Untersuchungen wurden durchgeführt, um die Wirkung von B. subtilis und deren Metaboliten auf den Meloidogyne- und Pratylenchus-Befall und ihre Vermehrung festzustellen sowie um die möglichen Wirkmechanismen zu studieren. Substratbehandlungen mit B. subtilis FZB 24® führten zu einem höherenMeloidogyne-Befall und einer verstärkten Nematodenvermehrung. Trotz verstärktem Befall wurde das Pflanzenwachstum verbessert (induzierte Toleranz). Weiterhin wurden durch "antibiotikafreie" Kulturfiltrate (KF) aus der bakterielen Übergangs- und stationären Phase ähnliche Wirkungen erzielt. Außerdem wurde eine systemische Wirkung von B. subtilis auf den Meloidogyne-Befall an Tomate nachgewiesen. B. subtilis bzw.die KF-behandelte Testpflanzen zeigten stärkere Anlockwirkung auf Meloidogyne-Larven (M. incognita, M. arenaria) als unbehandelte. KF (50, 10, 1%) von B. subtilis zeigten keine nematizide Wirkung auf die Meloidogyne-Larven. KNO3 als Trägersubstanz für das Bakterienpräparat besaß ähnliche Wirkungen auf den Meloidogyne-Befall und die Nematodenvermehrung.Ebenfalls wurde das Pflanzenwachstum durch KNO3-Zufuhr gefördert. Der Einsatz des nematodenfangenden Pilzes Arthrobotrys superba reduzierte den Meloidogyne-Befall (30% Gallenreduktion). Hingegen wurde durch die kombinierte Anwendung von A.superba und B. subtilis FZB 24® der Bekämpfungserfolg von A. superba reduziert. Die exogene Applikation von Phytohormonen bzw. Präkursoren zeigte keine Wirkung auf das Wachstum der Testpflanzen. Die Vermehrung von Meloidogyne wurde durch IAA und die Kombination von IAA und Kinetin gefördert. In den getesteten Konzentrationen der Phytohormone wurde keine Wirkung auf die Mortalität der Wurzelgallenälchen- (Meloidogyne-) Larven beobachtet. Der Gehalt von Enzymen (Chitinase, Glucanase und Peroxidase) aus dem Sproß behandelter Tomatenpflanzen wurde bestimmt. B. subtilis-Isolate (FZB 24® , S18) reduzierten die Population von Wurzelläsionsnematoden, Pratylenchus penetrans (nicht signifikant) [9 bzw. 15-20% pro Wurzelsystem bzw. g Wurzel]. Das Pflanzenwachstum wurde an befallenen Pflanzen durch beide Isolate verbessert (induzierte Resistenz). Es wurden keine Unterschiede zwischen den Bakterien-Isolaten festgestellt. KNO3 führte ebenfalls zu einer Verminderung der Nematodenpopulation. Die Ergebnisse werden hinsichtlich möglicher Wirkmechanismen des RhizobakteriumsB. subtilis und des nematodenfangendes PilzesA. superba zur Regulierung der Nematodenpopulation bei Tomate diskutiert. / Investigations were made to know about the effects of Bacillus subtilis and its metabolites on the infestation of tomato plants with root-knot and lesion nematodes. Further more, experiments were carried out to clear up the mode of actions ofB. subtilis and its culture filtrates on infestation of tomato seedlings and reproduction of root-knot nematodes. Substrate applications ofB. subtilis FZB 24® leaded to an increasement of infestations intensity and reproduction of root-knot nematodes (M. arenaria). Eventhough, bacterized and inoculated plants with root-knot nematodes showed better growth than with bacteria untreated plants (induced tolerance). In addition "antibiotica free" culture filtrates (CF) from transitional and stationary bacterial growth phase also promoted reproduction of root-knot nematodes. These CF elicitized tolerance of tomato plants towardsMeloidogyne too. It was proved that B. subtilis could induce a systemic reaction of tomato plants towards root-knot infestation. Besides that test plants treated with B. subtilis (cells) or CF were more attractive to Meloidogyne-Larvae (M.arenaria & M.incognita) than untreated once. CF in 50, 10 and 1% concentrations did not have a nematicidal effect on the root-knot larvae. KNO3 -the carrier of the bacterial preparate (B. subtilis FZB 24®) - had also the same effects on infestation and reproduction of root-knot nematodes. Plant growth was promoted due to application of KNO3. The use of nematode trapping fungus, Arthrobotrys superba gave some range of nematode (root-knot) control (30% gall reduction). Whereas, with the combination ofA. superba and B. subtilis FZB 24® the effect of the fungus was reduced. The application of exogenic phytohormones and precursores showed no effect on plant growth. Reproduction of Meloidogyne was promoted by IAA and combination of IAA and kinetin (not significant). In the tested concentrations of these phytohormones there was no direct mortality effect on root-knot larvae. Content of some enzymes (chitinase, glucanase and peroxidase) from shoot of treated tomato plants was determind. B. subtilis-Isolates (FZB 24® and S18) reduced the population of lesion nematodes,Pratylenchus penetrans in attacked plants (not significant) [9% per root system and 15-20% per g root]. The treatment improved the predisposition of the plants to lesion nematodes (induced resistance). Plant growth was also improved. There was no difference between the bacterial isolates in their effect. KNO3 reduced also nematode population. In general the results would be explained and discussed towards possible mode of actions of rhizobacterium B.subtilis and nematode trapping fungus A. superba.

Submitted by Maria Beatriz Vieira (mbeatriz.vieira@gmail.com) on 2017-04-06T12:36:48Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) dissertacao_adinara_maria_welter.pdf: 193527 bytes, checksum: f6ee9eb7b69a39c972690a65d2001e3f (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-04-19T19:22:34Z (GMT) No. of bitstreams: 2 dissertacao_adinara_maria_welter.pdf: 193527 bytes, checksum: f6ee9eb7b69a39c972690a65d2001e3f (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-04-19T20:06:49Z (GMT) No. of bitstreams: 2 dissertacao_adinara_maria_welter.pdf: 193527 bytes, checksum: f6ee9eb7b69a39c972690a65d2001e3f (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-04-19T20:10:47Z (GMT). No. of bitstreams: 2 dissertacao_adinara_maria_welter.pdf: 193527 bytes, checksum: f6ee9eb7b69a39c972690a65d2001e3f (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2015-06-01 / Sem bolsa / O presente trabalho objetivou avaliar a eficiência de produtos com princípio ativo biológico e químico no controle de Meloidogine javanica em ensaio de casa de vegetação e in vitro. O delineamento utilizado foi inteiramente casualizado com 15 repetições cada produto para casa de vegetação e 4 parcelas para teste in vitro. Os produtos testados foram: produto 1 (sem princípio ativo) dose 0; produto 2 (Trichoderma harzianum) com dose de 20g/100kg de semente; produto 3 (Pasteuria penetrans) com dose de 150ml/100kg de semente; produto 4 (Trichoderma harzianum) com dose de 20g/100kg de semente; produto 5 (Purpureocillium liliacinum) com dose de 20 l/ha; produto 6 (Bacillus megaterium/Bacillus subitilis) com dose de 60 ml/100kg de semente; produto 7 (Trichoderma asperellum/Bacillus subtilis) com dose de 1g/Kg de semente; produto 8 (Abamectina) com dose de 100ml/100kg de semente. Avaliou-se peso e comprimento de raiz, galha, massa de ovos, ovos, juvenis e fator de reprodução. Na avaliação de peso de raiz os tratamentos 2, 4 (Trichoderma harzianum), e 7 (Trichoderma asperellum/ Bacillus subtilis) não diferiram entre si, mantendo-se superiores aos demais tratamentos na avaliação de galhas os produtos 3 (Pasteuria penetrans) e 8 (Abamectina) foram os que apresentaram menor número de galhas por sistema radicular; para massas de ovos o produto 1 (sem princípio ativo) foi o que apresentou maior número; o resultado de juvenis os produtos com princípio ativo destacaram-se com exceção do 2 (Trichoderma harzianum) que não apresentou controle; na avaliação de ovos os produtos 1,6,7 e 8 foram os que menos multiplicaram; quanto ao fator de reprodução, os produtos com maior controle de M. javanica foram os produtos 3 (Pasteuria penetrans) e 8 (Abamectina). Na avaliação in vitro nas doses de 1 e 2 %, todos os produtos, biológicos e o químico, mostraram controle do nematoide das galhas. Conclui-se que, produtos biológicos são eficientes no controle de M. javanica. A Pasteuria penetrans e a Abamectina foram os produtos mais eficientes. / This study aimed to evaluate the efficiency of products with biological and chemical active ingredient in control Meloidogyne javanica under greenhouse and in vitro home test. The design was completely randomized with 15 repetitions each product for greenhouse and fourth installments in vitro test. The products tested were: 1 product (no active ingredient) dose 0; Product 2 (Trichoderma harzianum) at a dose of 20g / 100kg seed; 3 product (Pasteuria penetrans) with dose of 150ml / 100kg seed; Product 4 (Trichoderma harzianum) at a dose of 20g / 100kg seed; 5 product (Purpureocillium liliacinum) at a dose of 20 l / ha; 6 product (Bacillus megaterium /Bacillus subitilis.) with dose of 60 ml / 100kg seed; product 7 (Trichoderma asperellum / Bacillus subtilis) at a dose of 1 g / kg seed; 8 product (Abamectin) with dose of 100ml / 100kg seed. It evaluated weight and root length, gall, egg mass, eggs, juveniles and reproduction factor. For root weight the product 4 (Trichoderma harzianum) obtained the highest average; to root length the product 7 (. Trichoderma asperellum / Bacillus subtilis) was highest average; the gall evaluation Products 3 (Pasteuria penetrans) and 8 (Abamectin) showed the lowest number of galls per root system; for egg masses the product 1 (no active ingredient) showed the greatest number; the result of juvenile products with active ingredient stood out except 2 (Trichoderma harzianum) that did not show control; the eggs assessment Products 1,6,7 and 8 were the least multiplied; as the reproduction factor, products with greater control of M. javanica were the products 3 (Pasteuria penetrans) and 8 (Abamectin). When evaluated in vitro at doses of 1 and 2%, all organic products and the chemical showed control of nematode galls. We conclude that both, biological and chemicals can be effective in controlling M. javanica. Since for biological tested the bacteria Pasteuria penetrans showed greater efficiency.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / IntroduÃÃo: Existe pouco conhecimento sistemÃtico sobre a epidemiologia da tungÃase em Ãreas endÃmicas. SÃo necessÃrios mÃtodos de avaliaÃÃo rÃpida para delimitar Ãreas em risco e possibilitar a implementaÃÃo e avaliaÃÃo do impacto de intervenÃÃes. Objetivo: Desenvolver e avaliar um mÃtodo epidemiolÃgico rÃpido para estimar a prevalÃncia geral e a gravidade da tungÃase em comunidades endÃmicas com diferentes caracterÃsticas demogrÃficas, sÃcio-culturais e ambientais. Material e MÃtodos: AnÃlise de dez estudos transversais realizados em cinco comunidades, trÃs do Brasil (dados secundÃrios) e duas da NigÃria (dados primÃrios), no perÃodo de 2001-2008. A tungÃase foi diagnosticada clinicamente a partir da presenÃa de lesÃes de Tunga penetrans na epiderme dos indivÃduos. Para elaborar o mÃtodo rÃpido, seis localizaÃÃes topogrÃficas dos pÃs foram selecionadas como potenciais indicadores de infestaÃÃo em nÃvel comunitÃrio. Foram feitas regressÃes lineares, calculados coeficientes de determinaÃÃo (R2) e os valores p. CritÃrios operacionais como rapidez e facilidade do exame tambÃm foram utilizados para a escolha do mÃtodo mais adequado. Resultados: Ao todo, nos dez estudos transversais foram incluÃdos 7.121 indivÃduos. As prevalÃncias da tungÃase variaram entre 21% e 54%. Nas duas comunidades nigerianas a prevalÃncia geral da tungÃase nas 302 pessoas examinadas foi 47% (intervalo de confianÃa 95%: 41%-53%). Os homens foram mais infestados do que as mulheres (51% vs. 44%, p=0,2), assim como as crianÃas em comparaÃÃo aos indivÃduos >15 anos (60% vs. 33%, p<0,0001). Na anÃlise dos potencias mÃtodos rÃpidos, os coeficientes de determinaÃÃo foram altos, variaram entre 70% e 96% para as seis localizaÃÃes topogrÃficas, com valores p significantes (todos <0,002). A prevalÃncia na area periungueal dos pÃs apresentou o mais alto coeficiente de determinaÃÃo (96%), alÃm da maior rapidez e facilidade do exame. As prevalÃncias gerais estimadas a partir da equaÃÃo da reta Y= 1,12 x prevalÃncia na Ãrea periungueal + 5,0 apresentaram mÃdia do erro absoluto de 1,9%. PrevalÃncias graves (> 20 lesÃes), estimadas pela equaÃÃo da reta Y= 0,24 x prevalÃncia na Ãrea periungueal - 3,4, apresentaram erro absoluto mÃdio de 0,9%. ConclusÃo: A tungÃase à um problema de saÃde pÃblica em comunidades pesqueiras na NigÃria. A identificaÃÃo de T. penetrans na Ãrea periungueal dos pÃs pode ser usada como mÃtodo rÃpido e confiÃvel de avaliaÃÃo epidemiolÃgica. Sua aplicaÃÃo auxiliarà na delimitaÃÃo de Ãreas endÃmicas, bem como no planejamento de medidas que visem à reduÃÃo da tungÃase em Ãreas endÃmicas. . / . Rapid assessment methods are needed to delimitation of risk communities and to enable implementation and evaluation of impact interventions. Objective: To develop and assess a rapid epidemiologic method to estimate the overall prevalence of tungiasis and severity of disease in endemic communities with distinct demographic, socio-cultural and environment characteristics. Material and Methods: Analysis data from ten population-based surveys on tungiasis, performed in five endemic communities â three in Brazil (secondary data) and two Nigeria (primary data) â between 2001 and 2008. In all surveys, tungiasis was clinically diagnostic by presence of Tunga penetrans into epidermis of participants. To elaborate rapid assessment method six topographic sites of the feet were selected as potential infestation indicator in community level. Linear regression analyses were performed as well strength of associations (R2) and p values were calculated. Estimated prevalences were calculated for each of the ten surveys and compared to true prevalences. The most useful topographic localization to define a rapid assessment method was select based on the strength of association and operational aspects. Result: In total, 7121 individuals of the five communities were examined. Prevalence of tungiasis varied between 21.1% and 54.4%. In the two Nigerian communities the combined overall prevalence was 47% (142/302; 95% confidente intervale: 41%-53%). Tungiasis were more common in males than in females (51,5% vs. 46,3%; p=0,2). Children prevalence (<14 years) was statistically