivamente, as atividades antifúngicas dessas células. No entanto, os estudos têm mostrado que além de induzir a produção de eicosanóides, os próprios fungos podem produzir esses mediadores, revelando um mecanismo de virulência que tem potencialmente grandes implicações para o entendimento dos mecanismos das infecções fúngicas. Baseados nesses estudos detectamos que o P. brasiliensis produz tanto PGs como LTs, usando fontes endógenas e exógenas de ácido araquidônico, e que ambos são importantes para a manutenção da viabilidade do fungo. Adicionalmente, os resultados sugeriram que o fungo utiliza as vias metabólicas da ciclooxigenase (COX) para a produção de PGs e da lipoxigenase (5-LO) para LTs. No presente trabalho, aprofundamos esses estudos objetivando avaliar se o fungo produz outras classes de eicosanóides, além da PGE2 e LTB4, se as PGs e LTs liberados pelo P. brasiliensis podem ser considerados como PGE2 e LTB4 autênticos, isto é semelhantes aos detectados para os mamíferos, assim como estabelecer de uma forma definitiva se a COX e a 5-LO são as vias metabólicas utilizadas...Paracoccidioidomycosis (PCM) is a systemic mycosis caused by the dimorphic fungus Paracoccidioides brasiliensis. The establishment of paracoccidioidomycosis-infection or the evolution to paracoccidioidomycosis-disease in its more or less severe forms, primarily depends on complex host-parasite interaction, in which immune response mechanisms play an essential role. While many of these mechanisms have been elucidated, studies on the participation of other potential factors can greatly expand our understanding of PCM immunopathogenesis. Accordingly, eicosanoids such as prostaglandins (PGs) and leukotrienes (LTs) should be evaluated. In previous studies we found that the fungus induces the production of prostaglandins and leukotrienes by human monocytes that in an autocrine way suppress and increase, respectively, the antifungal activities of these cells. However, studies have shown that in addition to inducing the production of eicosanoids, fungi themselves can produce these mediators, showing a mechanism for virulence that has potentially important implications for understanding the mechanisms of fungal pathogenicity. Based on these studies, we found that P. brasiliensis produces both PGs and LTs, using exogenous and endogenous sources of arachidonic acid and that both are important for fungus viability. Additionally, the results suggested that the fungus uses cyclooxygenase (COX) and lipoxygenase (5-LO) metabolic pathways for PGs and LTs production, respectively. Here, we profound these studies evaluating whether PGs and LTs released by P. brasiliensis can be considered as authentic LTB4 and PGE2, i.e. similar to those detected for mammals. We also asked whether the fungus, in addition to PGE2 and LTB4, produces other classes of eicosanoids. Finally, we intended to definitively determine whether COX and 5-LO are the metabolic pathways used for this production. We found that fungus PGE2 and LTB4 ..

The influence of hypothyroidism on liver regeneration: an experimental study in rats A influência do hipotireoidismo na regeneração hepatica: estudo experimental em ratos

BACKGROUND: The influence of hypothyroidism in liver regeneration has been a controversial opinions. PURPOSE: The aim of this study is to identify the relationship between hypothyroidism and liver regeneration in rats. METHODS: Forty male Wistar rats divided into two groups of 20 specimens each. One group (C) consisted of euthyroid rats, and the other (H) of hypothyroid rats. All the animals were anesthetized with xylazine and ketamine and subjected to a longitudinal incision in the anterior cervical region. The thyroid was completely resected in group H and left intact in group C. Ten days after the first surgery, both groups of rats were weighed and submitted to partial hepatectomy, in which the left lateral and median lobes were resected and weighed. Examinations were carried out after 24 hours and, on day 7, using 3 methods: KWON et al.'s formula to identify increase in volume; mitotic figure count in five fields; and the percentage of PCNA-positive nuclei in five fields. RESULTS: Using KWON's formula, the regeneration rate for Group C after 24 hours was 58.49% whereas that for Group H was 50.42% (p=0.0165). After 7 days, the regeneration rate for Group C was 93.04% and Group H 93.74% (p=0.2165). The average number of mitotic figures after 24 hours was 14 &plusmn; 1.5 for Group C and 9.8 &plusmn; 2.2 for Group H (p=0,00016). After 7 days the corresponding figures were 5.4 &plusmn; 1.1 and 5.1 &plusmn; 1.2 (p=0,6343). The average number of PCNA-positive nuclei after 24 hours was 13.55 &plusmn; 3.84 in Group C and 7.7 &plusmn; 2.11 in Group H (p =0,0006)). The corresponding figures after 7 days were 3.5 &plusmn; 2.39 for Group C and 4.11 &plusmn; 1.90 for Group H (p>0.05). CONCLUSION: We conclude that hypothyroidism in rats causes a delay in hepatic regeneration in the first 24 hours, but that after seven days the rate of regeneration is equal to that in euthyroid rats.<br>BACKGROUND: A influência do hipotireoidismo na regeneração hepatica tem opiniões controvérsas. OBJETIVO: Identificar a relação entre o hipotireoidismo e a regeneração hepatica, em ratos. MÉTODOS: Quarenta ratos Wistar, machos, divididos em dois grupos de 20 animais. Um grupo (C) constituído de ratos eutireoideanos e outro (H) de ratos hipotireoideanos. Todos os animais foram anestesiados com xilasina e quetamina e submetidos a uma incisão cervical longitudinal. A tireóide foi completamente ressecada no animais do grupo H e deixada intacta no grupo C. Dez dias após a primeira intervenção ambos os grupos de ratos foram pesados e submetidos à hepatectomia parcial, retirando-se os lobos lateral esquerdo e mediano, que foram pesados. Foram examinados com 24 horas e com 7 dias usando-se 3 métodos: formula de KWON et al. que identifica o aumento de volume; contagem de figures de mitose em cinco campos e de núcleos PCNA positivos em cinco campos. RESULTOS: Usando a formula de KWON et al. a taxa de regeneração do grupo C, após 24 horas, foi de 58.49% enquanto que a do grupo H foi de 50.42% (p=0.0165). Após 7 dias, a taxa de regeneração no grupo C foi de 93.04% e do grupo H 93.74% (p=0.2165). A média de figures de mitose após 24 horas foi de 14 &plusmn; 1.5 no grupo C e de 9.8 &plusmn; 2.2 no groupo H (p=0,00016). Após 7 dias foi de 5.4 &plusmn; 1.1 e 5.1 &plusmn; 1.2 (p=0,6343). A média de núcleos PCA positivos, após 24 horas, foi de 13.55 &plusmn; 3.84 no grupo C e 7.7 &plusmn; 2.11 no grupo H (p =0,0006)). Com 7 dias foi de 3.5 &plusmn; 2.39 no groupo C e de 4.11 &plusmn; 1.90 for Group H (p>0.05). CONCLUSÃO: Concluiu-se que o hipotireoidismo, em ratos, causa atraso da regeneração hepatica nas primeiras 24 horas, mas após 7 dias a regeneração se equivale à dos ratos eutireoideanos

