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La presente tesis tiene como título Eficiencia de la implementación de la gestión por procesos en la Municipalidad Distrital de los Olivos - 2018 y tiene como objetivo general determinar la relación entre la eficiencia en la implementación y la gestión por procesos. Es una investigación descriptiva correlacional, de diseño no exsperimental con un enfoque cuantitativo. La población estuvo conformada por 200 trabajadores de la Municipalidad Distrital de los Olivos – 2018. Se utilizaron como instrumento para la recolección de datos dos cuestionarios, uno para la variable Eficiencia en la Implementación con tres dimensiones y otro para la variable Gestión por Procesois con tres dimensiones, los resultados muestran una correlación alta con un rho = 0.751 y significativa con un p = 0.000. Universidad Nacional de Educación Enrique Guzmán y Valle Tesis

Tropical forests are converted at an alarming rate for agricultural use and pastureland, but also regrow naturally through secondary succession. For successful forest restoration, it is essential to understand the mechanisms of secondary succession. These mechanisms may vary across forest types, but analyses across broad spatial scales are lacking. Here, we analyse forest recovery using 1,403 plots that differ in age since agricultural abandonment from 50 sites across the Neotropics. We analyse changes in community composition using species-specific stem wood density (WD), which is a key trait for plant growth, survival and forest carbon storage. In wet forest, succession proceeds from low towards high community WD (acquisitive towards conservative trait values), in line with standard successional theory. However, in dry forest, succession proceeds from high towards low community WD (conservative towards acquisitive trait values), probably because high WD reflects drought tolerance in harsh early successional environments. Dry season intensity drives WD recovery by influencing the start and trajectory of succession, resulting in convergence of the community WD over time as vegetation cover builds up. These ecological insights can be used to improve species selection for reforestation. Reforestation species selected to establish a first protective canopy layer should, among other criteria, ideally have a similar WD to the early successional communities that dominate under the prevailing macroclimatic conditions.

Trabajo presentado en el European Congress of Virology, celebrado en Rotterdam (Países Bajos), del 28 de abril al 1 de mayo de 2019, Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel human coronavirus that emerged in 2012 and is still active in the Middle East, causing severe pneumonia and acute respiratory distress syndrome (ARDS), with a case fatality rate of ~36%. CoV accessory proteins are not essential for replication in cell culture, although they have been involved in virus pathogenicity in vivo. When these proteins are expressed in isolation, they interfere with innate antiviral signaling pathways. However, there is limited information on the specific contribution of MERS-CoV accessory protein 4b to the repression of the innate antiviral response in the context of infection. We found that MERS-CoV 4b was required to prevent a robust NF-κB-dependent response during infection. In wild-type virus infected cells, 4b localized to the nucleus, while NF-κB was retained in the cytoplasm. In contrast, in the absence of 4b or in the presence of cytoplasmic 4b proteins mutated in its nuclear localization signal (NLS), NF-κB was translocated to the nucleus leading to a significant high expression of pro-inflammatory cytokines, as compared to the wild-type infection. This indicates that NF-κB repression required the nuclear import of 4b mediated by a specific NLS. Interestingly, we also found that 4b interacted with α-karyopherin proteins in an NLS-dependent manner. In particular, 4b had a strong preference for binding karyopherin-α4 (KPNA4), which is known to translocate the NF-κB protein complex into the nucleus. Binding of 4b to KPNA4 during infection inhibited its interaction with NF-κB-p65 subunit, thus preventing NF-κB nuclear import. As a consequence of 4b nuclear translocation, KPNA4 protein was also retained in the nucleus. Thereby we propose a model where 4b outcompetes NF-κB for KPNA4 binding and translocation into the nucleus as a mechanism of interference with the NF-κB-mediated innate immune response.