higher then adolescents and adults prevalence (60% vs. 33%, p<0,0001). In the ten surveys, all strength coefficients were high for the six localizations (ranged between 70% and 96%) and p values were significant (all <0,002). The presence of periungual lesions on the toes showed the highest strength coefficient (R2=96%; P<0.0001) and it was identified as the most useful and rapid localization to estimate the prevalence of tungiasis. Estimating prevalence of tungiasis by equation [estimated prevalence] = 1,12 x [prevalence on periungual sites] + 5,0), the mean absolute error was 1,9%. Tungiasis on this topographic site also reliably estimated prevalence of severe disease (>20 lesions) using the equation [estimated prevalence] = 0,24 x [prevalence on periungual sites] â 3,4); mean absolute error was 0,9%. Conslusion: Tungiasis is a public health problem in the fishing communities in Nigeria. Identification of T. penetrans in the periungual area of the feet can be used as a rapid and reliable method to assess the epidemiological situation of endemic areas. This approach will help to delimit endemic communities and to plan control measures aimed at the reduction of tungiasis. Key-words: Tungiasis; Tunga penetrans; Epidemiology; Rapid assessment method; Public Health; Africa; Brazil

Submitted by Marco Antônio de Ramos Chagas (mchagas@ufv.br) on 2017-04-27T18:33:44Z No. of bitstreams: 1 texto completo.pdf: 878289 bytes, checksum: 70c4ac466fdd682cb943dce260aec8df (MD5) / Made available in DSpace on 2017-04-27T18:33:44Z (GMT). No. of bitstreams: 1 texto completo.pdf: 878289 bytes, checksum: 70c4ac466fdd682cb943dce260aec8df (MD5) Previous issue date: 2004-08-27 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Experimentos foram conduzidos em laboratório e casa de vegetação objetivando o aprimoramento do método clássico de multiplicação de Pasteuria penetrans ‘in vivo’, proposto em 1980 por Stirling e Wachtel. No primeiro experimento comparou-se a produção de endósporos da P. penetrans em raízes de tomateiro de crescimento indeterminado cv. Santa Clara e determinado cv. TRural 1. Maiores pesos de matéria fresca e seca e número de endósporos foram observados em tomateiro cv. Santa Clara. O segundo experimento foi realizado para determinar a concentração ideal de endósporos de P. penetrans na suspensão e o tempo de agitação necessário para adesão adequada de endósporos aos nematóides para multiplicação da bactéria. Para que se obtenham seis endósporos, em média, por juvenil de segundo estádio (J2) de Meloidogyne javanica, verificou-se serem necessárias suspensões contendo 3,3 x 10 5 endósporos/mL, agitadas por 10 a 20 minutos, ou 3,3 x 10 4 endósporos/mL por 50 minutos. Em outro ensaio, comparou-se a multiplicação de P. penetrans em população pura de M. incognita ou em população composta de M. incognita e M. javanica oriunda de um campo de cultivo de tomate. A produção de endósporos de P. penetrans em tomateiro inoculado com M. incognita foi aproximadamente três vezes superior àquela em plantas inoculadas com população mista. Realizou-se também um teste de adesão de P. penetrans às duas populações do nematóide. Foi observado maior número de endósporos aderidos aos J2 de M. incognita. A produção de endósporos da P. penetrans em plantas de tomateiro cv. Santa Clara com 15, 30, 45 ou 60 dias de idade e inoculadas com 5.000, 15.000 ou 25.000 J2 foi avaliada. As plantas com 30 e 45 dias de idade inoculadas com 25.000 J2 permitiram a multiplicação de P. penetrans cerca de 19 vezes maior àquela obtida em plantas inoculadas com 5.000 J2. Com o objetivo de determinar se maiores níveis de matéria orgânica adicionada ao substrato provocavam alterações fisiológicas nos nematóides ou nas plantas de tomate, estudou-se a influência de diferentes proporções de solo, areia e esterco de curral (1:1:0, 2:2:1, 1:1:1 ou 1:1:2 (V:V:V), respectivamente) e três níveis de inóculo de espécies de Meloidogyne (3.000, 6.000 e 9.000 J2) sobre a concentração de fenóis em raízes de tomateiro, no teor de lipídios de espécies de Meloidogyne, e em possíveis alterações em células gigantes induzidas por esses nematóides. Não se observou efeito dos tratamentos no teor de lipídios dos nematóides. A concentração de fenóis nas raízes aumentou à medida que se acrescentou mais esterco de curral ao substrato ou quando as plantas foram inoculadas com mais nematóides (9.000 J2). As células gigantes em raízes de plantas cultivadas nos substratos 1:1:0 e 2:2:1 (solo:areia:esterco) foram mais numerosas, maiores e com maior número de núcleos. Por