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Scedosporium/Lomentospora complex is composed of filamentous fungi, including some clinically relevant species, such as Pseudallescheria boydii , Scedosporium aurantiacum, and Scedosporium apiospermum . Glucosylceramide (GlcCer), a conserved neutral glycosphingolipid, has been described as an important cell surface molecule playing a role in fungal morphological transition and pathogenesis. The present work aimed at the evaluation of GlcCer structures in S. aurantiacum and Pseudallescheria minutispora , a clinical and an environmental isolate, respectively, in order to determine their participation in fungal growth and host-pathogen interactions. Structural analysis by positive ion-mode ESI-MS (electrospray ionization mass spectrometer) revealed the presence of different ceramide moieties in GlcCer in these species. Monoclonal antibodies against Aspergillus fumigatus GlcCer could recognize S. aurantiacum and P. minutispora conidia, suggesting a conserved epitope in fungal GlcCer. In addition, these antibodies reduced fungal viability, enhanced conidia phagocytosis by macrophages, and decreased fungal survival inside phagocytic cells. Purified GlcCer from both species led to macrophage activation, increasing cell viability as well as nitric oxide and superoxide production in different proportions between the two species. These results evidenced some important properties of GlcCer from species of the Scedosporium/Lomentospora complex, as well as the effects of monoclonal anti-GlcCer antibodies on fungal cells and host-pathogen interaction. The differences between the two species regarding the observed biological properties suggest that variation in GlcCer structures and strain origin could interfere in the role of GlcCer in host-pathogen interaction., Competing Interests: The authors declare no conflict of interest.

Secondary metabolites isolated from Simira eleiezeriana and Simira glaziovii were evaluated against herpes simplex virus (HSV-1) and (HSV-2). The 50% effective concentrations values (EC 50 ) were calculated from the dose-response curve and the selectivity index (SI) against the virus. The physicochemical data LogP, (PSA), (NRB), (HBA) and (HBD) were obtained using Marvin Sketch. Among the tested compounds, conipheraldeyde, harman and simirane A showed better results with EC 50 6.39; 4.90; 4.61 µg/mL and SI 78.3; 11.8; 7.01, respectively, for HSV-1, and EC 50 41.2; 71.8; 3.73 µg/mL and SI 12.1; 24.7; 8.7, respectively, for HSV-2. The percentage of inhibition (PI) obtained for HSV-1 were higher than 60%, and for HSV-2 these compounds showed PI > 90%. The physical chemical data showed that the most active compounds satisfy the attributes for drugs with good oral bioavailability.

Context: Ocimum basilicum L. (Lamiaceae) has been used in folk medicine to treat headaches, kidney disorders, and intestinal worms., Objective: This study evaluates the anti-cryptococcal activity of ethanol crude extract and hexane fraction obtained from O. basilicum var. Maria Bonita leaves., Materials and Methods: The MIC values for Cryptococcus sp. were obtained according to Clinical and Laboratory Standards Institute in a range of 0.3-2500 μg/mL. The checkerboard assay evaluated the association of the substances tested (in a range of 0.099-2500 μg/mL) with amphotericin B and O. basilicum essential oil for 48 h. The ethanol extract, hexane fraction and associations in a range of 0.3-2500 μg/mL were tested for pigmentation inhibition after 7 days of treatment. The inhibition of ergosterol synthesis and reduction of capsule size were evaluated after the treatment with ethanol extract (312 μg/mL), hexane fraction (78 μg/mL) and the combinations of essential oil + ethanol extract (78 μg/mL + 19.5 μg/mL, respectively) and essential oil + hexane fraction (39.36 μg/mL + 10 μg/mL, respectively) for 24 and 48 h, respectively., Results: The hexane fraction presented better results than the ethanol extract, with a low MIC (156 μg/mL against C. neoformans T 444 and 312 μg/mL against C. neoformans H99 serotype A and C. gattii WM779 serotype C). The combination of the ethanol extract and hexane fraction with amphotericin B and essential oil enhanced their antifungal activity, reducing the concentration of each substance needed to kill 100% of the inoculum. The substances tested were able to reduce the pigmentation, capsule size and ergosterol synthesis, which suggest they have important mechanisms of action., Conclusions: These results provide further support for the use of ethanol extracts of O. basilicum as a potential source of antifungal agents.

Kalanchoe daigremontiana (Crassulaceae) is a medicinal plant native to Madagascar. The aim of this study was to investigate the flavonoid content of an aqueous leaf extract from K. daigremontiana (Kd), and assess its antiherpetic potential. The major flavonoid, kaempferol 3-O-β-d-xylopyranosyl-(1 → 2)-α-l-rhamnopyranoside (1), was isolated from the AcOEt fraction (Kd-AC). The BuOH-soluble fraction afforded quercetin 3-O-β-d-xylopyranosyl-(1 → 2)-α-l-rhamnopyranoside (2) and the new kaempferol 3-O-β-d-xylopyranosyl-(1 → 2)-α-l-rhamnopyranoside-7-O-β-d-glucopyranoside (3), named daigremontrioside. The crude extract, Kd-AC fraction, flavonoids 1 and 2 were evaluated using acyclovir-sensitive strains of HSV-1 and HSV-2. Kd-AC was highly active against HSV-1 (EC 50  = 0.97 μg/ml, SI > 206.1) and HSV-2 (EC 50  = 0.72 μg/ml, SI > 277.7). Flavonoids 1 and 2 showed anti-HSV-1 (EC 50  = 7.4 μg/ml; SI > 27 and EC 50  = 5.8 μg/ml; SI > 8.6, respectively) and anti-HSV-2 (EC 50  = 9.0 μg/ml; SI > 22.2 and EC 50  = 36.2 μg/ml; SI > 5.5, respectively) activities, suggesting the contribution of additional substances to the antiviral activity., (© 2016 Wiley-VHCA AG, Zurich, Switzerland.)

Aureocin A70 is the only four-component bacteriocin described to date. As it inhibits the growth of a wide range of Gram-positive bacteria, including Listeria monocytogenes strains isolated from food, its potential for improving food safety was investigated in this study. Aureocin A70 (10,240AU/mL) proved to be bactericidal, but not extensively lytic, against listerial strains. The antibacterial activity of aureocin A70 (16AU/mL) was then tested in UHT-treated skimmed milk inoculated with the food-associated L. monocytogenes L12 strain (4-log CFU/mL) during storage at 4°C for one week. Aureocin A70 caused a time-dependent reduction in the listerial viable cell counts (5.51-log units) up to 7days of incubation. Aureocin A70 was neither toxic to the Vero and the L-929 cell lines nor exhibited a hemolytic activity against sheep red blood cells. Aureocin A70 proved to be completely stable for one month at 25°C, 16weeks at 4°C and 20weeks at -20°C. Aureocin A70 exhibited a time-dependent susceptibility to simulated gastric juice and bile salts mimicking gastrointestinal conditions. The entrapment of aureocin A70 in an alginate/gelatin matrix revealed that this bacteriocin can be released from this matrix. Moreover, it remained adsorbed to and active on a low-density polyethylene plastic surface suggesting that aureocin A70 may be employed in bioactive packaging to control the growth of undesirable bacteria. Taken together these results suggest that aureocin A70 is a promising alternative to be used in food applications., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Aims: The aim of this study was to investigate the potential of aureocin A53, a staphylococcal antimicrobial peptide, for improving food safety., Methods and Results: The antimicrobial activity of aureocin A53 against strains of Listeria monocytogenes isolated from food was tested and the bacteriocin proved to be bactericidal and bacteriolytic against the listerial strains. Aureocin A53 was neither toxic to eukaryotic cell lines nor haemolytic against sheep erythrocytes. It also exhibited a remarkable stability during storage at different temperatures and sensitivity to both simulated gastric juice and bile salts. When the antibacterial activity of aureocin A53 (256 AU ml(-1) ) was tested in skimmed milk artificially inoculated with a L. monocytogenes strain (1·0 × 10(4)  CFU ml(-1) ) isolated from food, during storage at 4°C, the bacteriocin reduced the viable counts by 7·7-log10 units up to 7 days of incubation, when compared with the controls not treated with the bacteriocin., Conclusions: Aureocin A53 exhibited several features considered important for biopreservation and remained fully active in a food matrix., Significance and Impact of the Study: Taken together, the results confirmed that aureocin A53 has potential to be used as a food preservative, representing an alternative to the use of nisin in biopreservation of dairy products., (© 2016 The Society for Applied Microbiology.)

The aim of this study was to evaluate the activity of the EO and its major components of Ocimum basilicum var. Maria Bonita, a genetically improved cultivar, against the fluconazole sensitive and resistant strains of Candida albicans and Cryptococcus neoformans. Geraniol presented better results than the EO, with a low MIC (76 μg/mL against C. neoformans and 152 μg/mL against both Candida strains). The combination of EO, linalool, or geraniol with fluconazole enhanced their antifungal activity, especially against the resistant strain (MIC reduced to 156, 197, and 38 μg/mL, resp.). The ergosterol assay showed that subinhibitory concentrations of the substances were able to reduce the amount of sterol extracted. The substances tested were able to reduce the capsule size which suggests they have an important mechanism of action. Transmission electron microscopy demonstrated cell wall destruction of C. neoformans after treatment with subinhibitory concentrations. In C. albicans ultrastructure alterations such as irregularities in the membrane, presence of vesicles, and cell wall thickening were observed. The biofilm formation was inhibited in both C. albicans strains at MIC and twice MIC. These results provide further support for the use of O. basilicum EO and its major components as a potential source of antifungal agents.

In this study, we analyze the importance of O-linked oligosaccharides present in peptidorhamnomannan (PRM) from the cell wall of the fungus Scedosporium prolificans for recognition and phagocytosis of conidia by macrophages. Adding PRM led to a dose-dependent inhibition of conidia phagocytosis, whereas de-O-glycosylated PRM did not show any effect. PRM induced the release of macrophage-derived antimicrobial compounds. However, O-linked oligosaccharides do not appear to be required for such induction. The effect of PRM on conidia-induced macrophage killing was examined using latex beads coated with PRM or de-O-glycosylated PRM. A decrease in macrophage viability similar to that caused by conidia was detected. However, macrophage killing was unaffected when beads coated with de-O-glycosylated PRM were used, indicating the toxic effect of O-linked oligosaccharides on macrophages. In addition, PRM triggered TNF-α release by macrophages. Chemical removal of O-linked oligosaccharides from PRM abolished cytokine induction, suggesting that the O-linked oligosaccharidic chains are important moieties involved in inflammatory responses through the induction of TNF-α secretion. In summary, we show that O-glycosylation plays a role in the recognition and uptake of S. prolificans by macrophages, killing of macrophages and production of pro- inflammatory cytokines.

Objective: The aim of this study was to assess the in vitro cytotoxicity of acrylic resins of different colors over time., Methods: Specimens were divided into 4 groups (n = 6) according to the color of the acrylic resin (Orto Class, Clássico, Campinas, São Paulo, Brazil): Group 1, clear acrylic resin; Group 2, pink acrylic resin; Group 3, blue acrylic resin; and Group 4, green acrylic resin. All specimens were fabricated according to the mass manipulation technique and submitted to mechanical polishing protocol. The control was performed with an amalgam specimen (C+), a glass specimen (C-) and cell control (CC). Specimens were immersed in Minimum Eagle's Medium (MEM) and incubated for 24 h at 37ºC. The extracts from the experimental material were filtered and mixed with L929 fibroblast. Cytotoxicity was evaluated at four different times, 24, 48, 72 and 168 h. After contact, cells were incubated for 24 h and added to 100 µ of 0.01% neutral red dye. The cells were incubated for 3 h for pigment incorporation and fixed. Cells viability was determined by a spectroscopic (BioTek, Winooski, Vermont, USA) with a 492-nm wavelength λ=492 nm)., Results: There were no statistical differences between the experimental groups and the CC and C- groups., Conclusion: Clear, pink, blue and green self-curing acrylic resins fabricated by means of the mass manipulation technique and mechanically polished are not cytotoxic. Neither the pigment added to the self-curing acrylic resin nor the factor of time influenced the cytotoxicity of the material.

The anti-HMPV (human metapneumovirus) activity was determined for sulfated dl-hybrid galactans obtained from the red seaweed Cryptonemia seminervis and their depolymerized products obtained by reductive partial hydrolysis. Structural studies carried out in three homogeneous depolymerized fractions DS-1, DS-2e and DS-3 (Mw of 51.6-63.8 kDa) showed that these galactans present different chemical characteristics, as monosaccharide composition, content of sulfate groups (14.1-29.9%) and agaran:carrageenan molar ratio diads, 2.7:1 for DS-1 and DS-2e and 1:1 for DS-3. The sulfate groups are located principally on C-2 of β-d-galactopyranose and 4,6-O-(1'-carboxyethylidene)-β-d-galactopyranose residues and on C-6 of α-galactose residues. Sulfated dl-galactans and their depolymerized products exhibited antiviral activity at a very early stage of the viral infection cycle. All fractions, except DS-2e inhibited HMPV replication by binding to the viral particle. Besides depolymerized galactans DS-2e and DS-3 inhibited the recognition of cell receptor by HMPV and penetration to the host cell, respectively., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Ethnopharmacological Relevance: In traditional medicine, teas made from leaves and bark of Gallesia gorazema are used as antispasmodic, anthelmintic, antihemorrhagic and febrifuge agents. Crude leaves of this plant are also employed as a remedy in the treatment of abscesses, orchitis, gonorrhea and for rheumatic pain relief. this study investigates the presumed antinociceptive and anti-inflammatory activities of leaves and roots Gallesia gorazema (Phytolaccaceae) extracts. The most active extract and its isolated compound, a new natural product, are also evaluated against viruses HSV-1 and HSV-2., Materials and Methods: In vivo experiments with mice were used to assess the analgesic and anti-inflammatory activities of Gallesia gorazema extracts. Antiviral activity of extracts and the new natural product was investigated by in vitro experiments., Results: Results show that dichloromethanic root (DRE) and ethanolic leaf (ELE) extracts displayed significant antinociceptive and anti-inflammatory activities in in vivo experiments with mice. Both extracts were also assayed against the herpes simplex viruses HSV-1 and HSV-2, but only DRE was highly active, showing a selective antiviral effect against HSV-1. Phytochemical fractionation of DRE led to the isolation of 28-hydroxyoctacosyl ferulate, a novel natural product, which displayed strong antiviral activity against HSV-1 (EC₅₀=21.6 μg/mL) with a selective index above 9, justifying, at least in part, the high selective antiviral activity observed for DRE., Conclusion: These results suggest that the plant Gallesia gorazema is a potential candidate for the development of novel anti-herpetic phytomedicines., (© 2013 Elsevier Ireland Ltd. All rights reserved.)

Total lipids from the Brazilian brown seaweed Sargassum vulgare were extracted with chloroform/methanol 2:1 and 1:2 (v/v) at room temperature. After performing Folch partition of the crude lipid extract, the lipids recovered from the Folch lower layer were fractionated on a silica gel column eluted with chloroform, acetone and methanol. The fraction eluted with methanol, presented a strong orcinol-positive band characteristic of the presence of sulfatides when examined by TLC. This fraction was then purified by two successive silica gel column chromatography giving rise to fractions F4I86 and F4II90 that exhibited strong activity against herpes simplex virus type 1 and 2. The chemical structures present in both fractions were elucidated by ESI-MS and ¹H/¹³C NMR analysis HSQC fingerprints based on their tandem-MS behavior as Sulfoquinovosyldiacylglycerols (SQDGs). The main SQDG present in both fractions and responsible for the anti-herpes activity observed was identified as 1,2-di-O-palmitoyl-3-O-(6-sulfo-α-D-quinovopyranosyl)-glycerol.

This study evaluated the influence of the manipulation technique and polishing method on the flexural strength and cytotoxicity of acrylic resins. Two manipulation techniques and three polishing methods were used in the fabrication of acrylic plates that were divided into 6 groups (n=10). Groups MM, MC and MW: mass technique with mechanical polishing, chemical polishing and without polishing, respectively; and Groups SM, SC and SW: Saturation technique with mechanical polishing, chemical polishing and without polishing, respectively). Flexural strength was tested in a universal testing machine and the cytotoxicity assay used cell cultures (L-929) for periods of 24 h to 168 h. Flexural strength and cytotoxicity data were assessed using two-way and three-way ANOVA, respectively (α=0.05), followed by post hoc Bonferroni test for multiple comparisons. The effect of combinations of manipulation techniques and polishing methods on flexural strength showed significant differences only between Group SC and Groups MW, MM and MC (p<0.01). Cell viability ranged from 51% (3.9%) to 87,6% (3.2) in the 24-h time interval, and from 87.8% (5.0) to 95.7% (3.1%) in the 168-h time interval. With the increase of cell viability, from the third day (72 h), there was no significant difference among the groups, except between MM and SC (p<0.01) at 72 h. In conclusion, the manipulation technique and polishing method had more influence on the cytotoxicity than on flexural strength.

Objectives: An aqueous extract and fraction from the marine sponge Petromica citrina have antibacterial activity. We performed a chemical and biological characterization of the antibiotic substance from P. citrina and investigated its mode of action on Staphylococcus aureus cells., Methods: The inhibitory activity of the aqueous extract of P. citrina was determined against 14 bacteria belonging to type strains and clinical antibiotic-resistant strains. The aqueous extract was fractionated under bioassay guidance and the bioactive substance was identified by its (1)H-NMR, (13)C-NMR and mass spectra. The MIC and the MBC of this substance were determined. This substance was also subjected to cytotoxic bioassays. The mode of action on S. aureus cells was investigated by light and transmission electron microscopy analysis., Results: P. citrina showed a large spectrum of activity against type strains and resistant-bacteria such as S. aureus, Staphylococcus epidermidis, Enterococcus faecalis, Mycobacterium fortuitum and Neisseria gonorrhoeae. The aqueous extract was fractionated and halistanol trisulphate (24ε,25-dimethylcholestane-2β,3α,6α-triol trisodium sulphate) was isolated for the first time from P. citrina. Halistanol trisulphate had a bactericidal effect on exponentially growing S. aureus cells at the MIC (512 mg/L). Cytotoxicity biossays showed moderate toxicity against cancer cell line L929 (fibrosarcoma). This substance apparently acts by damaging the cell membrane, with subsequent cell lysis., Conclusions: Halistanol trisulphate is a broad-spectrum antibiotic isolated from P. citrina with a mode of action involving disruption of the cytoplasmic membrane. It is a new candidate for research on antibacterial substances.

Objective: To evaluate the influence of pH levels on interarch elastics with regard to force decay and cytotoxicity., Materials and Methods: One nonlatex (NLAO) group and one latex (LAO) group were tested (n  =  10). Elastics were stretched to 25 mm and were held for 1, 6, 12, and 24 hours in artificial saliva solutions with pH levels of 5.0, 6.0, and 7.5. Force magnitudes were measured at 25 mm of activation. The cytotoxicity assay was performed using cell cultures (L929 mouse fibroblast cell line), which were subjected to the cell viability test with neutral red ("dye-uptake"). Force decay and cytotoxicity were assessed using analysis of variance, the Sidak method, and a Tukey's test., Results: The interactions between group, pH, and time showed no statistically significant differences (P  =  .29). When pH per time (P  =  .032) and group per time (P  =  .0009) were considered, these interactions showed statistically significant differences (P < .05). The pH did not interfere directly in the degradation results of the tested elastics. The cytotoxicity test showed that group LAO presented lower cell viability when compared with group NLAO over the course of the entire experiment. There was a gradual reduction in cell viability from 1 hour to 24 hours. A significant difference (P < .05) was found between the interactions group pH and the control group of cells, except between group NLAO at the time point of 1 hour at different pH values and at the time points of 6 and 12 hours with pH 5 (P > .05)., Conclusions: No significant correlation between pH, force decay, and cytotoxicity was observed.

