Functional Characterization of an Aspergillus fumigatus Calcium Transporter (PmcA) that Is Essential for Fungal Infection

Aspergillus fumigatus is a primary and opportunistic pathogen, as well as a major allergen, of mammals. The Ca+2-calcineurin pathway affects virulence, morphogenesis and antifungal drug action in A. fumigatus. Here, we investigated three components of the A. fumigatus Ca+2-calcineurin pathway, pmcA,-B, and -C, which encode calcium transporters. We demonstrated that CrzA can directly control the mRNA accumulation of the pmcA-C genes by binding to their promoter regions. CrzA-binding experiments suggested that the 5'-CACAGCCAC-3' and 5'-CCCTGCCCC-3' sequences upstream of pmcA and pmcC genes, respectively, are possible calcineurin-dependent response elements (CDREs)-like consensus motifs. Null mutants were constructed for pmcA and -beta and a conditional mutant for pmcC demonstrating pmcC is an essential gene. The Delta pmcA and Delta pmcB mutants were more sensitive to calcium and resistant to manganese and cyclosporin was able to modulate the sensitivity or resistance of these mutants to these salts, supporting the interaction between calcineurin and the function of these transporters. The pmcA-C genes have decreased mRNA abundance into the alveoli in the Delta calA and Delta crzA mutant strains. However, only the A. fumigatus Delta pmcA was avirulent in the murine model of invasive pulmonary aspergillosis.

PKC signaling regulates drug resistance of the fungal pathogen Candida albicans via circuitry comprised of Mkc1, calcineurin, and Hsp90.

Fungal pathogens exploit diverse mechanisms to survive exposure to antifungal drugs. This poses concern given the limited number of clinically useful antifungals and the growing population of immunocompromised individuals vulnerable to life-threatening fungal infection. To identify molecules that abrogate resistance to the most widely deployed class of antifungals, the azoles, we conducted a screen of 1,280 pharmacologically active compounds. Three out of seven hits that abolished azole resistance of a resistant mutant of the model yeast Saccharomyces cerevisiae and a clinical isolate of the leading human fungal pathogen Candida albicans were inhibitors of protein kinase C (PKC), which regulates cell wall integrity during growth, morphogenesis, and response to cell wall stress. Pharmacological or genetic impairment of Pkc1 conferred hypersensitivity to multiple drugs that target synthesis of the key cell membrane sterol ergosterol, including azoles, allylamines, and morpholines. Pkc1 enabled survival of cell membrane stress at least in part via the mitogen activated protein kinase (MAPK) cascade in both species, though through distinct downstream effectors. Strikingly, inhibition of Pkc1 phenocopied inhibition of the molecular chaperone Hsp90 or its client protein calcineurin. PKC signaling was required for calcineurin activation in response to drug exposure in S. cerevisiae. In contrast, Pkc1 and calcineurin independently regulate drug resistance via a common target in C. albicans. We identified an additional level of regulatory control in the C. albicans circuitry linking PKC signaling, Hsp90, and calcineurin as genetic reduction of Hsp90 led to depletion of the terminal MAPK, Mkc1. Deletion of C. albicans PKC1 rendered fungistatic ergosterol biosynthesis inhibitors fungicidal and attenuated virulence in a murine model of systemic candidiasis. This work establishes a new role for PKC signaling in drug resistance, novel circuitry through which Hsp90 regulates drug resistance, and that targeting stress response signaling provides a promising strategy for treating life-threatening fungal infections.

Recurrent non-melanoma skin cancer: remission of field cancerization after conversion from calcineurin inhibitor- to proliferation signal inhibitor-based immunosuppression in a cardiac transplant recipient

Non-melanoma skin cancers (NMSCs) are the most common malignancies after solid organ transplantation. Their incidence increases with time after transplantation. Calcineurin-inhibitors (CNIs) and azathioprine are known as skin neoplasia-initiating and -enhancing immunosuppressants. In contrast, increasing clinical experience suggests a relevant antiproliferative effect of mammalian target of rapamycin inhibitors, also named proliferation signal inhibitors (PSIs). We report the case of a cardiac allograft recipient with an impressive and consolidated reduction of recurrent NMSC, observed after conversion from CNI-therapy to a PSI-based protocol.

Everolimus-based, calcineurin-inhibitor-free regimen in recipients of de-novo kidney transplants: an open-label, randomised, controlled trial

Non-nephrotoxic immunosuppressive strategies that allow reduction of calcineurin-inhibitor exposure without compromising safety or efficacy remain a goal in kidney transplantation. Immunosuppression based on the mammalian-target-of-rapamycin inhibitor everolimus was assessed as a strategy for elimination of calcineurin-inhibitor exposure and optimisation of renal-graft function while maintaining efficacy.

