MUEGANO: A divide and conquer algorithm to overcome memory limitations when assembling shotgun projects

When assembling a large quantity of reads in a genomic shotgun project a serious limitation is the amount of random access memory (RAM) of the computers used in the project. This arises because all assembling programs must look at all the overlaps between reads at the same time, using RAM in order to construct contigs, and the memory of the computer can be filled up during this step, causing the abortion of the assembling process.
Here we propose an algorithm that is capable of overcoming any memory limitation by using redundancy of processing and thus producing an increase in computing time but overcoming the memory limitation.
The proposed algorithm consists in dividing the reads in a set of groups which size is half the maximum capability in memory of the computer used and performing assembling for all the possible combination pairs of such groups. After eliminating the redundancy of the set of contigs obtained in the previous step, the process is iterated until a set of contigs of manageable size is obtained such that the set can be handled by the assembler in a final step.
Each step of the procedure increases the time of computing from k to approximately k + k(k-1)/2, but in many practical cases only one step is needed to finish the assembling process. The procedure is suitable for any kind of assembler and was successfully applied to the assembly of a very large set of reads from the maize genome

Energy-time uncertainty principle and lower bounds on sojourn time

One manifestation of quantum resonances is a large sojourn time, or autocorrelation, for states which are initially localized. We elaborate on Lavine's time-energy uncertainty principle and give an estimate on the sojourn time. For the case of perturbed embedded eigenstates the bound is explicit and involves Fermi's Golden Rule. It is valid for a very general class of systems. We illustrate the theory by applications to resonances for time dependent systems including the AC Stark effect as well as multistate systems.Comment: Version to appear in Annales Henri Poincar\'

Profile of residents: attitude towards tourism in Benalmádena (Costa del Sol, Spain)

Tourism development and evolution along the time causes multiple impacts. Based on characteristic profiles of resident, these impacts may be assessed differently. The attitude of residents and the identification of profiles is a tool that allows to asses the status of destination and propose future strategies for improving the destination. This study aims to analyse the role of residents regarding the perception of tourism.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Así mismo, Esta investigación ha sido subvencionada por el programa Nacional de Investigación Básica 2012, CS2010-30840. "Geografías de la crisis: analisis urbano turístico de las Islas Baleares, Costa del Sol y principales destinos en el Caribe

Residents´ perceptions of tourism development in Benalmádena (Spain)

DOI: 10.1016/j.tourman.2015.11.007This study examines the residents´ perceptions of the impact of tourism in Benalmádena, and the profiles of the residents according to socio-demographic characteristics. A questionnaire assessed how these characteristics influence the residents' perceptions towards the environment, economy, and socio-cultural aspects. The survey was administered to a stratified sample of 770 residents in Benalmádena. Results show a significant effect of socio-demographic variables on perception of tourism impact. The educational background, place of birth and how long respondents had been living in the community explain a significant amount of the variance in overall attitudes. Interaction analyses revealed that place of birth moderated the relationship between the tourism dimensions and the years of residence. For instance, the respondents with less than five years of residence showed more positive attitude towards the impact of tourism. We offer a profile of these residents according to their perceptions of the impact of tourism in their community.This research has received funding from the Spanish Government, Fundamental Research Program (R+D) (CSO2012-30840), “Geographies of crisis: analysis of urban and tourist territories of the Balearic Islands, Costa del Sol and main tourist destinations of the Caribbean and Central America”. Andalucia Tech, Universidad de Málaga, Spain

Hamiltonization of Nonholonomic Systems and the Inverse Problem of the Calculus of Variations

We introduce a method which allows one to recover the equations of motion of a class of nonholonomic systems by finding instead an unconstrained Hamiltonian system on the full phase space, and to restrict the resulting canonical equations to an appropriate submanifold of phase space. We focus first on the Lagrangian picture of the method and deduce the corresponding Hamiltonian from the Legendre transformation. We illustrate the method with several examples and we discuss its relationship to the Pontryagin maximum principle.Comment: 23 pages, accepted for publication in Rep. Math. Phy

