In vitro susceptibilities of Neoscytalidium spp. sequence types to antifungal agents and antimicrobial photodynamic treatment with phenothiazinium photosensitizers

Neoscytalidium spp. are ascomycetous fungi consisting of pigmented and hyaline varieties both able to cause skin and nail infection. Their colour-based identification is inaccurate and may compromise the outcome of the studies with these fungi. The aim of this study was to genotype 32 isolates morphologically identified as Neoscytalidium dimidiatum or Neoscytalidium dimidiatum var. hyalinum by multilocus sequence typing (MLST), differentiate the two varieties by their sequence types (STs), evaluate their susceptibility to seven commercial antifungal drugs [amphotericin B (AMB), voriconazole (VOR), terbinafine (TER), 5-flucytosine (5FC), ketoconazole (KET), fluconazole (FLU), and caspofungin (CAS)], and also to the antimicrobial photodynamic treatment (APDT) with the phenothiazinium photosensitizers (PSs) methylene blue (MB), new methylene blue (NMBN), toluidine blue O (TBO), and the pentacyclic derivative S137. The efficacy of each PS was determined, initially, based on its minimal inhibitory concentration (MIC). Additionally, the APDT effects with each PS on the survival of ungerminated and germinated arthroconidia of both varieties were evaluated. Seven loci of Neoscytalidium spp. were sequenced on MLST revealing eight polymorphic sites and six STs. All N. dimidiatum var. hyalinum isolates were clustered in a single ST. AMB, VOR, and TER were the most effective antifungal agents against both varieties. The hyaline variety isolates were much less tolerant to the azoles than the isolates of the pigmented variety. APDT with S137 showed the lowest MIC for all the isolates of both varieties. APDT with all the PSs killed both ungerminated and germinated arthroconidia of both varieties reducing the survival up to 5 logs. Isolates of the hyaline variety were also less tolerant to APDT. APDT with the four PSs also increased the plasma membrane permeability of arthroconidia of both varieties but only NMBN and S137 caused peroxidation of the membrane lipids. © 2017 British Mycological Society

Cylindrically symmetric inhomogeneous dust collapse with a zero expansion component

We investigate a class of cylindrically symmetric inhomogeneous Λ-dust spacetimes which have a regular axis and some zero expansion component. For Λ \neq 0, we obtain new exact solutions to the Einstein equations and show that they are unique, within that class. For Λ = 0, we recover the Senovilla– Vera metric and show that it can be locally matched to an Einstein–Rosen type of exterior. Finally, we explore some consequences of the matching, such as trapped surface formation and gravitational radiation in the exterior.We thank Jose Senovilla for suggesting solutions of the form (8) and (9) and for other very useful comments which led to substancial improvements in the paper. IB and FM are supported by Portuguese Funds through Fundacao para a Cincia e Tecnologia (FCT), within the Projects UID/MAT/00013/2013 and PTDC/MAT-ANA/1275/2014. FM and NOS acknowledge a grant received from UERJ and thank the warm hospitality from Instituto de Fisica, UERJ, Rio de Janeiro, where a great part of this work was completed. MFAdaSilva acknowledges the financial support from FAPERJ (no. E-26/171.754/2000, E-26/171.533.2002, E-26/170.951/2006, E-26/110.432/2009 and E-26/111.714/2010), Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq) Brazil (no. 450572/2009-9, 301973/2009-1 and 477268/2010-2) and Financiadora de Estudos e Projetos (FINEP) Brazil.info:eu-repo/semantics/publishedVersio

Completion of fit notes by GPs: a mixed methods study

Aims: The aim of this study was to investigate the completion of fit notes by UK general practitioners (GPs). A series of actual fit notes issued to employed patients were examined, and their GPs’ reflections and experiences of fit note completion explored. Methods: A mixed-methods design was used. Data were collected from copies of 94 fit notes issued to employed patients by 11 GPs, and from 86 questionnaires completed by these GPs reflecting on the fit notes they had issued. Face-to-face interviews were then conducted with each GP. Results: Fit note completion is not meeting expectations for a number of reasons. These include the following: limited knowledge and awareness of the guidance in fit note completion; problems with the fit note format; lack of mandatory training in completing fit notes; lack of incentive to change practice; incomplete implementation of the electronic fit note; GPs’ lack of confidence in, and doubts about the appropriateness of performing this role. Conclusion: If UK GPs are to continue their contractual responsibility for completing fit notes, further consideration of their education and training needs is urgently required. Weaknesses in the design and format of the fit note and the availability of the electronic version also need to be addressed

The Presence of IL-17A and T Helper 17 Cells in Experimental Mouse Brain Tumors and Human Glioma