The brain-computer interface research group of the University of Malaga (UMA-BCI) has developed a speller application based on the wellknown P300 potential which can be easily installed, configured and used. The application supports the common P300 paradigms: the Row-Column Paradigm and the Rapid Serial Visual Presentation Paradigm. The inner core of the application is implemented with a widely used and studied platform, BCI2000, which ensures its reliability and allows other researchers to apply modifications at will in order to test new features. There are many studies regarding brain-controlled spellers; however, these systems do not usually leap out of the lab because of technical and economic requirements. As a consequence, the potential end users do not benefit from these scientific advances in their daily life. The objective of this paper is to present a novel brain-controlled speller designed to be used by patients due to its versatility and ease of use. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech.

Trabajo presentado en el European Congress of Virology, celebrado en Rotterdam (Países Bajos), del 28 de abril al 1 de mayo de 2019, Introduction: With the ongoing outbreaks and 35% case fatality rate, Middle East respiratory syndrome coronavirus (MERS-CoV), continues to pose a global threat. However, licensed vaccines and therapeutics for MERS-CoV, including monoclonal antibodies, are not currently available. Camelids have unique heavy-chain-only antibodies with long finger-like complementary determining regions allowing accessibility to epitopes inaccessible to conventional antibodies. The variable domain of these antibodies (VHHs), known as nanobodies, have small size, high antigen affinity and high homology to human counterparts, making them potential candidates for clinical use. Aim: We aimed at generating MERS-CoV-specific VHHs targeting different epitopes within the MERS-CoV spike protein to prevent viral neutralization escape and provide efficient protection against viral infection. Methods: Using bone marrow from MERS-CoV vaccinated dromedary camels, we cloned and expressed the VHH repertoire in E. coli. From this library, we screened for and characterized MERS-CoV-specific VHHs targeting different domains on the vial spike protein. We selected potent VHHs and fused them to a human Fc, generating camel/human monoand bi-specific chimeric heavy-chain-only antibodies (cHCAbs). Results: We identified several potent MERS-CoV-specific VHHs targeting the viral spike receptor binding domain (RBD) and the sialic acid binding (S1A ) domain, thereby blocking virus entry and attachment, respectively. Those with the highest potencies were selected and fused to human Fc to extend their half-life in vivo. The cHCAbs directed against the RBD were capable of blocking MERS-CoV entry in vitro and in vivo. S1A -directed cHCAbs blocked virus attachment to sialic acids and reduced infection in vitro. Different combinations of cHCAbs and bi-specific cHCAbs were evaluated for their protective efficacy against different virus variants and escape mutants. Conclusion: The pathogenesis of a viral disease is the biological mechanism, or mechanisms, that lead to the diseased state. There are several possibilities for comparing the pathogenesis of viral diseases among host species: a viral disease, e.g. rabies, that naturally infects different host species; a viral disease in an experimentally infected laboratory animal species, e.g. avian influenza in ferrets, as a model for the disease in humans; an emerging viral disease, e.g. Marburg haemorrhagic fever, in its reservoir species and its new host species. We here focus on the latter possibility. Many emerging viral diseases in a given host species originate from another host species, which acts as a reservoir. While the clinical outcome of emerging virus infections ranges from subclinical to fatal, we usually focus on those virus infections that cause severe disease in the new host species. Currently, research concentrates on the pathogenesis of such a viral disease in its new host species or, if that is not possible, in a laboratory animal species acting as a model. However, research on the pathogenesis of emerging viral diseases in their reservoir host species is neglected. This is a pity, because there are several good reasons to compare the pathogenesis of emerging viral diseases between the new host species and the reservoir species. First, pathogenesis in the reservoir host species may predict several types of virus-host interaction in the new host species. Second, differences in the pathogenesis between reservoir host species and new host species may provide clues for the mechanism of increased severity of disease in the new host species. Third, pathogenesis in the reservoir host species may predict the future evolution of the emerging virus in the new host species. Therefore, we advocate an increased emphasis on understanding the pathogenesis of emerging viral diseases in their reservoir species.

