Effects of antimicrobial photodynamic therapy with photodithazine® on methicillin-resistant Staphylococcus aureus (MRSA): Studies in biofilms and experimental model with Galleria mellonella.

Staphylococcus aureus infections are a severe health problem due to the high mortality rate. Conventional treatment of these infections is via the administration of antibiotics. However, its indiscriminate use can select resistant microorganisms. Thus, it is necessary to develop alternatives for antibiotic therapy. Antimicrobial Photodynamic Therapy (aPDT), a therapeutic method that associates a photosensitizer (PS), a light source with adequate wavelength to the PS, interacts with molecular oxygen generating reactive oxygen species responsible for cell inactivation, is a viable alternative. This work aimed to analyze, in vitro and in vivo, the action of aPDT with PS Photodithazine® (PDZ) on the methicillin-resistant S. aureus (MRSA) strain. In the in vitro method, the S. aureus biofilm was incubated with PDZ at 50 and 75 µg.mL-1 for 15 min, adopting the light dose of 25, 50, and 100 J/cm2. In addition, PS interaction, formation of reactive oxygen species (ROS), bacterial metabolism, adhesion, bacterial viability, and biofilm structure were evaluated by scanning electron microscopy. Subsequently, the strain was inoculated into models of Galleria mellonella, and the survival curve, health scale, blood cell analysis, and CFU/mL of S. aureus in the hemolymph were analyzed after aPDT. In the in vitro results, bacterial reduction was observed in the different PDZ concentrations, highlighting the parameters of 75 µg.mL-1 of PDZ and 100 J/cm2. As for in vivo results, aPDT increased survival and stimulated the immune system of G. mellonella infected by S. aureus. aPDT proved effective in both models, demonstrating its potential as an alternative therapy in treating MRSA bacterial infections.

Potencialidades fitoquímicas do melão (Cucumis melo L. ) na região Noroeste do Rio Grande do Sul - Brasil / Phytochemical potentialities of melon (Cucumis melo L. ) in the northwest region of Rio Grande do Sul - Brazil

O melão (Cucumis melo L.) é uma fruta muito apreciada por suas qualidades e sua produção vem crescendo e ganhando espaço no mercado nacional e internacional. Em regiões como o Noroeste do Rio Grande do Sul, destaca-se como uma nova alternativa de renda para vários agricultores. Neste contexto, o presente trabalho teve como objetivo analisar o potencial fitoquímico de alguns cultivares de melão da região Noroeste do Rio Grande do Sul. A análise fitoquímica utilizando como farmacógeno as folhas, foi realizada para a verificação da presença de metabólitos secundários, tais como: saponinas, cumarinas, cardiotônicos, cianogenéticos, alcalóides, taninos, antraquinonas, flavonoides, e óleos voláteis. Também foi avaliado o teor de suco a partir dos frutos. Dentre os cinco cultivares analisados, Gaúcho, Imperial, Hy Mark, Magelan, e Cantaloupe, o cultivar Gaucho apresentou a maior variedade em metabólitos secundários. Na avaliação do teor de suco a cultivar Magelan se destacou em comparação às demais cultivares testadas.

The melon (Cucumis melo L.) is a fruit highly appreciated for its qualities and its production has been growing and gaining space in the national and in the international market. In regions like the northwest of Rio Grande do Sul - Brazil, it stands out as a new income alternative for farmers. In this context, this study aimed to analyze the phytochemical potential of some melon cultivars in the northwest region of Rio Grande do Sul. The phytochemical analysis, using the leaves as pharmacogen, was performed to verify the presence of secondary metabolites such as saponins, coumarins, cardiac glycosides, cyanogenetic glicosides, alkaloids, tannins, anthraquinones, flavonoids and volatile oils. The juice content from the fruits was also evaluated. Among the five analyzed cultivars, Gaucho, Imperial, Hy Mark, Magelan and Cantaloupe, cultivar Gaucho had the greatest variety of secondary metabolites. In the assessment of the juice content, cultivar Magelan was superior compared to the remaining tested cultivars.

CD4+ T cells from HIV-1-infected patients recognize wild-type and mutant human immunodeficiency virus-1 protease epitopes.

Human immunodeficiency virus (HIV)-1 protease is a known target of CD8+ T cell responses, but it is the only HIV-1 protein in which no fully characterized HIV-1 protease CD4 epitopes have been identified to date. We investigated the recognition of HIV-1 protease by CD4+ T cells from 75 HIV-1-infected, protease inhibitor (PI)-treated patients, using the 5,6-carboxyfluorescein diacetate succinimidyl ester-based proliferation assay. In order to identify putative promiscuous CD4+ T cell epitopes, we used the TEPITOPE algorithm to scan the sequence of the HXB2 HIV-1 protease. Protease regions 4-23, 45-64 and 73-95 were identified; 32 sequence variants of the mentioned regions, encoding frequent PI-induced mutations and polymorphisms, were also tested. On average, each peptide bound to five of 15 tested common human leucocyte antigen D-related (HLA-DR) molecules. More than 80% of the patients displayed CD4+ as well as CD8+ T cell recognition of at least one of the protease peptides. All 35 peptides were recognized. The response was not associated with particular HLA-DR or -DQ alleles. Our results thus indicate that protease is a frequent target of CD4+ along with CD8+ proliferative T cell responses by the majority of HIV-1-infected patients under PI therapy. The frequent finding of matching CD4(+) and CD8+ T cell responses to the same peptides may indicate that CD4+ T cells provide cognate T cell help for the maintenance of long-living protease-specific functional CD8+ T cells.

Expression and diversity of Echinococcus multilocularis AgB genes in secondarily infected mice: evaluating the influence of T-cell immune selection on antigenic variation.

The T-cell-mediated immune response exhibits a crucial function in the control of the intrahepatic proliferation of Echinococcus multilocularis larvae in mice and humans, both being natural intermediate hosts of the parasite. Antigen B (AgB), a metabolized Echinococcus spp. lipoprotein, contributes to the modulation of the T-cell immune response, and distinct sites of the corresponding AgB1, AgB3 and AgB4 genes were shown to be under positive selection pressure. Since several AgB gene variants are present in a single Echinococcus metacestode, we used secondary E. multilocularis infections in BALB/c and in athymic nude mice (devoid of T-cell responses) to analyze the effect of the cellular immune response on the expression and diversity of EmAgB1-EmAgB4 genes. We demonstrated hereby that EmAgB transcripts were less abundant in nude mice during the early phase of infection (at one month post-infection), and that EmAgB2 is simultaneously down-regulated when compared to the other three genes. A negative relationship exists between the level of transcription and diversity of EmAgB genes. Moreover, no excess of non-synonymous substitutions was found among the distinct EmAgB alleles from a single host. Together, these results pointed to the effect of purifying selection, which seemed to eliminate the detrimental AgB variants generated during the development of the metacestode within the peritoneal cavity of its intermediate host.

Redundancy and recombination in the Echinococcus AgB multigene family: is there any similarity with protozoan contingency genes?

Numerous genetic variants of the Echinococcus antigen B (AgB) are encountered within a single metacestode. This could be a reflection of gene redundancy or the result of a somatic hypermutation process. We evaluate the complexity of the AgB multigene family by characterizing the upstream promoter regions of the 4 already known genes (EgAgB1-EgAgB4) and evaluating their redundancy in the genome of 3 Echinococcus species (E. granulosus, E. ortleppi and E. multilocularis) using PCR-based approaches. We have ascertained that the number of AgB gene copies is quite variable, both within and between species. The most repetitive gene seems to be AgB3, of which there are more than 110 copies in E. ortleppi. For E. granulosus, we have cloned and characterized 10 distinct upstream promoter regions of AgB3 from a single metacestode. Our sequences suggest that AgB1 and AgB3 are involved in gene conversion. These results are discussed in light of the role of gene redundancy and recombination in parasite evasion mechanisms of host immunity, which at present are known for protozoan organisms, but virtually unknown for multicellular parasites.

Rhodococcus equi pneumonia in AIDS: high-resolution CT findings in five patients.

The aim of this study was to describe the high-resolution CT scan findings in five patients with AIDS and pulmonary infection due to Rhodococcus equi. The study included five patients with AIDS and proven R. equi infection. The CT scans were reviewed by two observers. The patients included four men and one woman ranging from 39 years to 49 years in age (mean 42 years). The findings included areas of consolidation (n=5) with single (n=1) or multiple cavitation (n=4), ground-glass opacities (n=5), centrilobular nodules (n=3), small centrilobular nodular opacities (n=3) and "tree in bud" opacities (n=3). None of the patients had pleural effusion or lymph node enlargement. The most common high-resolution CT manifestations of R. equi infection consist of areas of consolidation with cavitation, ground-glass opacities, nodules and a tree-in-bud pattern.

Silicoproteinosis: high-resolution CT and histologic findings.

The high-resolution CT findings of silicoproteinosis consist of numerous bilateral centrilobular nodular opacities, focal ground glass opacities, and patchy areas of consolidation. These findings reflect the presence of intra-alveolar accumulation of proteinaceous material.

Chronic pulmonary paracoccidioidomycosis (South American blastomycosis): high-resolution CT findings in 41 patients.

OBJECTIVE: To assess the pulmonary parenchymal findings on high-resolution CT in 41 patients with the chronic form of paracoccidioidomycosis (South American blastomycosis). SUBJECTS AND METHODS: The study included 41 consecutive patients in whom chronic paracoccidioidomycosis had been proven. All patients underwent high-resolution CT (1-mm collimation, high-spatial-frequency reconstruction algorithm) at 12 equally spaced intervals through the chest. The images were analyzed by two radiologists, and each final decision was reached by consensus. RESULTS: Thirty-eight (93%) of the 41 patients had CT scans with abnormal findings. The findings included interlobular septal thickening in 36 patients (88%), 1-25 mm diameter nodules in 34 (83%), peribronchovascular interstitial thickening in 32 (78%), centrilobular opacities in 26 (63%), intralobular lines in 24 (59%), ground-glass opacities in 14 (34%), cavitation in seven (17%), air-space consolidation in five (12%), traction bronchiectasis in 34 (83%), and paracicatricial emphysema in 28 (68%). In approximately 90% of patients, the abnormalities were bilateral and symmetrical and involved all lung zones. CONCLUSION: High-resolution CT findings of paracoccidioidomycosis consist predominantly of interstitial abnormalities and nodules associated with traction bronchiectasis and paracicatricial emphysema in a bilaterally symmetrical distribution.

Distinct prevalence of antibodies to the E2 protein of GB virus C/hepatitis G virus in different parts of the world.

Since the identification of the new human virus, GB virus C (GBV-C)/hepatitis G-virus (HGV), in 1995/1996, reverse transcription polymerase chain reaction remained the sole available diagnostic tool for GBV-C/HGV infection. Recently, a serologic test based on the detection of antibodies to the putative envelope protein 2 (anti-E2) has been introduced. We used this assay for a seroepidemiological survey including 3,314 healthy individuals from different parts of the world, 123 patients from Germany who were suspected to have an increased risk of acquiring GBV-C/HGV infection, 128 multiple organ donors, and 90 GBV-C/HGV RNA positive persons. In European countries, anti-E2 seropositivity ranged from 10.9% (Germany) to 15.3% (Austria). In South Africa (20.3%) and Brazil (19.5%), even higher anti-E2 prevalence rates were recorded. In Asian countries like Bhutan (3.9%), Malaysia (6.3%), and the Philippines (2.7%), anti-E2 positivity was significantly lower. GBV-C/HGV anti-E2 prevalence in potential "risk groups," i.e., patients on hemodialysis and renal transplant recipients, did not vary significantly from anti-E2 seroprevalence in German blood donors. Anti-E2 and GBV-C/HGV RNA were found to be mutually exclusive, confirming the notion that anti-E2 has to be considered as a marker of past infection.

Measurement of ventilation in children using the respiratory inductive plethysmograph.

A new technique to measure ventilation, based on the separate contributions of rib cage and abdomen to tidal volume, the respiratory inductive plethysmograph was evaluated in 20 healthy children. The accuracy of the method was determined by simultaneously measuring tidal volume with a pneumotachograph in the standing, sitting, supine, left lateral decubitus, and prone postures. Comparison of these two techniques showed mean correlation coefficients greater than 0.96, mean slopes between 0.98 and 1.11, and mean SEE of less than 8% in all postures studied. Breathing through a mouthpiece connected to a pneumotachograph resulted in a substantial change in the pattern of breathing and a mean increase in tidal volume of 32% (P less than 0.05). In the standing and sitting postures, rib cage contribution to tidal volume was predominant (greater than 65%) whereas in the recumbent postures abdominal contribution was predominant (greater than 61%). We conclude that the RIP is an accurate means of measuring ventilation in children and that it avoids the artifacts caused by using a conventional respiratory measuring apparatus.

The consequences of diaphragmatic muscle fatigue in the newborn infant.

We have previously demonstrated that diaphragmatic muscle fatigue can be diagnosed in infants from spectral frequency analysis of the surface diaphragmatic electromyogram. This requires a digital computer, but the analysis takes several days. Spectral frequency changes, however, can be accurately reflected by band pass filtering and expressing the ratio of high-frequency power to low-frequency power. A fall in this ratio of greater than 20% indicates muscle fatigue. Using a simple analog device to obtain this ratio permits the results to be immediately available; we have used this method to study weaning from mechanical ventilators in ten infants. With a successful weaning step there is no significant change in the ratio, whereas an unsuccessful weaning step invariably leads to a decrease in the ratio of greater than 20%, which precedes CO2 retention and clinical deterioration. These data indicate that diaphragmatic muscle fatigue plays an important role in the infant's response to lung disease. Monitoring of the high/low frequency ratio may be helpful in weaning infants from assisted ventilation.