Cellular determinants of Herpes simplex virus 1 (HSV-1) invasion in human skin: Unravelling conditions that allow HSV-1 to reach its receptor and internalize

Herpes simplex virus 1 (HSV-1), which is among the most prevalent of human pathogens, targets mucosa, skin or cornea. The overall aim of this thesis was to explore how HSV-1 is able to invade the highly-protected tissue structure of the human skin, access its cellular receptors on target skin cells for entry, and initiate infection in the epithelium. On the cellular level, the focus was on elucidating the impact of cellular entry mechanisms in successful infection. Previous studies have demonstrated that HSV-1 exploits dual modes of uptake—direct fusion at the plasma membrane and endocytic internalization—to gain entry in keratinocytes, however, attributing the contribution of either pathway is still open. Here, infection of human keratinocytes at low temperature (7°C) was used as a tool to selectively block energy-requiring endocytic processes while permitting plasma membrane fusion. Although the uptake of the endocytic markers was inhibited at low temperature, ultrastructural analyses revealed the presence of free capsids in the cytoplasm as well as enveloped virus particles in vesicles after infection at 7°C. In addition to virus particles undergoing direct fusion at the plasma membrane, characterization of virus-containing vesicles revealed the release of capsids from vesicles by fusion with vesicle membranes. These results strengthen the role of endocytic internalization in successful infection. At the tissue level, the emphasis was on elucidating how HSV-1 overcomes the highly protective barrier function of the human skin to engage a cellular receptor for entry. To gain insights on the early entry events during HSV-1 invasion, ex vivo infection studies using human skin explants were implemented. As expected, the barrier function of full-thickness skin prevented HSV-1 invasion from the apical skin surface. Once the epidermis was separated from the underlying dermis, HSV-1 efficiently infected basal keratinocytes and later gained access to the suprabasal layers. Viral replication inhibitor experiments as well as uptake of labelled latex beads (500 nm) demonstrated the role of virus-induced tissue damage in enabling HSV-1 access to all epidermal layers. In contrast, only single infected cells were detected in the most apical part of the papillary dermis demonstrating that the extracellular matrix acts as a barrier against HSV-1 invasion. Strikingly, while partly open skin lesions of wounded full-thickness skin allowed the enhanced uptake of latex beads, nearly no infected cells were identified at the sites of wounds. Intriguingly, when wounds reached through the dermis, HSV-1 infected epidermal keratinocytes via the damaged dermal layer. To further elucidate the impact of epidermal barrier function in restricting HSV-1 invasion, pathological skin conditions characterized by impaired epidermal barriers were explored. As ex vivo infection of lesional atopic dermatitis (AD) demonstrated viral penetration from the skin surface, Th2 cytokines interleukin (IL)-4 and IL-13 were employed to induce AD-like phenotypes in skin without pre-existing barrier defects. Indeed, infected cells in the epidermis of IL-stimulated skin were found indicating that Th2 cytokine-driven inflammatory responses induced modifications that facilitate HSV-1 invasion via the skin surface. Overcoming epidermal barriers must be accompanied by viral engagement of a cellular receptor to initiate infection in the skin. Thus, the distribution of nectin-1, the major receptor on keratinocytes was investigated. To dissect how barrier formation correlated with receptor accessibility, a human epidermal equivalent (HEE) based on primary human keratinocytes was adopted. In undifferentiated keratinocytes, nectin-1 is readily accessible at apical and basolateral surfaces correlating with a high susceptibility to HSV-1. In fully differentiated HEEs, nectin-1 is expressed at the lateral membranes of most epidermal layers although accessibility to the receptor is restricted by functional tight junctions (TJs) in the upper granular layer thus correlating with no infection. Intriguingly, while IL-4/IL-13 stimulation of HEEs resulted in redistributed nectin-1 and TJ components, the IL-induced modifications had very minor effects on facilitated HSV-1 invasion in fully differentiated epidermal equivalents in contrast to human skin explants where IL-stimulation allowed HSV-1 invasion. Moreover, redistributed TJ components in IL-4/IL-13-stimulated skin and AD skin, both of which promote HSV-1 invasion, support the role of TJ barrier defects in the facilitated access of HSV-1 to nectin-1

Logística Reversa: Como Reaproveitar Placas de Petri no Fluxo de Trabalho de Um Laboratório de Microbiologia

A cadeia de suprimentos de instituições de ensino públicas ou privadas, voltadas para o diagnóstico laboratorial microbiológico na área de saúde, deve ser gerenciada seguindo normas de qualidade e legislações pertinentes à biossegurança, onde o ambiente e uso de materiais estéreis são fundamentais. O custo elevado pela grande demanda de insumos laboratoriais plásticos estéreis é um fator importante nas organizações, com foco em gestão da saúde, sendo relevante a logística reversa de sobras de insumos das embalagens abertas, que não podem voltar ao fluxo de trabalho, por não serem mais estéreis. O objetivo deste relato técnico é descrever como foi possível realizar o reaproveitamento de sobras de placas de Petri (não mais estéreis), advindas de embalagens abertas, através do emprego do tratamento por radiação ionizante, em parceria com o Instituto de Pesquisas Energéticas e Nucleares. O relato técnico apresenta, através da pesquisa-ação, como foi possível reduzir custos e empregar a logística reversa no reaproveitamento de 400 placas de Petri irradiadas com a dose de 20 kGy, contribuindo para a sustentabilidade

Rapid detection of group B streptococcus and Escherichia coli in amniotic fluid using real-time fluorescent PCR.

OBJECTIVE: To establish reliability and validity of real-time fluorescent PCR for early detection of bacterial invasion of the amniotic cavity. METHODS: Amniotic fluid samples from 40 patients undergoing mid-trimester genetic amniocentesis were incubated for 6 h at 37 degrees C and were cultured on media specific for group B streptococcus (GBS) and E. coli. Concurrently, samples were analyzed with real-time fluorescent PCR (Roche LightCycler) using DNA primers and probes designed to detect the CAMP factor encoding cfb gene and uidA gene of GBS and E. coli, respectively. For positive control and to simulate amniotic fluid colonization, 104 cfu/ml of GBS and E. coli were inoculated on sterile amniotic fluid and incubated for 6 h. Bacterial genomic DNA for the two organisms was extracted and purified via the two-step precipitation method using a commercial kit. The real-time PCR assays were also tested against 25 non-GBS and non-E. coli bacterial species. The lower limit of detection for each pathogen was established using serial dilution of bacterial genomic DNA. RESULTS: All patient samples were negative for evidence of GBS and E. coli with both culture and real-time PCR methods. Amniotic fluid samples inoculated with GBS and E. coli were positive with real-time PCR whereas the 25 bacterial species other than GBS or E. coli tested negative with the assay. Average total sample processing time including the pre-enrichment step was 7 h 40 min. The average cost for DNA extraction and PCR testing was 8.50 dollars per test. CONCLUSION: Real-time fluorescent PCR is a valid and reliable method for detection of specific pathogens in amniotic fluid. This technique is sensitive for low inoculation levels. Real-time fluorescent PCR has potential to impact clinical management as a rapid, reliable detection method for GBS and E. coli in chorioamnionitis

Comparative Evaluation of the Twisted File™ and Revo-S® Rotary Systems Using Cone Beam Computed Tomography

Svrha: Korištenjem konične kompjutorizirane tomografije (CBCT-a) željela se odrediti mogućnost centriranja Twisted Fileom™ u usporedbi sa strojnim sustavom Revo-S®. Materijali i metode: Četrdeset bukomezijalnih kanala maksilarnog prvog kutnjaka sa zakrivljenošću od 25° do 30° podijeljeni su u dvije skupine po 20, ovisno o korištenom sustavu (skupina 1: Twisted File™; skupina 2: strojni sustav Revo-S®). Svi zubi slikani su CBCT-om te im je određen oblik korijenskih kanala prije instrumentacije i nakon toga postupka. Zatim su slike digitalno obrađene Image Tools Softwareom i izračunat je omjer centriranja kanala. Rezultati su statistički analizirani t- i Mann-Whitneyjevim testom. Stupanj statističke značajnosti postavljen je na ,05. Rezultati: Ni kod jednog sustava centriranje nije bilo besprijekorno te između dviju eksperimentalnih skupina nije bilo statistički značajnih razlika (p 0.05) between the two groups in terms of centering ability. Conclusion: None of the instruments evaluated in this study were totally effective in performing biomechanical preparation of the root canals, because each of them produced canal deviation

Conhecimento e interesse dos estudantes de medicina sobre as práticas integrativas e complementares de saúde

O uso de Práticas Integrativas e Complementares (PICs) no tratamento de doenças pelo Sistema Único de Saúde (SUS) tem aumentado consideravelmente e hoje 29 procedimentos são disponibilizados, sendo que 88% deles são oferecidos na Atenção Básica. Segundo dados do Ministério da Saúde, foram realizados aproximadamente 1,5 milhões de atendimentos em 2017, entre as práticas mais utilizadas destacam-se a homeopatia e acupuntura. Embora existam evidências científicas que demostrem os benefícios do tratamento da medicina alopata integrada às PICs, ainda existe um desconhecimento tanto da população como dos estudantes e profissionais da saúde sobre o assunto, gerando controvérsias com a busca crescente dos pacientes por esses tratamentos. Objetivo: Este estudo teve como objetivo analisar o conhecimento e o interesse dos acadêmicos de medicina à respeito das PICs. Metodologia: Os dados foram coletados em 2018 a partir de questionário elaborado pelos autores e aplicado a 224 estudantes matriculados no curso de Medicina da Universidade de Mogi das Cruzes (UMC) - SP. Resultados: Do total de entrevistados, verificou-se que 75,8% dos estudantes desconhecem que estas práticas já estão incorporadas ao SUS, contudo, 68,3% demonstraram interesse em inserir conteúdos correlacionados às PICs como componentes curriculares durante a graduação. Conclusão: Apesar do nível de conhecimento da amostra apresentar-se abaixo do esperado, os participantes da pesquisa se mostraram interessados em explorar o assunto. Além disso, foi evidenciada a crescente demanda dos pacientes por esse tipo de tratamento, indicando a necessidade da inserção de tais modalidades nos currículos médicos

Quão verde é a energia azul? Uma análise do desacoplamento e da ecoeficiência na geração de energia hidroelétrica no Brasil

O setor de energia no Brasil se baseia fortemente nas fontes de energia renováveis. As Usinas de Energia Hidrelétrica (UHE) são a principal geradora de eletricidade no país. Entendemos que as UHE devem receber maior atenção quanto ao desempenho produtivo versus seus impactos ambientais. Por esta razão, este breve artigo tem por objetivo analisar se as usinas hidrelétricas brasileiras apresentam evidências de Desacoplamento entre as variáveisárea inundada pelos reservatórios (impacto ambiental) vs PIB Setorial (vetor econômico), examinando se seus resultados apontam comportamento de ecoeficiência. Adotou-se a metodologia desenhada pela Organization for Economic Co-operation and Development. Esta metodologia está focada na descrição da relação entre dois primeiros elementos do quadro conceitual Driving Force – Pressure – State – Impact – Response (DPSIR). Para a análise de Decoupling, constatamos que predominou o Decoupling Fraco, ocorrendo em 37 dos 62 anos estudados. As UHE’s foram bem sucedidas no objetivo de ampliar oferta de energia, mas essa sua expansão vem ocorrendo a taxas marginais decrescentes, fortemente explicada pelo baixo Fator de Capacidade. E por fim, em termos de Ecoeficiência, as UHE tem apresentado taxa estável de produção de energia GWh por Km2 de área alagada desde 1972

PRESENÇA DE POLUENTES DE LIXIVIADO NO SOLO E ÁGUAS SUBTERRÂNEAS EM LOCAIS DE DISPOSIÇÃO INADEQUADA DE RESÍDUOS SÓLIDOS NO BRASIL: REVISÃO DA LITERATURA: PRESENCE OF LEACHATE POLLUTANTS IN SOIL AND GROUND WATER AT SITES OF INADEQUATE SOLID WASTE DISPOSAL IN BRAZIL: LITERATURE REVIEW

