ciliaFA : a research tool for automated, high-throughput measurement of ciliary beat frequency using freely available software

Background: Analysis of ciliary function for assessment of patients suspected of primary ciliary dyskinesia (PCD) and for research studies of respiratory and ependymal cilia requires assessment of both ciliary beat pattern and beat frequency. While direct measurement of beat frequency from high-speed video recordings is the most accurate and reproducible technique it is extremely time consuming. The aim of this study was to develop a freely available automated method of ciliary beat frequency analysis from digital video (AVI) files that runs on open-source software (ImageJ) coupled to Microsoft Excel, and to validate this by comparison to the direct measuring high-speed video recordings of respiratory and ependymal cilia. These models allowed comparison to cilia beating between 3 and 52 Hz. Methods: Digital video files of motile ciliated ependymal (frequency range 34 to 52 Hz) and respiratory epithelial cells (frequency 3 to 18 Hz) were captured using a high-speed digital video recorder. To cover the range above between 18 and 37 Hz the frequency of ependymal cilia were slowed by the addition of the pneumococcal toxin pneumolysin. Measurements made directly by timing a given number of individual ciliary beat cycles were compared with those obtained using the automated ciliaFA system. Results: The overall mean difference (± SD) between the ciliaFA and direct measurement high-speed digital imaging methods was −0.05 ± 1.25 Hz, the correlation coefficient was shown to be 0.991 and the Bland-Altman limits of agreement were from −1.99 to 1.49 Hz for respiratory and from −2.55 to 3.25 Hz for ependymal cilia. Conclusions: A plugin for ImageJ was developed that extracts pixel intensities and performs fast Fourier transformation (FFT) using Microsoft Excel. The ciliaFA software allowed automated, high throughput measurement of respiratory and ependymal ciliary beat frequency (range 3 to 52 Hz) and avoids operator error due to selection bias. We have included free access to the ciliaFA plugin and installation instructions in Additional file 1 accompanying this manuscript that other researchers may use

<i>Trypanosoma brucei rhodesiense</i> transmitted by a single tsetse fly bite in vervet monkeys as a model of human African trypanosomiasis

Sleeping sickness is caused by a species of trypanosome blood parasite that is transmitted by tsetse flies. To understand better how infection with this parasite leads to disease, we provide here the most detailed description yet of the course of infection and disease onset in vervet monkeys. One infected tsetse fly was allowed to feed on each host individual, and in all cases infections were successful. The characteristics of infection and disease were similar in all hosts, but the rate of progression varied considerably. Parasites were first detected in the blood 4-10 days after infection, showing that migration of parasites from the site of fly bite was very rapid. Anaemia was a key feature of disease, with a reduction in the numbers and average size of red blood cells and associated decline in numbers of platelets and white blood cells. One to six weeks after infection, parasites were observed in the cerebrospinal fluid (CSF), indicating that they had moved from the blood into the brain; this was associated with a white cell infiltration. This study shows that fly-transmitted infection in vervets accurately mimics human disease and provides a robust model to understand better how sleeping sickness develops

A Pre-clinical Animal Model of Trypanosoma brucei Infection Demonstrating Cardiac Dysfunction

African trypanosomiasis (AT), caused by Trypanosoma brucei species, results in both neurological and cardiac dysfunction and can be fatal if untreated. Research on the pathogenesis and treatment of the disease has centred to date on the characteristic neurological symptoms, whereas cardiac dysfunction (e.g. ventricular arrhythmias) in AT remains largely unstudied. Animal models of AT demonstrating cardiac dysfunction similar to that described in field cases of AT are critically required to transform our understanding of AT-induced cardiac pathophysiology and identify future treatment strategies. We have previously shown that T. brucei can interact with heart muscle cells (cardiomyocytes) to induce ventricular arrhythmias in ex vivo adult rat hearts. However, it is unknown whether the arrhythmias observed ex vivo are also present during in vivo infection in experimental animal models. Here we show for the first time the characterisation of ventricular arrhythmias in vivo in two animal models of AT infection using electrocardiographic (ECG) monitoring. The first model utilised a commonly used monomorphic laboratory strain, Trypanosoma brucei brucei Lister 427, whilst the second model used a pleomorphic laboratory strain, T. b. brucei TREU 927, which demonstrates a similar chronic infection profile to clinical cases. The frequency of ventricular arrhythmias and heart rate (HR) was significantly increased at the endpoint of infection in the TREU 927 infection model, but not in the Lister 427 infection model. At the end of infection, hearts from both models were isolated and Langendorff perfused ex vivo with increasing concentrations of the β-adrenergic agonist isoproterenol (ISO). Interestingly, the increased frequency of arrhythmias observed in vivo in the TREU 927 infection model was lost upon isolation of the heart ex vivo, but re-emerged with the addition of ISO. Our results demonstrate that TREU 927 infection modifies the substrate of the myocardium in such a way as to increase the propensity for ventricular arrhythmias in response to a circulating factor in vivo or β-adrenergic stimulation ex vivo. The TREU 927 infection model provides a new opportunity to accelerate our understanding of AT-related cardiac pathophysiology and importantly has the required sensitivity to monitor adverse cardiac-related electrical dysfunction when testing new therapeutic treatments for AT