Trabajo presentado en el European Congress of Virology, celebrado en Rotterdam (Países Bajos), del 28 de abril al 1 de mayo de 2019, Middle East respiratory syndrome coronavirus (MERS-CoV) is a life-threatening human virus that emerged in 2012. To date, more than 2,200 cases have been reported with 35% fatality rate. There is limited information on MERSCoV pathogenesis, as data from patients are scarce and most animal models do not reproduce the clinical manifestations. A mouse knock-in (KI) model and mouse-adapted MERS-CoV (MERS-MA) were recently obtained. Infection with MERS-MA was lethal and the animals showed pathological sings of respiratory disease and high viral titers in the lung. The 13 non-synonymous mutations contained in MERS-MA genome were introduced in our MERSCoV infectious cDNA, leading to a recombinant mouse-adapted virus (rMERS-MA) that was virulent in the KI mice. MERS-CoV adaptation to cell culture or mouse leads to mutations and deletions in genus-specific gene 5, avoiding full-length protein 5 expression. In contrast, analysis of 476 MERS-CoV field isolates showed that gene 5 is highly stable in camels and humans. To study the role of protein 5, two additional viruses were engineered expressing a full-length gene 5 (rMERS-MA-5FL), or containing a complete gene 5 deletion (rMERS-MA-∆5). rMERS-MA-5FL virus was unstable, as partial deletions within gene 5 sequence appeared from the first passages, highlighting MERS-CoV instability in cell cultures. The virulence of rMERS-MA-∆5 was analyzed in a sublethal KI mouse model. Gene 5 was not required for replication in the lung of KI mice, as both mutant and parental viruses reached similar titers. Interestingly, all mice infected with rMERS-MA-∆5 died while all the animals infected with the parental virus survived, indicating that rMERS-MA-∆5 was more virulent than the parental one. Lung damage, interferon and proinflammatory cytokines expression analysis is in progress. Overall, these data indicated that the gene 5 sequence present in the mouse-adapted virus had a function modulating MERS-CoV pathogenesis.

Trabajo presentado en el Keystone Symposia on Molecular and Cellular Biology. Positive-Strand RNA Viruses, celebrado en Killarney, Kerry (Irlanda), del 9 al 13 de junio de 2019, Coronaviruses (CoV) are pathogens responsible for a wide range of existing and emerging diseases. CoVs of genera, and encode proteins that have a PDZ-binding motif (PBM), which may bind over 400 cellular proteins containing PDZ domains, making them relevant for the control of cell function. Two highly pathogenic human CoVs have been identified to date: SARS-CoV and MERS-CoV. In this communication, the relevance of these CoV PBMs in virulence has been studied. First, recombinant SARS-CoV and MERS-CoV variants missing each of these PBMs were generated. Their pathogenicity was analyzed in mice showing that the PBMs of the E proteins of these two viruses were virulence factors, in contrast to the PBMs of proteins 3a and 5 of these viruses, respectively. To study the mechanisms of virulence induced by these PBMs, the cellular PDZ domains interacting with them were identified using a specific yeast-two-hybrid platform restricted to the 256 known human PDZ. More than 20 cellular proteins were identified, and four (syntenin, PTPN13, DLG1 and LNX2) were selected for further study as they interacted with SARS-CoV and MERS-CoV E protein PBMs and are involved in the pro-inflammatory response. It was shown that syntenin activated p38MAPK which induced an exacerbated proinflammatory response leading to virus pathogenesis. In view of the published data, the four cellular PDZ proteins may act by promoting the activation of p38MAPK indicating that these four pathways converged to produce the same effect. In fact, inhibition of p38MAPK lead to the survival of at least 80% of the infected mice, confirming the relevance of this signaling pathway. Preliminary data showed that syntenin PDZ had significantly higher avidity for SARS-CoV and MERS-CoV E protein PBMs than for the corresponding motifs of proteins 3a and 5 of these viruses, suggesting a potential correlation between the avidity of cellular PDZs for viral PBMs, and virus virulence. These results strongly suggest that CoVs PBMs are potential targets for antiviral therapy., This work was supported by grants of the Science Department (Spain), European Commission and NIH (USA).