Orientador: Lourenço Bozzo / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-07-16T05:19:31Z (GMT). No. of bitstreams: 1 Rocha_Walter_M.pdf: 2253124 bytes, checksum: 660dd639d36a903232fd35d7a137f0ea (MD5) Previous issue date: 1979 / Resumo: Não informado / Abstract: Not informed. / Mestrado / Farmacologia / Mestre em Ciências

Equine Protozoal Myeloencephalitis (EPM) is arguably the most common and costly equine neurologic diseases nationwide. The national seroprevalence is >50%, but only 0.5-1% of all horses develops disease during their lifetimes. Some EPM affected horses have decreased immune response. A cell-mediated immune response has been shown to be protective for development of EPM after infection with Sarcocystis neurona in mouse models. Levamisole has been proposed as an adjunctive therapy for EPM to upregulate the cell-mediated immune response based on positive results in other species, but there are very limited studies in equids. We hypothesized that levamisole will upregulate the equine cell-mediated macrophage (M1) dendritic cell (DC1) CD4 T-helper 1 (Th1) CD8 Tc1 immune response in vitro. The first aim was to determine optimal conditions and effects of levamisole on cellular proliferation. Equine PBMCs were harvested from ten horses seronegative for S. neurona. The cells were cultured alone, or with one of the mitogens: concanavalin A (ConA) or phorbol 12-myristate 13-acetate and ionomycin (PMA/I), or with a combination of the above mitogens and levamisole at several conditions. Cellular proliferation was assessed using a colorimetric bromodeoxyuridine ELISA assay. The second aim was to determine the ability of levamisole, under optimized conditions, to upregulate the M1 DC1 CD4Th1 CD8 Tc1 response in vitro based on activation and function. PBMCs from the same 10 horses were cultured with each of the following: no stimulation, conA, and levamisole with and without ConA. To determine proliferation of each specific subset, cells were labeled with a fluorescent dye, CellTrace. Proliferation was determined based on dye dilution using flow cytometry. To determine the effects of levamisole on the specific immune response, cell subsets were labeled with fluorescent antibodies for cell surface markers (CD4, CD8, CD21, CD172a, CD14) and dendritic and macrophage activations markers (MHC Class II, CD86). Induction of T-regs was based on FoxP3 expression. Immune phenotypes were determined based on intracellular cytokine expression (IFNɣ, IL4, IL10). Study results indicate that levamisole alone did not significantly alter PBMC proliferation compared to the response of unstimulated cells. Cells cultured with either ConA or PMA/I resulted in a statistically significant increase (P<0.05) in proliferation compared to unstimulated cells. Cells cultured with ConA and levamisole at 1µg/mL resulted in a significant decrease (P<0.05) in proliferation compared with cells cultured with ConA alone. Flow cytometry data failed to elucidate the specific immune phenotype that is affected by levamisole. Subjectively, there appeared to be a trend for inceased IFNɣ production by CD14 and CD172a positive cells (macrophages and dendritic cells) and a decrease in IFNɣ production by CD4 and CD8 positive cells (T-lymphocytes). These results demonstrate that levamisole downregulates ConA stimulated PBMC proliferation. Based on these in vitro results, further studies to determine the effectiveness of levamisole on modulating the equine immune system in vivo and to more specifically evaluate the immune cell subets affected by levamisole are warranted. / Master of Science

Orientador: Flávio Ruas de Moraes / Banca: Adjair Antonio do Nascimento / Banca: Gilson Pereira de Oliveira / Banca: Laura Satiko Okada Nakaghi / Banca: Oelinton Ferreira Barbosa / Resumo: Avaliou-se no presente trabalho, a eficácia anti-parasitária do praziquantel, levamisol e diflubenzuron administrados via oral, adicionados à ração e administrados a pacus (Piaractus mesopotamicus) infectados por Anacanthorus penilabiatus e Dolops carvalhoi. Foram utilizadas 19 caixas d’água de 300 litros de capacidade, comportando 28 peixes em cada uma. O alimento dos peixes foi feito misturando as drogas na ração. O experimento foi conduzido em quatro colheitas realizadas, um dia antes e três, sete e 15 dias após a aplicação dos medicamentos. A alimentação dos peixes com ração contendo diflubenzuron, levamisol e praziquantel isolado ou associados em diferentes concentrações foi feita durante sete dias. Os resultados da eficácia terapêutica sugerem que simples ou associado com levamisol e praziquantel, o diflubenzuron é eficiente contra o crustáceo D. carvalhoi, demonstrando que a eficácia dos tratamentos nos dias três, sete e 15 foi de 96,2 a 100%. Contra os monogenóides as drogas não apresentaram eficácia satisfatória. Os resultados sugerem o uso do diflubenzuron para o controle de D. cavalhoi em peixes de cativeiro nas condições deste ensaio. / Abstract: This assay evaluated the control efficacy of diflubenzuron, praziquantel and levamisole added to the diet of pacu (Piaractus mesoptamicus) infected with Anacanthorus penilabiatus and Dolops carvalhoi. Nineteen water tanks of 300 liters capacity were utilized with 28 fish in each one. The treatments were made by mixing the active principles in the diet. The experiment was evaluated in four harvests done one day before and three, seven and 15 days after the treatment. The medicated feeding was applied for seven days. The results of efficacy suggest that the diflubenzuron alone or associated with levamisole and praziquantel was efficient against the crustacean D. carvalhoi and the efficacy in the three, seven and 15 days evaluations ranged from 96,2 to 100%. Against the monogenean the drugs did not present efficacy. The results suggest the use of diflubenzuron for the control of D. carvalhoi in captive fishes in the conditions of this trial. / Doutor

Nematodes of the genus Ascaris are large gastrointestinal parasites. Ascaris lumbricoides infects ~1 billion people globally; causing malnutrition and general morbidity, and can block the gut or bile duct causing fatal complications. Ascaris suum is a parasite of pigs; in addition to its veterinary significance, it can occasionally be zoonotic, and is a good model of the human parasite. One of the main classes of drugs used to treat parasitic nematode infections are the cholinergic anthelmintics, such as levamisole and pyrantel, which act as agonists of nicotinic acetylcholine receptors at the nematode neuromuscular junction.

Made available in DSpace on 2014-06-11T19:30:31Z (GMT). No. of bitstreams: 0 Previous issue date: 2006-10-30Bitstream added on 2014-06-13T19:19:26Z : No. of bitstreams: 1 schalch_shc_dr_jabo.pdf: 1229379 bytes, checksum: b8e2410e3b10eb15e380a61c57c898cc (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Avaliou-se no presente trabalho, a eficácia anti-parasitária do praziquantel, levamisol e diflubenzuron administrados via oral, adicionados à ração e administrados a pacus (Piaractus mesopotamicus) infectados por Anacanthorus penilabiatus e Dolops carvalhoi. Foram utilizadas 19 caixas d água de 300 litros de capacidade, comportando 28 peixes em cada uma. O alimento dos peixes foi feito misturando as drogas na ração. O experimento foi conduzido em quatro colheitas realizadas, um dia antes e três, sete e 15 dias após a aplicação dos medicamentos. A alimentação dos peixes com ração contendo diflubenzuron, levamisol e praziquantel isolado ou associados em diferentes concentrações foi feita durante sete dias. Os resultados da eficácia terapêutica sugerem que simples ou associado com levamisol e praziquantel, o diflubenzuron é eficiente contra o crustáceo D. carvalhoi, demonstrando que a eficácia dos tratamentos nos dias três, sete e 15 foi de 96,2 a 100%. Contra os monogenóides as drogas não apresentaram eficácia satisfatória. Os resultados sugerem o uso do diflubenzuron para o controle de D. cavalhoi em peixes de cativeiro nas condições deste ensaio. / This assay evaluated the control efficacy of diflubenzuron, praziquantel and levamisole added to the diet of pacu (Piaractus mesoptamicus) infected with Anacanthorus penilabiatus and Dolops carvalhoi. Nineteen water tanks of 300 liters capacity were utilized with 28 fish in each one. The treatments were made by mixing the active principles in the diet. The experiment was evaluated in four harvests done one day before and three, seven and 15 days after the treatment. The medicated feeding was applied for seven days. The results of efficacy suggest that the diflubenzuron alone or associated with levamisole and praziquantel was efficient against the crustacean D. carvalhoi and the efficacy in the three, seven and 15 days evaluations ranged from 96,2 to 100%. Against the monogenean the drugs did not present efficacy. The results suggest the use of diflubenzuron for the control of D. carvalhoi in captive fishes in the conditions of this trial.