The role of oxidative stress in renal injury related to obesity

The mechanisms linking obesity to kidney damage are unknown. AGEs are responsible for renal damage in obese individuals. The receptor AGEs (RAGE) contributes to nuclear transcription factors that result in the production of proinflammatory cytokines and this seems to contribute to the development of renal disease. Thus, intervention with antioxidant can have an important effect in the prevention and treatment of pro-oxidant and pro-inflammatory state in the kidneys resulting from obesity

Paracoccidioides brasiliensis Uses Endogenous and Exogenous Arachidonic Acid for PGE(x) Production

Paracoccidioides brasiliensis is the agent of paracoccidioidomycosis, the most prevalent deep mycosis in Latin America. Production of eicosanoids during fungal infections plays a critical role on fungal biology as well as on host immune response modulation. The purpose of our study was to assess whether P. brasiliensis strains with different degree of virulence (Pb18, Pb265, Bt79, Pb192) produce prostaglandin E-x (PGE(x)). Moreover, we asked if P. brasiliensis could use exogenous sources of arachidonic acid (AA), as well as metabolic pathways dependent on cyclooxygenase (COX) enzyme, as reported for mammalian cells. A possible association between this prostanoid and fungus viability was also assessed. Our results showed that all strains, independently of their virulence, produce high PGE(x) levels on 4 h culture that were reduced after 8 h. However, in both culture times, higher prostanoid levels were detected after supplementation of medium with exogenous AA. Treatment with indomethacin, a COX inhibitor, induced a reduction on PGEx, as well as in fungus viability. The data provide evidence that P. brasiliensis produces prostaglandin-like molecules by metabolizing either endogenous or exogenous AA. Moreover, the results suggest the involvement of these mediators on fungal viability

Production of leukotriene B4 by Paracoccidioides brasiliensis

Paracoccidioides brasiliensis is the agent of paracoccidioidomycosis, the most prevalent deep mycosis in Latin America. The production of eicosanoids during fungal infection has been associated with the biology of these microorganisms and modulation of host immune response. The aim of our study was to evaluate whether P. brasiliensis strains with high or low virulence produce leukotriene B4 (LTB4), using endogenous and/or exogenous sources of arachidonic acid (AA). Moreover, we assessed whether this fungus might use the same metabolic pathway, described for mammalian cells, that involves the lipoxygenase (LOX) enzyme. The association between the production of this eicosanoid and fungus survival and growth was also evaluated. Our results showed that P. brasiliensis, irrespective of its virulence, produces high levels of LTB4 using endogenous AA. In addition, in cultures treated with exogenous AA, LTB4 levels were significantly higher, showing that this fungus also uses exogenous sources of fatty acids. Treatment with MK886, which blocks the activity of lipoxygenase, by inhibiting five-lipoxygenase-activating protein (FLAP) or with nordihydroguaiaretic acid (NDGA), a non-selective lipoxygenase inhibitor, resulted in a significant reduction in LTB4 levels, indicating that the fungus produces this eicosanoid by using the LOX pathway or an enzyme with biochemically similar function. The significant reduction in viability detected in cultures treated with these inhibitors was, however, restored by adding exogenous LTB4, confirming the role of this eicosanoid in fungus survival. Moreover, the addition of LTB4 to cultures capable of producing LTs induces fungal growth. These results provide a foundation for additional studies on the contributions of LTB4 in P. brasiliensis virulence. Copyright (c) 2012 John Wiley & Sons, Ltd.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP