Inhibitory Effect of the Ethanol Extract of a Rice Bran Mixture Comprising Angelica gigas, Cnidium officinale, Artemisia princeps, and Camellia sinensis on Brucella abortus Uptake by Professional and Nonprofessional Phagocytes.

In this study, we evaluated the inhibitory effect of a rice bran mixture extract (RBE) on Brucella abortus pathogenesis in professional (RAW 264.7) and nonprofessional (HeLa) phagocytes. We fermented the rice bran mixture and then extracted it with 50% ethanol followed by gas chromatography-mass spectrometry to identify the components in RBE. Our results clearly showed that RBE caused a significant reduction in the adherence of B. abortus in both cell lines. Furthermore, analysis of phagocytic signaling proteins by western blot assay revealed that RBE pretreatment resulted in inhibition of phosphorylation of JNK, ERK, and p38, leading to decline of internalization compared with the controls. Additionally, the intensity of F-actin observed by fluorescence microscopy and FACS was remarkably reduced in RBE-pretreated cells compared with control cells. However, the intracellular replication of B. abortus within phagocytes was not affected by RBE. Taken together, these findings suggest that the phagocytic receptor blocking and suppressive effects of RBE on the MAPK-linked phagocytic signaling pathway could negatively affect the invasion of B. abortus into phagocytes.

The first description of complete invertebrate arginine metabolism pathways implies dose-dependent pathogen regulation in Apostichopus japonicus.

In this study, three typical members representative of different arginine metabolic pathways were firstly identified from Apostichopus japonicus, including nitric oxide synthase (NOS), arginase, and agmatinase. Spatial expression analysis revealed that the AjNOS transcript presented negative expression patterns relative to those of Ajarginase or Ajagmatinase in most detected tissues. Furthermore, Vibrio splendidus-challenged coelomocytes and intestine, and LPS-exposed primary coelomocytes could significantly induce AjNOS expression, followed by obviously inhibited Arginase and AjAgmatinase transcripts at the most detected time points. Silencing the three members with two specific siRNAs in vivo and in vitro collectively indicated that AjNOS not only compete with Ajarginase but also with Ajagmatinase in arginine metabolism. Interestingly, Ajarginase and Ajagmatinase displayed cooperative expression profiles in arginine utilization. More importantly, live pathogens of V. splendidus and Vibrio parahaemolyticus co-incubated with primary cells also induced NO production and suppressed arginase activity in a time-dependent at an appropriate multiplicity of infection (MOI) of 10, without non-pathogen Escherichia coli. When increasing the pathogen dose (MOI = 100), arginase activity was significantly elevated, and NO production was depressed, with a larger magnitude in V. splendidus co-incubation. The present study expands our understanding of the connection between arginine's metabolic and immune responses in non-model invertebrates.

A repurposing approach identifies off-patent drugs with fungicidal cryptococcal activity, a common structural chemotype, and pharmacological properties relevant to the treatment of cryptococcosis.

New, more accessible therapies for cryptococcosis represent an unmet clinical need of global importance. We took a repurposing approach to identify previously developed drugs with fungicidal activity toward Cryptococcus neoformans, using a high-throughput screening assay designed to detect drugs that directly kill fungi. From a set of 1,120 off-patent medications and bioactive molecules, we identified 31 drugs/molecules with fungicidal activity, including 15 drugs for which direct antifungal activity had not previously been reported. A significant portion of the drugs are orally bioavailable and cross the blood-brain barrier, features key to the development of a widely applicable anticryptococcal agent. Structural analysis of this set revealed a common chemotype consisting of a hydrophobic moiety linked to a basic amine, features that are common to drugs that cross the blood-brain barrier and access the phagolysosome, two important niches of C. neoformans. Consistent with their fungicidal activity, the set contains eight drugs that are either additive or synergistic in combination with fluconazole. Importantly, we identified two drugs, amiodarone and thioridazine, with activity against intraphagocytic C. neoformans. Finally, the set of drugs is also enriched for molecules that inhibit calmodulin, and we have confirmed that seven drugs directly bind C. neoformans calmodulin, providing a molecular target that may contribute to the mechanism of antifungal activity. Taken together, these studies provide a foundation for the optimization of the antifungal properties of a set of pharmacologically attractive scaffolds for the development of novel anticryptococcal therapies.

Effect of dietary administration of Porphyridium cruentum on the respiratory burst activity of sole, Solea senegalensis (Kaup), phagocytes.

The stimulatory effect of the red microalga Porphyridium cruentum on respiratory burst activity of sole phagocytes was evaluated in vivo. Oral administration of a diet supplemented with lyophilized P. cruentum cells (10 g kg(-1)) stimulated respiratory burst activity after 4 weeks feeding in sole vaccinated with Photobacterium damselae subsp. piscicida bacterin.

The co-ordinated regulation of iron homeostasis in murine macrophages limits the availability of iron for intracellular Salmonella typhimurium.

In being both, a modifier of cellular immune effector pathways and an essential nutrient for microbes, iron is a critical determinant in host-pathogen interaction. Here, we investigated the metabolic changes of macrophage iron homeostasis and immune function following the infection of RAW264.7 murine macrophages with Salmonella typhimurium. We observed an enhanced expression of the principal iron export protein, ferroportin 1, and a subsequent increase of iron efflux in Salmonella-infected phagocytes. In parallel, the expression of haem oxygenase 1 and of the siderophore-binding peptide lipocalin 2 was markedly enhanced following pathogen entry. Collectively, these modulations reduced both the cytoplasmatic labile iron and the ferritin storage iron pool within macrophages, thus restricting the acquisition of iron by intramacrophage Salmonella. Correspondingly, limitation of macrophage iron decreased microbial survival, whereas iron supplementation impaired immune response pathways in Salmonella-infected macrophages (nitric oxide formation and tumour necrosis factor-alpha production) and promoted intracellular bacterial proliferation. Our findings suggest that the enhancement of ferroportin 1-mediated iron efflux, the upregulation of the haem-degrading enzyme haem oxygenase 1 and the induction of lipocalin 2 following infection concordantly aim at withholding iron from intracellular S. typhimurium and to increase antimicrobial immune effector pathways thus limiting pathogen proliferation.

Paracoccidioidomicose na criança / Paracoccidioidomycosis in children

No presente trabalho estuda-se a Paracoccidioidomicose na criança, que tem sido assunto frequente de estudo nomeio universitário, devido a sua incidência relativamente elevada, constituindo-se, mesmo numa patologia regional de destaque. O objetivo principal foi situar a real participação da criança segundo a concepção atual da história natural da doença. Para tanto fez-se uma abrangente revisão da literatura, comparando-se os aspectos epidemiológicos, clínicos e imunopatológicos com o estudo retrospectivo dos prontuários de 30 pacientes menores de 12 anos de idade, portadores de paracoccidioidomicose, confirmada pelo encontro do parasito em exames diretos e internados em 3 hospitais de referência de Goiânia, no período de janeiro de 1970 a julho de 1990. Os dados dos prontuários foram sintetizados em 17 tabelas. Dos resultados definiram-se 3 grupos de formas clínicas: Linfático abdominal; Linfático hepatoesplênica e Linfática; dos exames específicos: no hemograma a anemia e a eosinofilia foram achados marcantes, na eletroforese de proteínas séricas a hipoalbuminemia e hiperglobulinemia às custas de gama foram evidenciadas; as sorologias e as intradermorreações foram provas que denunciaram o envolvimento do sistema imune com elevação da imunidade humoral e depleção da imunidade celular; dos exames radiológicos (torax, trânsito e enema)e ultrossonográficos do abdomen verifica-se que o pulmão é raramente comprometido dos RX do torax realizados; o trânsito intestinal detecta lesões da mucosa, sugestivas do envolvimento dos linfáticos da parede; a USG é um recurso diagnóstico não invasivo as massas intrabdominais e pode sugerir fibrose hepática

Reactive oxygen intermediates inactivate Mycobacterium leprae in the phagocytes from human peripheral blood.

Reactive oxygen intermediates such as hydrogen peroxide, superoxide, and hydroxyl radicals are important microbicidal components, and they could also play a role in an infection with Mycobacterium leprae. A comparative study of the level of hydrogen peroxide and superoxide produced by peripheral blood phagocytes from normal healthy individuals and lepromatous leprosy patients showed a deficiency in superoxide production in the patients. In the phagocytes from normal healthy individuals, there was good release of superoxide ions, and this mediated the killing of M. leprae. The lack of superoxide production allowed the viability of M. leprae inside the macrophages from leprosy patients. This deficiency could be rectified by the use of an immunomodulator, the delipidified cell wall of M. leprae. This modulation resulted in the ability of the patients' phagocytes to respond to M. leprae, to produce reactive oxygen intermediates such as superoxide, and also to kill the bacteria. These observations indicate that delipidified cell wall could have significant potential to positively modulate the immune-deficient cells of leprosy patients.

The effect of antibiotics on the growth of Legionella pneumophila in guinea-pig alveolar phagocytes infected in vivo by an aerosol.

The likely efficacy of four antibiotics of possible use in the treatment of Legionnaires' disease was assessed in terms of their capacity to inhibit the replication of Legionella pneumophila within guinea-pig alveolar macrophages and polymorphonuclear leukocytes (PMN) compared with their minimum inhibitory concentrations (MIC) in vitro. All the antibiotics used had similar MIC values with regard to L. pneumophila (0.032-0.062 mg/l), but differences of up to 100 fold in the concentration required to eliminate viable intracellular organisms were observed. The most effective antibiotics were found to be rifampicin and ciprofloxacin. These eliminated viable L. pneumophila from alveolar macrophages and PMN at concentrations of 0.005 and 0.01 mg/l respectively, whereas erythromycin and gentamicin required higher concentrations of 0.1 and 0.5 mg/l respectively. It is suggested that assays performed in cultures of relevant cells may provide useful additional means of assessing antibiotic efficacy against intracellular pathogens and help to explain discrepancies otherwise observed between in vitro and in vivo findings.

Erythromycin: a microbial and clinical perspective after 30 years of clinical use (1).

Erythromycin is a macrolide that acts by inhibiting the translocation reaction during protein synthesis. Erythromycin is inactive against the Enterobacteriaceae and Pseudomonas aeruginosa except under alkaline conditions. Erythromycin is active against most gram-positive bacteria; some gram-negative bacteria, including Neisseria, Bordetella, Brucella, Campylobacter, and Legionella; and Treponema, Chlamydia, and Mycoplasma. The emergence of resistance to erythromycin is closely associated with its use and is often plasmid mediated. After its oral or parenteral administration, erythromycin diffuses readily into intracellular fluids and is actively concentrated intracellularly by polymorphonuclear leukocytes and alveolar macrophages.

The antimicrobial activity of rifampin: emphasis on the relation to phagocytes.

Rifampin appears to be a uniquely effective antibiotic for the treatment of certain infections. Only rifampin, of nine antimicrobial agents studied, killed staphylococci inside polymorphonuclear neutrophils or Escherichia coli inside macrophages. Rifampin, unlike penicillin, penetrated living, intact neutrophils. Rifampin was more effective than other antistaphylococcal antibiotics for the treatment of staphylococcal infections induced by subcutaneous, intraperitoneal, or intravenous injection of the organisms into mice. Rifampin-resistant variants occur in low numbers among clinical isolates (frequency, 10(-8)), and some of these variants are diminished in virulence for mice. Treatment of infected mice with rifampin plus another antimicrobial agent prevented the emergence of resistant variants. Thus, there is firm in vitro and in vivo support for studies of the clinical efficacy of rifampin for staphylococcal infections.