The objective of the present study was to evaluate the cytotoxicity and degree of monomer conversion of resin-reinforced glass ionomer cements (RGIC) over different time periods. Four RGICs: Fuji Ortho LC (FOLC), Fuji Ortho Band (FOB), Orthoglass (OGL), and Multicure Glass Ionomer (MCI) were evaluated for cytotoxicity in fibroblastic L929 cells and for their degree of monomer conversion over different time periods. Three control groups were also analysed: positive control (C+), consisting of Tween 80 cell detergent; negative control (C-), consisting of phosphate-buffered saline; and cell control (CC), consisting of cells exposed to any material. To evaluate the cytotoxicity, the dye-uptake technique was used and the degree of conversion was evaluated using infrared spectroscopy. The data obtained were analysed by analysis of variance and the Tukey's test. The results showed cytotoxicity of the RGICs at 1 and 24 hours; the viability values of these materials were statistically different from the C- and CC groups (P < 0.05). After 48 hours, the FOLC group was statistically similar to the CC and C- groups but different from the others. At 1 hour, there was no difference in the degree of conversion between the FOLC and OGL groups (P > 0.05) or between the FOB and MCI (P < 0.05) groups. However, at 48 hours, the FOLC group had greater conversion values than the other groups (P < 0.05). There is a direct relationship between the degree of conversion and RGIC cytotoxicity. Following initial polymerization, cytotoxicity decreases and, consequently, the degree of conversion of the material increases.

Bovine mastitis remains worldwide a major challenge for the dairy industry despite the widespread implementation of control strategies. The increasing number of coagulase-negative staphylococci (CNS) causing mastitis and of bacteria resistant to conventional antibiotics has become a serious problem in recent years. Marine sponges are a rich source of bioactive compounds, and many species can be useful for the development of new antimicrobial drugs. In the present study, 49 CNS strains were isolated from bovine mastitis cases from 21 different dairy herds kept at farms in Southeast Brazil. Strains were analyzed for antimicrobial susceptibility and mecA gene detection. Fifty-nine percent of the CNS strains were resistant to at least one of the drugs tested and 12.2% were classified as multiresistant. Three strains carried the mecA gene, confering resistance to the beta-lactamic antibiotics. In addition, the CNS strains were submitted to in vitro screening for antimicrobial activities of extracts from marine sponges. Extracts from the sponge species Cinachyrella sp., Haliclona sp. and Petromica citrina showed antibacterial activity against 61% of the CNS strains, including strains resistant to conventional antibiotics. Extracts from P. citrina showed the largest spectrum of inhibitory activity. The aqueous extract inhibited 51% of the CNS strains and presented a bactericidal effect over susceptible and multiresistant-bacteria at a minimal inhibitory concentration of 1.024μg/ml. This study shows the potential of marine sponges as new sources of antibiotics and disinfectants for the control of CNS involved in bovine mastitis., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Pseudallescheria boydii is a filamentous fungus that causes a wide array of infections that can affect practically all the organs of the human body. The treatment of pseudallescheriosis is difficult since P. boydii exhibits intrinsic resistance to the majority of antifungal drugs used in the clinic and the virulence attributes expressed by this fungus are unknown. The study of the secretion of molecules is an important approach for understanding the pathogenicity of fungi. With this task in mind, we have shown that mycelial cells of P. boydii were able to actively secrete proteins into the extracellular environment; some of them were recognized by antibodies present in the serum of a patient with pseudallescheriosis. Additionally, molecules secreted by P. boydii induced in vitro irreversible damage in pulmonary epithelial cells. Subsequently, two-dimensional gel electrophoresis combined with mass spectrometry was carried out in order to start the construction of a map of secreted proteins from P. boydii mycelial cells. The two-dimensional map showed that most of the proteins (around 100 spots) were focused at pH ranging from 4 to 7 with molecular masses ranging from 14 to >117 kDa. Fifty spots were randomly selected, of which 30 (60%) were consistently identified, while 20 (40%) spots generated peptides that showed no resemblance to any known protein from other fungi and/or MS with low quality. Notably, we identified proteins involved in metabolic pathways (energy/carbohydrate, nucleotide, and fatty acid), cell wall remodeling, RNA processing, signaling, protein degradation/nutrition, translation machinery, drug elimination and/or detoxification, protection against environmental stress, cytoskeleton/movement proteins, and immunogenic molecules. Since the genome of this fungus is not sequenced, we performed enzymatic and immunodetection assays in order to corroborate the presence of some released proteins. The identification of proteins actively secreted by P. boydii provides important new information for understanding immune modulation and provides important new perspectives on the biology of this intriguing fungus.

