Current status of the whitefly Aleurodicus dispersus as an invasive pest in the CapeVerte Islands

In the last three decades, the spiralling whitefly (Aleurodicus dispersus) has become an important international pest. The movement of plants and parts of plants (such as fruits) in international trade and tourism, and by natural dispersal, has favoured its introduction to new areas. In common with others whiteflies of economic importance, the immature and adult stages cause direct feeding damage by piercing and sucking of sap from foliage, and indirect damage following the accumulation all over host plants of honeydew and waxy flocculent material produced by the insects. Spiralling whitefly is a pest of tropical and subtropical crops, and highly polyphagous. Up to the 1970s, it had been recorded on 44 genera of plants, belonging to 26 botanical families (Mound & Halsey, 1978). This situation changed with the dispersal of the pest to new areas. Nowadays, the spiralling whitefly is one of the major pest of vegetable, ornamental and fruit crops around the globe (Lambkin, 1999). Important host crops include: banana (Musa sapientum), Citrus spp., coconut (Cocos nocifera), eggplant (Solanum melanogena), guava (Psidium guajava), Hibiscus rosa sinensis, Indian almond (Terminalia catappa), papya (Carica papaya), Rosa sp. and tomato (Lycopersicon esculentum) (Saminathan & Jayaraj, 2001). Spiralling whitefly has its origin in the tropical Americas, including Brazil. Although the pest has been recorded only once in Brasil, in the 1920s in the state of Bahia (Bondar, 1923), it now has official quarantine status because of its economic importance. In the Cape Verte Islands, on the West African coast, the pest was initially introduced in the first half of 2000; it has since become established, reaching urban, natural and agricultural areas of the islands that constitute the archipelago. Since then, the pest has been causing damage to many native plants, ornamentals and cultivated food crops (Anon., 2001; Monteiro, 2004). The present study was done in order to produce an inventory of the most common host plants of spiralling whitefly in this new habitat.

O Processo de Intrusão Salina a jusante da Bacia Hidrográfica da Ribeira Seca. A Inversão do Processo e Impactos da Barragem

Os recursos hídricos na Bacia hidrográfica da Ribeira Seca não têm sido explorados numa base auto-sustentável. Como consequência, verifica-se a deterioração da qualidade da água dos poços e dos furos como resultado da intrusão salina que chega a avançar à volta dos quatro quilómetros para o interior da bacia hidrográfica. Na tentativa de mitigar os efeitos de uma exploração desequilibrada dos recursos em água, muitos projectos de conservação do solo e água foram levados a cabo, destacando-se o Projecto de Desenvolvimento de Bacias Hidrográficas, “Watershed Development Project” – financiado no quadro da USAID na década de 80, (mais concretamente em 1984) e que contemplava todas as bacias hidrográficas da ilha de Santiago. As principais actividades levadas a cabo no âmbito do “Watershed Development Project” foram essencialmente as seguintes: 1. Infra-estruturas hidráulicas de captação, armazenamento, adução e distribuição de água para a rega e consumo doméstico; 2. Infra-estruturas de correcção do leito da ribeira, de espalhamento e recarga das águas de escoamento superficial; 3. Instalação de bombas eólicas para bombagem da água dos poços; 4. Adaptação ao regadio de terrenos de cultura e construção de terraços; 5. Arborização das encostas em dispositivos anti-erosivos (banquetas, muretes, caldeiras, etc.) 6. Sistemas de “water harvesting/runoff farming” (incluindo construção de grandes reservatórios, diques de captação de águas de escoamento superficial); 7. Plantação de feijão Congo (Cajanus cajan) em muretes e banquetas nas encostas. No quadro do projecto de Desenvolvimento de Bacias Hidrográficas, “Watershed Development Project”, dois projectos de luta contra a intrusão salina foram elaborados. Um dos projectos contemplou a Ribeira de Saltos e outro na Ribeira Seca. Só o da Ribeira de Saltos foi executado sob a orientação do coordenador técnico do projecto e os resultados, embora evidentes, estão sendo quantificados no âmbito de um trabalho por nós conduzido. O projecto de luta contra a intrusão salina na Ribeira Seca não foi implementado e bem assim, os projectos considerados no PDH-Ribeira Seca financiados no quadro da Cooperação Austríaca.Pretende-se com este trabalho de fim de curso não só analisar os efeitos da degradação ecológica dos terrenos de cultura localizados a jusante e submetidos à intrusão salina, como também, quantificar os impactos das infra-estruturas hidráulicas executadas a jusante da Ribeira Seca, mais concretamente, o efeito da construção da Barragem de Poilão na melhoria das condições hidrológicas do troço da bacia hidrográfica a jusante.