The Fecal Viral Flora of Wild Rodents

The frequent interactions of rodents with humans make them a common source of zoonotic infections. To obtain an initial unbiased measure of the viral diversity in the enteric tract of wild rodents we sequenced partially purified, randomly amplified viral RNA and DNA in the feces of 105 wild rodents (mouse, vole, and rat) collected in California and Virginia. We identified in decreasing frequency sequences related to the mammalian viruses families Circoviridae, Picobirnaviridae, Picornaviridae, Astroviridae, Parvoviridae, Papillomaviridae, Adenoviridae, and Coronaviridae. Seventeen small circular DNA genomes containing one or two replicase genes distantly related to the Circoviridae representing several potentially new viral families were characterized. In the Picornaviridae family two new candidate genera as well as a close genetic relative of the human pathogen Aichi virus were characterized. Fragments of the first mouse sapelovirus and picobirnaviruses were identified and the first murine astrovirus genome was characterized. A mouse papillomavirus genome and fragments of a novel adenovirus and adenovirus-associated virus were also sequenced. The next largest fraction of the rodent fecal virome was related to insect viruses of the Densoviridae, Iridoviridae, Polydnaviridae, Dicistroviriade, Bromoviridae, and Virgaviridae families followed by plant virus-related sequences in the Nanoviridae, Geminiviridae, Phycodnaviridae, Secoviridae, Partitiviridae, Tymoviridae, Alphaflexiviridae, and Tombusviridae families reflecting the largely insect and plant rodent diet. Phylogenetic analyses of full and partial viral genomes therefore revealed many previously unreported viral species, genera, and families. The close genetic similarities noted between some rodent and human viruses might reflect past zoonoses. This study increases our understanding of the viral diversity in wild rodents and highlights the large number of still uncharacterized viruses in mammals

ICAR: endoscopic skull‐base surgery

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Measurement of the t(t)over-bar production cross section in pp collisions at √s=7 TeV with lepton plus jets final states

This is the pre-print version of the Article. The official published can be accessed from the link below. Copyright @ 2013 ElsevierA measurement of the tt¯ production cross section in pp collisions at √s=7 TeV is presented. The results are based on data corresponding to an integrated luminosity of 2.3 fb−1 collected by the CMS detector at the LHC. Selected events are required to have one isolated, high transverse momentum electron or muon, large missing transverse energy, and hadronic jets, at least one of which must be consistent with having originated from a b quark. The measured cross section is 158.1 ± 2.1 (stat.) ± 10.2(syst.) ± 3.5 (lum.) pb, in agreement with standard model predictions.This study is funded by the: BMWF and FWF (Austria); FNRS and FWO (Belgium); CNPq, CAPES, FAPERJ, and FAPESP (Brazil); MEYS (Bulgaria); CERN; CAS, MoST, and NSFC (China); COLCIENCIAS (Colombia); MSES (Croatia); RPF (Cyprus); MoER, SF0690030s09 and ERDF (Estonia); Academy of Finland, MEC, and HIP (Finland); CEA and CNRS/IN2P3 (France); BMBF, DFG, and HGF (Germany); GSRT (Greece); OTKA and NKTH (Hungary); DAE and DST (India); IPM (Iran); SFI (Ireland); INFN (Italy); NRF and WCU (Republic of Korea); LAS (Lithuania); CINVESTAV, CONACYT, SEP, and UASLP-FAI (Mexico); MSI (New Zealand); PAEC (Pakistan); MSHE and NSC (Poland); FCT (Portugal); JINR (Armenia, Belarus, Georgia, Ukraine, Uzbekistan); MON, RosAtom, RAS and RFBR (Russia); MSTD (Serbia); SEIDI and CPAN (Spain); Swiss Funding Agencies (Switzerland); NSC (Taipei); ThEPCenter, IPST and NSTDA (Thailand); TUBITAK and TAEK (Turkey); NASU (Ukraine); STFC (United Kingdom); DOE and NSF (USA)

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe