Genotyping of South American clinical isolates of Pythium insidiosum based on single nucleotide polymorphism-based multiplex PCR / Genotipagem de isolados clínicos de Pythium insidiosum da América do Sul utilizando polimorfismos de nucleotídeo único baseado em PCR multiplex

ABSTRACT: We aimed to genotype the South American clinical isolates of Pythium insidiosum using the single nucleotide polymorphisms (SNP) of the ribosomal DNA sequences (rDNA). Previously, an SNP-based multiplex-PCR was able to distinguish three different clades of P. insidiosum isolates. Thus, we used this assay to evaluate South American clinical isolates of P. insidiosum (n=32), standard strains from Costa Rica (n=4), Thailand (n=3), Japan (n=1), and India (n=1), a standard strain of Pythium aphanidermatum, and Brazilian environmental isolates of Pythium torulosum, Pythium rhizo-oryzae and Pythium pachycaule voucher (n=3). It was possible to allocate each American P. insidiosum isolate to clade I, the isolates of India, Japan, and Thailand to clade II, and the Thai isolate to clade III. P. aphanidermatum, P.torulosum, P.rhizo-oryzae and P.pachycaule voucher isolates were not amplified. For the first time, a P. insidiosum isolate from Uruguay, South America, was included in molecular analyzes. By SNP-based multiplex-PCR, it was possible to perform the identification and genotyping of the South American isolates of P. insidiosum, demonstrating similar genetic characteristics of these isolates.

RESUMO: O objetivo deste estudo foi genotipar isolados clínicos de Pythium insidiosum da América do Sul utilizando polimorfismos de nucleotídeo único (SNP) de sequências de rDNA. Anteriormente, um multiplex-PCR baseado em SNP foi capaz de distinguir P. insidiosum em três diferentes clados. Dessa forma, utilizamos este método para avaliar isolados clínicos de P. insidiosum da América do Sul (n=32), cepas padrão da Costa Rica (n=4), Tailândia (n=3), Japão (n=1) e Índia (n=1), uma cepa padrão de Pythium aphanidermatum e isolados ambientais brasileiros de Pythium torulosum; Pythium rhizo-oryzae e Pythium pachycaule voucher (n=3). Os isolados analisados foram alocados aos clados: I (americanos), II (isolados da Índia, Japão e Tailândia), e III (um isolado tailandês). P. aphanidermatum, P.torulosum, P.rhizo-oryzae e P.pachycaule voucher não foram amplificados. Pela primeira vez, um isolado de P. insidiosum do Uruguai foi incluído em análises moleculares. Através da multiplex-PCR baseada em SNP, foi possível realizar a identificação e genotipagem dos isolados sul-americanos de P. insidiosum, demonstrando características genéticas semelhantes entre esses isolados.

In vitro antifungal susceptibility of clinical and environmental isolates of Aspergillus fumigatus and Aspergillus flavus in Brazil

ABSTRACT The in vitro susceptibility of 105 clinical and environmental strains of Aspergillus fumigatus and Aspergillus flavus to antifungal drugs, such as amphotericin B, azoles, and echinocandins was evaluated by the broth microdilution method proposed by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Following the EUCAST-proposed breakpoints, 20% and 25% of the clinical and environmental isolates of A. fumigatus, respectively, were found to be resistant to itraconazole (Minimal Inhibitory Concentration, MIC > 2.0 mg/L). Voriconazole showed good activity against A. fumigatus and A. flavus strains, except for one clinical strain of A. fumigatus whose MIC was 4.0 mg/L. Posaconazole (≤0.25 mg/L) also showed appreciable activity against both species of Aspergillus, except for six A. fumigatus strains with relatively higher MICs (0.5 mg/L). The MICs for Amphotericin B ranged from 0.06 to 1.0 mg/L for A. fumigatus, but were much higher (0.5-8.0 mg/L) for A. flavus. Among the echinocandins, caspofungin showed a geometric mean of 0.078 and 0.113 against the clinical and environmental strains of A. flavus, respectively, but had elevated minimal effective concentrations (MECs) for seven of the A. fumigatus strains. Anidulafungin and micafungin exhibited considerable activity against both A. fumigatus and A. flavus isolates, except for one environmental isolate of A. fumigatus that showed an MEC of 1 mg/L to micafungin. Our study proposes that a detailed investigation of the antifungal susceptibility of the genus Aspergillus from different regions of Brazil is necessary for establishing a response profile against the different classes of antifungal agents used in the treatment of aspergillosis.

Antifungal activity of synthetic antiseptics and natural compounds against Candida dubliniensis before and after in vitro fluconazole exposure

ABSTRACT INTRODUCTION: This study evaluated the susceptibilities of oral candidiasis-derived Candida albicans, fluconazole-resistant (FR) Candida dubliniensis, and fluconazole-susceptible (FS) C. dubliniensis to synthetic antiseptics [chlorhexidine gluconate (CHX), cetylpyridinium chloride (CPC), and triclosan (TRC)] and natural compounds (carvacrol, eugenol and thymol). METHODS: Susceptibility tests were performed based on the M27-A3 reference method. The fluconazole-resistant C. dubliniensis strains were obtained after prolonged in vitro exposure to increasing fluconazole concentrations. The geometric mean values for minimum inhibitory concentrations and minimum fungicidal concentrations were compared among the groups. RESULTS: Fluconazole-susceptible C. dubliniensis was more sensitive to CPC and TRC than FR C. dubliniensis and C. albicans were. However, eugenol and thymol were more active against FR C. dubliniensis. The fungicidal activities of CHX and TRC were similar for the three groups, and FR C. dubliniensis and C. albicans had similar sensitivities to CPC. CONCLUSIONS: The resistance of C. dubliniensis to fluconazole affects its sensitivity the synthetic antiseptics and natural compounds that were tested.

Antifungal activities of tacrolimus in combination with antifungal agents against fluconazole-susceptible and fluconazole-resistant Trichosporon asahii isolates

ABSTRACT The antifungal activity of tacrolimus in combination with antifungal agents against different fungal species has been previously reported. Here we report the in vitro interactions between tacrolimus and amphotericin B, fluconazole, itraconazole, and caspofungin against 30 clinical isolates of both fluconazole-susceptible and fluconazole-resistant Trichosporon asahii. For these analyses, we used the broth microdilution method based on the M27-A3 technique and checkerboard microdilution method. Tacrolimus showed no activity against T. asahii strains (minimal inhibitory concentrations, MICs > 64.0 µg mL−1). However, a larger synergistic interaction was observed by the combinations tacrolimus + amphotericin B (96.67%) and tacrolimus + caspofungin (73.33%) against fluconazole-susceptible isolates. Combinations with azole antifungal agents resulted in low rates of synergism for this group (fluconazole + tacrolimus = 40% and itraconazole + tacrolimus = 10%). Antagonistic interactions were not observed. For the fluconazole-resistant T. asahii group, all tested combinations showed indifferent interactions. The synergism showed against fluconazole-susceptible T. asahii isolates suggests that the potential antifungal activity of tacrolimus deserves in vivo experimental investigation, notably, the combination of tacrolimus with amphotericin B or caspofungin.

Epidemiological, clinical and diagnostic aspects of sheep conidiobolomycosis in Brazil / Aspectos epidemiológicos, clínicos e de diagnóstico da conidiobolomicose ovina no Brasil

ABSTRACT: Conidiobolomycosis is an emerging disease caused by fungi of the cosmopolitan genus Conidiobolus. Particular strains of Conidiobolus coronatus, Conidiobolus incongruus and Conidiobolus lamprauges, mainly from tropical or sub-tropical origin, cause the mycosis in humans and animals, domestic or wild. Lesions are usually granulomatous and necrotic in character, presenting two clinical forms: rhinofacial and nasopharyngeal. This review includes the main features of the disease in sheep, with an emphasis on the epidemiology, clinical aspects, and diagnosis of infections caused by Conidiobolus spp. in Brazil. In this country, the disease is endemic in the Northeast and Midwest, affecting predominantly woolless sheep breeds and occasioning death in the majority of the studied cases. The species responsible for infections of sheep are C. coronatus and C. lamprauges and the predominant clinical presentation is nasopharyngeal. These fungal infections are very important, since they compromise the health status of the sheep flock and cause serious economic losses to the sheep industry. Thus, research is needed to investigate faster tools for diagnosis and effective methods for the control and treatment of conidiobolomycosis.

RESUMO: Conidiobolomicose é uma enfermidade emergente causada por fungos cosmopolitas do gênero Conidiobolus. Isolados de Conidiobolus coronatus, Conidiobolus incongruus e Conidiobolus lamprauges, principalmente de origem tropical e subtropical, afetam humanos, animais domésticos e silvestres. As lesões da conidiobolomicose normalmente são de caráter granulomatoso e necrótico, apresentando-se sob duas formas clínicas: rinofacial e nasofaríngea. O presente artigo tem como objetivo revisar as principais características da doença em ovinos, particularizando a epidemiologia, assim como os aspectos clínicos e o diagnóstico das infecções causadas por Conidiobolus spp. no Brasil. Neste País, a enfermidade é endêmica nas regiões nordeste e centro-oeste, afetando ovinos predominantemente de raças deslanadas, ocasionando a morte na grande maioria dos casos estudados. As espécies do fungo responsáveis pelas infecções em ovinos são C. coronatus e C. lamprauges e a forma clínica predominante é a nasofaríngea. Tais infecções fúngicas são muito importantes, uma vez que comprometem o status sanitário do rebanho ovino e acarretam sérios prejuízos econômicos à ovinocultura. Dessa forma, pesquisas são necessárias para investigar ferramentas rápidas de diagnóstico e métodos eficazes de controle e tratamento da conidiobolomicose.

Seroprevalence of Pythium insidiosum infection in equine in Rio Grande do Sul, Brazil / Soroprevalência da infecção por Pythium insidiosum em equinos no estado do Rio Grande do Sul

ABSTRACT: An epidemiological survey was carried out by performing an Enzyme Linked Immuno Sorbent Assay (ELISA) test to determine the seroprevalence of Pythium insidiosum infection in equine in Rio Grande do Sul State (RS), Brazil. The serological study covered seven geographical regions of RS, classified according to the Instituto Brasileiro de Geografia e Estatística (IBGE). The samples were obtained from official veterinary service (Serviço Veterinário Oficial, SVO) linked to the Secretaria da Agricultura, Pecuária e Agronegócio of RS (SEAPA-RS) to proceed the investigation of equine infectious anemia in 2014. Samples were collected during the months of September and October of 2013, covering the seven geographical regions of RS, and totalized 1,002 serum samples. The seroprevalence for P. insidiosum in RS was 11.1% (CI95% 9.23 to 13.22). The relative risk (RR) of the presence of antibodies anti-P. insidiosum was in the regions Southeast 11.17 (CI95%, 4.65 to 26.8), Porto Alegre 4.62 (CI95%, 1.70 to 12.55), Southwest 11.17 (CI95%, 4.65 to 26.8) and Northwestern 3.72 (CI95%, 1.52 to 9.09). The highest prevalence (69.1%) was observed in females with RR of 1.59 (CI95%, 1.11 to 2.27). When the presence of dams was evaluated, the seropositivity was evident in 74.4%, presenting an association of 2.13 (CI95%, 1.16 to 3.91) compared to farms without dams. In properties with veterinary assistance, the frequency of 72.7% and RR of 3.04 (CI95%,, 1,85 to 4,98) of seropositivity were observed. Due to the importance of pythiosis in horse herds, this study highlights the presence of anti-P. insidiosum antibodies in horses in RS, Brazil.

RESUMO: Um levantamento soroepidemiológico foi realizado através do teste de ELISA indireto para determinar a soroprevalência da infecção por Pythium insidiosum em equinos no estado do Rio Grande do Sul (RS), classificadas de acordo com o Instituto Brasileiro de Geografia e Estatística (IBGE). As amostras utilizadas eram provenientes do cadastro das propriedades do Serviço Veterinário Oficial (SVO), da Secretaria da Agricultura, Pecuária e Agronegócio do RS (SEAPA-RS), coletadas para o inquérito da anemia infecciosa equina de 2014. As coletas foram realizadas durante os meses de setembro e outubro de 2013, abrangendo as sete mesorregiões geográficas do RS, e totalizaram 1.002 amostras de soro. Do total das amostras testadas, 11.1% (CI95% 9.23 to 13.22) foram soropositivas para P. insidiosum. Constatou-se o risco relativo (RR) da presença de anticorpos anti-P. insidiosum nas regiões Sudeste 11,17(IC95%, 4,65-26,8), Porto Alegre 4,62 (IC95%, 1,70-12,55), Sudoeste 11,17 (IC95%, 4,65-26,8) e Noroeste 3,72 (IC95%, 1,52-9,09). Observou-se a maior soroprevalência (69,1%) em fêmeas com RR de 1,59 (IC95%, 1,11-2,27). Quanto à presença de açudes, evidenciou-se soropositividade em 74,4% das propriedades, apresentando associação de 2,13 (IC95%,1,16-3,91) em comparação com propriedades sem açude. Em propriedades com assistência veterinária, foi verificada a frequência de 72,7% e RR de 3,04 (IC95%,1,85-4,98). Diante da relevância da pitiose em rebanhos equinos, destaca-se a presença de anticorpos anti-P. insidiosum em equinos no estado do RS.

Sporothrix schenckii complex: susceptibilities to combined antifungal agents and characterization of enzymatic profiles / Complexo Sporothrix schenckii: susceptibilidade à combinação de antifúngicos e caracterização dos perfis enzimáticos

SUMMARYSporothrix schenckiiwas reclassified as a complex encompassing six cryptic species, which calls for the reassessment of clinical and epidemiological data of these new species. We evaluated the susceptibility of Sporothrix albicans(n = 1) , S. brasiliensis(n = 6) , S. globosa(n = 1), S. mexicana(n = 1) and S. schenckii(n = 36) to terbinafine (TRB) alone and in combination with itraconazole (ITZ), ketoconazole (KTZ), and voriconazole (VRZ) by a checkerboard microdilution method and determined the enzymatic profile of these species with the API-ZYM kit. Most interactions were additive (27.5%, 32.5% and 5%) or indifferent (70%, 50% and 52.5%) for TRB+KTZ, TRB+ITZ and TRB+VRZ, respectively. Antagonisms were observed in 42.5% of isolates for the TRB+VRZ combination. Based on enzymatic profiling, the Sporothrix schenckiistrains were categorized into 14 biotypes. Leucine arylamidase (LA) activity was observed only for S. albicansand S. mexicana. The species S. globosaand S. mexicanawere the only species without β-glucosidase (GS) activity. Our results may contribute to a better understanding of virulence and resistance among species of the genus Sporothrixin further studies.