Background: Recently, CD4 + IL-17A + T helper 17 (Th17) cells were identified and reported in several diseased states, including autoimmunity, infection and various peripheral nervous system tumors. However, the presence of Th17 in gliaderived tumors of the central nervous system has not been studied. Methodology/Principal Findings: In this report, we demonstrate that mRNA expression for the Th17 cell cytokine IL-17A, as well as Th17 cells, are present in human glioma. The mRNA expression for IL-17A in glioma was recapitulated in an immunocompetent mouse model of malignant glioma. Furthermore, the presence of Th17 cells was confirmed in both human and mouse glioma. Interestingly, some Th17 cells present in mouse glioma co-expressed the Th1 and Th2 lineage markers, IFN-c and IL-4, respectively, but predominantly co-expressed the Treg lineage marker FoxP3. Conclusions: These data confirm the presence of Th17 cells in glia-derived CNS tumors and provide the rationale for further investigation into the role of Th17 cells in malignant glioma

Protective Intestinal Effects of Pituitary Adenylate Cyclase Activating Polypeptide

Pituitary adenylate cyclase activating polypeptide (PACAP) is an endogenous neuropeptide widely distributed throughout the body, including the gastrointestinal tract. Several effects have been described in human and animal intestines. Among others, PACAP infl uences secretion of intestinal glands, blood fl ow, and smooth muscle contraction. PACAP is a well-known cytoprotective peptide with strong anti-apoptotic, anti-infl ammatory, and antioxidant effects. The present review gives an overview of the intestinal protective actions of this neuropeptide. Exogenous PACAP treatment was protective in a rat model of small bowel autotransplantation. Radioimmunoassay (RIA) analysis of the intestinal tissue showed that endogenous PACAP levels gradually decreased with longer-lasting ischemic periods, prevented by PACAP addition. PACAP counteracted deleterious effects of ischemia on oxidative stress markers and cytokines. Another series of experiments investigated the role of endogenous PACAP in intestines in PACAP knockout (KO) mice. Warm ischemia–reperfusion injury and cold preservation models showed that the lack of PACAP caused a higher vulnerability against ischemic periods. Changes were more severe in PACAP KO mice at all examined time points. This fi nding was supported by increased levels of oxidative stress markers and decreased expression of antioxidant molecules. PACAP was proven to be protective not only in ischemic but also in infl ammatory bowel diseases. A recent study showed that PACAP treatment prolonged survival of Toxoplasma gondii infected mice suffering from acute ileitis and was able to reduce the ileal expression of proinfl ammatory cytokines. We completed the present review with recent clinical results obtained in patients suffering from infl ammatory bowel diseases. It was found that PACAP levels were altered depending on the activity, type of the disease, and antibiotic therapy, suggesting its probable role in infl ammatory events of the intestine

Photodynamic and Antibiotic Therapy Impair the Pathogenesis of Enterococcus faecium in a Whole Animal Insect Model

Enterococcus faecium has emerged as one of the most important pathogens in healthcare-associated infections worldwide due to its intrinsic and acquired resistance to many antibiotics, including vancomycin. Antimicrobial photodynamic therapy (aPDT) is an alternative therapeutic platform that is currently under investigation for the control and treatment of infections. PDT is based on the use of photoactive dye molecules, widely known as photosensitizer (PS). PS, upon irradiation with visible light, produces reactive oxygen species that can destroy lipids and proteins causing cell death. We employed Galleria mellonella (the greater wax moth) caterpillar fatally infected with E. faecium to develop an invertebrate host model system that can be used to study the antimicrobial PDT (alone or combined with antibiotics). In the establishment of infection by E. faecium in G. mellonella, we found that the G. mellonella death rate was dependent on the number of bacterial cells injected into the insect hemocoel and all E. faecium strains tested were capable of infecting and killing G. mellonella. Antibiotic treatment with ampicillin, gentamicin or the combination of ampicillin and gentamicin prolonged caterpillar survival infected by E. faecium (P = 0.0003, P = 0.0001 and P = 0.0001, respectively). In the study of antimicrobial PDT, we verified that methylene blue (MB) injected into the insect followed by whole body illumination prolonged the caterpillar survival (P = 0.0192). Interestingly, combination therapy of larvae infected with vancomycin-resistant E. faecium, with antimicrobial PDT followed by vancomycin, significantly prolonged the survival of the caterpillars when compared to either antimicrobial PDT (P = 0.0095) or vancomycin treatment alone (P = 0.0025), suggesting that the aPDT made the vancomycin resistant E. faecium strain more susceptible to vancomycin action. In summary, G. mellonella provides an invertebrate model host to study the antimicrobial PDT and to explore combinatorial aPDT-based treatments

Efficient, non‐toxic anion transport by synthetic carriers in cells and epithelia

