Characterization and Biological Activities of Ocellatin Peptides from the Skin Secretion of the Frog Leptodactylus pustulatus

Eight new peptides were isolated from the skin secretion of the frog Leptodactylus pustulatus and their amino acid sequences determined by de novo sequencing and by cDNA cloning. Structural similarities between them and other antimicrobial peptides from the skin secretion of Leptodactylus genus frogs were found. Ocellatins-PT1 to -PT5 (25 amino acid residues) are amidated at the C-terminus, while ocellatins-PT6 to -PT8 (32 amino acid residues) have free carboxylates. Antimicrobial activity, hemolytic tests, and cytotoxicity against a murine fibroblast cell line were investigated. All peptides, except for ocellatin-PT2, have antimicrobial activity against at least one Gram negative strain. Ocellatin-PT8 inhibited the growth of Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, and Salmonella choleraesuis strains with MICs in the 60−240 μM range. No significant effect was observed in human erythrocytes and in a murine fibroblast cell line after exposure to the peptides at MICs. A comparison between sequences obtained by both direct HPLC-MS de novo sequencing and cDNA cloning demonstrates the secretion of mature peptides derived from a pre-pro-peptide structure.

How to teach mathematics with MOODLE

The aim of this article is to present a Project in the Oporto’s Institute of Accounting and Administration, which pretends to contribute for a change in the way of teaching and learning Mathematics. One of the main objectives of this project is to innovate the teaching and learning processes, exploring technologies as a pedagogical resource and to induce higher motivation to students, improve the rate of success and make available to students a set of materials adapted to their needs. This concern is justified due to the fact that students have a weak preparation, without consolidated basis. Since the year 2007/2008 the courses were adjusted to the Bologna process, which requires several changes in teacher’s and student’s roles, methodologies and assessment. The number of weekly classes has been reduced, so it was necessary to develop new strategies and methodologies to support the student. With the implementation of the Bologna Process in the Accounting degree, we felt a great need to provide other types of activities to students. To complement our theoretical and practical classes we have developed a project called MatActiva based on the Moodle platform offered by PAOL - Projecto de Apoio On-Line (Online Support Project). Moodle allows us to use the language TEX to create materials that use mathematical symbols. Using this functionality, we created a set of easy to use interactive resources. In MatActiva project, the students have access to a variety of different materials. We have followed a strategy that makes the project compatible with the theoretical and practical subjects/classes, complementing them. To do so, we created some resources, for instance multiple-choice tests, which are the most accessed by the students. These tests can be realized and corrected on-line and for each wrong answer there is a feedback with the resolution. We can find other types of resources: diagnostic tests, theoretical notes. There are not only the pre-requirements for subjects mathematics, but also materials to help students follow up the programs. We also developed several lessons. This activity consists of a number of pages, where each page has contents and leads to other pages, based on the student's progress. The teacher creates the choices and determines the next page that the student will see, based upon their knowledge. There is also an area of doubts, where the students can place all the mathematical doubts they have, and a teacher gives the answers or clues to help them in their work. MatActiva also offers an area where we can find some humour, curiosities, contests and games including mathematical contents to test the math skills, as well as links to pages about mathematical contents that could be useful for the study. Since ISCAP receives ERASMUS students and some of them attend mathematics, we developed some materials in English, so they can also use MatActiva. The main objectives of our project are not only to bring success in the subjects of mathematics, but also to motivate the students, encourage them to overcome theirs difficulties through an auto-study giving them more confidence and improve their relationship with the mathematics as well as the communication between students and teachers and among students.

A influência da hipertonia biomecânica VS neuronal na selecção das estratégias de intervenção em sujeitos com sequelas de acidente vascular encefálico no território da artéria cerebral média

Introdução: Após lesão do SNC o músculo pode perder a sua variabilidade e flexibilidade, tal como se verifica em indivíduos após AVE. A caracterização do tónus muscular tem sido um indicador a ter em atenção para o diagnóstico clínico. As alterações do tónus podem resultar de uma combinação de alterações neurais, como consequência dos processos inerentes à neuroplasticidade e alterações biomecânicas. Objectivo: Verificar quais as modificações no tónus muscular, segundo a escala de Tardieu, após a aplicação de um programa de reabilitação neuromotora baseado no conceito de Bobath em dois indivíduos com sequelas de AVE. Pretendeu-se também verificar as repercussões nas actividades funcionais. Participantes e métodos: Foram seleccionados dois indivíduos e aplicado um programa de reabilitação, durante onze semanas, e avaliados em dois momentos, antes da intervenção (PRE) e após a intervenção (APÓS). Aplicaram-se vários instrumentos de avaliação, nomeadamente a escala de Tardieu. O programa de reabilitação realizado baseou-se no conceito de Bobath. Resultados: Na escala de Tardieu, foi comum aos dois indivíduos melhorias a nível da qualidade de reacção muscular. Ambos os indivíduos apresentaram melhorias no controlo postural e equilíbrio, que se evidenciaram na CIF. Conclusão: Foi possível observar modificações no tónus muscular após aplicação de um programa de reabilitação e, consequentemente modificações na distribuição da carga na base de suporte, no alinhamento das estruturas articulares e musculares e na marcha. Ao longo da intervenção, observaram-se repercussões positivas em ambos os indivíduos, permitindo a estes realizar as AVDs com menor dificuldade

Avaliação da qualidade química e microbiológica de tintas didácticas do ensino pré-escolar

As tintas utilizadas nas actividades didácticas possuem na sua composição ingredientes que dada a sua natureza, modo de fabrico e de utilização, podem representar um risco para a saúde das crianças. Neste âmbito, procurou-se com este estudo avaliar a qualidade química e microbiológica das tintas utilizadas pelas crianças no ensino pré-escolar. Vinte e nove amostras de tintas, incluindo guaches, tintas de águas, digitintas e tintas para pinturas faciais foram recolhidas em oito estabelecimentos de ensino, nomeadamente, Jardins de Infância, do conselho de Vila Nova de Gaia. A avaliação microbiológica envolveu não só a determinação da concentração microbiana presente nas amostras, como também, a avaliação da estabilidade microbiana nas tintas das espécies S.aureus e E.coli. Na avaliação química procedeu-se à determinação da concentração dos metais chumbo (Pb), cádmio (Cd), crómio (Cr), cobalto (Co), níquel (Nq), manganês (Mn), cobre (Cu) e zinco (Zn) quer em algumas das amostras recolhidas nos estabelecimentos de ensino, quer em tintas adquiridas em três estabelecimentos comerciais. Os resultados obtidos da avaliação microbiológica revelam uma contaminação estática na generalidade das tintas. Três amostras de tintas apresentaram ainda elevada contaminação por fungos, nomeadamente Aspergillus spp. e Trichophyton spp. Da avaliação da estabilidade microbiana das espécies S.aureus e E.coli observou-se uma sensibilidade das mesmas às tintas, sendo evidenciado, em alguns casos, um decrescimento da concentração ao longo do tempo de exposição, e noutros, uma sensibilidade imediata. A espécie S.aureus revelou, contudo, maior capacidade de resistência que a E.coli. Os resultados obtidos da avaliação química revelaram a presença de Cr em todas as amostras, registando as tintas adquiridas em estabelecimentos comerciais concentrações mais elevadas para este metal. Os metais Cu e Zn foram detectados, em algumas amostras de tintas artísticas, em concentrações acima dos valores limites. Nas tintas para a cara foram encontrados os metais Pb, Cd, Cr e Nq, cuja utilização é interdita nestes produtos. O conhecimento das características químicas e microbiológicas das tintas utilizadas por crianças do ensino pré-escolar revelou-se de grande importância, nomeadamente, para a determinação dos riscos a que este grupo de indivíduos pode estar exposto no seu dia-a-dia quando utilizam estes produtos.

Estudo das propriedades psicofisiológicas da pintura como actividade criativa remediativa das manifestações de ansiedade

Ao longo das últimas duas décadas, as intervenções de Arte-Terapia baseada nas actividades criativas de Pintura têm sido implementadas em várias instituições e com variados objectivos e fins terapêuticos, diferindo nas suas abordagens (antroposófica, arte-pedagógica e apenas baseada na arte) e nos seus métodos e materiais aplicados. Constatamos que o recurso a actividades criativas, como a Pintura, provocam efeitos positivos nos indivíduos levando-os a resolver problemas e a reduzir ansiedades, melhorando a sua qualidade de vida. Deste modo é objectivo deste estudo pré-experimental efectuar uma análise das propriedades psicofisiológicas da Pintura como actividade criativa remediativa das manifestações de ansiedade em adultos, sem diagnóstico de perturbação ansiosa. De encontro a esse objectivo, perante uma amostra constituída por 7 indivíduos com níveis de ansiedade elevados, desenvolvemos um processo terapêutico, em que foram realizadas dez sessões de pintura, com a periodicidade de duas vezes por semana tendo cada sessão a duração de 45 min. Antes e depois da intervenção pela pintura, foi administrado o inventário para Avaliação do Traço da Ansiedade (STAY-Y), e durante essas sessões foram registadas as manifestações psicofisiológicas da ansiedade, mais especificamente a actividade electrodérmica da pele (EDA) e a frequência cardíaca. (FC), sendo efectuada a sua análise. Na sequência da análise dos dados obtidos nos dois instrumentos aplicados, verificamos que em relação aos marcadores psicofisiológicos EDA e FC não obtivemos diferenças estatísticas significativas para comprovar que de facto existe uma relação entre estes marcadores e a redução da ansiedade. Em relação ao STAY-Y referente à ansiedade traço, aplicado antes e depois das sessões de pintura, verificamos uma diminuição do score, o que evidência a ocorrência de uma diminuição da ansiedade traço nos indivíduos alvo do nosso estudo. Significa isto que apesar de existir uma variância nos resultados obtidos através das duas abordagens, é provavel que a pintura, enquanto actividade terapêutica, reduza o traço de ansiedade em indivíduos normais, com níveis de ansiedade elevados e sem manifestações de ansiedade diagnosticada.

Scheduling maintenance activities of electric power transmission networks using an hybrid constraint method

In this paper we present a Constraint Logic Programming (CLP) based model, and hybrid solving method for the Scheduling of Maintenance Activities in the Power Transmission Network. The model distinguishes from others not only because of its completeness but also by the way it models and solves the Electric Constraints. Specifically we present a efficient filtering algorithm for the Electrical Constraints. Furthermore, the solving method improves the pure CLP methods efficiency by integrating a type of Local Search technique with CLP. To test the approach we compare the method results with another method using a 24 bus network, which considerers 42 tasks and 24 maintenance periods.

Peculiarities of the professional education and qualification appraisal of assistants of pharmacists in Lithuania

Following the alterations and additions to the pharmaceutical law of Lithuanian Republic the pharmaceutical activity in Lithuanian pharmacies is carried out by pharmacy specialists – pharmacists and pharmacy assistants (pharmacy technicians). The ability of pharmacy specialists to adapt, to improve and to develop their competencies is one of the success preconditions for Lithuanian health care pharmacy sector. Work aim: to investigate the changes among the specialists employed in pharmacies and pharmacy sector as well as to highlight the necessity to develop the qualification of pharmacy assistants (pharmacy technicians) according to the modern pharmaceutical legal basis in Lithuania. In 2008–2011 the research in Lithuanian pharmacy sector was performed which identified the changes in the numbers of pharmacies and their staff: the number of pharmacies decreased, while the number of pharmacists increased and the number of the assistants of pharmacists (pharmacy technicians) decreased. The decrease in the number of the assistants of pharmacists (pharmacy technicians) and the danger of their extinction in the future has been identified. The qualification appraisal for pharmacy assistants (pharmacy technicians) is implemented through the leveling studies at the Pharmacy technique department at the Health care faculty of Kauno Kolegija University of applied sciences and pharmacy study programme at the faculty of Pharmacy of Medical Academy at Lithuanian University of Health Sciences. 75 % of respondents use their own money to pay for the qualification appraisal, and 25 % of respondents get financial support for the qualification appraisal activities from the pharmacy managers.

Synthesis, characterization and antiproliferative activity on cancer cell lines of new ionic liquids from ampicillin

Ionic Liquids (ILs) are ionic compounds that possess melting temperature below 100ºC and they have been a topic of great interest since the mid-1990s due to their unique properties. The range of IL uses has been broadened, due to a significant increase in the variety of physical, chemical and biological ILs properties. They are now used as Active Pharmaceutical Ingredients (APIs) and recent interests are focused on their application as innovative solutions in new medical treatment and delivery options.1 In this work, our principal objective was the synthesis and investigation of physicochemical and medical properties of ionic liquids (ILs) and organic salts from ampicillin. This approach is of huge interest in pharmaceutical industry as cation and anion composition of ILs and organic salts can greatly alter their desired properties, namely the melting temperature and even synergistic effects can be obtained.2,3 For the synthesis of these compounds we used a recently developed method proposed by Ohno et al.4 for the preparation of quaternary ammonium and phosphonium hydroxides, that were neutralized by ampicillin. After purification we obtained pure ILs and salts in good yields. These ILs shows good antimicrobial and antifungal activities. As it is well known that some ionic liquids containing phosphonium and ammonium cation also shows anti-cancer activity1,5 we also decided to study these compounds against some cancer cell lines.

Estágio profissional: Câmara Municipal da Trofa

Relatório de Estágio apresentado ao Instituto Superior de Contabilidade e Administração do Porto para a obtenção do Grau de Mestre em Auditoria Orientador: Rodrigo Mário Oliveira Carvalho, Dr. Coorientador: Vicente António Fernandes Seixas, Dr.

Influence of environmental temperature and humidity on the acute ventilatory response to exercise in asthmatic adolescents

Asthma is a chronic inflammatory disorder of the respiratory airways affecting people of all ages, and constitutes a serious public health problem worldwide (6). Such a chronic inflammation is invariably associated with injury and repair of the bronchial epithelium known as remodelling (11). Inflammation, remodelling, and altered neural control of the airways are responsible for both recurrent exacerbations of asthma and increasingly permanent airflow obstruction (11, 29, 34). Excessive airway narrowing is caused by altered smooth muscle behaviour, in close interaction with swelling of the airway walls, parenchyma retractile forces, and enhanced intraluminal secretions (29, 38). All these functional and structural changes are associated with the characteristic symptoms of asthma – cough, chest tightness, and wheezing –and have a significant impact on patients’ daily lives, on their families and also on society (1, 24, 29). Recent epidemiological studies show an increase in the prevalence of asthma, mainly in industrial countries (12, 25, 37). The reasons for this increase may depend on host factors (e.g., genetic disposition) or on environmental factors like air pollution or contact with allergens (6, 22, 29). Physical exercise is probably the most common trigger for brief episodes of symptoms, and is assumed to induce airflow limitations in most asthmatic children and young adults (16, 24, 29, 33). Exercise-induced asthma (EIA) is defined as an intermittent narrowing of the airways, generally associated with respiratory symptoms (chest tightness, cough, wheezing and dyspnoea), occurring after 3 to 10 minutes of vigorous exercise with a maximal severity during 5 to 15 minutes after the end of the exercise (9, 14, 16, 24, 33). The definitive diagnosis of EIA is confirmed by the measurement of pre- and post-exercise expiratory flows documenting either a 15% fall in the forced expiratory volume in 1 second (FEV1), or a ≥15 to 20% fall in peak expiratory flow (PEF) (9, 24, 29). Some types of physical exercise have been associated with the occurrence of bronchial symptoms and asthma (5, 15, 17). For instance, demanding activities such as basketball or soccer could cause more severe attacks than less vigorous ones such as baseball or jogging (33). The mechanisms of exercise-induced airflow limitations seem to be related to changes in the respiratory mucosa induced by hyperventilation (9, 29). The heat loss from the airways during exercise, and possibly its post-exercise rewarming may contribute to the exercise-induced bronchoconstriction (EIB) (27). Additionally, the concomitant dehydration from the respiratory mucosa during exercise leads to an increased interstitial osmolarity, which may also contribute to bronchoconstriction (4, 36). So, the risk of EIB in asthmatically predisposed subjects seems to be higher with greater ventilation rates and the cooler and drier the inspired air is (23). The incidence of EIA in physically demanding coldweather sports like competitive figure skating and ice hockey has been found to occur in up to 30 to 35% of the participants (32). In contrast, swimming is often recommended to asthmatic individuals, because it improves the functionality of respiratory muscles and, moreover, it seems to have a concomitant beneficial effect on the prevalence of asthma exacerbations (14, 26), supporting the idea that the risk of EIB would be smaller in warm and humid environments. This topic, however, remains controversial since the chlorified water of swimming pools has been suspected as a potential trigger factor for some asthmatic patients (7, 8, 20, 21). In fact, the higher asthma incidence observed in industrialised countries has recently been linked to the exposition to chloride (7, 8, 30). Although clinical and epidemiological data suggest an influence of humidity and temperature of the inspired air on the bronchial response of asthmatic subjects during exercise, some of those studies did not accurately control the intensity of the exercise (2, 13), raising speculation of whether the experienced exercise overload was comparable for all subjects. Additionally, most of the studies did not include a control group (2, 10, 19, 39), which may lead to doubts about whether asthma per se has conditioned the observed results. Moreover, since the main targeted age group of these studies has been adults (10, 19, 39), any extrapolation to childhood/adolescence might be questionable regarding the different lung maturation. Considering the higher incidence of asthma in youngsters (30) and the fact that only the works of Amirav and coworkers (2, 3) have focused on this age group, a scarcity of scientific data can be identified. Additionally, since the main environmental trigger factors, i.e., temperature and humidity, were tested separately (10, 28, 39) it would be useful to analyse these two variables simultaneously because of their synergic effect on water and heat loss by the airways (31, 33). It also appears important to estimate the airway responsiveness to exercise within moderate environmental ranges of temperature and humidity, trying to avoid extreme temperatures and humidity conditions used by others (2, 3). So, the aim of this study was to analyse the influence of moderate changes in air temperature and humidity simultaneously on the acute ventilatory response to exercise in asthmatic children. To overcome the above referred to methodological limitations, we used a 15 minute progressive exercise trial on a cycle ergometer at 3 different workload intensities, and we collected data related to heart rate, respiratory quotient, minute ventilation and oxygen uptake in order to ensure that physiological exercise repercussions were the same in both environments. The tests were done in a “normal” climatic environment (in a gymnasium) and in a hot and humid environment (swimming pool); for the latter, direct chloride exposition was avoided.