RESUMOAvaliou-se a susceptibilidade de Sporothrix albicans(n = 1), S. brasiliensis(n = 1), S. globosa(n = 1), S. mexicana(n = 1) e S. schenckii(n = 36) frente à terbinafina (TRB) e a TRB em combinação com itraconazol (ITZ), cetoconazol (KTZ) e voriconazol (VRZ) pelo método de microdiluição ( checkerboard); o perfil enzimático destas espécies foi também avaliado, com base no kit API-ZYM. A maioria das interações foram aditivas (27,5%, 32,5% e 5%) ou indiferentes (70%, 50% e 52,5%) para TRB+KTZ, TRB+ITZ e TRB+VRZ, respectivamente. Antagonismo foi observado em 42,5% dos isolados para a combinação TRB+VRZ. Com base nos perfis enzimáticos, as cepas de Sporothrix schenckiievidenciaram 14 biotipos distintos. A atividade da leucina arilamidase (LA) só foi observada em S. albicanse S. mexicana.As espécies S. globosae S. mexicanaforam as únicas que não evidenciaram atividade da enzima β-glucosidase (GS). Estes resultados poderão contribuir para um melhor entendimento da virulência e resistência entre as espécies do gênero Sporothrixem futuros estudos.

Synergistic effects of tacrolimus and azole antifungal compounds in fluconazole-susceptible and fluconazole-resistant Candida glabrata isolates

In vitro interaction between tacrolimus (FK506) and four azoles (fluconazole, ketoconazole, itraconazole and voriconazole) against thirty clinical isolates of both fluconazole susceptible and -resistant Candida glabrata were evaluated by the checkerboard microdilution method. Synergistic, indifferent or antagonism interactions were found for combinations of the antifungal agents and FK506. A larger synergistic effect was observed for the combinations of FK506 with itraconazole and voriconazole (43%), followed by that of the combination with ketoconazole (37%), against fluconazole-susceptible isolates. For fluconazole-resistant C. glabrata, a higher synergistic effect was obtained from FK506 combined with ketoconazole (77%), itraconazole (73%), voriconazole (63%) and fluconazole (60%). The synergisms that we observed in vitro, notably against fluconazole-resistant C. glabrata isolates, are promising and warrant further analysis of their applications in experimental in vivo studies.

Susceptibility variation of Malassezia pachydermatis to antifungal agents according to isolate source

Malassezia pachydermatisis associated with dermatomycoses and otomycosis in dogs and cats. This study compared the susceptibility of M. pachydermatis isolates from sick (G1) and healthy (G2) animals to azole and polyene antifungals using the M27-A3 protocol. Isolates from G1 animals were less sensitive to amphotericin B, nystatin, fluconazole, clotrimazole and miconazole.

The activity of echinocandins, amphotericin B and voriconazole against fluconazole-susceptible and fluconazole-resistant Brazilian Candida glabrata isolates

The extensive use of azole antifungal agents has promoted the resistance of Candida spp to these drugs. Candida glabrata is a problematic yeast because it presents a high degree of primary or secondary resistance to fluconazole. In Brazil, C. glabrata has been less studied than other species. In this paper, we compared the activity of three major classes of antifungal agents (azoles, echinocandins and polyenes) against fluconazole-susceptible (FS) and fluconazole-resistant (FR) C. glabrata strains. Cross-resistance between fluconazole and voriconazole was remarkable. Among the antifungal agents, the echinocandins were the most effective against FS and FR C. glabrata and micafungin showed the lowest minimal inhibitory concentrations.

In vitro activities of antifungal agents alone and in combination against fluconazole-susceptible and -resistant strains of Candida dubliniensis

In the present study we used two groups of Candida dubliniensis strains: one containing fluconazole-susceptible clinical isolates and another containing fluconazole-resistant laboratory derivative from the former to examine the changes on susceptibility accompanying the development of resistance to fluconazole. Our findings confirmed the ability of C. dubliniensis isolates to become resistant to fluconazole and indicated that this resistance was crossed with ketoconazole, itraconazole, ravuconazole and terbinafine. We also tested combinations of terbinafine, amphotericin B, itraconazole and voriconazole against both groups of isolates in a checkerboard assay. Surprisingly, most combinations evidenced indifferent interactions, and the best synergism appeared when terbinafine and itraconazole were combined against the fluconazole-resistant group.

Pitiose em animais de produção no Pantanal Matogrossense / Pythiosis of livestock in the Pantanal, Mato Grosso, Brazil

An epidemiological study (2009-2010) included cattle and horses affected with pythiosis from farms in Pantanal, Mato Grosso, Brazil. The disease occurred predominantly from November to March annually, period of the local rainy season. The mean annual incidence of pythiosis was 0.22 percent and 12.5 percent in cattle and horses, respectively. In cattle, the case distribution occurred during the peak of the floods and was restricted to 6 to 18 months of age heifers. Mild perilesional edema and lameness evolved to spontaneous resolution within until 90 days. In horses, pythiosis affected animals of both sexes with 3 to 8 years of age. Reinfection afflicted one horse. The lesions evolved to extensive granulation tissue and ‘kunkers' in horses that experienced marked cachexy. Death occurred three to seven months after the beginning of symptoms. The mean mortality rate was 5.88 percent and the case fatality rate was 45.45 percent. Diagnosis was confirmed by ELISA test, PCR, histopathology (HE and Grocott) and isolation of Pythium insidiosum. In the endemic area studied, the disease in cattle did not represent an economic impact; on the other hand, the majority of cases in horses progressed unsatisfactorily. The incidence of pythiosis in horses was 57.23 times the incidence recorded in cattle, with statistical significance. While environmental flooded conditions were the same, such difference might be associated with species susceptibility, behavior and management.

Realizou-se, em quatro propriedades rurais no Pantanal Matogrossense, em 2009 e 2010, um estudo clínico e epidemiológico da pitiose em bovinos e equinos. A enfermidade ocorreu predominantemente entre os meses de novembro e março, correspondendo ao período chuvoso na região. A incidência média anual foi de 0,22 por cento e 12,5 por cento em bovinos e equinos, respectivamente. Nos bovinos, a distribuição dos casos ocorreu no ápice das cheias e restringiu-se a novilhas de 6 a 18 meses de idade, nas quais as lesões cutâneas estiveram associadas com edemas perilesionais discretos e claudicações, mas curaram espontaneamente, em um período máximo de 90 dias. Nos equinos, a pitiose acometeu animais de ambos os sexos, de três a oito anos de idade e registrou-se um caso de reinfecção. A doença evoluiu com agravos no sítio lesional, com desenvolvimento de extenso tecido de granulação, kunkersem permeio à lesão, acentuada caquexia e mortes, as quais ocorreram entre três e sete meses após o início dos sinais. A mortalidade média foi 5,88 por cento e a letalidade 45,45 por cento. A confirmação do diagnóstico incluiu ELISA-teste, PCR, histopatologia (HE e Grocott) e isolamento de P. insidiosum. Na área endêmica estudada, a enfermidade não causou impacto econômico em bovinos, a despeito da evolução insatisfatória registrada na maioria dos equinos. Nesse estudo, a incidência de pitiose em equinos foi 57,23 vezes a observada em bovinos, com significância estatística. Apesar das mesmas condições ambientais, tal diferença foi provavelmente associada com susceptibilidade, comportamento e manejo das espécies nos campos alagados.

Óleos essenciais como substituintes de antibióticos promotores de crescimento em frangos de corte: perfil de soroproteínas e peroxidação lipídica / Essential oils as substitutes for antibiotic growth promoters in broilers: soroproteins profile and lipid peroxidation

Os óleos essenciais são alternativas ao uso de promotores de crescimento antibióticos na avicultura, devido à sua ação antimicrobiana, além de possuírem propriedades antioxidante e imunomoduladora. Este estudo foi realizado com o objetivo de avaliar o efeito da suplementação dietética de três doses de óleos essenciais (OLES) de orégano (Origanum vulgare L.), sálvia (Salvia officinalis L.), alecrim (Rosmarinus officinalis L.) e extrato de pimenta (Capsicum frutescens L.) em frangos de corte, pela análise do perfil eletroforético de soroproteínas e da peroxidação lipídica plasmática. Os animais (n=910) foram alocados de forma aleatória em cinco tratamentos, com sete repetições de 26 aves cada: o grupo controle (Tc), que recebeu dieta basal sem aditivos; o grupo que recebeu promotor de crescimento antibiótico na dieta (Tatb); e os grupos T50, T100 e T150, alimentados com OLES na doses de 50, 100 e 150mg kg-1, respectivamente. Aos 42 dias de idade, sete animais foram aleatoriamente selecionados (um de cada repetição) para o estudo do perfil eletroforético de soroproteínas e para a avaliação da peroxidação plasmática de lipídeos, pelo teste de formação de substâncias reativas ao ácido tiobarbitúrico (TBARS). Houve diminuição na concentração de globulinas totais no T150 e na fração betaglobulina nos grupos Tatb e T150 em relação ao grupo controle e ao T50 (P<0,05). Além disso, os níveis de TBARS plasmático foram menores nos grupos que receberam OLES em comparação ao Tc (P<0,05). Dessa forma, pode-se inferir que o efeito dos OLES, na maior dose administrada, sugere menor estímulo ao sistema inume humoral de frangos de corte, assim como acontece com a suplementação de promotores de crescimento antibióticos. Adicionalmente, ocorre menor peroxidação plasmática de lipídios e, consequentemente, menos dano oxidativo em frangos de corte, em resposta ao uso dos OLES.

Essential oils are an alternative to growth promoters based on antibiotics used in animal diets, due to its antimicrobial potential, and immunomodulatory properties. Serum proteins electrophoresis and plasma lipid peroxidation were evaluated in broilers fed with diets supplemented with antibiotics or essential oils from oregano (Origanum vulgare L.), sage (Salvia officinalis L.), rosemary (Rosmarinus officinalis L.) and pepper (Capsicum frutescens L.) crude extract (OLES). The animals (n=910) were distributed within five treatment groups and seven replicates containing 26 birds each one: control group (diet without additives); the group receiving an antibiotic growth promoter diet (Tatb); and the groups T50, T100 and T150 (supplemented with 50, 100 and 150mg kg-1 of OLES, respectively). After 42 days, seven animals were randomly selected for serum proteins electrophoretic fractionation and plasma lipid peroxidation evaluation by thiobarbituric acid reactive species (TBARS) test. Total globulins (T150), betaglobulin fraction (Tatb and T150) and plasma TBARS levels in the groups that received OLES (P<0.05) presented a decrease in relation to the control group. These results suggests lower stimulus to the humoral immune response at the higher dose of OLES, as occurred in the antibiotic growth promoter group. Moreover, it suggests lower lipid peroxidation and, consequently, lower oxidative damage caused by OLES use in broiler chickens.

Sporothrix schenckii associated with armadillo hunting in Southern Brazil: epidemiological and antifungal susceptibility profiles / Sporothrix schenckii relacionado à caça ao tatu no Sul do Brasil: aspectos epidemiológicos e suscetibilidade dos isolados aos antifúngicos

INTRODUCTION: Sporotrichosis is the most common subcutaneous mycosis observed in Brazil and it is generally consequent to a little trauma caused by vegetal particles or spines which inoculate the fungi in the subcutaneous area. Although sporotrichosis had been frequently mentioned with armadillo hunting this form has not been widely reported in Brazil until now. In this study we report ten cases of sporotrichosis evolving the armadillo's hunting diagnosed in some towns located in the central and west regions of Rio Grande do Sul State. METHODS: The cases were established based on clinical and classic mycological laboratorial techniques. The susceptibility tests were conducted by microdilution technique according to M38-A2 CLSI documents. RESULTS: Ten cases of sporotrichosis associated with armadillo hunting detected in the State of Rio Grande do Sul were diagnosed by mycological methods. The susceptibility tests of Sporothrix schenckii isolates to antifungal agents itraconazole, ketoconazole and terbinafine showed that all the isolates were susceptible. CONCLUSIONS: The paper discusses some cultural aspects related to hunting of this wild animal as well as possible causes of this unexpected occurrence in southern Brazil.

INTRODUÇÃO: A esporotricose constitui-se na micose subcutânea mais frequentemente observada e, na maioria dos casos, a infecção é decorrente de pequenos traumas envolvendo fragmentos vegetais ou espinhos que inoculam o fungo no tecido subcutâneo. Embora frequentemente relacionada a caça a tatus, esta ocorrência tem sido raramente relatada no Brasil. Neste estudo relatamos dez casos envolvendo esta prática, observados em várias cidades das regiões centro e oeste do Estado do Rio Grande do Sul. MÉTODOS: o diagnóstico clínico foi confirmado pelos métodos clássicos de cultura em ágar Mycobiotic, identificação micromorfológica seguida de reversão a fase leveduriforme em ágar BHI. Os testes de suscetibilidade foram realizados pela técnica de microdiluição em caldo, de acordo com as normas estabelecidas pelo documento CLSI M38-A2 (2008). RESULTADOS: A esporotricose, decorrente de lesões causadas pela caça ao tatu foi confirmada pelo métodos microbiológicos. Os testes de suscetibilidade indicaram que todos os isolados eram sensíveis ao itraconazol, cetoconazol e terbinafina. CONCLUSÕES: O artigo discute aspectos ambientais e culturais relacionados a caça a este animal silvestre bem como àqueles relacionados a esta inesperada ocorrência.

Differentiation of Candida dubliniensis from Candida albicans with the use of killer toxins / Avaliação das toxinas killer na diferenciação entre Candida albicans e Candida dubliniensis

The aim of this study was to report the ability of killer toxins, previously used as biotyping techniques, as a new tool to differentiate C. albicans from C. dubliniensis. The susceptibility of C. albicans and C. dubliniensis to killer toxins ranged from 33.9 to 93.3 percent and from 6.67 to 93.3 percent, respectively.

Avaliou-se a capacidade das toxinas killer, previamente utilizadas na biotipagem de C. albicans, como método para diferenciar C. albicans de C. dubliniensis. A susceptibilidade de C. albicans e C. dubliniensis às toxinas killer variou de 33,9 por cento a 93,3 por cento para C. albicans e de 6,67 por cento a 93,3 por cento para C. dubliniensis.

Candida dubliniensis does not show phospholipase activity: true or false? / Candida dubliniensis não evidencia atividade fosfolipásica: verdadeiro ou falso?

INTRODUCTION: The phospholipase activity in Candida albicans and Candida dubliniensis isolated from oral candidiasis cases were studied. METHODS: The phospholipase activity was evaluated in egg yolk agar. RESULTS: All the C. albicans isolates (n = 48) showed phospholipase activity (mean Pz = 0.66). However, none of the C. dubliniensis isolates (n = 24) showed this activity. CONCLUSIONS: The authors discuss whether these findings are a true characteristic of C. dubliniensis or a consequence of the methodology employed, which includes the possibility that NaCl may have inhibited the enzymatic activity of C. dubliniensis.

INTRODUÇÃO: Avaliou-se a atividade fosfolipásica em Candida albicans e Candida dubliniensis isoladas de casos de candidíase oral. MÉTODOS: A atividade de fosfolipase foi avaliada em ágar gema de ovo. RESULTADOS: Todos os isolados de C. albicans (nº = 48) evidenciaram atividade fosfolipásica (média Pz = 0.66). Todavia, nenhum isolado de C. dubliniensis (nº= 24) demonstrou esta atividade. CONCLUSÕES: Os autores discutem se estes achados são uma característica verdadeira de C. dubliniensis ou uma conseqüência da metodologia empregada, a qual inclui a possibilidade de que o NaCl seja inibidor da atividade enzimática de C. dubliniensis.

Mastite micótica em ruminantes causada por leveduras / Mycotic mastitis in ruminants caused by yeasts

A ocorrência de casos esporádicos de mastite causados por microrganismos de origem ambiental pode ser considerada como emergente e as leveduras, os fungos leveduriformes e os filamentosos são alguns dos principais agentes envolvidos. Em relação à mastite micótica, as leveduras são freqüentemente causas de infecções da glândula mamária em animais produtores de leite. A maior parte dos casos ocorre sob a forma de surtos localizados e/ou após tratamento com antimicrobianos. Os principais gêneros envolvidos são Candida e Cryptococcus, além de outros como Geotrichum, Pichia e Trichosporon. O objetivo do trabalho é revisar os aspectos gerais sobre a mastite micótica causada por leveduras e fungos leveduriformes, tais como etiologia, causas predisponentes, patogenicidade, diagnóstico, tratamento e profilaxia por meio de uma abordagem cronológica dos primeiros relatos, mas com enfoque principal nos resultados descritos recentemente na literatura veterinária nacional e internacional. O controle da mastite micótica é fundamentado em métodos preventivos, como adequado manejo da ordenha, correta higienização dos equipamentos e das instalações, com a finalidade de reduzir o número de animais acometidos, além de garantir a qualidade e a inocuidade dos produtos lácteos.

The veterinary literature registers sporadic cases of infections caused by environmental microorganisms in which the main agents involved are yeasts, yeasts-like and filamentous fungi. In relation to dairy animals, the yeasts are more frequently incriminated as the aetiological agents of mycotic mastitis. Most cases occur under the form of localized outbreaks and / or after treatment with antimicrobial agents. The main genera involved in mastitis are Candida and Cryptococcus, besides others such as Geotrichum, Pichia and Trichosporon. The purpose of this review is to point out the main aspects of mastitis caused by yeasts and yeast-like fungi, such as etiology, predisposing factors, pathogenicity, diagnosis, treatment and prophylaxis, through a chronological approach of the first reports, with main focus on results described recently in national and international veterinary literature. The mycotic mastitis control should be mainly focused on preventive methods, especially based on adequate management of essential factors like proper milking procedures and a high level of hygiene of the environment and equipments, with the aim of reducing the number of affected animals, assuring the quality and innocuity of dairy food safety.

Duddingtonia flagrans: controle biológico de nematodeos de bovinos a campo / Duddingtonia flagrans: biological control of cattle nematodes in the field

O controle biológico é um método para diminuir uma população pela utilização de antagonista natural. No presente estudo, testou-se a eficácia do fungo nematófago Duddingtonia flagrans no controle de nematódeos parasitos gastrintestinais de bovinos criados à campo no município de Júlio de Castilhos. Foram utilizados 20 bezerros, distribuídos igualmente em duas áreas formadas por pastagem nativa. O grupo A foi tratado com o fungo D. flagrans, cultivado em sorgo, numa concentração de 1x10(6)clamidósporos kg-1 de peso animal, misturados em ração de manutenção, diariamente, durante oito meses. O grupo B serviu como controle e não recebeu fungo, apenas ração. Foram coletadas amostras para contagem de ovos por grama de fezes (OPG) semanalmente. Mensalmente, foram realizadas coproculturas para identificar as espécies de larvas de nematódeos, a pesagem dos animais e a coleta de pasto para contagem das larvas na pastagem. Dados de temperatura e índice pluviométrico foram registrados diariamente. O OPG foi reduzido no grupo tratado, em média 56,8, por cento nos últimos três meses de experimento, variando entre 40,4 e 67,1 por cento no grupo tratado (P<0,001). A coprocultura demonstrou que os principais nematódeos encontrados em ambos os grupos foram dos gêneros Cooperia e Haemonchus. A contagem de larvas na pastagem obteve um percentual de redução 77,1 por cento no grupo tratado ao final do experimento (P<0,01). Pôde-se concluir com este estudo que o papel do fungo Duddingtonia flagrans, é, sem dúvida, importante, principalmente, na diminuição do OPG e na redução significativa de larvas na pastagem. Portanto, este fungo nematófago é uma ferramenta biológica eficaz para ser empregado em um controle integrado de nematódeos de bovinos criados a campo.

Biological control is an alternative method to reduce parasite's population by the use of natural antagonist. In the present study, efficacy of nematophagous fungus Duddingtonia flagrans was tested to control gastrointestinal nematodes parasites of cattle livestock in the field. Twenty calves were used, distributed equally in two distinct plots formed by native pasture. Group A was treated with D. flagrans fungus, cultivated in sorghum (1x10(6) clamidospores kg-1 body weight) mixed with maintenance ration, each day, during eight months. Group B served as a control and did not receive the fungus. Samples for faecal egg count (FEC), were collected each week. There were monthly counts in faecal cultures to identify the species of nematodes larvae, animals weight, blood collection to determine red cell counts and collection of pasture to larvae counting. Temperature and rainfall data were registered daily. The FEC reduced around 56.8 percent in the last three months of the experiment, with a variation between 40.4 and 67.1 percent in the treated group (P<0.001). The faecal cultures demonstrated that the main nematodes found in both the groups were Cooperia and Haemonchus. Larvae counting in the pasture showed a reduction percentage around 77.1 percent in treated group at the end of experiment (P<0.01). It could be concluded with this study, that Duddingtonia flagrans has an important role in the reduction of FEC and significant reduction of larvae in the pasture. Therefore, this nematophagous fungus is efficient as a biological tool to be used in an integrated nematodes control of bovine raised in the field.

