Molecular detection of Paracoccidioides in soil from an urban area of southern Brazil

Abstract INTRODUCTION: Previous studies that detected Paracoccidioides spp. DNA in soil taken from rural areas have shown this to be a valuable tool for ecological and epidemiological studies. This study reports the detection of Paracoccidioides spp. DNA in soil samples from an urban area of southern Brazil. METHODS: Sixteen soil samples were submitted to nested-PCR and the amplicons of a representative number of positive samples were sequenced. RESULTS: Paracoccidioides spp. DNA was found in 44% of samples. Four DNA amplicons were sequenced, showing 100% homology with P. brasiliensis. CONCLUSIONS: The southern Brazilian urban population is commonly exposed to the Paracoccidioides fungus.

Can Aspergillus fumigatus conidia cause false-positive results in the galactomannan enzyme immunoassay test?

Abstract INTRODUCTION: Several factors can cause false-positive results in the galactomannan (GM) test; however, others remain unknown. Presently, the impact of airborne contamination by Aspergillus conidia during enzyme-linked immunosorbent assay (ELISA) remains uninvestigated. METHODS: We studied 12 A. fumigatus isolates. Fungal conidia were serially diluted and tested for GM detection using the Platelia® Aspergillus enzyme immunoassay (EIA). RESULTS: The conidia concentration required for an EIA-positive result was 4.8 × 103 (median). CONCLUSIONS: This is the first study to evaluate the impact of environmental contamination on the Platelia® Aspergillus EIA assay. Only massive contamination can interfere with GM optical readings, suggesting that environmental contamination does not cause false-positive test results.

Prevalence of Candida spp. in cervical-vaginal samples and the in vitro susceptibility of isolates

Abstract Vulvovaginal candidiasis (VVC) is an infection of the genital mucosa caused by different species of the genus Candida. Considering the lack of data on this topic in the south of Brazil, this study aimed to assess the prevalence of Candida spp. in the cervical-vaginal mucosa of patients treated at a university hospital in southern Rio Grande do Sul, as well as the etiology and the susceptibility of the isolates against fluconazole, itraconazole, miconazole and nystatin. Samples were collected at the gynecology clinic of the Federal Hospital of the University of Rio Grande, and the isolates were identified using phenotypic and biochemical tests. The susceptibility analysis was performed according to the CLSI M27-A2 protocol. Of the 263 patients included, Candida spp. was isolated in 27%, corresponding to a prevalence of approximately 15% for both VVC and colonization. More than 60% of the isolates were identified as Candida albicans; C. non-albicans was isolated at a rate of 8.6% in symptomatic patients and 14.3% in asymptomatic patients. The prevalence of resistance against fluconazole and itraconazole was 42% and 48%, respectively; the minimal inhibitory concentration of miconazole ranged from 0.031 to 8 µg/mL, and that of nystatin ranged from 2 to >16 µg/mL. The high rate of resistance to triazoles observed in our study suggests the necessity of the association of laboratory exams to clinical diagnosis to minimize the practice of empirical treatments that can contribute to the development of resistance in the isolates.

Cytotoxicity and anti-Sporothrix brasiliensis activity of the Origanum majorana Linn. oil

Abstract The study aimed to evaluate the anti-Sporothrix sp. activity of the essential oil of Origanum majorana Linn. (marjoram), its chemical analysis, and its cytotoxic activity. A total of 18 fungal isolates of Sporothrix brasiliensis (n: 17) from humans, dogs and cats, and a standard strain of Sporothrix schenckii (n: 1) were tested using the broth microdilution technique (Clinical and Laboratory Standard Institute - CLSI M27-A3) and the results were expressed in minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC). The MIC50 and MIC90 of itraconazole against S. brasiliensis were 2 µg/mL and 8 µg/mL, respectively, and the MFC50 and MFC90 were 2 µg/mL and >16 µg/mL, respectively, with three S. brasiliensis isolates resistant to antifungal. S. schenckii was sensitive at MIC of 1 µg/mL and MFC of 8 µg/mL. For the oil of O. majorana L., all isolates were susceptible to MIC of ≤2.25-9 mg/mL and MFC of ≤2.25-18 mg/mL. The MIC50 and MIC90 were ≤2.25 mg/mL and 4.5 mg/mL, respectively, and the MFC50/90 values were twice more than the MIC. Twenty-two compounds were identified by gas chromatography with a flame ionization detector (CG-FID) and 1,8-cineole and 4-terpineol were the majority. Through the colorimetric (MTT) assay, the toxicity was observed in 70-80% of VERO cells between 0.078 and 5 mg/mL. For the first time, the study demonstrated the satisfactory in vitro anti-Sporothrix sp. activity of marjoram oil and further studies are needed to ensure its safe and effective use.

Anti- Sporothrix spp. activity of medicinal plants

ABSTRACT Cases of sporotrichosis in humans and animals without satisfactory clinical response have increased, a warning sign of strains resistant to conventional antifungal agents. The urgent search for alternative therapies was an incentive for research on medicinal plants with anti-Sporothrix spp. properties. A bibliographic survey was performed based on scientific papers about in vitro and in vivo antifungal activity of essential oils and extracts of plants in differents solvents against the fungal of the Sporothrix schenckii complex. The study methodology consisted of a literature review in Google Scholar, Science Direct, Pubmed, Bireme and Springer link with papers from 1986 to 2015. We found 141 species of plants that were investigated, of which 100 species were concentrated in 39 botanical families that had confirmed anti-Sporothrix activity. Combretaceae, Asteraceae and Lamiaceae represented the botanical families with the greatest number of plants species with antifungal potential, using different methodologies. However, there are few studies with medicinal plants in experimental infection in animals that prove their activity in the treatment of sporotrichosis. It reinforces the need for further research related to standardization of in vitro methodologies and in vivo studies related to safety and to toxicity potential of these plants with anti-Sporothrix spp. activity.

RESUMO Casos de esporotricose em humanos e animais sem resposta clínica satisfatória têm aumentado, sinal de alarme para o surgimento de cepas resistentes aos antifúngicos convencionais. A urgente busca por alternativas terapêuticas tem incentivado as pesquisas em plantas medicinais com atividade anti-Sporothrix spp. Um levantamento bibliográfico foi realizado com base em artigos científicos sobre a atividade antifúngica in vitro e in vivo de óleos essenciais e extratos de plantas preparados em diferentes solventes contra o complexo Sporothrix schenckii. A metodologia do estudo consistiu em uma revisão bibliográfica em Google Scholar, Science Direct, Pubmed, Bireme e Springer link com artigos desde 1986 até 2015. Foram encontradas 141 espécies de plantas já investigadas, das quais 100 espécies concentradas em 39 famílias botânicas apresentaram atividade anti-Sporothrix spp. confirmada. Combretaceae, Asteraceae e Lamiaceae representaram as famílias botânicas com maior número de espécies vegetais com potencial antifúngico, empregando diferentes metodologias. Entretanto, há poucos estudos com plantas medicinais em infecção experimental animal comprovando sua atividade no tratamento da esporotricose. Reforça-se a necessidade de mais pesquisas relacionadas à padronização de metodologias in vitro e a estudos in vivo relacionados à segurança e potencial tóxico dessas plantas com atividade anti-Sporothrix spp.

Seroepidemiology of Paracoccidioides brasiliensis infection in horses from Rio Grande do Sul, Brazil

Paracoccidioides brasiliensis is the etiological agent of the major systemic mycosis in Brazil, called paracoccidioidomycosis. Although the Rio Grande do Sul is considered an endemic area of the disease, there are few studies on the ecology of P. brasiliensis in the state. Therefore, this study aimed to evaluate the infection of P. brasiliensis in horses from the mesoregion of Southwest Riograndense, using these animals as sentinels. Serological techniques, such as double immunodiffusion in agar gel (AGID) and indirect ELISA, were performed to detect the anti-gp43 P. brasiliensis antibody in horses from five different farms in the region of Bagé, RS, Brazil. Serology was performed in 200 Pure Blood English horses up to two years of age that were born and raised exclusively at the farms. Of these horses, 12% had anti-gp43 antibodies according to the ELISA results, with rates ranging from 0 to 30% according to the farm of origin (p < 0.001). Based on the immunodiffusion results, all equine serum samples were negative. These results indicate the presence of the fungus P. brasiliensis in the middle region of the southwestern state of Rio Grande do Sul, Brazil.

