Skin is an organ with a complex structure which plays a crucial role in thebody’s defence against external threats and in maintaining major homeostatic functions. The need for in vitro models that mimic the in vivo milieu is therefore high and relevant with various applications including, among others, penetration, absorption, and toxicity studies. In this context, the choice of the biomaterial that will provide a 3D scaffold to the cultured cells is defining the model’s success. The FN-4RepCT silk is here suggested as a potent biomaterial for skin tissue engineering applications. This recombinantly produced spider silk protein (FN-4RepCT), which can self-assemble into fibrils, creates a robust and elastic matrice with high bioactivity, due to its functionalization with the fibronectin derived RGD-containing peptide. Hence it overcomes the drawbacks of other available biomaterials either synthetic or based on animal derived proteins. Additionally, the FN-4RepCT silk protein can be cast in various 3D formats, two of which are utilized within this project. We herein present a bilayered skin tissue equivalent supported by the FN-4RepCT silk. This is constructed by the combination of a foam format, integrated with dermal fibroblasts and endothelial cells, and a membrane format supporting epidermal keratinocytes. As a result, a vascularized dermal layer that contains ECM components (Collagen I, Collagen III, and Elastin) is constructed and attached to an epidermal layer of differentiated keratinocytes.The protocol presented in this project offers a successful method of evenly integrating cells in the FN-4RepCT silk scaffold, while preserving their ability to resume some of their major in vivo functions like proliferation, ECM secretion, construction of vascular networks, and differentiation. The obtained results were evaluated with immunofluorescence stainings of various markers of interest and further analysed, when necessary, with image processing tools. The results that ensued from the herein presented protocol strongly suggest that the FN-4RepCT silk is a promising biomaterial for skin tissue engineering applications.

The composition of gut microbes affects host weight, immune function, and disease status, and is sensitive to diet, environment, and pharmaceutical exposure. The gut microbiome modulates the toxicity and bioavailability of chemical stressors, however the effects of chemicals on the gut microbiome of aquatic biota are largely unknown. The Waterloo and Kitchener wastewater treatment plants (WWTPs) release effluents containing antibiotics, pharmaceuticals, and other contaminants into the Grand River (ON) that may negatively affect the gut microbiome of downstream organisms. In this study done in Fall 2018, I collected freshwater mussels (Lasmigona costata), several species of insect larvae, and riparian spiders (Tetragnathidae) from sites upstream and downstream of these WWTPs. The gut microbiome was analyzed following the extraction, PCR amplification, and sequencing of bacterial DNA using the V3-V4 hypervariable regions of the 16S rRNA genetic barcode. Changes in the relative abundance of major gut microbiome phyla were observed in all targeted aquatic organisms downstream of WWTPs except Hydropsychidae. Shannon alpha diversity, a measure of bacterial abundance and evenness, differed significantly among sites for mussels (one-way ANOVA: F=7.894, p=0.001), spiders (F=4.788, p=0.01), Perlidae (F=3.1, p=0.0056), Hydropsychidae (F=3.674, p=0.0014), and Heptageniidae (F=2.715, p=0.0143), but not for Baetidae and Ephemerellidae. In sites downstream of the Waterloo WWTP, alpha diversity decreased in spiders, while in sites downstream of the Kitchener WWTP diversity decreased in mussels and Perlidae, while increasing for spiders. Bray-Curtis beta diversity, a measure of dissimilarity between bacterial communities, was significantly dissimilar among sites in all invertebrate taxa (Permanova: p<0.02). Upstream sites differed from downstream Waterloo sites in spiders, Perlidae, and Hydropsychidae (Adonis pairwise: p<0.05), while upstream mussels, spiders, Perlidae, and Hydropsychidae differed from downstream Kitchener sites (p<0.05). Additionally, effluent-derived bacteria were found in the microbiomes of aquatic invertebrates downstream of the WWTPs and not upstream. Taxa was also a significant driver of bacterial composition and diversity in invertebrates. These results indicate that the gut microbiome of downstream organisms differed from the bacterial composition observed in the same invertebrate taxa upstream of the WWTPs, potentially leading to altered host health. This adds to our understanding of how chemical stressors impact the gut microbiome of aquatic and riparian biota; however, future studies are needed to investigate linkages between the gut microbiome and health of these species. / Thesis / Master of Science (MSc)