Actuellement, les infections des humains, des plantes et des animaux par les nématodes parasites ont un impact économique majeur. En l’absence de vaccin efficace, les anthelminthiques sont les principaux agents chimiothérapeutiques utilisés pour le traitement et la prophylaxie des infections à nématodes. Cependant, la résistance est apparue pour la plupart des anthelminthiques. Par conséquent il est urgent de comprendre la génétique des récepteurs ciblés par ces anthelminthiques et de trouver des cibles alternatives afin de développer de nouveaux anthelminthiques. Nous avons démontré les effets du nouvel anthelminthique cyclooctadepsipeptide, emodepside, sur le potentiel de membrane et les courants voltagedépendants chez le parasite du porc Ascaris suum. Enfin, nous avons cloné quatre gènes codant des sous-unités du récepteur de l'acétylcholine d'un autre parasite du porc, Oesophagostomum dentatum et caractérisé dans des ovocytes de Xenopus laevis quatre sous-types de récepteurs au lévamisole. / Parasitic nematode infections of humans, plants and animals are of major economic impact. Anthelmintics are the main chemotherapeutic agents used for treatment and prophylaxis of nematode infections because there is presently no effective vaccine on the market. However, resistance has been reported to the mainstay anthelmintics. There is therefore the urgent need to understand the genetics of the receptors targeted by these anthelmintics and to find alternative targets for developing new anthelmintics. We have demonstrated the effects of the new novelacting cyclooctadepsipeptide anthelmintic, emodepside, on the membrane potential and voltageactivated currents in the pig parasite Ascaris suum. Finally, we show the cloning of four acetylcholine receptor subunit genes from another pig parasite, Oesophagostomum dentatum and the expression and characterization of four levamisole receptor subtypes in Xenopus laevis oocytes.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / O monitoramento da sensibilidade antifÃngica em espÃcies de Candida de origem veterinÃria à uma prÃtica recente e os mecanismos envolvidos ainda nÃo foram completamente elucidados. Considerando que o arsenal de drogas antifÃngicas à limitado e que o fenÃmeno de resistÃncia tem se tornado mais freqÃente, a compreensÃo deste fenÃmeno e a busca por alternativas terapÃuticas se fazem necessÃrias. Dessa forma, o presente trabalho teve como objetivo monitorar a sensibilidade in vitro de Candida spp. oriundas de animais, com Ãnfase na resistÃncia aos azÃlicos mediada por bombas de efluxo e na avaliaÃÃo da atividade do imidazÃlico levamisol sobre o crescimento destas leveduras. Para tanto, em um primeiro momento, foram avaliados 126 isolados de Candida, sendo 19 C. albicans, 17 C. famata, 5 C. guilliermondii, 8 C. krusei, 29 C. parapsilosis, 48 C. tropicalis, dos quais 22 foram isolados de rapinantes, 32 de periquitos do sertÃo, 56 de papagaios, 7 de araras canindà e 3 de um ouriÃo-cacheiro. Todas as cepas foram submetidas ao teste de microdiluiÃÃo em caldo, ante a anfotericina B, itraconazol, fluconazol, segundo metodologia padronizada pelo Clinical Laboratory Standards Institute (documento M27-A3). As concentraÃÃes inibitÃrias mÃnimas (CIMs) variaram de 0,03125 a 2 Âg/mL, 0,125 a 250 Âg/mL e de 0,03125 a 125 Âg/mL para anfotericina B, fluconazol e itraconazol, respectivamente. Dos 126 isolados avaliados por microdiluiÃÃo, 33 (26,2%) foram resistentes aos azÃlicos, sendo 7 (5,6%) resistentes somente a fluconazol, 1 (0.8%) resistente somente ao itraconazol e 24 (19%) resistentes a ambas as drogas. Em um segundo momento, todas estas cepas resistentes aos derivados azÃlicos, mais 20 C. albicans e 3 C. tropicalis resgatadas da nossa coleÃÃo de leveduras resistentes de origem veterinÃria, foram submetidas ao teste de inibiÃÃo de bomba de efluxo com prometazina, perfazendo um total de 56 cepas resistentes a azÃlicos. Dessa forma, as CIMs de fluconazol e itraconazol sofreram reduÃÃes significativas de 2 a 250 e de 16 a 4000 vezes, respectivamente. Investigou-se, ainda, a atividade antifÃngica do levamisol contra 12 C. albicans, 12 C. krusei, 12 C. parapsilosis e 12 C. tropicalis, sendo obtidas CIMs e concentraÃÃes fungicidas mÃnimas que variaram de 0,58 a 2,34 mg/mL e de 2,34 a 9,37 mg/mL, respectivamente. Paralelamente, foi demonstrado que o levamisol inibe a formaÃÃo do biofilme de C. albicans e C. tropicalis, bem como interfere na manutenÃÃo do biofilme maduro. Os dados apontam que a resistÃncia aos derivados azÃlicos Ã, pelo menos em parte, mediada por bombas de efluxo, bem como demonstram o potencial antifÃngico do imidazÃlico levamisol e sua capacidade de inibir o biofilme de cepas de Candida spp. oriundas de animais. / Monitoring of the antifungal susceptibility of Candida species from veterinary sources is a recent practice and the mechanisms involved in antifungal resistance have not been completely elucidated. Considering that the antifungal arsenal is limited and that antifungal resistance has become more frequent, the comprehension of this phenomenon and the pursuit for therapeutic alternatives are necessary. Thus, the present work aimed at monitoring the in vitro susceptibility of Candida spp. isolated from animals, with emphasis on efflux pump-mediated azole resistance and on the evaluation of the effect of the imidazole levamisole on the growth of these yeasts. For such, in a first approach, 126 Candida isolates (19 C. albicans, 17 C. famata, 5 C. guilliermondii, 8 C. krusei, 29 C. parapsilosis, 48 C. tropicalis), out of which 22 were recovered from raptors, 32 from cactus parakeets, 56 from Amazon parrots, 7 from blue-and-gold macaws and 3 from a Brazilian porcupine. All isolates were submitted to broth microdilution test against amphotericin B, itraconazole and fluconazole, according to the methodology recommended by the Clinical Laboratory Standards Institute (document M27-A3). The MICs ranged from 0.03125 to 2 Âg/mL, 0.125 to 250 Âg/mL and 0.03125 to 125 Âg/mL for amphotericin B, fluconazole and itraconazole, respectively. Out of 126 evaluated isolates, 33 (26.2%) were resistant to azoles, with 7 (5.6%) isolates resistant to fluconazole, 1 (0.8%) isolate resistant to itraconazole and 24 (19%) resistant to both drugs. In a second approach, all these azole resistant isolates, plus 20 C. albicans and 3 C. tropicalis that were recovered from our collection of resistant yeasts from veterinary sources, were submitted to the efflux pump inhibition assay with promethazine, with a total of 56 azole resistant isolates. Thus, MICs for fluconazole and itraconazole significantly reduced from 2 to 250 times for fluconazole and from 16 to 4000 times for itraconazole. The antifungal activity of levamisole against 12 C. albicans, 12 C. krusei, 12 C. parapsilosis and 12 C. tropicalis, was also evaluated, and MICs and minimum fungicidal concentrations varying from 0.58 to 2.34 mg/mL and from 2.34 to 9.37 mg/mL were obtained, respectively. Parallelly, it was demonstrated that levamisole significantly inhibits biofilm formation and interferes with the maintenance of mature biofilms. These data show that azole resistance is partially mediated by efflux-pumps and demonstrate the antifungal potential of the imidazole levamisole and its capacity of inhibiting the biofilm of strains of Candida spp. from animals.

Made available in DSpace on 2016-01-26T18:55:27Z (GMT). No. of bitstreams: 1 ARTIGO FINAL_02_02_09.pdf: 347998 bytes, checksum: 56e6a67c83644f58e3352638f56c4d37 (MD5) Previous issue date: 2009-02-02 / The aim of this study was to evaluate the effect of levamisole on the antirabies humoral immune response in bovines primovaccinated against rabies. Forty-two bovines, about twelve months old, were randomly divided into three groups. Animals from VL group received one dose of levamisole and antirabies vaccine on day zero. Cattle from group V7L received one dose of antirabies vaccine on day zero and, on day seven, one dose of levamisole. Bovines from control group (GC) were immunized with just one dose of antirabies vaccine on day zero. A lyophilized, inactivated rabies vaccine, developed at Instituto Butantan was used. Blood samples were taken on days zero, 30 and 60. The neutralizing antibody titers were determined by serum neutralization in BHK21 cells, based on the Rapid Fluorescent Focus Inhibition Test (RFFIT) and on the Fluorescent Inhibition Microtest (FMIT). The results obtained showed that there was not significant difference in neutralizing-antibody titers and neither in the frequency of immunized animals between groups treated and not treated with levamisole 30 and 60 days after antirabies vaccination. In conclusion, levamisole administration (6.0 mg/kg of body weight) in primovaccinated bovines against rabies did not influence the antirabies humoral immune response. / O objetivo desse estudo foi avaliar o efeito do levamisol na resposta imune humoral anti-rábica de bovinos primovacinados contra a raiva. Quarenta e dois bovinos, com idade média de 12 meses, foram divididos aleatoriamente em três grupos. Os animais do grupo VL receberam uma dose de levamisol e vacina anti-rábica no dia zero. Nos bovinos do grupo V7L foi aplicada uma dose de vacina no dia zero e, no dia 7, uma dose de levamisol. Nos bovinos do grupo controle (GC), somente uma dose de vacina no dia zero. Utilizou-se uma vacina anti-rábica inativada, liofilizada, desenvolvida pelo Instituto Butantan. Colheram-se amostras de sangue nos dias zero, 30 e 60. Os títulos de anticorpos neutralizantes anti-rábicos foram determinados por meio da técnica de soroneutralização em células BHK21, baseado no Rapid Fluorescent Focus Inhibition Test (RFFIT) e no Fluorescent Inhibition Microtest (FIMT). Os resultados obtidos mostraram que não houve diferença significativa nos títulos de anticorpos anti-rábicos e nem na freqüência de animais imunizados entre os grupos tratados ou não com levamisol 30 e 60 dias após a vacinação anti-rábica. Conclui-se que a administração de levamisol (6,0 mg/Kg de peso vivo) em bovinos primovacinados contra a raiva, não influenciou a resposta imune humoral anti-rábica.

Made available in DSpace on 2016-07-18T17:53:05Z (GMT). No. of bitstreams: 1 ARTIGO FINAL_02_02_09.pdf: 347998 bytes, checksum: 56e6a67c83644f58e3352638f56c4d37 (MD5) Previous issue date: 2009-02-02 / The aim of this study was to evaluate the effect of levamisole on the antirabies humoral immune response in bovines primovaccinated against rabies. Forty-two bovines, about twelve months old, were randomly divided into three groups. Animals from VL group received one dose of levamisole and antirabies vaccine on day zero. Cattle from group V7L received one dose of antirabies vaccine on day zero and, on day seven, one dose of levamisole. Bovines from control group (GC) were immunized with just one dose of antirabies vaccine on day zero. A lyophilized, inactivated rabies vaccine, developed at Instituto Butantan was used. Blood samples were taken on days zero, 30 and 60. The neutralizing antibody titers were determined by serum neutralization in BHK21 cells, based on the Rapid Fluorescent Focus Inhibition Test (RFFIT) and on the Fluorescent Inhibition Microtest (FMIT). The results obtained showed that there was not significant difference in neutralizing-antibody titers and neither in the frequency of immunized animals between groups treated and not treated with levamisole 30 and 60 days after antirabies vaccination. In conclusion, levamisole administration (6.0 mg/kg of body weight) in primovaccinated bovines against rabies did not influence the antirabies humoral immune response. / O objetivo desse estudo foi avaliar o efeito do levamisol na resposta imune humoral anti-rábica de bovinos primovacinados contra a raiva. Quarenta e dois bovinos, com idade média de 12 meses, foram divididos aleatoriamente em três grupos. Os animais do grupo VL receberam uma dose de levamisol e vacina anti-rábica no dia zero. Nos bovinos do grupo V7L foi aplicada uma dose de vacina no dia zero e, no dia 7, uma dose de levamisol. Nos bovinos do grupo controle (GC), somente uma dose de vacina no dia zero. Utilizou-se uma vacina anti-rábica inativada, liofilizada, desenvolvida pelo Instituto Butantan. Colheram-se amostras de sangue nos dias zero, 30 e 60. Os títulos de anticorpos neutralizantes anti-rábicos foram determinados por meio da técnica de soroneutralização em células BHK21, baseado no Rapid Fluorescent Focus Inhibition Test (RFFIT) e no Fluorescent Inhibition Microtest (FIMT). Os resultados obtidos mostraram que não houve diferença significativa nos títulos de anticorpos anti-rábicos e nem na freqüência de animais imunizados entre os grupos tratados ou não com levamisol 30 e 60 dias após a vacinação anti-rábica. Conclui-se que a administração de levamisol (6,0 mg/Kg de peso vivo) em bovinos primovacinados contra a raiva, não influenciou a resposta imune humoral anti-rábica.