A disposição de resíduos sólidos no solo de forma inadequada ainda é uma realidade em muitos municípios brasileiros. Segundo dados no SNIS de 2021, 54,9% das áreas de disposição de resíduos são lixões. A falta de remediação dessas áreas pós encerramento pode causar danos à saúde pública ao contaminar as águas subterrâneas através da infiltração de lixiviado pelo solo. Esse trabalho teve como objetivo analisar e organizar dados referentes a trabalhos de campo realizados em áreas de disposição de resíduos sólido. A consulta partiu de um material suplementar publicado em artigo em 2021 e foi complementada com busca nas bases de dados Science Direct, SciELO, Biblioteca Brasileira de Teses e Dissertações e Google Acadêmico, para o período de 01 de janeiro de 2020, a 01 janeiro de 2022. Ao todo foram 111 trabalhos selecionados, que geraram 254 grupos de dados referentes a água superficial, água subterrânea, lixiviado e solos superficiais, em profundidade e sedimentos. Foram selecionados os trabalhos que realizaram análises em mais de um meio (lixiviado, solo e/ou água subterrânea), isso reduziu o número de publicações para 31 trabalhos. Em seguida foi aplicado um novo filtro selecionando apenas os que realizaram mais de quatro campanhas de coletas de amostras, foram discutidos os dados gerados por 11 trabalhos. Os parâmetros que mais se destacaram nas análises de lixiviado, predominantemente alcalino, foram a condutividade, demanda bioquímica de oxigênio (DBO), demanda química de oxigênio (DQO), o nitrogênio amoniacal e o cloreto. As amostras de água subterrânea apresentaram altos valores de condutividade e cloreto. Alguns íons de interesse para a identificação gerada pela contaminação por lixiviado como Na, K, Mg e Ca não possuem valores de referência de qualidade (VRQ) orientados pela Legislação, o que compromete a discussão dos resultados. Os diferentes solos de cada sítio contaminado foram amostrados nas áreas de influência da contaminação por lixiviado e apresentaram concentrações acima do permitido pela legislação para alguns metais como Cd, Cr, Cu e Zn. É possível notar que o lixiviado exerceu influência na natureza do solo e água subterrânea amostrados

Barriers and facilitators of purchasing from short food supply chains: evidence from consumer focus groups in Germany, Spain, Greece and Hungary

This study aimed to identify the barriers and facilitators of consumers purchasing from short food supply chains (SFSC). Eight focus groups were conducted with consumers in the rural and urban areas of Germany, Spain, Hungary and Greece. Participants generally felt that increasing the convenience of purchasing SFSC products (in terms of a proximal location and being able to purchase a wide range of produce in one place) was a prerequisite for them to buy such products. Food quality in terms of taste, freshness and organic status were also taken into account in purchase decisions, and there appears to be a greater focus on health rather than the environmental implications of organic production, although the environmental aspects are also appreciated. This study aimed to identify the barriers and facilitators of consumers purchasing from short food supply chains (SFSC). Eight focus groups were conducted with consumers in the rural and urban areas of Germany, Spain, Hungary and Greece. Participants generally felt that increasing the convenience of purchasing SFSC products (in terms of a proximal location and being able to purchase a wide range of produce in one place) was a prerequisite for them to buy such products. Food quality in terms of taste, freshness and organic status were also taken into account in purchase decisions, and there appears to be a greater focus on health rather than the environmental implications of organic production, although the environmental aspects are also appreciated. Some participants also like the idea of supporting their local community through purchasing from local producers and/or retailers. It was believed that small-scale production and SFSC result in better quality food, but participants had less confidence in the hygiene and food safety standards of SFSC compared to longer chains. Participants thought that consumers would purchase local food if they could more easily access a variety of local food in one place, such as through supermarkets, cooperatives, farm shops and markets, or an online platform that aggregates producers.info:eu-repo/semantics/publishedVersio

Claiming Equality: Puerto Rican Farmworkers in Western New York

n July of 1966, a group of Puerto Rican migrant workers protested against police brutality and discrimination in North Collins, a small farm community of western NewYork. Puerto Rican farmworkers made up a substantial part of the population, and had transformed the ethnic, racial, and gender landscape of the town. Local officials and residents produced and reproduced images of Puerto Ricans as inferior subjects within US racial and ethnic hierarchies. Those negative images of Puerto Ricans shaped the way in which local authorities elaborated policies of social control against these farmworkers in North Collins. At the same time, Puerto Rican farmworkers challenged those existing images and power relations that attempted to stigmatize them as inferior. They affirmed their presence in western New York and, in effect, stood up for their rights as citizens, as Puerto Ricans, and as Latinos