A rational quest for drug targets in the protein kinome of Trypanosoma brucei

Trypanosoma brucei is the protozoan parasite causing African trypanosomiasis, a neurological disease that affects humans and farm-stock in the impoverished sub-Saharan areas where tsetse fly transmission vector is endemic. Although it has great impact on public health and local economies, it has been neglected in drug discovery for almost a century. Current treatments are either toxic or of difficult administration, besides having serious risks of inducing resistance. Protein kinases are the primary set of signaling proteins in eukaryotes, including Trypanosoma brucei. Their druggability has been widely exploited in cancer research, and has been established in the parasite too. A recent kinome-wide RNAi screen with 176 individual cell lines of mammalian infective bloodstream forms of Trypanosoma brucei identified protein kinases required for proliferation in vitro. In order to investigate which protein kinases are also essential virulence factors in vivo, lines were pooled, inoculated into mice and screened for loss of fitness after 48 hours RNAi compared to uninduced controls. The presence of trypanosomes in the bloodstream was assessed using RNAi target sequencing (RITseq) and compared to an in vitro control. This revealed 49 protein kinases with a significant loss of fitness in vivo in two independent experiments, and a strong correlation between in vitro and in vivo loss of fitness for the majority. However, depletion of nine protein kinases affected more pronouncedly the growth in vivo than in vitro. Amongst these protein kinases were several with putative functions related with stress responses mediated through the PI3K/TOR or MAPK signaling cascades including CK2A2, a promiscuous protein kinase whose activity can be stress-induced; two MAP3Ks, involved in cell integrity upon osmotic shock; VPS15, component of the PI3K complex with roles in autophagosome formation and vesicular trafficking; BUD32, transducer of the PI3K/TOR pathway involved in translational regulation; and FAZ20, a parasite-specific pseudo-kinase localizing to the flagellum attachment zone. The other three have been implicated in repair of alkylation-induced cellular damage: SRPK1, a stress response RNA splicing regulator; AUK2, which acts during mitosis; and CAMKL, an AMPK with calcium-binding domains putatively involved in metabolic regulation. Identification of these virulence-associated protein kinases provides new insights in T. brucei-host interaction and reveals novel potential drug targets for protein kinase inhibitors. This RNAi screen revealed that the evolutionary divergent NEK kinase Repressor of Differentiation Kinase 2 (RDK2) has severe loss of fitness both in vitro and in vivo. Depletion of RDK2 had been shown previously to promote differentiation from bloodstream to procyclic-like forms causing the parasite’s death. Further investigation showed RDK2 to be an active protein kinase capable both of phosphorylating a substrate and to autophosphorylate. Protein kinase activity could be ablated by mutation of lysine 70 to methionine. Mutation of both serine residues (195 and 197), identified as sites of phosphorylation by phosphoproteomics, to alanine or glutamic acid, preventing and mimicking phosphorylation respectively, had no effect on protein kinase activity, suggesting they do not have a direct regulatory role on protein kinase activity. Introducing in the RNAi line a recoded RDK2 whose transcript eluded interference, permitted to some extent the rescue of the induced phenotype, while introducing a recoded inactive mutant did not. This may suggest that the lack of kinase activity was responsible for the RNAi phenotype and not depletion of the protein alone. RDK2 RNAi-differentiated cells could be maintained in conditioned procyclic form media for more than a week. However, they were unable to proliferate. Overexpression of RDK2 blocked the differentiation mediated by sequential treatment with 8-pCPT-cAMP and citrate/cis-aconitate (CCA). RNAi experiments in combination with known differentiation cues, suggested that when differentiation is triggered by the CCA signalling pathway, RDK2 inactivation happens downstream of the phosphatase TbPTP1. Differentiation caused by RDK2 inactivation could be traced in flow cytometry by the detection of EP procyclin expression. This was exploited in a cell-based mechanism-directed phenotypic screen for RDK2 inhibitors. A preliminary run with 518 drug-like molecules that had shown protein kinase inhibition, trypanocidal activity and/or activation of the EP procyclin promoter, unveiled 6 compounds triggering EP procyclin expression and parasite death in the low micromolar range. These compounds can be investigated further to assess whether RDK2 is their in vivo target

A multi-parametric analysis of Trypanosoma cruzi infection: common pathophysiologic patterns beyond extreme heterogeneity of host responses

The extreme genetic diversity of the protozoan Trypanosoma cruzi has been proposed to be associated with the clinical outcomes of the disease it provokes: Chagas disease (CD). To address this question, we analysed the similarities and differences in the CD pathophysiogenesis caused by different parasite strains. Using syngeneic mice infected acutely or chronically with 6 distant parasite strains, we integrated simultaneously 66 parameters: parasite tropism (7 parameters), organ and immune responses (local and systemic; 57 parameters), and clinical presentations of CD (2 parameters). While the parasite genetic background consistently impacts most of these parameters, they remain highly variable, as observed in patients, impeding reliable one-dimensional association with phases, strains, and damage. However, multi-dimensional statistics overcame this extreme intra-group variability for each individual parameter and revealed some pathophysiological patterns that accurately allow defining (i) the infection phase, (ii) the infecting parasite strains, and (iii) organ damage type and intensity. Our results demonstrated a greater variability of clinical outcomes and host responses to T. cruzi infection than previously thought, while our multi-parametric analysis defined common pathophysiological patterns linked to clinical outcome of CD, conserved among the genetically diverse infecting strains

High-throughput mass spectrometry analysis of N -glycans and protein markers after FUT8 knockdown in the syngeneic SW480/SW620 colorectal cancer cell model

Disruption of the glycosylation machinery is a common feature in many types of cancer, and colorectal cancer (CRC) is no exception. Core fucosylation is mediated by the enzyme fucosyltransferase 8 (FucT-8), which catalyzes the addition of α1,6-l-fucose to the innermost GlcNAc residue of N-glycans. We and others have documented the involvement of FucT-8 and core-fucosylated proteins in CRC progression, in which we addressed core fucosylation in the syngeneic CRC model formed by SW480 and SW620 tumor cell lines from the perspective of alterations in their N-glycosylation profile and protein expression as an effect of the knockdown of the FUT8 gene that encodes FucT-8. Using label-free, semiquantitative mass spectrometry (MS) analysis, we found noticeable differences in N-glycosylation patterns in FUT8-knockdown cells, affecting core fucosylation and sialylation, the Hex/HexNAc ratio, and antennarity. Furthermore, stable isotopic labeling of amino acids in cell culture (SILAC)-based proteomic screening detected the alteration of species involved in protein folding, endoplasmic reticulum (ER) and Golgi post-translational stabilization, epithelial polarity, and cellular response to damage and therapy. This data is available via ProteomeXchange with identifier PXD050012. Overall, the results obtained merit further investigation to validate their feasibility as biomarkers of progression and malignization in CRC, as well as their potential usefulness in clinical practiceXunta de Galicia | Ref. CN 2011/024Xunta de Galicia | Ref. GRC 2014/019Ministerio de Educación, Cultura y Deporte | Ref. FPU12/03662Universidade de Vigo/CISUGInstituto de Salud Carlos III | Ref. PT13/000