[ES] El ácaro de Texas, Eutetranychus banksi McGregor, es nativo de América y está ampliamente distribuido por el cultivo de cítricos de este continente. En 2013, esta especie se detectó en el sur de la provincia de Valencia, afectando a la principal zona citrícola de España. El ácaro produce graves daños reduciendo la fotosíntesis, causando defoliación y decoloración de los frutos, lo que podría afectar a su valor económico. En primer lugar, este trabajo evaluó el impacto ecológico producido por la especie invasora sobre las especies de ácaros tetraníquidos residentes en los cítricos valencianos Panonychus citri (McGregor) y Eutetranychus orientalis (Klein). Desde su llegada en 2013, E. banksi se ha convertido en el tetraníquido más frecuente y abundante en cítricos desplazando competitivamente a las otras especies, reduciendo su presencia y distribución geográfica, a su vez influenciada por su historia de colonización. En segundo lugar, este trabajo estudió la distribución dentro del árbol y las tendencias estacionales de la plaga y de los ácaros fitoseidos asociados, que pueden contribuir a su control. El ácaro de Texas se alimentó principalmente en la cara adaxial (haz) de las hojas en la periferia de la copa del árbol, mostrando un único pico de población a finales de verano-principios de otoño, mientras que los fitoseidos prefirieron la cara abaxial (envés) de las hojas del interior de la copa, mostrando dos picos, uno principal en primavera y otro menos abundante en otoño. Euseius stipulatus (Athias-Henriot) fue el fitoseido más frecuente y abundante, y cuando E. banksi aumentó se desplazó a las caras adaxiales de las hojas de la parte exterior de la copa y los frutos para alimentarse de su presa, cambiando su coloración de blanco a marrón rojizo evidenciando su contribución al control biológico de la plaga. Sin embargo, no fue capaz de mantener las poblaciones de E. banksi bajo densidades tolerables debido a la desfavorable relación depredador/presa que existe en verano y principios de otoño. En tercer lugar, este trabajo pretendió describir la estructura de población del ácaro de Texas y definir un plan de muestreo. Existieron diferencias en la estructura de edad en frutos y hojas, así como entre hojas de diferentes brotes. Además, a lo largo del tiempo se observaron fluctuaciones en su composición correlacionadas con variaciones en la proporción de sexos. No se observaron diferencias de agregación entre estratos vegetales, pero sí entre estadios inmaduros y adultos, siendo las hembras las menos agregadas. La alta correlación de la población total con las formas móviles y las hembras permitió utilizar ambas como estadio de referencia para el desarrollo del plan de muestreo, estableciendo un muestreo de presencia/ausencia de 100 hojas para las hembras o 400 hojas para las formas móviles. Finalmente, un ácaro fitoseido recientemente descrito, Neoseiulus madeirensis Papadoulis & Kapaxidi, se encontró asociado a E. banksi, sugiriendo que podría ser un candidato prometedor para su control biológico. En cuarto lugar, este trabajo pretendió evaluar el potencial de este depredador para controlar las poblaciones de la plaga. Neoseiulus madeirensis mostró un rápido desarrollo y altas tasas de supervivencia y reproducción alimentándose de E. banksi. El depredador se alimentó principalmente de estadios inmaduros, siendo los huevos el estadio preferido, y mostrando una respuesta funcional de tipo II para todos los estadios de presa ensayados, que se estabilizó a altas densidades de presa con una elevada puesta de huevos. Los valores de supervivencia, reproducción y depredación han sido los mejores obtenidos hasta el momento para cualquier fitoseido ensayado previamente contra E. banksi, lo convierte en un candidato idóneo para el desarrollo de un programa de control biológico basado en sueltas aumentativas, o en la importación con vistas al establecimiento de poblaciones permanentes en cítricos. / [CA] L'àcar de Texas, Eutetranychus banksi McGregor, és nadiu d' Amèrica i està àmpliament distribuït pel cultiu de cítrics d'aquest continent. El 2013, esta espècie es va detectar al sud de la província de València, afectant la principal zona citrícola d'Espanya. L'àcar produeix greus danys reduint la fotosíntesi, causant defoliació i decoloració dels fruits, cosa que podria afectar el seu valor econòmic. En primer lloc, aquest treball va avaluar l'impacte ecològic produït per l'espècie invasora sobre les espècies d'àcars tetraníquids residents als cítrics valencians Panonychus citri (McGregor) i Eutetranychus orientalis (Klein). Des de la seva arribada el 2013, E. banksi s'ha convertit en el tetraníquid més freqüent i abundant en cítrics desplaçant competitivament les altres espècies, reduint la seva presència i distribució geogràfica, alhora influenciat per la seva història de colonització. En segon lloc, aquest treball va estudiar la distribució dins de l'arbre i les tendències estacionals de la plaga i dels àcars fitoseids associats, que poden contribuir al seu control. L'àcar de Texas es va alimentar principalment en la cara adaxial (fes) de les fulles a la perifèria de la copa de l'arbre, mostrant un únic pic de població a finals d'estiu-principis de tardor, mentre que els fitoseids van preferir la cara abaxial (revers) de les fulles de l'interior de la copa, mostrant dos pics, un de principal a la primavera i un altre menys abundant a la tardor. Euseius stipulatus (Athias-Henriot) va ser el fitoseid més freqüent i abundant, i quan E. banksi va augmentar es va desplaçar a les cares adaxials de les fulles de la part exterior de la copa i els fruits per alimentar-se de la presa, canviant la seva coloració de blanc a marró vermellós evidenciant la seva contribució al control biològic de la plaga. Tot i això, no va ser capaç de mantenir les poblacions d'E. banksi sota densitats tolerables a causa de la desfavorable relació depredador/presa que hi ha a l'estiu i principis de tardor. En tercer lloc, aquest treball va voler descriure l'estructura de població de l'àcar de Texas i definir un pla de mostreig. Hi hagué diferències en l'estructura d'edat de fruits i fulles, així com entre fulles de diferents brots. A més, al llarg del temps es van observar fluctuacions en la composició correlacionades amb variacions en la proporció de sexes. No es van observar diferències d'agregació entre estrats vegetals, però sí entre estadis immadurs i adults, sent les femelles les menys agregades. L'alta correlació de la població total amb les formes mòbils i les femelles va permetre utilitzar totes dues com a estadi de referència per al desenvolupament del pla de mostreig, establint un mostreig de presència/absència de 100 fulls per a les femelles o 400 fulls per a les formes mòbils. Finalment, una espècie fitoseid recentment descrit, Neoseiulus madeirensis Papadoulis & Kapaxidi, es va trobar associat a E. banksi, suggerint que podria ser un candidat prometedor per al seu control biològic. En quart lloc, aquest treball va voler avaluar el potencial d'aquest depredador per controlar les poblacions de la plaga. Neoseiulus madeirensis va mostrar un desenvolupament ràpid i altes taxes de supervivència i reproducció alimentant-se d'E. banksi. El depredador es va alimentar principalment d'estadis immadurs, sent els ous l'estadi preferit, i mostrant una resposta funcional de tipus II per a tots els estadis de presa assajats, que es va estabilitzar a altes densitats de presa amb una posta d'ous elevada. Els valors de supervivencia, depredació i reproducció han estat els millors obtinguts fins ara per a qualsevol espècie de fitoseid assajat prèviament contra E. banksi, cosa que el converteix en un candidat idoni per al desenvolupament d'un programa de control biològic basat en soltes augmentatives, o en la importació amb vista a l'establiment de poblacions permanents en cítrics. / [EN] The Texas citrus mite, Eutetranychus banksi McGregor, is native to the Americas and widely distributed across this continent. In 2013 it was detected in the south of the province of Valencia, affecting the main citrus-growing area in Spain. The mite produces severe damage, reducing photosynthesis, causing defoliation, and producing a lack in fruit pigmentation, which could affect its economic value. Firstly, this work evaluated the ecological impact produced by the invasive species on the resident spider mites Panonychus citri (McGregor) and Eutetranychus orientalis (Klein). Since its arrival in 2013, E. banksi has become the most frequent and abundant spider mite on citrus, competitively displacing the other species and reducing their presence and geographic range, which is influenced by its colonisation history. Secondly, this work studies the within-tree distribution and seasonal trends of the pest and associated phytoseiid mites, which may contribute to its control. The Texas citrus mite was feeding mainly on the adaxial (upper) side of leaves in the periphery of the tree canopy showing a single population peak in late summer-early autumn, while phytoseiids preferred the abaxial (lower) sides inside the canopy showing two peaks, a main spring peak and a second, less abundant, in autumn. Euseius stipulatus (Athias-Henriot) was the most frequent and abundant phytoseiid, and when E. banksi