outro lado, as células gigantes de plantas cultivadas no substrato 1:1:1 e 1:1:2, além de menos numerosas, apresentaram alterações no tamanho e formato, demonstrando o efeito deletério das maiores doses de esterco sobre esses sítios de alimentação. O último ensaio foi conduzido para avaliar o efeito de crescentes quantidades de esterco de curral nos substratos e níveis de inóculo de espécies de Meloidogyne na reprodução de P. penetrans. Maior percentual de fêmeas infectadas por P. penetrans foi observado quando se utilizou o substrato 1:1:0 em relação aos substratos 1:1:1 e 1:1:2 ou quando as plantas foram inoculadas com 3.000 J2. O experimento foi repetido uma vez e na primeira condução do experimento, plantas cultivadas no substrato 1:1:0 ou inoculadas com 9.000 J2 apresentaram maior número de endósporos; entretanto, na segunda condução do experimento as plantas inoculadas com 9.000 J2 e cultivadas no substrato 2:2:1 foram as que permitiram maior reprodução de P. penetrans. / Greenhouse and laboratory experiments were conducted to improve the classical method of multiplying P. penetrans ‘in vivo’, proposed by Stirling & Wachtel in 1980. In the first experiment, the mass production of P. penetrans in tomatoes of indeterminated and determinated growth, cv. Santa Clara and Trural I, respectively, was compared. Higher fresh and dry root weight and endospore number were observed in ‘Santa Clara’ tomato. The second experiment was conducted to determine the best endospore concentration of P. penetrans in the aqueous suspension and the time of shaking necessary to obtain adequate attachment of endospores on the nematodes, as the first step for P. penetrans multiplication. To obtain an average of six endospores per second stage juvenile (J2) of M. javanica, it is necessary to shake suspension of 3,3 x 10 5 endospores/mL per 10 to 20 minutes or to shake 3,3 x 10 4 endospores/mL for 50 minutes. In another experiment, the reproduction of P. penetrans in pure population of M. incognita and in a mixed population of M. incognita and M. javanica, originated from the field, was studied. The endospore production of P. penetrans in tomato plants inoculated with M. incognita was approximately three times higher than in plants inoculated with the field population. An attachment test of P. penetrans on the two populations was performed and higher number of endospores attached to J2 of M. incognita was observed. The multiplication of P. penetrans in 15, 30, 45 or 60 day-old ‘Santa Clara’ tomato plants inoculated with 5,000, 15,000 or 25,000 J2, was also evaluated. The 30 and 45 days old plants inoculated with 25,000 J2 provided P. penetrans multiplication up to nineteen times more than plants inoculated with 5,000 J2 in the first experimental run. To determine if high cow manure levels added to substrate promote physiological changes on the nematodes or on the tomato plants, the influence of cow manure amendment to mixtures of soil and sand giving rates of 1:1:0; 2:2:1; 1:1:1 and 1:1:2 (V:V:V) of soil, sand and manure, respectively, and three inoculum levels of Meloidogyne species, i.e., 3,000; 6,000 and 9,000 J2 on the phenolic content in the tomato roots, changes in nematode lipid content and possible alterations in the giant cells induced by the nematodes, were studied. No conclusion could be drawn about the effect of manure on the nematode lipid content. The phenolic content in the roots increased as more cow manure was added to the substrate or when the plants were inoculated with more nematodes (9,000 J2). The giant cells in the roots of plants cultivated in the substrates 1:1:0 and 2:2:1 were more numerous, bigger and with more nuclei. On the other hand, the giant cells of plants cultivated on 1:1:1 and 1:1:2 substrates were less numerous, showed changes on their format and were smaller, demonstrating the deleterious effect of organic amendments to these feeding sites. A subsequent experiment was carried out to evaluate the effect of the interaction of increasing rates of cow manure in the substrates and of nematode levels on the reproduction of Pasteuria penetrans. Higher perceptual of infected females by P. penetrans was observed when the plants grew in the substrate 1:1:0 than in the substrates 1:1:1 and 1:1:2, or when plants were inoculated with 3,000 J2. The experiment was repeated once and in the first run, plants cultivated on substrate 1:1:0 or inoculated with 9,000 J2 had higher endospore number. However, in the second run, plants inoculated with 9,000 J2 and cultivated on substrate 2:2:1 yielded more endospores per root system. / Tese importada do Alexandria

Submitted by MARCOS LEANDRO TEIXEIRA DE OLIVEIRA (marcosteixeira@ufv.br) on 2018-09-18T13:33:07Z No. of bitstreams: 1 texto completo.pdf: 2023955 bytes, checksum: c00bada9d7cdc721128642c40a1fc2a1 (MD5) / Made available in DSpace on 2018-09-18T13:33:07Z (GMT). No. of bitstreams: 1 texto completo.pdf: 2023955 bytes, checksum: c00bada9d7cdc721128642c40a1fc2a1 (MD5) Previous issue date: 2017-03-31 / Na natureza, um fitopatógeno geralmente está sob influência de um complexo de microrganismos, que associados garantem o equilibrio ecológico estável. Com foco no fungo nematófago Pochonia chlamydosporia, começamos investigando sua compatibilidade com Bradyrhizobium japonicum em soja e a influência dessa interação sobre o controle do nematoide das galhas e sobre a absorção de nutrientes pela planta. Constatou-se que a aplicação conjunta desses dois organismos não prejudica a ação de controle de nematoides por P. chlamydosporia e nem a fixação de N 2 por B. japonicum. Adicionalmente, quando os dois agentes estavam juntos, ocorreu maior produção de nódulos da bactéria nas raízes e aumento do conteúdo de Fe na parte aérea das plantas de soja. No segundo capítulo, a compatibilidade dos agentes de controle biológico de nematoides, P. chlamydosporia e P. penetrans foi avaliada. A co-aplicação dos microrganismos possibilitou maior redução do número de ovos do nematoide do que a aplicação em separado e foi possível observar a associação do fungo nas raízes infectadas por nematoides colonizados por P. penetrans. Pesquisamos ainda, no terceiro capítulo, a presença de vírus em isolados de P. chlamydosporia. Dos dezoito isolados avaliados, apenas um (Pc-M4) estava infectado por dois virus, um com genoma de RNA fita dupla (dsRNA) e outro de RNA fita simples sentido negativo (-ssRNA). Os nomes propostos para os micovírus são Pochonia chlamydosporia chrysovirus 1 (PcCV1) e Pochonia chlamydosporia negative-stranded RNA virus 1 (PcNSRV1). Ensaios biológicos do isolado fúngico Pc-M4 revelaram que este foi capaz de parasitar ovos, produzir metabólitos e proteases letais aos juvenis e reduzir a reprodução do nematoide de galhas M. javanica. Os resultados apresentados indicam que o fungo P. chlamydosporia é capaz de interagir com diferentes organismos sem perder a capacidade de controlar o nematoide de galhas e de promover o crescimento vegetal, o que faz dele um excelente agente de biocontrole de nematoides. / In nature, a plant pathogen is usually under the influence of a complex of microorganisms, which guarantees stable ecological balance. Focusing on the nematophagous fungus Pochonia chlamydosporia, we started investigating its compatibility with Bradyrhizobium japonicum in soybean. We also looked at the influence of this interaction on the control of the root knot nematode and on the nutrient uptake by the plant. It was found that the joint application of these two organisms does not affect the action of control of nematodes by P. chlamydosporia nor the fixation of N 2 by B. japonicum. In addition, when the two agents were together, there was a higher production of nodules of the bacteria in the roots and an increase in the Fe content in the aerial part of the soybean plants. In the second chapter, the compatibility of biological control agents of nematodes, P. chlamydosporia and P. penetrans was evaluated. The co-application of the microorganisms allowed a greater reduction in the number of nematode eggs than the separate application. It was possible to observe the association of the fungus in the roots infected by nematodes colonized by P. penetrans. We also investigated the presence of viruses in isolates of P. chlamydosporia in the third chapter. Of the eighteen isolates evaluated, only one (Pc-M4) was infected by two viruses, one with double-stranded RNA genome (dsRNA) and the other with RNA single-stranded sense negative (-ssRNA). The proposed names for the mycoviruses are Pochonia chlamydosporia chrysovirus 1 (PcCV1) and Pochonia chlamydosporia negative-stranded RNA virus 1 (PcNSRV1). Biological assays of the fungal isolate Pc-M4 revealed that it was able to parasitize eggs, produce metabolites, lethal proteases to juveniles and reduce reproduction of M. javanica. The results indicated that the fungus P. chlamydosporia is capable of interacting with different organisms without losing the ability to control the root knot nematode and to promote plant growth. This makes it an excellent biocontrol agent for plant parasitic nematodes.

No description available.

Submitted by Marco Antônio de Ramos Chagas (mchagas@ufv.br) on 2017-04-27T13:11:40Z No. of bitstreams: 1 texto completo.pdf: 832503 bytes, checksum: c9ddb50df4ac046edfb1b18f02406400 (MD5) / Made available in DSpace on 2017-04-27T13:11:40Z (GMT). No. of bitstreams: 1 texto completo.pdf: 832503 bytes, checksum: c9ddb50df4ac046edfb1b18f02406400 (MD5) Previous issue date: 2004-03-23 / Fundação de Amparo à Pesquisa do Estado de Minas Gerais / Extratos enzimáticos de diferentes fungos foram avaliados quanto à maceração de raízes, a fim de se extrair mais fácil e rapidamente fêmeas de Meloidogyne spp. para estudos de controle biológico e para a identificação de espécies por eletroforese de isoenzimas. Raízes de tomateiro infectadas por Meloidogyne javanica foram imersas em extrato enzimático produzidas pelos fungos Aspergillus niger, Penicillium griseoroseum ou Penicillium italicum, separadamente ou em mistura de 50% de enzimas de cada fungo, em combinações de dois a dois, ou em tampão fosfato (testemunha), e incubadas a 30oC/24h. As enzimas produzidas pelo fungo P. griseoroseum foram as mais eficientes na degradação da parede das células das raízes de tomateiro, equiparando-se às enzimas comerciais importadas, pectinase + celulase (Clarex, Miles do Brasil - 15.000 ajdv/g; Sigma Chemical Ltda- 5.000 un.), utilizadas para a extração de fêmeas de Meloidogyne spp. Raízes com galhas parasitadas com as espécies M. incognita, M. javanica ou M. arenaria foram imersas em tampão de acetato de sódio apenas ou com enzimas liofilizadas de P. griseoroseum, em diferentes concentrações, e incubadas a 40°C/24h. A atividade de 200 U/mL proporcionou maior liberação de fêmeas, e essas foram submetidas à eletroforese de isoenzimas. Não foram observadas bandas, em gel de poliacrilamida, de fêmeas retiradas pela incubação em enzimas fúngicas, mas bandas foram vistas quando as fêmeas foram retiradas manualmente. Para avaliar o efeito das enzimas na adesão de endósporos aos juvenis de M. javanica, fêmeas foram retiradas manualmente e por maceração enzimática a 200 U/mL. Os endósporos de fêmeas extraídas por digestão enzimática apresentaram maior adesão a J2 de M. javanica em relação à testemunha. A reprodução dos endósporos de P. penetrans em fêmeas de M. javanica não foi afetada pela enzima. Para melhorar a quantificação de endósporos de P. penetrans em pó de raiz utilizado como inóculo para aplicação da bactéria no campo, as enzimas a 2% e 20% foram adicionadas a 2g do pó de raiz com P. penetrans e incubados a 40°C. O extrato enzimático na concentração de 20%, independente do tempo de incubação, possibilitou observar maior número de endósporos. O uso de enzimas fúngicas para a extração de fêmeas de Meloidogyne spp., demonstrou ser bastante eficiente e vantajoso, sem prejudicar a adesão e multiplicação de P. penetrans no nematóide; porém, não é recomendada para a identificação de M. javanica, M. incognita e M. arenaria por eletroforese de isoenzimas. / Enzymatic extracts from different fungi were evaluated for root maceration as an easier and faster way of extracting Meloidogyne spp. females from roots, to be used in biological control studies and in species identification by isozyme electrophoresis. Tomato roots infected by M. javanica were immersed in enzymatic extracts produced by Aspergillus niger, Penicillium griseoroseum or P. italicum, separately or in mixtures of 50% of enzymatic extracts of each fungus, two by two, or in phosphate buffer (control), and incubated at 30oC/24h. The enzymes produced by the fungus P. griseoroseum were the most efficient in the degradation of the tomato root cell walls, being comparable to the imported commercial enzymes pectinase + celulase (Clarex, Miles do Brasil - 15.000 ajdv/g; Sigma Chemical Ltda- 5.000 un.) used for Meloidogyne spp. females extraction. Galled roots parasitized by M. incognita, M. javanica or M. arenaria were immersed in sodium acetate buffer alone or with lyophilized enzymes of P. griseoroseum in different concentrations and incubated at 40°C/24h. The number of 200 U/mL of enzyme preparation provided the best female release, and the females attained using this concentration were subjected to isozyme electrophoresis analysis. No bands could be seen from the females extracted by incubation in fungal enzymes, in the poliacrilamide gel, but typic bands formed from females extracted manually. In order to evaluate the effect of enzymatic extraction on the attachment of endospores on second-stage juveniles (J2) of M. javanica, nematode females parasitized with Pasteuria penetrans were extracted from the roots manually or by maceration in enzyme extract at 200 U/mL. The endospores from females extracted by enzimatic digestion attached in higher number to the M. javanica J2 when compared to the control treatment. The endospore production in M. javanica females was not affected by the enzymes. To improve the endospore counting in root powder used as carrier for field applications, the enzymes at 2% and 20% were added to 2g of root powder containing P. penetrans endospores, and incubated at 40 °C. The enzyme extract at 20%, independent of the incubation time, allowed to observe a higher number of endospores. The use of fungal enzymes to extract Meloidogyne spp. showed to be very efficient and advantageous, without disturbing attachment or reproduction of P. penetrans in the nematode, however, it is not recommended for the identification of M. javanica, M. incognita nor M. arenaria by isozyme electrophoresis. / Dissertação importada do Alexandria

Au Canada, les pertes de rendement en agriculture attribuées aux nématodes sont généralement associées aux nématodes des lésions du genre Pratylenchus. En 2011, la découverte d’une nouvelle espèce exotique au Canada et qualifiée de rare dans le Nord-Est américain, soit Pratylenchus alleni Ferris, a soulevé de nouvelles inquiétudes. Afin de déterminer si cette espèce représente une menace pour les productions agricoles du Québec, mon projet de maîtrise visait à recueillir des informations sur sa virulence. Dans un premier temps, le spectre d’hôtes de P. alleni a été étudié et les résultats ont montré que ce nématode se développe très bien sur la pomme de terre, mais non sur la luzerne et le trèfle rouge. Ensuite, la reproduction de P. alleni dans un contexte de changements climatiques a été étudiée. L’augmentation prévue des températures et CO2 devrait favoriser le développement de P. alleni puisqu’il possède un meilleur taux de reproduction sur le soya lorsque soumis à un régime de températures de 17/28 ˚C et à une concentration en CO2 de 1200 ppm comparativement à 12/23 ˚C (400 ppm) et 15/26 ˚C (800 ppm). Dans cette même étude, une réduction de 19 à 58 % du poids sec racinaire des plants de soya inoculés avec P. alleni a été observée comparativement aux plants témoins. De plus, une méthode moléculaire de détection et de quantification simultanée de P. alleni et P. penetrans, l’espèce de Pratylenchus la plus répandu dans l’Est canadien, par qPCR a également été développée. Pour chacune des deux espèces, une sonde TaqMan associée avec le fluorophore CY5 pour P. alleni et FAM pour P. penetrans ciblant la région D2/D3 de la grande sous-unité ribosomale (28S) ont été développées et celles-ci se sont avérées spécifiques à chaque espèce. Ces résultats amènent de nouvelles connaissances sur ce ravageur et mettent en lumière sa pathogénicité. / In Canada, yield losses attributed to nematodes are generally associated with root-lesion nematodes from the genus Pratylenchus. In 2011, a new exotic species was detected in Canada and identified as Pratylenchus alleni Ferris. Pratylenchus alleni is rare in the Northeastern U.S. and its discovery has raised new concerns. To determine whether this species is a threat to agricultural production in Québec, my project aims to collect information about its pathogenicity. First, the host range of P. alleni was studied and the results showed that the nematode was developing well on potato, but poorly performed on alfalfa and red clover. The reproduction of P. alleni has also been studied in a context of climate change. The results showed that anticipated temperature and CO2 increases should favor P. alleni since it has a better reproduction rate on soybeans subjected to a night/day temperature regime of 17/28°C and a CO2 concentration of 1200 ppm compared to 12/23˚C (400 ppm) and 15/26°C (800 ppm) regimes. In the same study, a reduction of 19 to 58 % of roots dry weight of soybeans inoculated with P. alleni was observed compared to control plants. A simultaneous molecular detection and quantification method by qPCR of P. alleni and P. penetrans, the most widespread Pratylenchus species in Eastern Canada, was also developed. For each species, a TaqMan probe associated with the CY5 fluorophore for P. alleni and FAM for P. penetrans targeting the D2/D3 expansion segments of the large ribosomal subunit (28S) were developed and proved to be specific to each species. These results bring new insights into this new pest and highlight its pathogenicity.