Females of Tapinotaspoides, a genus in the oil-collecting bee tribe Tapinotaspidini, collect secretions from non-floral trichomes (Hymenoptera, Apidae)

Harvesting of secretions from non-floral trichomes by females of Tapinotaspoides serraticornis is reported for the first time. The females exhibit a type of mopping behavior using the fringes of long, wavy setae along the posterior margins of their metasomal sterna. Our observations indicated a wide range of host plants used as sources for these secretions, including Waltheria (Sterculiaceae), Tibouchina (Melastomataceae), Sida (Malvaceae), Jacquemontia (Convolvulaceae), and unidentified species of Commelinaceae and Cyperaceae.

Ejaculation failure on the day of oocyte retrieval for IVF: case report.

Unexpected ejaculation failure on the day of oocyte retrieval for IVF occurs once or twice a year in our Reproductive Medicine Unit, where approximately 500 oocyte retrievals are performed each year. Two clinical situations which occurred in 2001 are presented. In the first case, sperm were finally obtained by epididymal aspiration and resulted in the fertilization of five oocytes by ICSI. The transfer of two fresh embryos did not result in a pregnancy and the three supernumerary zygotes were cryopreserved. The male patient presented an anxio-depressive episode necessitating psychiatric hospitalization 1 week after the oocyte retrieval. In the second case, no sperm were obtained and the four oocytes were therefore lost. The couple went through a crisis in their relationship and tried another cycle of IVF 10 months later, after the preventive cryopreservation of a sperm sample. On the day of oocyte retrieval the patient was unable to produce a fresh sample but three zygotes were obtained through ICSI using the back-up cryopreserved sperm. Two embryos were transferred but no pregnancy ensued. The clinical decision-making processes for these two cases are described, as well as the measures employed to help prevent these unfortunate situations.

Implementação de um Sistema MPPT em Circuito Integrado CMOS

Ao longo dos últimos anos a indústria microelectrónica tem evoluído no sentido de reduzir o consumo energético dos seus dispositivos no sentido de estes serem alimentados por fontes energéticas diversas, nomeadamente fontes renováveis. A crescente demanda por componentes energeticamente eficientes e pela miniaturização dos componentes eletrónicos exigem a conceção de fontes de alimentação com potência reduzida na ordem das dezenas de μW ás centenas de mW. Atualmente, com os crescentes avanços tecnológicos é possível obter componentes energéticamente eficientes e com tamanhos reduzidos capazes de colmatar as restrições energéticas das mais variadas aplicações. A utilização de fontes energia elétrica que tirão proveito da energia existente no meio onde se inserem os diversos dispositivos eletrónicos ou simplesmente para recarregar as baterias, apresenta-se como um dos principais objetivos a alcançar. Entre todas as energias renováveis, a energia fotovoltaica surge como umas das que proporciona um maior potencial. A sua disponibilidade global e os constantes desenvolvimentos tecnológicos no âmbito do fotovoltaico permitem o desenvolvimento de sistemas de alimentação com rendimento energético cada vez mais elevado. A presente dissertação tem como objetivo o estudo, a simulação e implementação de um conversor CC-CC, step-up, com algoritmo de controlo MPPT integrado designado por método de correlação de ripple (Ripple Correlation Control – RCC), para aplicações de baixa potência na ordem das dezenas de μW às centenas de mW. Desenvolveu-se um conversor CC-CC, autónomo, para carregamento de baterias com recurso a painéis fotovoltaicos, como fonte de energia, de forma contínua mesmo em situações de baixa luminosidade. Um circuito de gestão de energia devidamente dimensionado foi implementado com recurso a um algoritmo de procura do ponto de máxima potência (Maximum Power Point Tracking – MPPT). O objetivo desta implementação é extrair a máxima potência disponível da fonte de energia elétrica, neste caso o painel fotovoltaico, independentemente das condições meteorológicas e da potência requerida pela carga, sendo o excesso de energia redirecionado para a bateria. Nesta dissertação apresentam-se os resultados das simulações, assim como os resultados experimentais de todos os circuitos desenvolvidos de forma a validar todo o sistema implementado.

Extracção de areia na Praia de Calhetona (Ilha de Santiago, Cabo Verde) : causas, processos e consequências

A extracção de areia nas praias do mar, nas bacias hidrográficas e nas encostas montanhosas, tem sido prática de muitas famílias cabo-verdianas. A Praia de Calhetona foi uma das que nos últimos anos sofreu degradação significativa, em função da apanha clandestina de areia, sem qualquer plano de extracção, destinada a construção civil. A extracção de areia, para vender ou para a autoconstrução, é efectuada em família por indivíduos, maioritariamente do sexo feminino, de baixa renda ou sem profissão remunerada. O processo de extracção de areia na Praia de Calhetona teve o seu ponto alto nos anos 80, diminuindo consideravelmente com a escassez de areia e implementação do Decreto – Lei nº 69/97, de 3 de Novembro, que visava disciplinar a exploração de inertes nas praias. Em face da pesquisa feita, baseada no estudo de caso, ficou expresso que os inquiridos implicados na actividade de extracção clandestina de areia correm riscos elevados para obterem lucros reduzidos. Quem efectivamente beneficia são os camionistas que compram esse inerte e o vendem pelo dobro do preço. Qualquer actividade de exploração de recursos naturais causa impactes sobre o espaço de actuação. No caso da Praia de Calhetona, observa-se o recuo da linha de costa, a quase ausência de areia e a desertificação de propriedades nas suas proximidades, para além dos impactes negativos sobre o turismo, a função balnear e a desova de tartaruga. Apesar de tudo, constata-se uma consciência generalizada dos impactes ambientais causados pelos inquiridos, os quais, no entanto, alegam que a extracção de areia é uma das alternativas para garantir a sobrevivência individual e das suas famílias.

Extracção de areia na Praia de Calhetona (Ilha de Santiago, Cabo Verde) : causas, processos e consequências

A extracção de areia nas praias do mar, nas bacias hidrográficas e nas encostas montanhosas, tem sido prática de muitas famílias cabo-verdianas. A Praia de Calhetona foi uma das que nos últimos anos sofreu degradação significativa, em função da apanha clandestina de areia, sem qualquer plano de extracção, destinada a construção civil. A extracção de areia, para vender ou para a autoconstrução, é efectuada em família por indivíduos, maioritariamente do sexo feminino, de baixa renda ou sem profissão remunerada. O processo de extracção de areia na Praia de Calhetona teve o seu ponto alto nos anos 80, diminuindo consideravelmente com a escassez de areia e implementação do Decreto – Lei nº 69/97, de 3 de Novembro, que visava disciplinar a exploração de inertes nas praias. Em face da pesquisa feita, baseada no estudo de caso, ficou expresso que os inquiridos implicados na actividade de extracção clandestina de areia correm riscos elevados para obterem lucros reduzidos. Quem efectivamente beneficia são os camionistas que compram esse inerte e o vendem pelo dobro do preço. Qualquer actividade de exploração de recursos naturais causa impactes sobre o espaço de actuação. No caso da Praia de Calhetona, observa-se o recuo da linha de costa, a quase ausência de areia e a desertificação de propriedades nas suas proximidades, para além dos impactes negativos sobre o turismo, a função balnear e a desova de tartaruga. Apesar de tudo, constata-se uma consciência generalizada dos impactes ambientais causados pelos inquiridos, os quais, no entanto, alegam que a extracção de areia é uma das alternativas para garantir a sobrevivência individual e das suas famílias.

Extracção de areia na Praia de Calhetona (Ilha de Santiago, Cabo Verde) : causas, processos e consequências

A extracção de areia nas praias do mar, nas bacias hidrográficas e nas encostas montanhosas, tem sido prática de muitas famílias cabo-verdianas. A Praia de Calhetona foi uma das que nos últimos anos sofreu degradação significativa, em função da apanha clandestina de areia, sem qualquer plano de extracção, destinada a construção civil. A extracção de areia, para vender ou para a autoconstrução, é efectuada em família por indivíduos, maioritariamente do sexo feminino, de baixa renda ou sem profissão remunerada. O processo de extracção de areia na Praia de Calhetona teve o seu ponto alto nos anos 80, diminuindo consideravelmente com a escassez de areia e implementação do Decreto – Lei nº 69/97, de 3 de Novembro, que visava disciplinar a exploração de inertes nas praias. Em face da pesquisa feita, baseada no estudo de caso, ficou expresso que os inquiridos implicados na actividade de extracção clandestina de areia correm riscos elevados para obterem lucros reduzidos. Quem efectivamente beneficia são os camionistas que compram esse inerte e o vendem pelo dobro do preço. Qualquer actividade de exploração de recursos naturais causa impactes sobre o espaço de actuação. No caso da Praia de Calhetona, observa-se o recuo da linha de costa, a quase ausência de areia e a desertificação de propriedades nas suas proximidades, para além dos impactes negativos sobre o turismo, a função balnear e a desova de tartaruga. Apesar de tudo, constata-se uma consciência generalizada dos impactes ambientais causados pelos inquiridos, os quais, no entanto, alegam que a extracção de areia é uma das alternativas para garantir a sobrevivência individual e das suas famílias.

Extracção de Areia na Praia de Calhetona (Ilha de Santiago, Cabo Verde) - Causas, Processos e Consequências

A extracção de areia nas praias do mar, nas bacias hidrográficas e nas encostas montanhosas, tem sido prática de muitas famílias cabo-verdianas. A Praia de Calhetona foi uma das que nos últimos anos sofreu degradação significativa, em função da apanha clandestina de areia, sem qualquer plano de extracção, destinada a construção civil. A extracção de areia, para vender ou para a autoconstrução, é efectuada em família por indivíduos, maioritariamente do sexo feminino, de baixa renda ou sem profissão remunerada. O processo de extracção de areia na Praia de Calhetona teve o seu ponto alto nos anos 80, diminuindo consideravelmente com a escassez de areia e implementação do Decreto – Lei nº 69/97, de 3 de Novembro, que visava disciplinar a exploração de inertes nas praias. Em face da pesquisa feita, baseada no estudo de caso, ficou expresso que os inquiridos implicados na actividade de extracção clandestina de areia correm riscos elevados para obterem lucros reduzidos. Quem efectivamente beneficia são os camionistas que compram esse inerte e o vendem pelo dobro do preço. Qualquer actividade de exploração de recursos naturais causa impactes sobre o espaço de actuação. No caso da Praia de Calhetona, observa-se o recuo da linha de costa, a quase ausência de areia e a desertificação de propriedades nas suas proximidades, para além dos impactes negativos sobre o turismo, a função balnear e a desova de tartaruga. Apesar de tudo, constata-se uma consciência generalizada dos impactes ambientais causados pelos inquiridos, os quais, no entanto, alegam que a extracção de areia é uma das alternativas para garantir a sobrevivência individual e das suas famílias.

Spatial distribution of a spherical gall (Hymenoptera, Eulophidae) on Caryocar brasiliense (Caryocaraceae)

We obtained the first data on spatial distribution of a spherical galling insect (Hymenoptera, Eulophidae) at the Caryocar brasiliense Camb. (Caryocaraceae) tree level. This work was developed in two pastures in Montes Claros, Minas Gerais State, Brazil. The areas studied were: pasture 1 (in activity) and pasture 2 (abandoned pasture = savanna in recovery). We evaluated the distribution of spherical galls in: foliage orientation (slope), among leaves (border and interior of the tree crown), among leaflets (right, central, left), distal, median, and proximal as well as border, central area, and adjacent to the mid leaf vein of the leaflet, and difference between areas in 10 infested trees per area. The smaller number of spherical gall/leaflet was observed in pasture 1 than in pasture 2. More spherical galls were found on the northern in pasture 1, but in the pasture 2, the lower spherical galls were observed on the northeast than other slopes. The average number of spherical galls did not differ statistically among the three leaflets of C. brasiliense in pasture 2. However, in pasture 1, we observed highest number of spherical galls in the central leaflet. More spherical galls were found in the border than interior of the tree crown. The average number of spherical galls did not differ statistically among the longitudinal region on leaflet of C. brasiliense. The spherical gall insect preferred to colonize the leaf margin than the central portion or near mid vein on transversal regions on a leaflet.