increased, it moved to the adaxial sides on outer leaves and fruits to feed on its prey and changed its colouring from white to reddish-brown, evidencing its contribution to biological pest control. However, it was not capable of maintaining E. banksi populations under tolerable densities due to the unfavourable predator/prey ratios in summer and early autumn. Thirdly, this work aimed to describe the pest population structure and define a sampling plan. There were differences in the age structure on fruits and leaves, as well as between leaves from different flushes. Furthermore, over time, there were fluctuations in its composition correlated with variations in sex-ratio. No aggregation differences among plant strata were found, but there were significant differences between immature and adult stages, the females being the less aggregated. The high correlation of the total population with the motile forms and females allowed the use of both as a reference stage in the sampling plan, establishing a presence/absence sampling of 100 leaves for females or 400 leaves for motile forms. Finally, a recently described phytoseiid mite, Neoseiulus madeirensis Papadoulis & Kapaxidi, was found to be associated with E. banksi, suggesting that it could be a promising candidate for pest suppression. Fourthly, this work aimed to evaluate the potential of this predator to control pest populations. Neoseiulus madeirensis exhibited a short developmental time, high survival and reproductive rates feeding on E. banksi. The predator was fed mainly on immature stages, with eggs being the preferred stage, showing a type II functional response for all the prey stages tested, that stabilises at high prey densities with high egg laying. Survival, predation and reproduction values were the best obtained so far for any phytoseiid previously tested against E. banksi, making it a suitable candidate for the development of a biological control program based on augmentative releases, or importation aiming for the establishment of permanent populations on citrus. / López Olmos, S. (2023). Towards the Integrated Management of the Texas Citrus Mite Eutetranychus Banksi (Acari: Tetranychidae) in Spain [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/201554

Forests are more than a stand of trees in the landscape. They represent a complex, functional system of interacting and often interdependent biological, physical and chemical components (Kimmins 1997). In the past, complex interactions were increasingly recognized over time as food webs, abiotic processes and biotic feedbacks since then defined as the forest ecosystem. Trees grow in a world of multitrophic interactions (van der Putten et al. 2001). One component of this functional system is represented in several aspects by spiders and insects, as they contribute considerably to the biodiversity and ecosystem functioning in forest habitats (Watt et al. 1997). There is knowledge on the community composition of several forests of different stand type or tree species composition referring to soil dwelling arthropods. Moreover, studies often highlight the orientation of single arthropod species on abiotic factors or the composition of species assemblages in case studies; these represent ecologically well described groups that can be used as indicators of habitat quality (Pearce and Venier 2006; Cardoso et al. 2004). Evidence on the scale of interactions between the species and their environment are rare. This applies particularly to examples based on fine spatial and temporal scales.

Cytogenetics of spiders of the infraorder Mygalomorphae is largely unknown. My thesis is focused on the karyotype evolution of spiders of families Hexathelidae Dipluridae, which are basal groups of the superfamily Avicularioidea. In this thesis the evolution of diploid chromosome number, chromosome morfology, and also sex chromosomes are studied. The karyotypes of the mygalomorphs are quite interesting for their diversity. Many members of the family Hexathelidae exhibit extreme systems of a chromosomal sex determination which are composed by many gonosomes X. The meiotic division of males is analyzed. Some species exhibit modifications of this division such as achiazmatic meiosis. Selected markers have been detected by fluorescence in situ hybridization. Keywords: evolution, fluorescent in situ hybridization, karyotype, nucleolus organizer region, sex chromosomes, spider

INTRODUÇÃO: O aracnídeo Phoneutria nigriventer, também conhecido por aranha-armadeira, possui um veneno complexo, contendo vários peptídeos que ativam canais iônicos nas células. Dentre estes, só dois neuropeptídeos, Tx2-5 e Tx2-6, destacam-se por relaxar o músculo liso trabecular do corpo cavernoso, induzindo ereção peniana em camundongos e ratos. Este efeito tem sido associado à produção de oxido nítrico pela ativação de óxido nítrico sintases. No entanto, faltam estudos mais amplos para determinar o papel de Tx2-6 na indução da ereção. OBJETIVOS: Identificar os genes diferencialmente expressos no tecido erétil de camundongos após indução da ereção pela Tx2-6 utilizando microarranjos de oligonucleotídeos. Validação dos resultados obtidos nos microarranjos por PCR quantitativa e imuno-histoquímica. MATERIAIS E MÉTODOS: Camundongos machos e adultos da linhagem Swiss foram divididos em dois grupos: controle (n=10), inoculados pela via intracavernosa com 20 l de solução salina; e tratado (n=10), os quais receberam 0,006gg/animal do peptídeo Tx2-6 diluído em 20 l de salina pela via intracavernosa. Uma hora após o início da ereção no grupo tratado todos os animais foram sacrificados e retirou-se o pênis. Este último foi dividido em dois fragmentos, uma parte do material foi congelada em nitrogênio líquido e mantida a 80°C até a extração do RNA para os experimentos de microarranjos e PCR quantitativa; outra parte foi utilizada para avaliação imuno-histoquímica. RESULTADOS: No grupo tratado a ereção foi observada 30-45 minutos após aplicação de Tx2-6 e mantida durante 120 minutos. Os camundongos de grupo controle não apresentaram nenhum indício de ereção. Nos experimentos de microarranjos, onde foram analisados 34.000 genes representando o genoma total do camundongo, identificou-se 3.803 (12,3%) genes com expressão diferencial de pelo menos ±1,5 vez entre os grupos (1.823 genes superexpressos e 1.980 genes subexpressos no grupo tratado comparado ao controle). Os genes ednrb, sparc, fn1, sstr2, pdgfr foram selecionados para validação dos microarranjos por PCR quantitativa e confirmaram a superexpressão em relação aos controles. As proteínas Fn1, Sstr2 e Pdgfr resultaram aumentadas no grupo tratado após avaliação imuno-histoquímica. CONCLUSÕES: A inoculação de Tx2-6 pela via intracavernosa alterou o perfil de expressão gênica no tecido erétil de camundongos. O número de genes superexpressos foi similar ao de genes subexpressos. Serão necessários outros estudos para entender melhor as vias moleculares que Tx2-6 afeta na indução da ereção peniana. / INTRODUCTION: The Phoneutria nigriventer arachnid, also known as armed-spider, has a complex venom, composed by several peptides that affect cellular ionic channels. Among these, only two neuropeptides, Tx2-5 and Tx2-6 induce penile erection in mice and rats and this effect has been associated with the production of nitric oxide by the activation of nitric oxide synthases. Moreover, there is a scarcity of studies focusing on the role of Tx2-6 in the induction of erection. OBJECTIVES: To identify the differently expressed genes in the erectile tissue of mice after erection induction by Tx2-6 using oligonucleotide microarrays. To validate microarray results by quantitative PCR and immunohistochemistry. MATERIALS AND METHODS: Swiss adult male mice were divided in two groups: control (n=10) were injected intracavernously with 20 gl of saline solution; and treated (n=10) were injected intracavernously with 0.006gg/mouse of the Tx2-6 peptide diluted in 20 gl of saline solution. After checking the penile erection in the treated group, all mice were sacrificed one hour after the beginning of erection for the removal of the penis. Penile organ was divided into two fragments, one piece was immediately frozen in liquid-nitrogen and stored at -80°C until RNA extraction to make the microarray and quantitative PCR experiments; the other was reserved for immunohistochemistry analysis. RESULTS: In the treated group, erection was noticed 30-45 minutes after Tx2-6 inoculation and lasted for 120 minutes. Control mice did not present any sign of erection. Considering as differentially expressed genes with a ±1.5 fold expression difference, of the 34,000 genes on the microarray we identified 3,803 (12.3%) genes differentially expressed between the groups (1,823 genes up-regulated and 1,980 genes down-regulated in the treated group compared to controls). The ednrb, sparc, fn1, sstr2, pdgfr genes were selected for validation of microarray results by using quantitative PCR and confirmed the up-regulation when compared to controls. After immunohistochemistry analysis the Fn1, Sstr2 and Pdgfr proteins were found increased in the treated group. CONCLUSIONS: The intracavernous inoculation of Tx2-6 modified the gene expression profile of erectile tissue of mice. The number of upregulated and down-regulated genes was similar. Further studies are needed to understand the molecular pathways that Tx2-6 affect to induce penile erection.

Made available in DSpace on 2017-07-21T19:59:48Z (GMT). No. of bitstreams: 1 Alinemargraf.pdf: 7420254 bytes, checksum: 591624049aa55aa68ad2d024197a427a (MD5) Previous issue date: 2009-09-30 / Loxosceles intermedia is a poisonous species widely distributed in the region of Curitiba and Campos Gerais of the Paraná State. There are descriptions in the literature concerning the reproductive behavior of this species and, more recently, studies have been carried out in order to describe the post-embryonic development of the female reproductive system. The adult male reproductive system is composed by two testicles connected to the vas deferens by two structures, a duct and an ampulla, not yet described for other spider species. Then in this work were described the morphological aspects of the development of the male reproductive system, the morphology of the duct and ampulla in the adult male and discussed possible functional and evolutionary implications of this segments on reproduction of L. intermedia. For the analysis in light microcopy, male animals at different stages of the development were etherized and sacrificed. Their abdomens were removed and fixed in 4% paraformaldehyde/ 0.1 M phosphate buffer, pH 7.4, and embedded in historesin (Leica). Subsequently, the serial 5 Tm sections were submitted to HE staining and PAS reaction. For the analysis in SEM, adult male animals were dissected and their whole testis-duct-ampulla was removed and fixed in 0.2 M sodium cacodilate, 2.5% glutaraldehyde and 2% paraformaldehyde, pH 7.4. The tissues were dehydrated and dried in alcohol series and critical point, respectively. Subsequently, the tissues were coated with gold and analyzed by SEM. Considering the fact that the organs of the male reproductive system of L. intermedia are tiny and fragile, which makes them difficult to dissect for observation in SEM by the usual methods, in this work was also evaluated some protocols, using paraffin, paraplast or PEG (for short and long time of infiltration) as infiltration medium, for obtaining serial sections, as well as three adherence substances to slides (5.5% albumin, 0.1% gelatin and 1% gelatin, the tissues samples were submitted of infiltration and 20 Tm sections were obtained, placed on glass slides, washed with solvent or water, dried at room temperature, coated with gold and analyzed in SEM. Infiltration with PEG for a short time and adhesion with 1% gelatin was found to be the best protocol resulting in the maintenance of the cell shape and tissue characteristics. Following the development of the male reproductive system of the brown spider it was observed that the maturation of the testis is the first event to take place. After the appearance of synspermium inside the testicles, the differentiation of the duct, ampulla and vas deferens is observed, and finally the external modification of the palps in copulatory organs occurs. In adult male, the tissue of the duct is composed of a non-secretory epithelium, without a defined lumen. The function of the duct is probably the selection of viable sperm cells. The ampulla wall is composed of a secretory epithelium whose glycoprotein secretion involves the synspermium. The secretion released by the ampulla could be involved in distinct aspects of the functionality of the gametes. The ampulla secretion could also be related to mechanisms of sexual selection, not yet established for this species of spider. In conclusion, this morphological novelty in L. intermedia male reproductive system appears to constitute an important evolutionary event on spider reproduction. / Loxosceles intermedia é uma espécie de aranha venenosa amplamente distribuída na região de Curitiba e Campos Gerais do Paraná. Há descrições na literatura sobre o comportamento reprodutivo dessa espécie e, mais recentemente, estudos têm sido realizados a fim de descrever o desenvolvimento pós-embrionário do sistema reprodutor masculino. O sistema reprodutor masculino adulto é composto por dois testículos, ligado ao vas deferens por duas estruturas, um ducto e uma ampola, ainda não descritas para outras espécies de aranhas. Neste trabalho foram descritos os aspectos morfológicos do desenvolvimento do sistema reprodutor masculino, a morfologia do ducto e da ampola no macho adulto e discutidas as possíveis implicações funcionais e evolutivas destes segmentos sobre a reprodução de L. intermedia. Para a análise em microscopia óptica, os machos em diferentes fases do desenvolvimento foram anestesiados e sacrificados. Seus abdomens foram removidos e fixados em 4% paraformaldeído / tampão fosfato 0,1 M, pH 7,4, e incorporado em historesina (Leica). Posteriormente, os cortes seriados a 5 Tm foram submetidos à coloração HE e reação PAS. Para a análise em MEV, machos adultos foram dissecados e o conjunto testículo-ducto-ampola foi removido e fixado em cacodilato de sódio 0,2 M, glutaraldeído 2,5% e paraformaldeído 2%, pH 7,4. Os tecidos foram desidratados e secos em série alcoólica e ponto crítico. Posteriormente, os tecidos foram cobertos com ouro e analisadas por MEV. Considerando-se o fato de que os órgãos do sistema reprodutor masculino de L. intermedia são pequenos e frágeis, o que torna difícil a dissecção para observação em MEV pelos métodos usuais, neste trabalho também foram avaliados alguns protocolos de infiltração, utilizando parafina, paraplast e PEG (por tempo curto e longo prazo de infiltração), para a obtenção de cortes seriados, bem como testou-se três substâncias de adesão às lamínulas (albumina 5,5%, gelatina 0,1% e gelatina 1%), as amostras de tecidos foram submetidos a infiltração e cortes a 20 μm foram obtidos. Estes foram colocados em lâminas de vidro, lavados com solvente ou água, secos à temperatura ambiente, revestidos com ouro e analisados em MEV. Infiltração com PEG por um tempo curto e aderência com gelatina 1% foi o melhor protocolo resultando na manutenção da forma das células e das características do tecido. No desenvolvimento do sistema reprodutor masculino da aranha marrom foi observado que a maturação do testículo é o primeiro evento a acontecer. Após o aparecimento da synspermium dentro dos testículos, a diferenciação do ducto, ampola e vas deferens é observada e, finalmente, a modificação externa do palpos em órgãos copulatórios ocorre. No macho adulto, o tecido do ducto é composto de um epitélio não secretor, sem uma luz definida. A função do ducto é, provavelmente, a seleção de espermatozóides viáveis. A parede da ampola é composta por um epitélio secretor, cuja secreção glicoproteíca envolve o synspermium. A secreção liberada pelo ampola pode esta envolvida em aspectos distintos da funcionalidade dos gametas, bem como pode também estar relacionada a mecanismos de seleção sexual, ainda não definidos para esta espécie de aranha. Conclui-se que, esta novidade morfológica no sistema reprodutor masculino de L. intermedia parece constituir um importante evento evolutivo na reprodução de aranha.

O sul de Minas Gerais é a principal região produtora de morangos (Fragaria x ananassa Duch.) no Brasil. O ácaro-rajado, Tetranychus urticae Koch (Acari: Tetranychidae), é considerado uma das principais pragas do morangueiro no Brasil e em vários países. Uma prática importante na cultura do morango refere-se à cobertura de solo, usualmente realizada com filme de polietileno, porém além de ser caro gera resíduos muito persistentes no ambiente. Objetivou-se com este trabalho comparar a resistência de oito genótipos de morango (\'Albion\', \'IAC Guarani\', \'IAC Princesa Isabel\', \'Oso Grande\', IAC T-0104, IAC 12, IAC 4 e IAC 1.13) ao ácaro-rajado; determinar o efeito da cobertura de solo (com a polpa de café desidratada, conhecida como palha de café) nas pragas/ patógenos e ácaros predadores desta cultura; comparar a ocorrência de pragas, patógenos e inimigos naturais entre um cultivo orgânico e outro convencional de morango. Os resultados obtidos indicaram que \'IAC Princesa Isabel\', IAC T-0104 e IAC 12 são resistentes, IAC 1.13 e IAC 4 apresentam resistência moderada e \'IAC Guarani\', \'Oso Grande\' e \'Albion\' são suscetíveis ao ácaro-rajado. \'Oso Grande\' e \'Albion\' são bastante utilizadas pelos produtores do sul de Minas Gerais. O uso de palha de café aumentou o número de predadores edáficos, tanto no solo dos canteiros (campo) e em vasos (laboratório) como em folíolos de morangueiro. O Gamasina Proctolaelaps pygmaeus (Müller) (Melicharidae) foi visto sobre morangueiros, principalmente no período noturno. Maior número de ácaro-rajado e maior severidade de doenças foram observados em plantas cultivadas em solo coberto com polietileno. Além disso, foi maior o nível de infecção de ácaro-rajado pelo fungo Neozygites floridana (Weiser e Muma) em plantas cultivadas em solo coberto com palha de café. Não foi observada diferença significativa entre a produtividade da cultura em solo coberto com polietileno e em solo coberto com palha seca de café. Os resultados da comparação da ocorrência de ártropodes e patógenos entre sistema orgânico e convencional mostraram menor ocorrência de ácaro-rajado e predadores em cultivo de morangueiro convencional, porém ocorrência considerável de tripes, mosca-branca e mofo cinzento. A incidência de dendrofoma e mancha de pestalotia foi a mesma nos dois sistemas de cultivos. Os resultados sugerem a conveniência de se continuar o processo de desenvolvimento dos genótipos que se mostraram menos afetados pelo ácaro-rajado, para que estes possam no futuro ser utilizados