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Introduction: Onychomycosis is a debilitating, difficult-to-treat nail fungal infection with increasing prevalence worldwide. The main etiological agents are dermatophytes, which are common causative pathogens in superficial fungal mycoses. Conventional detection methods such as fungal culture have low sensitivity and specificity and are time-consuming.Aim: The main objective of this study was to design, develop and validate a real-time probe-based multiplex qPCR assay for the detection of dermatophytes and Fusarium species.Methodology: The performance characteristics of the qPCR assays were evaluated. The multiplex qPCR assays targeted four genes (assay 1: pan-dermatophytes/Fusarium spp.; assay 2: Trichophyton rubrum/Microsporum spp.). Analytical validation was accomplished using 150 fungal isolates and clinical validation was done on 204 nail specimens. The performance parameters were compared against the gold standard (fungal culture) and expanded gold standard (culture in conjunction with sequencing).Results: Both the single-plex and multiplex qPCR assays performed well especially when compared against the expanded gold standard. Among the 204 tested nail specimens, the culture method showed that 125 (61.3 %) were infected with at least one organism, of which 40 yielded positive results for dermatophytes and Fusarium spp. These target organisms detected include 20 dermatophytes and 22 Fusarium spp. The developed qPCR assays demonstrated excellent limit of detection, efficiency, coefficient of determination, analytical and clinical sensitivity and specificity.Conclusion: The multiplex qPCR assays were reliable for the diagnosis of onychomycosis, with shorter turn-around time as compared to culture method. This aids in the planning of treatment strategies to achieve optimal therapeutic outcome.

Competing demands in coaching duties hinder coaches from effecting Positive Youth Development (PYD) among athletes. While research has documented how supporting staff in youth sport teams can help ...

The purpose of the present study was to investigate the level of physical activity (PA) in an all-boys school and, using a mixed method design, to identify factors that facilitate or hinder PA engagement. Participants ( n = 382) completed the Three Day Physical Activity Recall (3DPAR) questionnaire to enable examination of the pattern of their PA and the amount of time spent on sedentary activities. To understand further the motivation behind their level of PA, 24 participants with the highest and lowest 3DPAR scores were selected for focus groups. Quantitative data of the PA levels of the all-boys sample of this study and those of a mixed sample from a study by Lee and Trost were analysed using percentage means with 95% confidence intervals, with a 2x2 analysis of variance being used for the comparisons. The focus group data were transcribed and thematic analysis was performed on the transcripts. The quantitative results revealed evidence that male students in the current sample were more active in comparison with the students from the mixed sample. The qualitative results indicated that PA participation was facilitated by factors such as intention to maintain health and fitness levels, a strong school sports culture, regular exercise with friends and family, and an individual’s exercise history. In contrast, academic workload, poor exercise history and after-school non-PA related activities were identified as factors that hindered PA participation. The findings are discussed in relation to wider school decisions and policies that have an effect on PA among adolescent students.

This study examines perceptions of learners on pronunciation acquisition, learning phonetic transcriptions and their sounds associations amongst Polytechnic Kuala Terengganu Learners. This study was carried-out amongst semester one diploma of Electronics and Electrical Engineering learners in Kuala Terengganu Polytechnic, Terengganu, Malaysia. This study also hopes to show that by learning to recognise and read phonetic symbols learners will be able to overcome any pronunciation disability using the dictionary. This research follows the quasi- experimental design of a control-group A and treatment group B selected randomly doing a pre-test and post-test. The treatment of explicit instruction of phonetic transcriptions for 45 hours was given only to the experimental group. This study focuses on the analysis of oral test answers by learners. Some of the challenges identified in the acquisition of pronunciation are mother tongue interference, mispronunciation because spelling does not reflect pronunciation, unintelligibility and miscommunication and the lack of confidence to speak the English Language. Whilst factors that affect pronunciation acquisition are mother tongue, exposure, motivation, students’ perception, ways of learning, which English to speak and others. However, it was discovered that phonetic instruction of transcriptions and their sounds’ association improves pronunciation competence. Some of the implications of this study are aimed at ELT academicians and practitioners to apply the method used in this study to up-grade the standard of pronunciation proficiency and a suggestion that the learning of pronunciation using phonetic symbols should be taught earlier in their education. Keywords: Pronunciation, unintelligibility, phonetic transcriptions instruction, oral tests, polytechnic learners

The innovations of the twenty-first century in digital technology and media have had major influences in the way young urbanites and future city designers think as well as experience places. In embracing this conceptual shift of how new communication technologies are changing people’s perceptions, experiences and ways of knowing the city, Oxford Brookes University continues to develop pedagogical methods which use new media in the teaching, learning and production of contemporary urban design.These methods are examined in this paper in the context of three undergraduate modules:• Taking the urban design learning experience out of the classroom and into the field using mobile lectures;• Using digital media in research and engaging with oral, visual and sensory experiences;• Developing design communication skills using video, animation, interactive PDFs and website design;• Bringing new perspectives to classic theoretical concepts by engaging with different media: Cullen’s Townscape from serial expe...

Economic growth in African cities has attracted the attention of international property developers and architecture firms seeking new opportunities as demand has waned in the shrinking economies of the Global North. Their interest has produced new computer generated master plans for African cities depicting modern utopias which would not look out of place in Dubai or Shanghai. The incongruity of these plans for cities where 70% or more of urban residents live on the margins in unplanned settlements raises important questions about the processes shaping the future of African cities. Urban theories, in particular neoliberal ideologies of urban entrepreneurialism, offer compelling explanations for how these visions have come to be. What is less well understood is the local dynamics at play in the structuring of African cities and the role of local actors in the construction of these new master plans. This paper documents the attitudes of residents of Lusaka to market driven development in their city and new master plans for African cities. Making use of concepts of policy mobility – it explores the aspirations of those that live in the city and the extent to which these correspond to the promises of these grand new visions. The findings highlight ways in which these new visions of African cities are at odds with lives and concerns of residents in Lusaka from across the social spectrum.

Excitotoxicity mediated via the (S)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) subtype of receptor for l-glutamate contributes to various neuropathologies involving acute brain injury and chronic degenerative disorders. In this study, AMPA-induced neuronal injury and staurosporine (STS)-mediated apoptosis were compared in primary neuronal cultures of murine cerebral cortex by analyzing indices up- and downstream of mitochondrial activation. AMPA-mediated apoptosis involved induction of Bax, loss of mitochondrial transmembrane potential (deltapsi(m)), early release of cytochrome c (cyt c), and more delayed release of second mitochondrial activator of caspases (SMAC), Omi, and apoptosis-inducing factor (AIF) with early calpain and minor late activation of caspase 3. STS-induced apoptosis was characterized by a number of differences, a more rapid time course, non-involvement of deltapsi(m), and relatively early recruitment of SMAC and caspase 3. The AMPA-induced rise in intracellular calcium appeared insufficient to evoke feltapsi(m) as release of cyt c preceded mitochondrial depolarization, which was followed by the cytosolic translocation of SMAC, Omi, and AIF. Bax translocation preceded cyt c release for both stimuli inferring its involvement in apoptotic induction. Inclusion of the broad spectrum caspase inhibitor zVAD-fmk reduced the AMPA-induced release of cyt c, SMAC, and AIF, while only affecting the redistribution of Omi and AIF in the STS-treated neurons. Only AIF release was affected by a calpain inhibitor (calpastatin) which exerted relatively minor effects on the progression of cellular injury. AMPA-mediated release of apoptogenic proteins was more hierarchical relative to STS with its calpain activation and caspase-dependent AIF redistribution arguing for a model with cross-talk between caspase-dependent/independent apoptosis.

Rotenone and 1-methyl-4-phenylpyridinium produce parkinsonian models and we determined whether their mitochondrially mediated actions differentially redistributed the apoptogenic proteins, apoptosis-inducing factor and cytochrome c. Cultured rat mesencephalic dopamine neurons were exposed to rotenone (30 nM) and 1-methyl-4-phenylpyridinium (300 muM, 24 and 48 h) and apoptosis and mitochondrial redistribution of cytochrome c or apoptosis-inducing factor were quantified. Tyrosine hydroxylase-positive dopamine neurons underwent apoptosis (shrinkage, less neurites) and 40% released apoptosis-inducing factor with rotenone (24 h), whereas cytochrome c release reached this value at 48 h when 70% of cells had released apoptosis-inducing factor-positive. 1-Methyl-4-phenylpyridinium produced similar redistribution patterns for both proteins. Preferential redistribution of apoptosis-inducing factor before cytochrome c in dopamine neurons indicates caspase-independent mitochondrial proapoptotic signalling predominates in these parkinsonian models.

Here we report the complete, accurate 1.89-Mb genome sequence of Francisella tularensis subsp. holarctica strain FSC200, isolated in 1998 in the Swedish municipality Ljusdal, which is in an area where tularemia is highly endemic. This genome is important because strain FSC200 has been extensively used for functional and genetic studies of Francisella and is well-characterized.

Sequencing of the non-pathogenic Francisella tularensis sub-species novicida U112, and comparison with two pathogenic sub-species, provides insights into the evolution of pathogenicity in these species., Background Francisella tularensis subspecies tularensis and holarctica are pathogenic to humans, whereas the two other subspecies, novicida and mediasiatica, rarely cause disease. To uncover the factors that allow subspecies tularensis and holarctica to be pathogenic to humans, we compared their genome sequences with the genome sequence of Francisella tularensis subspecies novicida U112, which is nonpathogenic to humans. Results Comparison of the genomes of human pathogenic Francisella strains with the genome of U112 identifies genes specific to the human pathogenic strains and reveals pseudogenes that previously were unidentified. In addition, this analysis provides a coarse chronology of the evolutionary events that took place during the emergence of the human pathogenic strains. Genomic rearrangements at the level of insertion sequences (IS elements), point mutations, and small indels took place in the human pathogenic strains during and after differentiation from the nonpathogenic strain, resulting in gene inactivation. Conclusion The chronology of events suggests a substantial role for genetic drift in the formation of pseudogenes in Francisella genomes. Mutations that occurred early in the evolution, however, might have been fixed in the population either because of evolutionary bottlenecks or because they were pathoadaptive (beneficial in the context of infection). Because the structure of Francisella genomes is similar to that of the genomes of other emerging or highly pathogenic bacteria, this evolutionary scenario may be shared by pathogens from other species.

Redistribution of cytochrome c and Smac/DIABLO from mitochondria occurs during apoptosis, although the relative timing of their release is not well characterized. Double immunocytochemistry was utilized here to study quantitatively the patterns of release of cytochrome c and Smac/DIABLO from mitochondria in single cells. Human osteosarcoma cells and murine embryonic cortical neurons were analyzed during apoptosis induced by staurosporine. In osteosarcoma cells treated with staurosporine for 24 h, a substantial proportion of cells (36%) released cytochrome c from the mitochondria before Smac/DIABLO. In contrast, these proteins were released mostly concordantly in neurons; only a minority of cells (or = 15%) released cytochrome c without Smac/DIABLO (or vice versa) from mitochondria. Patterns of release in either cell type were unaltered by addition of the caspase inhibitor, zVAD-fmk. The double immunocytochemistry procedure facilitated clear definition of the temporal release of cytochrome c and Smac/DIABLO from mitochondria in intact apoptotic cells, enabling us to demonstrate for the first time that their mutual redistribution during apoptosis varies between different cell types.

An approach to restriction-site mapping and contig building that uses fragment-size data from multiple complete digests of a set of clones that oversample a genomic region is presented. Maps containing both fragment-length data and clone-end data are maintained for each restriction enzyme. Synchronization between the maps for the different enzymes is achieved by requiring the clone-end maps for all enzymes to be compatible. Basic concepts that underlie multiple-complete-digest mapping - including the match/merge approach to map incorporation, extension vs. assimilation, ambiguity, and clone-end compatibility - are presented. An initial application of multiple-complete-digest mapping to real data on a set of cosmid clones suggests that this mapping method has exceptional power to produce accurate maps that are well suited to the needs of large-scale DNA-sequencing projects. 31 refs., 7 figs., 2 tabs.

The paradigm of sequence-tagged site (STS)-content mapping involves the systematic assignment of STSs to individual cloned DNA segments. To date, yeast artificial chromosomes (YACs) represent the most commonly employed cloning system for constructing STS maps of large genomic intervals, such as whole human chromosomes. For developing a complete YAC-based STS-content map of human chromosome 7, we wished to utilize a limited set of YAC clones that were highly enriched for chromosome 7 DNA. Toward that end, we have assembled a human chromosome 7 YAC resource that consists of three major components: (1) a newly constructed library derived from a human-hamster hybrid cell line containing chromosome 7 as its only human DNA; (2) a chromosome 7-enriched sublibrary derived from the CEPH mega-YAC collection by Alu-polymerase chain reaction (Alu-PCR)-based hybridization; and (3) a set of YACs isolated from several total genomic libraries by screening for >125 chromosome 7 STSs. In particular, the hybrid cell line-derived YACs, which comprise the majority of the clones in the resource, have a relatively low chimera frequency (10-20%) based on mapping isolated insert ends to panels of human-hamster hybrid cell lines and analyzing individual clones by fluorescence in situ hybridization. An efficient strategy for polymerase chain reactionmore » (PCR)-based screening of this YAC resource, which totals 4190 clones, has been developed and utilized to identify corresponding YACs for >600 STSs. The results of this initial screening effort indicate that the human chromosome 7 YAC resource provides an average of 6.9 positive clones per STS, a level of redundancy that should support the assembly of large YAC contigs and the construction of a high-resolution STS-content map of the chromosome. 72 refs., 4 figs., 3 tabs.« less