Influence of the pre-treatments on the properties of biodegradable films from bovine hair

The replacement of conventional synthetic films and coatings by biodegradable alternatives reduces the use of non-renewable resources and waste disposal problems. Considering that Portugal is a major producer of leather, and consequently a large producer of related wastes, in this research, bovine hair was tested for the production of biodegradable films directly by thermo-compression, allowing waste valorisation and reduction of environmental pollution. The aim of this study was to determine the influence of the different pre-treatments performed by two processes (removal by mechanical action and removal by chemical process), applied to bovine hair, in order to obtain a biodegradable film with appropriate properties. Mechanical properties for these films were evaluated, namely strain at break, stress at break and Young modulus. Additionally colour, solubility and swelling in water were also studied. The mechanical removal hair only produced films with Na2S treatment. Chemical removed hair (immunization) depends of the pre-treatment and the degreasing with petroleum ether or sodium sulphide pre-treatment leads better mechanical properties. The results obtained indicated that the pre-treatments have an important role in the final properties of biodegradable films.

Cyclotech - contribution to solve the Technetium shortage by using low energy medical cyclotrons

This paper presents work in progress, to develop an efficient and economic way to directly produce Technetium 99metastable (99mTc) using low-energy cyclotrons. Its importance is well established and relates with the increased global trouble in delivering 99mTc to Nuclear Medicine Departments relying on this radioisotope. Since the present delivery strategy has clearly demonstrated its intrinsic limits, our group decided to follow a distinct approach that uses the broad distribution of the low energy cyclotrons and the accessibility of Molybdenum 100 (100Mo) as the Target material. This is indeed an important issue to consider, since the system here presented, named CYCLOTECH, it is not based on the use of Highly Enriched (or even Low Enriched) Uranium 235 (235U), so entirely complying with the actual international trends and directives concerning the use of this potential highly critical material. The production technique is based on the nuclear reaction 100Mo (p,2n) 99mTc whose production yields have already been documented. Until this moment two Patent requests have already been submitted (the first at the INPI, in Portugal, and the second at the USPTO, in the USA); others are being prepared for submission on a near future. The object of the CYCLOTECH system is to present 99mTc to Nuclear Medicine radiopharmacists in a routine, reliable and efficient manner that, remaining always flexible, entirely blends with established protocols. To facilitate workflow and Radiation Protection measures, it has been developed a Target Station that can be installed on most of the existing PET cyclotrons and that will tolerate up to 400 μA of beam by allowing the beam to strike the Target material at an adequately oblique angle. The Target Station permits the remote and automatic loading and discharge of the Targets from a carriage of 10 Target bodies. On other hand, several methods of Target material deposition and Target substrates are presented. The object was to create a cost effective means of depositing and intermediate the target material thickness (25 - 100μm) with a minimum of loss on a substrate that is able to easily transport the heat associated with high beam currents. Finally, the separation techniques presented are a combination of both physical and column chemistry. The object was to extract and deliver 99mTc in the identical form now in use in radiopharmacies worldwide. In addition, the Target material is recovered and can be recycled.