Transmembrane anion transporters (anionophores) have potential for new modes of biological activity, including therapeutic applications. In particular they might replace the activity of defective anion channels in conditions such as cystic fibrosis. However, data on the biological effects of anionophores are scarce, and it remains uncertain whether such molecules are fundamentally toxic. Here, we report a biological study of an extensive series of powerful anion carriers. Fifteen anionophores were assayed in single cells by monitoring anion transport in real time through fluorescence emission from halide-sensitive yellow fluorescent protein. A bis-(p-nitrophenyl)ureidodecalin shows especially promising activity, including deliverability, potency and persistence. Electrophysiological tests show strong effects in epithelia, close to those of natural anion channels. Toxicity assays yield negative results in three cell lines, suggesting that promotion of anion transport may not be deleterious to cells. We therefore conclude that synthetic anion carriers are realistic candidates for further investigation as treatments for cystic fibrosis.info:eu-repo/semantics/publishe

Exacerbation of facial motoneuron loss after facial nerve axotomy in CCR3-deficient mice

We have previously demonstrated a neuroprotective mechanism of FMN (facial motoneuron) survival after facial nerve axotomy that is dependent on CD4+ Th2 cell interaction with peripheral antigen-presenting cells, as well as CNS (central nervous system)-resident microglia. PACAP (pituitary adenylate cyclase-activating polypeptide) is expressed by injured FMN and increases Th2-associated chemokine expression in cultured murine microglia. Collectively, these results suggest a model involving CD4+ Th2 cell migration to the facial motor nucleus after injury via microglial expression of Th2-associated chemokines. However, to respond to Th2-associated chemokines, Th2 cells must express the appropriate Th2-associated chemokine receptors. In the present study, we tested the hypothesis that Th2-associated chemokine receptors increase in the facial motor nucleus after facial nerve axotomy at timepoints consistent with significant T-cell infiltration. Microarray analysis of Th2-associated chemokine receptors was followed up with real-time PCR for CCR3, which indicated that facial nerve injury increases CCR3 mRNA levels in mouse facial motor nucleus. Unexpectedly, quantitative- and co-immunofluorescence revealed increased CCR3 expression localizing to FMN in the facial motor nucleus after facial nerve axotomy. Compared with WT (wild-type), a significant decrease in FMN survival 4 weeks after axotomy was observed in CCR3−/− mice. Additionally, compared with WT, a significant decrease in FMN survival 4 weeks after axotomy was observed in Rag2−/− (recombination activating gene-2-deficient) mice adoptively transferred CD4+ T-cells isolated from CCR3−/− mice, but not in CCR3−/− mice adoptively transferred CD4+ T-cells derived from WT mice. These results provide a basis for further investigation into the co-operation between CD4+ T-cell- and CCR3-mediated neuroprotection after FMN injury

Inhibitory action of phenothiazinium dyes against Neospora caninum.

Neospora caninum is an Apicomplexan parasite related to important losses in livestock, causing abortions and decreased fertility in affected cows. Several chemotherapeutic strategies have been developed for disease control; however, no commercial treatment is available. Among the candidate drugs against neosporosis, phenothiazinium dyes, offer a low cost-efficient approach to parasite control. We report the anti-parasitic effects of the phenothiaziums Methylene Blue (MB), New Methylene Blue (NMB), 1,9-Dimethyl Methylene Blue (DMMB) and Toluidine Blue O (TBO) on N. caninum, using in vitro and in vivo models. The dyes inhibited parasite proliferation at nanomolar concentrations (0.019-1.83 μM) and a synergistic effect was achieved when Methylene Blue was combined with New Methylene Blue (Combination Index = 0.84). Moreover, the phenothiazinium dyes improved parasite clearance when combined with Pyrimethamine (Pyr). Combination of Methylene Blue + 1,9-Dimethyl Methylene Blue demonstrated superior efficacy compared to Pyrimethamine based counterparts in an in vivo model of infection. We also observed that Methylene Blue, New Methylene Blue and 1,9-Dimethyl Methylene Blue increased by 5000% the reactive oxygen species (ROS) levels in N. caninum tachyzoites. Phenothiazinium dyes represent an accessible group of candidates with the potential to compound future formulations for neosporosis control

Exploring new physics frontiers through numerical relativity

The demand to obtain answers to highly complex problems within strong-field gravity has been met with significant progress in the numerical solution of Einstein's equations - along with some spectacular results - in various setups. We review techniques for solving Einstein's equations in generic spacetimes, focusing on fully nonlinear evolutions but also on how to benchmark those results with perturbative approaches. The results address problems in high-energy physics, holography, mathematical physics, fundamental physics, astrophysics and cosmology