Calcineurin inhibition or mutation enhances cell wall inhibitors against Aspergillus fumigatus

Calcineurin mutation or inhibition enhanced the antifungal morphological effect of cell wall inhibitors caspofungin or nikkomycin Z against Aspergillus fumigatus. Quantification of 1,3-beta-d-glucan revealed decreased amounts in the calcineurin A (DeltacnaA) mutant. Calcineurin can be an excellent adjunct therapeutic target in combination with other cell wall inhibitors against A. fumigatus.

Improved maritime target tracker using colour fusion

Searching for humans lost in vast stretches of ocean has always been a difficult task. This paper investigates a machine vision system that addresses this problem by exploiting the useful properties of alternate colour spaces. In particular, the paper investigates the fusion of colour information from the HSV, RGB, YCbCr and YIQ colour spaces within the emission matrix of a Hidden Markov Model tracker to enhance video based maritime target detection. The system has shown promising results. The paper also identifies challenges still needing to be met.

A Vision Based Target Detection System for Docking of an Autonomous Underwater Vehicle

This paper describes the development and preliminary experimental evaluation of a visionbased docking system to allow an Autonomous Underwater Vehicle (AUV) to identify and attach itself to a set of uniquely identifiable targets. These targets, docking poles, are detected using Haar rectangular features and rotation of integral images. A non-holonomic controller allows the Starbug AUV to orient itself with respect to the target whilst maintaining visual contact during the manoeuvre. Experimental results show the proposed vision system is capable of robustly identifying a pair of docking poles simultaneously in a variety of orientations and lighting conditions. Experiments in an outdoor pool show that this vision system enables the AUV to dock autonomously from a distance of up to 4m with relatively low visibility.

Evaluating the Java Virtual Machine as a Target for Languages Other Than Java

The portability and runtime safety of programs which are executed on the Java Virtual Machine (JVM) makes the JVM an attractive target for compilers of languages other than Java. Unfortunately, the JVM was designed with language Java in mind, and lacks many of the primitives required for a straighforward implementation of other languages. Here, we discuss how the JVM may be used to implement other object-oriented languages. As a practical example of the possibilities, we report on a comprehensive case study. The open source Gardens Point Component Pascal compiler compiles the entire Component Pascal language, a dialect of Oberon-2, to JVM bytecodes. This compiler achieves runtime efficiencies which are comparable to native-code implementations of procedural languages.

Involvement of target contrast in visual masking location tasks deficients in myopia

Purpose: There have been few studies of visual temporal processing of myopic eyes. This study investigated the visual performance of emmetropic and myopic eyes using a backward visual masking location task. Methods: Data were collected for 39 subjects (15 emmetropes, 12 stable myopes, 12 progressing myopes). In backward visual masking, a target’s visibility is reduced by a mask presented in quick succession ‘after’ the target. The target and mask stimuli were presented at different interstimulus intervals (from 12 to 300 ms). The task involved locating the position of a target letter with both a higher (seven per cent) and a lower (five per cent) contrast. Results: Emmetropic subjects had significantly better performance for the lower contrast location task than the myopes (F2,36 = 22.88; p < 0.001) but there was no difference between the progressing and stable myopic groups (p = 0.911). There were no differences between the groups for the higher contrast location task (F2,36 = 0.72, p = 0.495). No relationship between task performance and either the magnitude of myopia or axial length was found for either task. Conclusions: A location task deficit was observed in myopes only for lower contrast stimuli. Both emmetropic and myopic groups had better performance for the higher contrast task compared to the lower contrast task, with myopes showing considerable improvement. This suggests that five per cent contrast may be the contrast threshold required to bias the task towards the magnocellular system (where myopes have a temporal processing deficit). Alternatively, the task may be sensitive to the contrast sensitivity of the observer.

Contributions of vision-proprioception interactions to the estimation of time-varying hand and target locations

We investigated the relative importance of vision and proprioception in estimating target and hand locations in a dynamic environment. Subjects performed a position estimation task in which a target moved horizontally on a screen at a constant velocity and then disappeared. They were asked to estimate the position of the invisible target under two conditions: passively observing and manually tracking. The tracking trials included three visual conditions with a cursor representing the hand position: always visible, disappearing simultaneously with target disappearance, and always invisible. The target’s invisible displacement was systematically underestimated during passive observation. In active conditions, tracking with the visible cursor significantly decreased the extent of underestimation. Tracking of the invisible target became much more accurate under this condition and was not affected by cursor disappearance. In a second experiment, subjects were asked to judge the position of their unseen hand instead of the target during tracking movements. Invisible hand displacements were also underestimated when compared with the actual displacement. Continuous or brief presentation of the cursor reduced the extent of underestimation. These results suggest that vision–proprioception interactions are critical for representing exact target–hand spatial relationships, and that such sensorimotor representation of hand kinematics serves a cognitive function in predicting target position. We propose a hypothesis that the central nervous system can utilize information derived from proprioception and/or efference copy for sensorimotor prediction of dynamic target and hand positions, but that effective use of this information for conscious estimation requires that it be presented in a form that corresponds to that used for the estimations.