Translation to success of surgical innovation.

Contemporary thoracic and cardiovascular surgery uses extensive equipment and devices to enable its performance. As the specialties develop and new frontiers are crossed, the technology needs to advance in a parallel fashion. Strokes of genius or problem-solving brain-storming may generate great ideas, but the metamorphosis of an idea into a physical functioning tool requires a lot more than just a thinking process. A modern surgical device is the end-point of a sophisticated, complicated and potentially treacherous route, which incorporates new skills and knowledge acquisition. Processes including technology transfer, commercialisation, corporate and product development, intellectual property and regulatory routes all play pivotal roles in this voyage. Many good ideas may fall by the wayside for a multitude of reasons as they may not be marketable or may be badly marketed. In this article, we attempt to illuminate the components required in the process of surgical innovation, which we believe must remain in the remit of the modern-day thoracic and cardiovascular surgeon.

Evaluation of an online, case-based interactive approach to teaching pathophysiology

Rapport de synthèse : L'article qui fait l'objet de ma thèse évalue une nouvelle approche pédagogique pour l'apprentissage de certains chapitres de physiopathologie. Le dispositif pédagogique se base sur l'alternance d'apprentissage ex-cathedra et de l'utilisation d'un site web comprenant des vignettes cliniques. Lors de la consultation de ces-dernières, l'étudiant est invité à demander des examens de laboratoire dont il pourrait justifier la pertinence selon le cas clinique étudié. La nouveauté du procédé réside dans le fait que, préalablement à son cours ex-cathedra, l'enseignant peut consulter les statistiques de demandes de laboratoire et ainsi orienter son cours selon les éléments mal compris par les étudiants. A la suite du cours ex-cathedra, les étudiants peuvent consulter sur internet la vignette clinique complète avec des explications. A l'issue de tout le cours, une évaluation auprès des étudiants a été conduite. Le procédé a été mis en place durant deux années consécutives et l'article en discute notamment les résultats. Nous avons pu conclure que cette méthode innovatrice d'enseignement amène les étudiants à mieux se préparer pour les cours ex-cathedra tout en permettant à l'enseignant d'identifier plus précisément quelles thématiques étaient difficiles pour les étudiants et donc d'ajuster au mieux son cours. Mon travail de thèse a consisté à créer ce dispositif d'apprentissage, à créer l'application web des vignettes cliniques et à l'implanter durant deux années consécutives. J'ai ensuite analysé les données des évaluations et écrit l'article que j'ai présenté à la revue 'Medical Teacher'. Après quelques corrections et précisions demandées par le comité de lecture, l'article a été accepté et publié. Ce travail a débouché sur une seconde version de l'application web qui est actuellement utilisée lors du module 3.1 de 3è année à l'Ecole de Médecine à Lausanne. Summary : Since the early days of sexual selection, our understanding of the selective forces acting on males and females during reproduction has increased remarkably. However, despite a long tradition of experimental and theoretical work in this field and relentless effort, numerous questions remain unanswered and many results are conflicting. Moreover, the interface between sexual selection and conservation biology has to date received little attention, despite existing evidence for its importance. In the present thesis, I first used an empirical approach to test various sexual selection hypotheses in a population of whitefish of central Switzerland. This precise population is characterized by a high prevalence of gonadal alterations in males. In particular, I challenged the hypothesis that whitefish males displaying peculiar gonadal features are of lower genetic quality than other seemingly normal males. Additionally, I also worked on identifying important determinant of sperm behavior. During a second theoretical part of my work, which is part of a larger project on the evolution of female mate preferences in harvested fish populations, I developed an individual-based simulation model to estimate how different mate discrimination costs affect the demographical behavior of fish populations and the evolutionary trajectories of female mate preferences. This latter work provided me with some insight on a recently published article addressing the importance of sexual selection for harvesting-induced evolution. I built upon this insight in a short perspective paper. In parallel, I let some methodological questions drive my thoughts, and wrote an essay about possible synergies between the biological, the philosophical and the statistical approach to biological questions.