pelos produtores, e a condução de estudos complementares, que avaliem o desempenho de cultivos em sistema orgânico que incorporem o uso da palha de café para a cobertura do solo. / The southern of Minas Gerais is the main region producing strawberries (Fragaria x ananassa Duch.) in Brazil. The two-spotted spider mite, Tetranychus urticae Koch (Acari: Tetranychidae), is considered one of the main strawberry pests in Brazil and in several countries. An important practice in strawberry cultivation refers to soil cover, which is usually made with polyethylene film, but it is expensive and generates very persistent residues in the environment. The objective of this study was to compare the resistance of eight strawberry genotypes (\'Albion\', \'IAC Guarani\', \'IAC Princesa Isabel\', \'Oso Grande\', IAC T-0104, IAC 12, IAC 4 and IAC 1.13); to determine the effect of soil cover (with dehydrated coffee pulp, known as coffee straw) on the pests mites/ pathogens and predatory mites of this crop; to compare the occurrence of pests, pathogens and natural enemies between an organic and a conventional strawberry crop. The results indicated that \'IAC Princesa Isabel\', IAC T-0104 and IAC 12 are resistant, IAC 4 and IAC 1.13 are moderately resistant, and \'IAC Guarani\', \'Oso Grande\' and \'Albion\' are susceptible to the two-spotted spider mite. \'Oso Grande\' and \'Albion\' are widely used by producers in the southern of Minas Gerais. The use of coffee straw increased the number of edaphic predators, both in the soil beds (field) and in pots (laboratory) as well as in strawberry leaflets. The Gamasina Proctolaelaps pygmaeus (Müller) (Melicharidae) was seen on strawberry leaftlets, mainly in the nocturnal period. Higher numbers of the two-spotted spider mites and disease severity were observed in plants grown on soil covered with polyethylene. In addition, the level of the two-spotted spider mite infected by the fungi Neozygites floridana (Weiser and Muma) was higher in plants cultivated in soil covered with coffee straw. No significant difference was observed between yield in soil covered with polyethylene and in soil covered with dry coffee straw. The results of the comparison of the occurrence of arthropods and pathogens between the organic and conventional systems showed fewer occurrences of the two-spotted spider mites and predators in conventional strawberry cultivation, but considerable occurrence of thrips, whitefly and gray mold. The incidence of dendrophoma and leaf spots caused by pestalotia was the same in both cropping systems. The results suggest the convenience to continue the development process of the genotypes that were less affected by the two-spotted spider mite, which could be used in the future by producers, and the conduction of complementary studies to evaluate the performance of crops in organic system that incorporate the use of coffee straw as soil coverage.

The importance of social behaviour is evident in human society, but there are both costs and benefits associated with cooperation and sociality throughout the animal kingdom. At what point do the benefits outweigh the costs, and when do selective pressures favour sociality and colonization over solitude and independence? To investigate these questions, we have focused on an anomalous species of spider, Anelosimus studiosus, also known now as the northern social spider. Throughout its broad range, A. studiosus is solitary and aggressive, but recently, colonies of cooperative and social individuals have been observed at northern latitudes. This leads to two research questions: 1) what characteristics differentiate the two variants behaviourally, and, 2) how are they different physiologically? Colonies and individuals were collected from multiple populations throughout the Tennessee River watershed area and maintained in a laboratory environment for quantitative and qualitative assessment of behavioural traits as well as specific neurochemical analysis by high performance liquid chromatography with electrochemical detection. After classifying individuals as social or aggressive, I looked at the influence of factors such as age, reproductive state, nutritional state, and time of day on behaviour and neurophysiology. I found correlations between social behaviours and serotonin, aggressive behaviours and octopamine (invertebrate counterpart of norepinephrine), and several other compounds associated with an increase or decrease in aggression. These studies combine techniques from multiple disciplines to contribute to the greater understanding of the proximate control of social and aggressive behaviours as well as factors influencing the evolution of sociality.

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