A eficácia entre duas intervenções na Síndrome do Canal Cárpico

A Síndrome do Canal Cárpico (SCC) é a neuropatia compressiva mais comum do membro superior, causada pela compressão direta sobre o nervo mediano no interior do canal cárpico.Os resultados deste estudo mostram em cada um dos grupos, após a intervenção, uma melhoria estatisticamente significativa da sintomatologia no G-AFN (p=0,02) e no GTRN/ EAA (p=0,004) e uma melhoria estatisticamente significativa do estado funcional no G-AFN (p=0,022). Verificamos também em cada um dos grupos, após a intervenção, uma melhoria estatisticamente significativa na “Força de preensão” (p=0,005), na “Pinça polegar/dedo indicador” (p=0,021), na “Pinça polegar/dedo médio” (p=0,026) e “Pinça polegar/dedo anular” (p=0,026) no G-AFN, e uma melhoria estatisticamente significativa na “Pinça polegar/indicador” (p=0,016), na “Pinça polegar/dedo médio” (p=0,035), na “Pinça polegar/dedo anular” (p=0,010), na “Pinça trípode” (p=0,005) e na “Pinça lateral” (p=0,051) no G-TRN/EAA. Após a intervenção, não verificamos diferenças estatisticamente significativas nos valores das escalas de gravidade de sintomas (p=0,853) e de estado funcional (p=0,148) entre os grupos, mas diferenças estatisticamente significativas nos valores dos testes neurofisiológicos (p=0,047) e força de preensão da mão (p=0,005). Do estudo, concluímos que a utilização da intervenção articular/fascial/neural (AFN) e a intervenção com tala de repouso noturna e exercícios de auto alongamento (TRN/EAA), beneficia os indivíduos com SCC não severa, como nos casos incipientes, ligeiros ou moderados. Os indivíduos com esta condição clínica apresentam sintomatologia caraterística de dor, parestesia, especialmente noturna e disfunção muscular da mão. Tais manifestações originam perda funcional com implicações nas áreas de desempenho ocupacional, nomeadamente, nas atividades da vida diária, produtivas e de lazer. O tratamento conservador na SCC não severa, como nos casos incipientes, ligeiros e moderados, apesar de controverso, é recomendado. O tema suscita o nosso interesse, razão pela qual nos propomos realizar um estudo experimental em indivíduos com o diagnóstico clínico de SCC não severa e aplicar num grupo a intervenção articular, fascial e neural (AFN) e noutro grupo a intervenção com tala de repouso noturna e exercícios de auto alongamento (TRN/EAA). O estudo tem como principais objetivos, por um lado, verificar o impacto das intervenções em cada um dos grupos e, por outro lado, comparar o seu impacto entre os grupos, no que respeita à gravidade de sintomas, ao estado funcional, à força de preensão da mão e força de pinças finas. Fomos também comparar os resultados dos testes neurofisiológicos (Velocidade de Condução Motora) antes e depois da intervenção AFN e da intervenção com TRN/EAA, e averiguar o seu impacto nos valores da latência motora distal e da velocidade de condução sensitiva, entre os grupos. Identificamos também quais as variáveis sócio demográficas e as que caraterizam a patologia que estão relacionadas com o problema em estudo e com os valores obtidos com as escalas do Boston Carpal Tunnel Questionnaire (BCTQ), no grupo articular, fascial e neural (G-AFN) e no grupo com tala de repouso noturna e exercícios de auto alongamento (G-TRN/EAA). Para a concretização do estudo, recorremos a uma amostra de 23 indivíduos de ambos os sexos do Hospital Curry Cabral, Empresa Pública Empresarial -Centro Hospitalar de Lisboa Central (HCC, EPE -CHLC).