Factors promoting the exposure to bioaerosols among crop workers

Introduction. Agricultural workers are among the professional groups most at risk of developing acute or chronic respiratory problems. Despite this fact, the etiology of these occupational diseases is poorly known, even in important sectors of agriculture such as the crops sector. Cereals can be colonized by a large number of fungal species throughout the plants' growth, but also during grain storage. Some of these fungi deliver toxins that can have a serious impact on human health when they are ingested via wheat products. Although International and European legislation on contaminants in food, including mycotoxins, include measures to ensure protection of public health by setting down the maximum levels for certain contaminants, the risks associated with the inhalation of such molecules during grain handling remains poorly documented. Goal of study. This project's objective was to characterize worker exposure to pathogenic, irritative or allergenic microorganisms and to identify the abiotic or biotic factors that reduce the growth of these microorganisms in crops. Indeed, the proliferation of microorganisms on wheat is dependent on temperature, rainfall and human disturbance (e.g. usage of tillage, addition of fungicides). A change in the concentration of these microorganisms in the substrate will directly result in a change in the concentration of aerosolized particles of the same microorganisms. Therefore, the exposure of worker to bioaérosols will also change. The Vaud region of Switzerland is a perfect region for conduct such a project as weather conditions vary and agricultural land management programs are divers at a small geographic scale. Methods. Bioaerosols and wheat dust have been sampled during wheat harvesting of summer 2010 at 100 sites uniformly distributed in the Vaud region that are representative of the different agriculture practices. Personal exposure has been evaluated for different wheat related activities: harvesting, grain unload, baling straw, the cleaning of harvesters and silos. Aerosols have been sampled at a rate of 2L/min between 15 min to 4 hours (t) on a 5m PVC filter for estimating the total dust inhaled, on gelatine filter for the identification and quantification of molds, and on a 0.45um polycarbonate filter for endotoxin quantification. Altitude, temperature and annual average rainfall were considered for each site. The physical and chemical characteristics of soils were determined using the methods in effect at Sol Council (Nyon). Total dust has been quantified following NIOSH 0500 method. Reactive endotoxine activity has been determined with Limulus Amebocyte Lysate Assay. All molds have been identified by the pyrosequencing of ITS2 amplicons generated from bioaerosol or wheat dust genomic DNA. Results & Conclusions. Our results confirm the previous quantitative data on the worker exposure to wheat dust. In addition, they show that crop workers are systematically exposed to complex mixtures of allergens, irritants or cytotoxic components. The novelty of our study is the systematic detection of molds such as Fusarium - that is a mycotoxins producer - in the bioaerosols. The results are interpreted by taking in account the agriculture practice, the Phosphorus : Carbon : Nitrogen ratio of the soil, the altitude and the average of rainy days per year.

Soil compaction and eucalyptus growth in response to forwarder traffic intensity and load

During timber exploitation in forest stands harvesting machines pass repeatedly along the same track and can cause soil compaction, which leads to soil erosion and restricted tree root growth. The level of soil compaction depends on the number of passes and weight of the wood load. This paper aimed to evaluate soil compaction and eucalyptus growth as affected by the number of passes and wood load of a forwarder. The study was carried out in Santa Maria de Itabira county, Minas Gerais State - Brazil, on a seven-year-old eucalyptus stand planted on an Oxisol. The trees were felled by chainsaw and manually removed. Plots of 144 m² (four rows 12 m long in a 3 x 2 m spacing) were then marked off for the conduction of two trials. The first tested the traffic intensity of a forwarder which weighed 11,900 kg and carried 12 m³ wood (density of 480 kg m-3) and passed 2, 4, and 8 times along the same track. In the second trial, the forwarder carried loads of 4, 8, and 12 m³ of wood, and the machine was driven four times along the same track. In each plot, the passes affected four rows. Eucalyptus was planted in 30 x 30 x 30 cm holes on the compacted tracks. The soil in the area is clayey (470 clay and 440 g kg-1 sand content) and at depths of 0-5 cm and 5-10 cm, respectively, soil organic carbon was 406 and 272 g kg-1 and the moisture content during the trial 248 and 249 g kg-1. These layers were assessed for soil bulk density and water-stable aggregates. The infiltration rate was measured by a cylinder infiltrometer. After 441 days the measurements were repeated, with additional analyses of: soil organic carbon, total nitrogen, N-NH4+, N-NO3-, porosity, and penetration resistance. Tree height, stem diameter, and stem dry matter were measured. Forwarder traffic increased soil compaction, resistance to penetration and microporosity while it reduced the geometric mean diameter, total porosity, macroporosity and infiltration rate. Stem dry matter yield and tree height were not affected by soil compaction. Two passes of the forwarder were enough to cause the disturbances at the highest levels. The compaction effects were still persistent 441 days after forwarder traffic.

