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The ten most frequently isolated fungi from the wood of the dead branches of Acer pseudoplatanus L. were tested in dual cultures to evaluate their in vitro antagonistic activity against Eutypella parasitica R.W. Davidson and R.C. Lorenz, the causative agent of a destructive disease of maples in Europe and North America. The tested fungi, treated also as challenge isolates, were Diaporthe sp., Eutypa sp., Eu. maura, E. parasitica, Fusarium avenaceum, Neocucurbitaria acerina, Neonectria sp., Peniophora incarnata, Petrakia irregularis, and Phomopsis pustulata. The antagonistic ability of each challenge isolate was evaluated by calculating an index of antagonism (AI) based on the interaction type in the dual cultures. The results of competition between the fungal isolates were quantified after re-isolations from the interaction zone (s). The dual cultures revealed two main types of competitive interactions: Deadlock, consisting of mutual inhibition after mycelial contact or at a distance, and replacement, reflecting in the inhibition of E. parasitica, followed by partial overgrowth by the replacing fungus. Statistical analysis showed significant differences in average AI and s of challenge isolates between different dual culture assays. Based on the results of the antagonism index, Eutypa sp., Eu. maura, Neonectria sp., and P. incarnata had the highest inhibitory effect on E. parasitica growth and were recognized as the most promising candidates for further biocontrol studies of E. parasitica. The mycelium of E. parasitica at the interaction zones remained mostly viable, except in dual cultures with Eutypa sp., F. avenaceum, and Neonectria sp., where re-isolations did not yield any colony of the E. parasitica isolate. Based on the results, we assume that E. parasitica is a weak competitor, which invests less energy in direct mycelial competition. We discuss the potential of the observed antagonists as a possible biocontrol of Eutypella canker of maple. Nevertheless, additional experiments should be performed for a solid conclusion about competitive ability of E. parasitica and usefulness of antagonists as biocontrol.

Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2018-08-28T18:22:19Z No. of bitstreams: 1 2016 - Allan Felipe Marciano.pdf: 1368631 bytes, checksum: bba1066f388233008b1c3f41343d33e2 (MD5) Made available in DSpace on 2018-08-28T18:22:19Z (GMT). No. of bitstreams: 1 2016 - Allan Felipe Marciano.pdf: 1368631 bytes, checksum: bba1066f388233008b1c3f41343d33e2 (MD5) Previous issue date: 2016-10-03 CAPES - Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior CNPq - Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico The tick Rhipicephalus microplus has been considered the most important parasite in cattles. It causes economic lost in dairy and beef cattle. As the fungus Metarhizium spp, a promising biocontrol agent, with remarkable virulence and efficacy against this arthropod. Environmental conditions can directly influence entomopathogenic fungal activity when it is released on the field. Therefore, adjuvants should be used to enhance fungal pathogenicity, stability and viability under unfavorable environmental conditions. Accordingly, several oily vehicles have been studied as adjuvant components. The present study aimed at investigate under semi-natural conditions, the permanence of fungi in the soil, the pathogenicity and the efficacy of two isolates of Metarhizium anisopliae sensu latu (ARSEF 3641 and CG 148) formulated in oil (soy and mineral), on engorged females of R. microplus engorged. The fungus was mass produced on rice cultivation and the semi-natural environment was framed from pots with Brachiaria brizantha cv. Marandu. The experiment was compound of seven groups: Control OLV (treated with vegetable oil and 0,1% of tween 80), Control OLM (treated with mineral oil and 0,1% of Tween 80), Treated 3641 OLV (treated with isolated ARSEF 3641 + vegetable oil); Treated 3641 OLM (treated with ARSEF 3641 isolated + mineral oil); Treated CG 148 OLV (treated with CG148 isolated + vegetable oil); Treated CG 148 OLM (treated with CG 148 isolated + mineral oil). Each group contained ten pots. The soil of each pot was sprayed with 80 ml of formulation at 108 conidia/ml in accordance with the proposed group. Afterwards, 5 engorged females were put on each pot. All pots were daily inspected for evaluating the percent mortality of famales. The efficacy of formulations tested was obtained by comparison of the number of live larvae collected at the apex of forage leaves of the control groups with the number of larvae from the treated groups. The collection of larvae was held on the 7th, 14th and 21st after the hatch. To evaluate the presence of Metarhizium spp. in the soil, a methodology was established to quantify the colonies number found in samples collected from each pot before the spraying and at 3th, 7th, 21th and 60th day after treatment. For the standardization of the methodology set out in the quantification of colonies in the soil, data from three evaluators were confronted by Kaapa test, the statistical analysis of data were submitted to the distribution of normality by the Shapiro-Wilk test. Data were analyzed as nonparametric and we used the Kruskal-Wallis test for analysis of variance followed by the Student-Newman-Keuls to compare the mean. The tested oil fungal formulations showed daily efficacy ranging between 85,08 and 96,13% compared to their respective controls. The group trated with isolated ARSEF 3641 formulated both in mineral oil and vegetable oil showed the better efficacy when compared to the respective control groups. Also, during tick females? laying period, it was possible to observe Metarhizium spp. externalization on R. microplus engorged females cuticle in all groups that were exposed to fungal formulations, confirming the ability of these fungal isolates on develop all growth stages over this arthropod in semi-natural conditions. Both fungal isolates formulated in vegetable or mineral oil showed to be able to remain active in semi-natural environment as they were pathogenic and effective against engorged females, and able to reduce significantly future generations of the tick. O carrapato Rhipicephalus microplus ? considerado um dos principais agentes de parasitismo em bovinos por representar significativas perdas econ?micas na pecu?ria de leite e corte. Sendo o fungo Metarhizium spp, um promissor agente biocontrolador para este artr?pode, por apresentar grande virul?ncia e efic?cia contra este ectoparasita. Agentes microbianos quando lan?ados a campo, podem sofrer grande influ?ncia pelas intemp?ries ambientais, devendo estes receberem componentes que favore?am a sua patogenicidade, perman?ncia e viabilidade no ambiente, a fim de atingirem resultados satisfat?rios. Visto isto, diferentes ve?culos oleosos v?m sendo alvos de estudos como componentes adjuvantes. O objetivo deste trabalho foi investigar, sob condi??es seminaturais, a perman?ncia no solo, a virul?ncia e a efic?cia de dois isolados do fungo Metarhizium anisopliae s.l. (ARSEF 3641 e CG 148) formulados em ?leo (soja e mineral) sobre f?meas de R. microplus, assim como a taxa e recupera??o de larvas. O fungo foi produzido em massa a partir do cultivo dos isolados em arroz e para a forma??o do ambiente seminatural, sementes de Brachiaria brizantha, cultivar Marandu, foram plantadas em vasos de polipropileno. O experimento foi composto por sete grupos: Controle CTR (grupo sem tratamento); Controle OLV (tratado com ?leo vegetal e Tween 80 a 0,1%), Controle OLM (tratado com ?leo mineral e Tween 80 a 0,1%), Tratado 3641 OLV (tratado com o isolado ARSEF 3641 + ?leo vegetal + Tween 80 a 0,1%); Tratado 3641 OLM (tratado com o isolado f?ngico ARSEF 3641 + ?leo mineral + Tween 80 a 0,1%); Tratado CG 148 OLV (tratado com o isolado 148 + ?leo vegetal + Tween 80 a 0,1%); Tratado CG 148 OLM (tratado com o isolado CG 148 + ?leo mineral + Tween 80 a 0,1%). Cada grupo conteve dez vasos. Foram pulverizados sobre a superf?cie do solo de cada vaso, 80ml da formula??o a 108 con?dios/ml de acordo com o grupo proposto para cada formula??o e posteriormente 5 f?meas ingurgitadas de peso m?dio e homog?neo foram adicionadas em cada vaso. Os mesmos foram vistoriados diariamente para a avalia??o da mortalidade das f?meas. O percentual de efic?cia das formula??es dos isolados testados foi obtido a partir da compara??o entre o n?mero de larvas vivas coletadas no ?pice das folhas da forrageira dos grupos controles e dos grupos tratados. A coleta de larvas foi realizada no 7?, 14? e 21? ap?s a eclos?o das mesmas. Para a avalia??o da presen?a de Metarhizium spp. no solo, foi estabelecido e validada uma metodologia para a quantifica??o do n?mero de col?nias encontradas em amostras coletadas de cada vaso antes da pulveriza??o e no 3?, 7?, 21? e 60? dia ap?s o tratamento. A valida??o do m?todo se fez pelo confronto da avalia??o de tr?s observadores utilizando o teste Kaapa. Para a an?lise estat?stica dos demais dados, os mesmos foram submetidos ? distribui??o da normalidade pelo teste de Shapiro-Wilk sendo classificados como n?o param?tricos e foi utilizado o teste de Kruskal Wallis para an?lise de vari?ncia, seguido do Student-Newman-Keuls para compara??o entre as ordena??es m?dias. Os grupos tratados com as formula??es f?ngicas oleosas apresentaram efic?cia m?dia no controle de R. microplus variando entre 85,08 e 96,13% em rela??o aos seus respectivos grupos controles durante o per?odo de avalia??o do n?mero de larvas infestantes. O isolado ARSEF 3641 formulado tanto em ?leo vegetal quanto em ?leo mineral apresentou os melhores percentuais de efic?cia. Durante o per?odo de postura das f?meas, foi poss?vel encontrar a exterioriza??o de Metarhizium sp. nas mesmas em todos os grupos tratados com as formula??es f?ngicas, confirmando a capacidade dos isolados f?ngicos em se manterem e realizarem todas as fases de desenvolvimento sobre o artr?pode em condi??es seminaturais. Ambos os isolados testados, nas duas condi??es de formula??es propostas, mostraram-se aptos a permanecerem ativos no ambiente seminatural, virulentos e eficazes contra f?meas de R. microplus, sendo capazes de reduzir significativamente gera??es futuras do carrapato.

Submitted by Jorge Silva (jorgelmsilva@ufrrj.br) on 2019-07-04T19:49:03Z No. of bitstreams: 1 2015 - Caio Junior Balduino Coutinho Rodrigues.pdf: 2161917 bytes, checksum: a8e9dfc42ba79298765996c0d20bd488 (MD5) Made available in DSpace on 2019-07-04T19:49:03Z (GMT). No. of bitstreams: 1 2015 - Caio Junior Balduino Coutinho Rodrigues.pdf: 2161917 bytes, checksum: a8e9dfc42ba79298765996c0d20bd488 (MD5) Previous issue date: 2015-02-11 CNPq - Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico Beauveria bassiana sensu lato (s.l.) is a cosmopolitan fungus of great importance in agriculture. Good results are obtained when this microrganism is used as a biocontrol of Rhipicephalus microplus. In this context, studies involving the analysis of the development and fungal metabolism enable to select arthropodopatogenic isolates with better potencial. The aim of this study was to evaluate the morphology, virulence and enzymatic activities of ten B. bassiana s.l. isolates before (Micoteca group) and after reisolation (Reisolado group) through one passage (one cycle) on R. microplus. For morphology was analyzed: macromorphology, performed using the single point of inoculum; micromorphology, with cultivation between slide and cover slip and conidia production. The virulence before and after reisolation was evaluated by observing the biological parameters of R. microplus engorged females after immersion in B. bassiana s.l. aqueous suspensions (108 and 107 conidia mL-1). The proteolytic (semi-quantitative, total, Pr1 and Pr2) and lipolytic activities (?-nitrophenyl palmitate) were performed after 72 hours of fungal growth on minimal medium. The statistical analysis of the experiments was performed by comparing the data before and after reisolation for determining isolates with greater pathogenic potential and observing the effect of one passage to the potential host. Parametric data were evaluated by analysis of variance (ANOVA) followed by Tukey test and nonparametric data by Kruskal-Wallis test followed by Student-Newman-Keuls test (SNK) with 5% significance. When compared to each other before reisolation, morphological, virulence and enzymatic activities differences were observed among the isolates. CG 481 and CG 206 showed the highest control percentage (65.75% and 61.65%, respectively), although do not show the highest enzyme activity. After fungal reisolation, the process changed in small-scale the morphology and virulence. In enzymatic activity, most isolates increased the Pr2 and the lipase activities. Isolates as Bb 02 and CG 500 were those who suffered most influence after growth cycle. Bb 02 increased 1.9 times the control percentage (15.8% to 30.6%), incrementing the conidia production and making colonies that are more powdery. For this isolate was also evidenced an increase in all the enzyme activities (6.3 times to Pr1; 2.4 times to Pr2 and 9 times to lipase activity). Differently, CG 500 reduced 4.8 times the control percentage and changed the macromorphology. The enzymatic activities were also reduced by 3.6 times to Pr1; 1.2 times to Pr2 and 1.6 times to lipase activity. Therefore, the study concluded that one B. bassiana s.l. passage on R. microplus was able to change some of the isolates, although more passes by the host are necessary to provide relevant results with other isolates. Moreover, the study of enzymes such as chitinases, production of toxins and hydrophobicity of conidia should be considered to determine the fungal effectiveness. Beauveria bassiana sensu lato (s.l.) ? um fungo cosmopolita de grande import?ncia agr?cola. Seu uso na pecu?ria como biocontrolador de Rhipicephalus microplus apresenta bons resultados. Dentro deste contexto, estudos que associem a an?lise do desenvolvimento e do metabolismo f?ngico permitem selecionar isolados de melhor potencial artropodopag?nico. O presente estudo avaliou a morfologia, virul?ncia e atividades enzim?ticas de dez isolados brasileiros de B. bassiana s.l. antes (grupo Micoteca) e ap?s reisolamento (grupo Reisolado) atrav?s de um ciclo de crescimento sobre R. microplus. Para a morfologia, foram realizadas an?lises macromorfol?gicas, atrav?s de ponto ?nico de in?culo; micromorfol?gicas, a partir do cultivo entre l?mina e lam?nula, e avalia??o da produ??o de con?dios. A virul?ncia foi avaliada a partir da observa??o dos par?metros biol?gicos de f?meas ingurgitadas de R. microplus ap?s imers?o em suspens?es aquosas de B. bassiana s.l. (108 e 107 con?dios/mL). As atividades proteol?tica (semi-quantitativa, total, Pr1 e Pr2) e lipol?tica (?-nitrofenil palmitato) foram realizadas ap?s 72 horas de crescimento dos fungos em meio m?nimo. A estat?stica dos experimentos foi realizada comparando os dados antes e ap?s reisolamento para determinar os isolados de maior potencial patog?nico, al?m de observar o efeito de uma passagem sobre um hospedeiro em potencial. Os dados param?tricos foram avaliados pela an?lise de vari?ncia (ANOVA) seguida do teste de Tukey e os n?o param?tricos pelo teste de Kruskal Wallis seguido de Student-Newman-Keuls (SNK) com n?vel de signific?ncia de 5%. Quando comparados entre si antes do reisolamento, foram observadas diferen?as morfol?gicas, de virul?ncia e atividades enzim?ticas entre os isolados. CG 481 e CG 206 apresentaram os maiores percentuais de controle (65,75% e 61,65%, respectivamente), embora n?o tenham demonstrado alta atividade enzim?tica. Quanto ao reisolamento f?ngico, o processo alterou em pequena escala a morfologia e a virul?ncia. Em rela??o ? atividade enzim?tica, a maioria dos isolados elevaram a atividade proteol?tica espec?fica Pr2 e a atividade lipol?tica. Isolados como Bb 02 e CG 500 foram aqueles que mais sofreram influ?ncia ap?s um ciclo de crescimento. Bb 02 aumentou em 1,9 vezes o percentual de controle (15,8% para 30,6%), elevando a produ??o de con?dios e tornando as col?nias mais pulverulentas. Al?m do mais, foi evidenciado tamb?m o aumento de todas as atividades enzim?ticas (6,3 vezes para Pr1; 2,4 vezes para Pr2 e 9 vezes para atividade lip?sica). Em contrapartida, CG 500 reduziu em 4,8 vezes o percentual de controle, al?m de alterar macromorfologicamente as col?nias. As atividades enzim?ticas tamb?m foram reduzidas em 3,6 vezes para Pr1; 1,2 vezes para Pr2 e 1,6 vezes para a atividade lip?sica. Portanto, conclui-se que um ciclo de crescimento de B. bassiana s.l. sobre R. microplus foi capaz de alterar alguns dos isolados avaliados, embora mais passagens pelo hospedeiro sejam necess?rias para propiciar resultados mais relevantes. Al?m disto, o estudo de outras enzimas como as quitinases, a produ??o de toxinas e a hidrofobicidade dos con?dios devem ser considerados na determina??o da efic?cia f?ngica.

Salmonella Gallinarum is the agent of fowl typhoid in poultry and infects mainly adult galliforms, causing significant economic losses in poultry production. Because quails are susceptible to this disease and quail production is becoming increasingly important in Brazil, this study was carried out to evaluate the virulence of Salmonella Gallinarum strain to quails. The inoculum was prepared from S. Gallinarum strain resistant to nalidixic acid. Forty eight 16-week-old Japanese quails were randomly distributed in three groups. Before the experiment, cloacal swabs were collected from all birds in order to confirm they were free from Salmonella spp. Cloacal swabs and fecal samples were collected on days 03, 06, 09, 12, and 15 post-inoculation. Birds that died during the experiment were submitted to post-mortem examination, and had their organs aseptically collected for bacteriological examination. All eggs produced during the experiment were also examined. The mortality rate recorded during the experiment was 43.75% (21/48). S. Gallinarum was recovered from the organs of the birds that naturally died during the experiment, but the agent was not isolated from the organs of sacrificed birds. No egg sample was positive for Salmonella Gallinarum. It was concluded that S. Gallinarum may be recovered from the organs of experimentally-infected Japanese quails.

In Europe, the speciesLactobacillus rhamnosusis currently on the Qualified Presumption of Safety list used by the European Food Safety Authority (EFSA) for internal safety assessment, but according to the EFSA the species should remain a topic of surveillance. In the present study, the safety and tolerance of the potentially probiotic strainL. rhamnosusPRSF-L477 was investigated in a placebo-controlled double-blind volunteer trial following FAO/WHO guidelines. A total of thirty-four subjects received daily doses of 1 × 1011colony-forming units (cfu) ofL. rhamnosusPRSF-L477 (n17) or placebo (n17) for a period of 3 weeks, followed by a wash-out period of another 3 weeks. A questionnaire on gastrointestinal tolerance and a diary was kept daily to record compliance throughout these 6 weeks. Faecal and blood samples were collected for microbiological and haematological analysis. The recorded gastrointestinal symptoms, defecation frequency and stool consistency were not influenced indicating thatL. rhamnosusPRSF-L477 was well tolerated. The speciesL. rhamnosuswas detected in the faeces of sixteen out of seventeen subjects of the probiotic group during the intervention period. Using pulsed-field gel electrophoresis, re-isolates ofL. rhamnosusPRSF-L477 were confirmed in nine of these subjects. Antibiotic susceptibility profiles of these re-isolates were unchanged compared with PRSF-L477. No clinically relevant changes in blood parameters such as liver and kidney function and no serious adverse events appeared during and after administration. Therefore, we conclude thatL. rhamnosusPRSF-L477 can safely be administrated to healthy subjects at a daily dose of 1 × 1011 cfu.

AIMS: To define predictors of long-term outcome of a first repeat ablation solely consisting of re-isolation of reconnected pulmonary veins (PVs). METHODS: Three hundred seven patients (age 59 ± 9%, 77% males, non-paroxysmal AF 43%) with recurrent AF after first PVI were studied. Re-isolation of reconnected PVs was guided by a circular mapping catheter and 3D mapping system using RF ablations. A PV was defined as “triggering” in case of spontaneous ectopy or AF paroxysms originating from the PV. RESULTS: After a mean follow-up of 5.05 ± 2.21 years, 194 (63.2%) patients (73.0% in PAF vs 50.4% in non-PAF, log Rank