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Made available in DSpace on 2014-06-11T19:31:00Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-07-30Bitstream added on 2014-06-13T20:21:38Z : No. of bitstreams: 1 goncalves_hb_me_araiq.pdf: 1598700 bytes, checksum: 9f31d3b16656a31ddf594835f3745ebf (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A tanase (EC 3.1.1.20) é uma enzima induzível que age sobre os taninos hidrolisando suas ligações éster e depsídicas obtendo-se como produtos a glicose e o ácido elágico ou ácido gálico, sendo este último, um importante substrato para as indústrias farmacêutica e química. Entre os diferentes organismos capazes de produzir tanases, os microorganismos, de modo especial os fungos filamentosos, vêm se destacando uma vez que são mais versáteis na degradação de diferentes tipos de taninos. Neste contexto, o objetivo deste trabalho foi estudar as tanases intra e extracelulares do fungo filamentoso Emericella nivea produzidas em Fermentação Submersa (FSbm) e em Fermentação em Substrato Sólido (FSS), purificando-as e caracterizando-as bioquimicamente, além de imobilizá-las em suportes de agarose. Em princípio, foi realizada a seleção da melhor cepa produtora de tanases, submetendo-se 42 linhagens fúngicas a FSbm em meio de cultura Khanna com 2% de ácido tânico como fonte de carbono, por 3 a 4 dias a 30ºC, tendo sido o fungo Emericella nivea selecionado para prosseguimento do trabalho. Para este microorganismo os maiores nívies enzimáticos extracelulares foram obtidos em 3 dias de cultivo em FSbm e 8 dias em FSS, sendo para esta última utilizados produtos agroindustriais e folhas de vegetais de diferentes espécies secas trituradas umedecidas com água de torneira (1:1; p/v). As tanases extra e intracelular foram purificadas 61 e 2,5 vezes com recuperação de 30% e 8,8%, respectivamente. Eletroforese em condições não desnaturantes (PAGE 7%) mostrou a presença de uma única banda protéica revelada por prata e para atividade tanásica com a mesma mobilidade relativa. A forma extracelular possui massa molecular nativa de aproximadamente 322kDa com 50% de conteúdo de carboidratos. Já a enzima intracelular apresentou massa molecular nativa de 258kDa e 17% de... / Tannases (EC 3.1.1.20) are inducible enzymes that catalyze the hydrolysis of ester and depside bonds in hydrolysable tannins releasing glucose and ellagic acid or gallic acid, which is an important compound used in pharmaceutical and chemical industries. Among different organisms able to produce these enzymes, the microorganisms, especially filamentous fungi deserve attention since they can act on different tannins degradation ways. In this context, the aim of this work was to study the intra and extracellular tannases from the filamentous fungus Emericella nivea produced in Submerged Fermentation (SbmF) and Solid Substrate Fermentation (SSF), purifying and characterizing them biochemically, as well to immobilize the extracellular enzyme in agarose supports. First of all, it was selected the best tannase producer among 42 strains, in Khanna culture medium with 2% tannic acid as carbon source for 3-4 days at 30°C, and the fungus Emericella nivea was selected. This fungus produced high levels of extracellular enzyme at 3 and 8 days when cultivated in SbmF and SSF at 30°C, respectivally. FSS was performed with agroindustrial products or crushed dried leaves of different plants umidified with tap water (1:1, w/v). The extra and intracellular tannases were purified 61 times and 2.5-times, with recovery of 30% and 8.8%, respectivally. Non-denaturing electrophoresis (PAGE 7%), showed a unique proteic band stained by silver and for activity, both with the same relative mobility. The extracellular enzyme, probably, is a hetero-dimeric protein with native molecular mass of 322 kDa with 50% of carbohydrate content and the intracellular with native molecular mass of 258 kDa and 17% of carbohydrate. The optimum temperature were 45ºC and 50°C for the extra and intracellular enzymes, respectively and the optimum pH for both enzymes was 5.0. The soluble tannases were thermostable with... (Complete abstract click electronic access below)

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In its introduced southern hemisphere range, Sirex noctilio causes considerable mortality in non-native pine forests. In its native Eurasian range however, S. noctilio is of little concern perhaps due to interactions with a well-developed community of pine-inhabiting insects and their associated microorganisms. If such interactions occur, they may limit the woodwasp’s impact in its newly introduced range in North America. My research addresses two broad questions: 1) Does S. noctilio share its habitat with other insects and if so, with whom? 2) Is there evidence that co-habitants affect S. noctilio, and if so how might such interactions occur? Field studies undertaken to describe the woodwasp’s host-attack ecology in Pinus sylvestris showed S. noctilio activity occurred between mid-July and late August, and other phloem- and woodborers sometimes entered the tree after the woodwasp. Tree mortality occurred from two weeks to several months after initial woodwasp symptoms. Suppressed or intermediate trees, those with ≤ 25% residual foliage, or those with stem injury or previous woodwasp symptoms were most likely to have symptoms of woodwasp attack. A second field study conducted to identify associated insect species in S. noctilio-infested Pinus sp. showed the wasp was sometimes found alone, but usually shared the tree with other phloem- or woodboring insects, most commonly the curculionids Tomicus piniperda, Pissodes nemorensis and Ips grandicollis and the cerambycid Monochamus carolinensis. I found no indication that wasps were absent when beetles were present, but there was evidence that woodwasps were less abundant, but larger, when beetles were present. Experiments showed that indirect interactions can occur between the two insect groups via fungal associates of one or both. In the laboratory, the woodwasp symbiont was outcompeted by two beetle-associated fungi, Leptographium wingfieldii and Ophiostoma minus, over a range of temperatures. Under field conditions the woodwasp was able to detect and avoid ovipositing in P. sylvestris inoculated with L. wingfieldii, but its oviposition was unaffected by O. minus. My results show that insects co-habiting pine with S. noctilio have potential to exert a measure of biological control on the woodwasp and may help to limit its impact in North America.