Recently, defense peptides that are able to act against several targets have been characterized. The present work focuses on structural and functional evaluation of the peptide analogue Pa-MAP, previously isolated as an antifreeze peptide from Pleuronectes americanus. Pa-MAP showed activities against different targets such as tumoral cells in culture (CACO-2, MCF-7 and HCT-116), bacteria (Escherichia coli ATCC 8739 and Staphylococcus aureus ATCC 25923), viruses (HSV-1 and HSV-2) and fungi (Candida parapsilosis ATCC 22019, Trichophyton mentagrophytes (28d&E) and T. rubrum (327)). This peptide did not show toxicity against mammalian cells such as erythrocytes, Vero and RAW 264.7 cells. Molecular mechanism of action was related to hydrophobic residues, since only the terminal amino group is charged at pH 7 as confirmed by potentiometric titration. In order to shed some light on its structure-function relations, in vitro and in silico assays were carried out using circular dichroism and molecular dynamics. Furthermore, Pa-MAP showed partial unfolding of the peptide changes in a wide pH (3 to 11) and temperature (25 to 95°C) ranges, although it might not reach complete unfolding at 95°C, suggesting a high conformational stability. This peptide also showed a conformational transition with a partial α-helical fold in water and a full α-helical core in SDS and TFE environments. These results were corroborated by spectral data measured at 222 nm and by 50 ns dynamic simulation. In conclusion, data reported here show that Pa-MAP is a potential candidate for drug design against pathogenic microorganisms due to its structural stability and wide activity against a range of targets.

In this paper, we evaluated the antiviral activity against HMPV replication of crude extract of the marine algae Stypopodium zonale and of two meroditerpenoids obtained from it, atomaric acid and epitaondiol, and a methyl ester derivative of atomaric acid. Their selectivity indexes were 20.78, >56.81, 49.26 and 12.82, respectively. Compared to ribavirin, the substances showed a relatively low cytotoxicity on LLC-MK2 cells, with a significant antiviral activity, inhibiting at least 90% of viral replication in vitro, which demonstrates the potential of these marine natural products to combat infections caused by HMPV in vitro.

Background: The Ankle Brachial Index is a useful clinical test for establishing blood supply to the foot. However, there are limitations to this method when conducted on people with diabetes. As an alternative to the Ankle Brachial Index, measuring Toe Systolic Pressures and the Toe Brachial Index have been recommended to assess the arterial blood supply to the foot. This study aimed to determine the intra and inter-rater reliability of the measurement of Toe Systolic Pressure and the Toe Brachial Index in patients with diabetes using a manual measurement system., Methods: This was a repeated measures, reliability study. Three raters measured Toe Systolic Pressure and the Toe Brachial Index in thirty participants with diabetes. Measurement sessions occurred on two occasions, one week apart, using a manual photoplethysmography unit (Hadeco Smartdop 45) and a standardised measurement protocol., Results: The mean intra-class correlation for intra-rater reliability for toe systolic pressures was 0.87 (95% LOA: -25.97 to 26.06 mmHg) and the mean intra-class correlation for Toe Brachial Indices was 0.75 (95% LOA: -0.22 to 0.28). The intra-class correlation for inter-rater reliability was 0.88 for toe systolic pressures (95% LOA: -22.91 to 29.17.mmHg) and 0.77 for Toe Brachial Indices (95% LOA: -0.21 to 0.22)., Conclusion: Despite the reasonable intra-class correlation results, the range of error (95% LOA) was broad. This raises questions regarding the reliability of using a manual sphygmomanometer and PPG for the Toe Systolic Pressure and Toe Brachial Indice.

Using cigarettes to change client behaviour is a common, yet little studied, practice in mental health care. A questionnaire survey was used to explore mental health nursing student's experiences and attitudes to this practice. The sample was four cohorts of mental health nursing students (n= 151). Of them, 84% had experienced the practice of using cigarettes to change client behaviour in acute wards (73%), rehabilitation wards (28%) and elderly care (14%). Cigarettes were used to change client behaviour in areas such as attending to personal hygiene (57%) or engaging in the ward routine (39%). However, items such as leave (60%) or drinks (tea and coffee) (38%) were also reportedly used. Of the respondents, 54% inferred that the practice did not work well with 46% stating it was not written up in care plans; 52% felt it was an ad hoc practice, 60% inferred that at times it was used as a punishment while 55% intimated that they felt bad withholding cigarettes. There are ethical and moral dilemmas around using lifestyle risk factors as rewards or using client's nicotine addiction as a means of controlling behaviour. The question of whether this intervention should ever be used, given its associated health risk, requires more critical debate in clinical practice., (© 2010 Blackwell Publishing.)