Aceturato de diminazeno e dipropionato de imidocarb no controle de infecção por Trypanosoma evansi em Rattus norvegicus infectados experimentalmente / Diminazene aceturate and imidocarb dipropionate in the control of Trypanosoma evansi infection in Rattus norvegicus experimentally infected

Este estudo teve como objetivo avaliar o efeito do aceturato de diminazeno e do dipropionato de imidocarb no controle da infecção por Trypanosoma evansi em ratos (Rattus norvegicus) infectados experimentalmente. Cinqüenta e quatro ratos machos foram inoculados via intraperitonial com 104 tripomastigotas de T. evansi/animal. Os ratos foram monitorados diariamente por meio de esfregaço sanguíneo periférico. No momento em que se observassem oito protozoários por campo microscópico de 1000x, era iniciado o tratamento com as drogas (dia zero). O estudo foi dividido em dois protocolos terapêuticos e os fármacos foram administrados via intramuscular. O primeiro protocolo foi aplicado nos grupos A, B, C e D e o segundo protocolo nos grupos E, F, G e H. O grupo controle foi identificado como grupo I, não medicados. No primeiro protocolo, os ratos receberam uma dose única dos fármacos no dia zero e sempre que se observasse T. evansi na circulação periférica. No segundo protocolo, os roedores receberam as mesmas doses, no entanto, por cinco dias consecutivos. No primeiro protocolo, os dois princípios ativos não apresentaram eficácia curativa, ocorrendo reincidência da parasitemia após alguns dias do tratamento. No segundo protocolo, o aceturato de diminazeno eliminou a forma tripomastigota da circulação e os ratos foram eutanasiados após 90 dias do início do tratamento. Os roedores tratados com dipropionato de imidocarb apresentaram recidiva da infecção após 30 dias. Na histopatologia não se observou alteração renal e hepática relacionada à doença ou aos medicamentos testados. Com base nos resultados, foi concluído que o aceturato de diminazeno, quando administrado por cinco dias consecutivos, é efetivo no tratamento da tripanossomose em ratos.

The aim of this study was to evaluate the efficacy of diminazene aceturate and imidocarb dipropionate in the control of Trypanosoma evansi infection in rats (Rattus norvegicus) experimentally infected. Fifty-four male rats were inoculated through intraperitoneal route with 104 T. evansi trypomastigotes. The rats were evaluated daily by periferic blood smears examination and treated when eight flagellated parasites were observed in 1000x microscopic field. Two therapeutics protocols were used. The first one included Groups A, B, C, D in which the rats were submitted to a single dose of the testing drugs administered by intramuscular route at the day 0 and again when T. evansi was observed in the blood smears. The rats of the second protocol (Groups E, F, G, H) were submitted to the same treatment by five consecutive days. Four rats (Group I) were used as control and were not submitted to any treatment. Tested drugs did not show any curative effect when used in the first protocol, since parasitaemia was evident few days after treatment. The use of diminazene aceturate in the second protocol resulted in elimination of the trypomastigotes from circulation. In this case the rats were euthanized at the day 90. The infection recurred 30 days after the administration of imidocarb dipropionate. Histologically, no lesions were found in the liver or kidney. Diminazene aceturate is effective in treating trypanosomosis in rats when used five days consecutively.

Ocorrência de Trypanosoma evansi em eqüinos no município de Cruz Alta, RS, Brasil / Occurrence of Trypanosoma evansi in equines in Cruz Alta, RS, Brazil

O objetivo deste trabalho foi relatar a ocorrência de Trypanosoma evansi em eqüinos no município de Cruz Alta, Estado do Rio Grande do Sul, abordando aspectos epidemiológicos e sinais clínicos da infecção. A tripanosomose ocorreu em uma propriedade rural no município de Cruz Alta. Ao exame clínico, observou-se que quatro dos animais apresentavam marcha oscilante, com incoordenação dos membros posteriores. No entanto, eles estavam em bom estado nutricional, sem febre, bem hidratados e alimentavam-se normalmente. Foram coletadas amostras de sangue das éguas para hemograma, sendo identificado aumento das proteínas plasmáticas, leucocitose, eosinofilia e linfocitose em animais com sinais clínicos. No esfregaço sangüíneo periférico, observou-se a forma flagelada do T. evansi em três dos eqüinos.

This study aimed at describing the occurrence of Trypanosoma evansi in equines from the city of Cruz Alta, RS, Brazil, relating epidemiological aspects and clinical signs of the infection. The tripanosomiasis occurred in a rural area of Cruz Alta, RS. Clinical signs presented by four animals were stiff and incoordinated gait of the pelvic members, although they were in good nutritional status, without fever, well-hydrated and eating normally. Blood samples were collected from the mares for hemogram. Increased levels of plasmatic proteins, leukocytosis, eosinophilia, and limphocytosis were observed in animals with clinical signs. Flagellated forms of T. evansi were observed in the blood smear of three animals.