Coculture of bladder urothelial and smooth muscle cells on small intestinal submucosa: potential applications for tissue engineering technology.

PURPOSE: Small intestinal submucosa is a xenogenic, acellular, collagen rich membrane with inherent growth factors that has previously been shown to promote in vivo bladder regeneration. We evaluate in vitro use of small intestinal submucosa to support the individual and combined growth of bladder urothelial cells and smooth muscle cells for potential use in tissue engineering techniques, and in vitro study of the cellular mechanisms involved in bladder regeneration. MATERIALS AND METHODS: Primary cultures of human bladder urothelial cells and smooth muscle cells were established using standard enzymatic digestion or explant techniques. Cultured cells were then seeded on small intestinal submucosa at a density of 1 x 105 cells per cm.2, incubated and harvested at 3, 7, 14 and 28 days. The 5 separate culture methods evaluated were urothelial cells seeded alone on the mucosal surface of small intestinal submucosa, smooth muscle cells seeded alone on the mucosal surface, layered coculture of smooth muscle cells seeded on the mucosal surface followed by urothelial cells 1 hour later, sandwich coculture of smooth muscle cells seeded on the serosal surface followed by seeding of urothelial cells on the mucosal surface 24 hours later, and mixed coculture of urothelial cells and smooth muscle cells mixed and seeded together on the mucosal surface. Following harvesting at the designated time points small intestinal submucosa cell constructs were formalin fixed and processed for routine histology including Masson trichrome staining. Specific cell growth characteristics were studied with particular attention to cell morphology, cell proliferation and layering, cell sorting, presence of a pseudostratified urothelium and matrix penetrance. To aid in the identification of smooth muscle cells and urothelial cells in the coculture groups, immunohistochemical analysis was performed with antibodies to alpha-smooth muscle actin and cytokeratins AE1/AE3. RESULTS: Progressive 3-dimensional growth of urothelial cells and smooth muscle cells occurred in vitro on small intestinal submucosa. When seeded alone urothelial cells and smooth muscle cells grew in several layers with minimal to no matrix penetration. In contrast, layered, mixed and sandwich coculture methods demonstrated significant enhancement of smooth muscle cell penetration of the membrane. The layered and sandwich coculture techniques resulted in organized cell sorting, formation of a well-defined pseudostratified urothelium and multilayered smooth muscle cells with enhanced matrix penetration. With the mixed coculture technique there was no evidence of cell sorting although matrix penetrance by the smooth muscle cells was evident. Immunohistochemical studies demonstrated that urothelial cells and smooth muscle cells maintain the expression of the phenotypic markers of differentiation alpha-smooth muscle actin and cytokeratins AE1/AE3. CONCLUSIONS: Small intestinal submucosa supports the 3-dimensional growth of human bladder cells in vitro. Successful combined growth of bladder cells on small intestinal submucosa with different seeding techniques has important future clinical implications with respect to tissue engineering technology. The results of our study demonstrate that there are important smooth muscle cell-epithelial cell interactions involved in determining the type of in vitro cell growth that occurs on small intestinal submucosa. Small intestinal submucosa is a valuable tool for in vitro study of the cell-cell and cell-matrix interactions that are involved in regeneration and various disease processes of the bladder.