Objective: To test the hypothesis that a difference in cytotoxicity exists between latex and non-latex orthodontic separating elastics., Material and Methods: Five intra-oral separating elastics from different manufactures (four latex and one non-latex) were divided into five groups of 15 elastics each: Group MA (non-latex elastics, Masel), Group MO (natural latex, Morelli), Group DE (natural latex, Dentaurum), Group TP (natural latex, TP Orthodontics) and Group UN (natural latex, Unitek). The cytotoxicity assay was performed using cell cultures (epithelial HEp-2 cells originating from human laryngeal carcinoma) that were submitted to the cell viability test with neutral red (dye-uptake) at 24, 48, 72 and 168 h. Analysis of variance (anova) with multiple comparisons and Tukey's test were employed (p < 0.05)., Results: The results showed no statistically significant differences between groups MA, DE, TP and UN in relation to Group CC (cell control) for experimental times of 24, 48 and 168 h (p > 0.05). Morelli, Dentaurum, TP Orthodontics and Unitek elastics induced a great amount of cell lyses at 72 h., Conclusion: One can demonstrate that the Masel elastic induced less cell lysis compared with other elastics, but all trademarks were found to be clinically biocompatible., Clinical Relevance: Separating orthodontic elastics are used in the interdental subgingival region with the aim to separate the teeth for placement of orthodontic bands. However, latex has been known to cause allergy. As these materials are widely used in clinical orthodontics, care regarding the cytotoxicity of orthodontic elastics should be taken. Thus, clinically proven biocompatible materials should be acquired whenever possible.

This study investigated the cytotoxicity exists between latex and non-latex Orthodontic elastomeric ligatures. Six elastomeric ligatures (1 latex, 2 latex-free and 3 polyurethane) from different manufacturers were divided into 6 groups of 15 elastics each: A (Latex-free, American Orthodontics), M (Polyurethane, Morelli), G (Polyurethane,GAC International), Te (Polyurethane, Tecnident), TP (Natural latex,TP Orthodontics) and U (Latex-free,3M Unitek). The cytotoxicity assay was performed using cell cultures (L929 mouse fibroblast cell line), which were subjected to the cell viability test with neutral red ("dye-uptake") at 1, 2, 3, 7 and 28 days. Data were analyzed statistically by ANOVA and Tukey's test (α=0.05). No statistically significant differences (p>0.05) were observed between Groups M and Te in all experimental periods, except at 2 days. No significant differences (p>0.05) in cell viability were found either among Groups A, G, TP and U or between Groups M and Te at 24 h or among Groups CC, A, G, TP and U at 2 and 28 days. It may be concluded that latex-free elastomeric ligatures from American Orthodontics and Unitek trademarks induced less cell lysis compared to latex and polyurethane ligatures.

We evaluated the antiviral activity of the marine alga, Ulva fasciata, collected from Rasa beach and Forno beach, Búzios, Rio de Janeiro, Brazil on the replication of human metapneumovirus (HMPV). The algae extracts were prepared using three different methodologies to compare the activity of different groups of chemical composites obtained through these different methodologies. Four out of the six extracts inhibited nearly 100% of viral replication. The results demonstrated that the majority of the extracts (five out of six) possess virucidal activity and therefore have the ability to interact with the extracellular viral particles and prevent the infection. On the other hand, only two extracts (from Forno beach, obtained by maceration and maceration of the decoction) were able to interact with cell receptors, hindering the viral entry. Finally, only the extract of algae collected at Forno beach, obtained by maceration presented intracellular activity. To our knowledge, this is a pioneer study on antiviral activity of marine algae against HMPV. It is also the first on antiviral activity against HMPV ever done in Brazil. The study also shows the effect of different environment factors and different chemical procedures used to obtain the extract on its biological properties.

Verbascoside has been isolated form L. speciosa after several different chromatographic methods. After its purification, the structure has been unequivocally established using modern spectroscopic techniques. As for the antiviral activity, the maximum non toxic concentration has been established and this concentration has been used in the anti herpes assay, in vitro. Mechanism of action for this molecule regarding the anti-herpes activity has been studied encompassing the following assays: virucidal activity, cellular receptor assay, penetration assay and intracellular assay, in order to understand the activity for this natural product.

Objectives: Natural latex does not fall into the category of materials known to be entirely inoffensive. The purpose of this in vitro study was to test the hypothesis that there is no difference in the cytotoxicity between elastics of different colors and those from different manufacturers., Material and Methods: Different latex intraoral elastics of different colors (5/16 = 7.9 mm, mean load) were compared. The sample was divided into 7 groups of 24 elastics each: Group T (TP Orthodontics, natural latex elastics, control); Groups U1, U2, U3, U4, U5 and U6 (Uniden, natural latex elastics and colored elastics, namely, green, pink, yellow, red and purple, respectively). Cytotoxicity assays were performed by using cell culture medium containing epithelioid-type cells (Hep-2 line) derived from human laryngeal carcinoma. The cytotoxicity was evaluated by using the 'dye-uptake' test, which was employed at two different moments (0 and 24 h). Data were compared by analysis of variance (ANOVA) and Tukey's test (p<0.05)., Results: There was statistically significant difference (p<0.05) between Group T and all other groups (U1, U2, U3, U4, U5 and U6) at 0 and 24 h. No statistically significant difference (p<0.05) was found between Groups U1 and U5, U1 and U6, U2 and U3, U2 and U4, U2 and U5, U2 and U6, U3 and U4, U3 and U5, U3 and U6, U4 and U5, U4 and U6, and U5 and U6 at 0 and 24 h., Conclusions: The TP Orthodontics elastics promoted less cell lysis compared to the Uniden elastics regardless of their color.

The use of low-level laser to suppress infections caused by Herpes simplex viruses 1 and 2 was evaluated after one to five applications. A gradual reduction in replication of Herpes simplex viruses 1 and 2 was observed, with 68.4% and 57.3% inhibition, respectively, after five applications, thus favoring its clinical use.

Parasites belonging to the Leptomonas genus have been used as model organisms for studying biochemical, cellular, and genetic processes unique to members of the Trypanosomatidae family. In the present study, the cell-associated and extracellular peptidases of three Leptomonas species, Leptomonas collosoma, Leptomonas samueli, and Leptomonas wallacei, were assayed and characterized by gelatin-sodium dodecyl sulfate polyacrylamide gel electrophoresis. All parasites released metallopeptidases, whereas no cell-associated proteolytic activity could be detected in the cellular extracts from L. collosoma. Western blotting probed with a polyclonal antibody raised against gp63 from Leishmania amazonensis revealed two major reactive polypeptides of apparent molecular masses of 63 and 52 kDa, with different intensities in cellular extracts and released proteins from the studied trypanosomatids. Flow cytometry and fluorescence microscopy analyses showed that the gp63-like molecules have a surface location. This is the first report on the presence of gp63-like molecules in L. collosoma, L. samueli, and L. wallacei. The pretreatment of L. samueli and L. wallacei with anti-gp63 antibody significantly diminished their association index to Aedes albopictus cell line (C6/36), suggesting a potential involvement of the gp63-like molecules in the interaction process of these insect trypanosomatids with the vector.

Acute diarrhea, especially in children, is a very common disease with worldwide distribution and with a significant public health impact. Rotaviruses have been recognized as the major agents of diarrhea in infants and young children in developed as well as developing countries. In Brazil, diarrhea is one of the principal causes of death, mainly in the infant population. To fight diarrhea, traditional Brazilian medicine uses a great variety of plants. In this work, 12 medicinal plant species were screened for simian (SA-11) and human (HCR3) rotaviruses inhibition in vitro. At non-cytotoxic concentrations, the extracts from Artocarpus integrifolia L. (Moraceae) bark (480 microg/ml) and Spondias lutea L. (Anacardiaceae) leaves (160 microg/ml) had antiviral activity against both viruses. They showed inhibition of 99.2% and 97%, respectively, for human rotavirus, and 96.4% and 96.2% for simian rotavirus. The extracts from Myristica fragrans Houtt (Myristicaceae) seeds (160 microg/ml) and Spongias lutea bark (40 microg/ml) inhibited human rotavirus (90% and 82.2% inhibition, respectively), whereas the extracts from Anacardium occidentale L. (Anacardiaceae) leaves (4 microg/ml) and Psidium guajava L. (Myrtaceae) leaves (8 microg/ml) showed activity only against simian rotavirus (82.2% and 93.8% inhibition, respectively). Our results indicate that the extracts of Artocarpus integrifolia, Myristica fragrans and Spongias lutea can be useful in the treatment of human diarrhea if the etiologic agent is a rotavirus.

Incubation of acyclovir-resistant herpes simplex virus type 1 (ACVr-HSV1), during infection of the HEp-2 cell culture, with an extract prepared from the seeds of Licania tomentosa (Benth.) Fritsch (Chrysobalanaceae) species impaired the productive replication of this virus in a concentration-dependent manner. The extract was able to inhibit extracellular virus (virucidal effect) and also interfered with a very early event of cell infection, at a non-cytotoxic concentration.

This work evaluated the effect of a sulphated fucan extracted from the Laminaria abyssalis marine algae on the human T cell lymphotropic virus type 1 (HTLV-1)-induced syncytium formation. The experiments were carried out in HeLa cells cocultured with a HTLV-1-infected T cell line (C91/PL cells) in the presence of the sulphated polysaccharide at concentration below that corresponding to the ED50. The sulphated fucan inhibited almost 100% of the syncytium formation at concentration of 100 microg/mI and was still active (>95%) at a concentration of 25 microg/ml. It was also observed that the best inhibition occurred when the compound was added in the first 2 h of the cell-to-cell contact. This is the first report showing that a purified sulphated polysaccharide, extracted from marine algae, is able to inhibit the cell-to-cell contact essential for the spreading of the HTLV-1.

Extracts and fractions rich in flavonoids from fruits and leaves of Vitex polygama Cham. (Verbenaceae) were tested against acyclovir-resistant herpes simplex virus type 1 (ACV-HSV-1). Both fruit and leaf extracts exhibited a dose-dependent antiviral activity. The extract from the leaves showed intracellular antiviral activity while the extract from the fruits had virucidal effect. A fraction from the ethyl actetate extract of the leaves inhibited virus propagation by blocking HEp-2 cell receptors.