Susceptibility of dermatophytic fungi to commonly used disinfectants / Suscetibilidade de fungos dermatófitos a desinfetantes comumente utilizados

This study aimed evaluate the antidermatophytic activity of three commercial disinfectants commonly used for environmental control of microorganisms in veterinary medicine. Sodium hypochlorite at 40 µL/mL, chloro-phenol derived at 30 µL/mL and chlorhexidine digluconate at 66.7 µL/mL were tested against 14 strains of dermatophytes, identified as Microsporum canis (n: 3) and Microsporum gypseum (n: 11). The tests was performed in accordance with guidelines of the Clinical and Laboratory Standards Institute (CLSI), documents M38-A2 and M51-A, adapted to disinfectants. In the microdilution broth test, chlorhexidine digluconate had MIC values (Minimum Inhibitory Concentration) of 4.16 µL/mL and MCF (Minimum Fungicidal Concentration) from 4.16 to 8.33 µL/mL, while chloro-phenol derived obtained MIC and MCF of 1.87 µL/mL, and both disinfectants had fungicidal activity at concentrations below the recommended. Sodium hypochlorite obtained MIC from 10 to 80 µL/mL and MFC of 40 to 80 µL/mL, requiring at most isolates twice the recommended concentration to achieve same activity. In the disc diffusion test, the mean inhibition zones for chlorhexidine digluconate was 10.53 mm, for chloro-phenol of 9.9 mm and for sodium hypochlorite was 6.2 mm. Chlorhexidine digluconate and chloro-phenol presented a significant reduction in the growth of dermatophytes, while sodium hypochlorite in concentration recommended showed a low antifungal activity against tested isolates.

O objetivo do estudo foi avaliar a atividade antidermatofítica de três desinfetantes comerciais frequentemente utilizados no controle ambiental de micro-organismos em medicina veterinária. Hipoclorito de sódio a 40 µL/mL, derivado de clorofenol a 30 µL/mL e digluconato de clorexidine a 66.7 µL/mL foram testados contra 14 cepas de dermatófitos, identificados como Microsporum canis (n: 3) e Microsporum gypseum (n: 11). Foram utilizadas as diretrizes do Clinical and Laboratory Standard Institute (CLSI), documentos M38-A2 e M51-A, com adaptações para desinfetantes. Na microdiluição em caldo, digluconato de clorexidine apresentou valores de CIM (Concentração Inibitória Mínima) de 4.16 µL/mL e CFM (Concentração Fungicida Mínima) entre 4.16 a 8.33 µL/mL; derivado de clorofenol obteve CIM e CFM de 1.87 µL/mL, demonstrando que ambos os desinfetantes apresentaram atividade fungicida em concentrações inferiores às recomendadas. O hipoclorito de sódio demonstrou CIM entre 10 a 80 µL/mL e CFM de 40 a 80 µL/mL, requerendo duas vezes a concentração recomendada pelo fabricante para obter atividade fungicida frente a maioria dos isolados fúngicos testados. No teste de disco-difusão, a média das zonas de inibição do digluconato de clorexidine foi de 10.53 mm; do derivado clorofenol 9.9 mm e do hipoclorito de sódio 6.2 mm. O digluconato de clorexidine e o derivado cloro-fenol apresentaram redução significante no crescimento dos dermatófitos testados, enquanto o hipoclorito de sódio, na concentração recomendada, demonstrou baixa atividade antifúngica contra os dermatófitos testados.

First isolation of the Stephanoascus ciferrii in feline otitis in Brazil

Ear infections in cats are uncommon, especially involving yeasts. This report describes the first isolation of the Stephanoascus ciferrii, teleomorph of the Candida genus, in a case of feline otitis in Brazil. The identification and characterization of Stephanoascus ciferrii were confirmed by the Vitek2 System (BioMerieux ®).

Microsporose mista canina - relato e caso / Microporosis mixta canina ? relato de caso / Canine mcrosporose mixed - case report

Dermatofitoses são micoses causadas por fungos como: Epidermophyton, Microsporum e Trichophyton. M. canis e M. gypseum são os principais responsáveis pelas infecções dermatofíticas em pequenos animais e acometem principalmente animais jovens. Essas micoses cursam com lesões nas camadas queratinizadas da pele e pelos e em infecções mistas, a intensidade das lesões pode ser aumentada, pois os agentes causais atuam potencializando o quadro clínico. Descreve-se um caso de microsporose mista canina em um filhote de cão da raça Rottweiler. Um canino da raça Rottweiller atendido em uma clínica particular, pois apresentava lesões alopécicas e descamativas em diferentes sítios do corpo. Foi realizada coleta por meio de raspado cutâneo e o material foi encaminhado para exame micológico. O exame direto do pelo revelou estruturas características de dermatofitose com hifas artrosporadas e aglomerados de artroconídeos. O cultivo micológico resultou em crescimento de colônias fúngicas sugestivas de M. gypseum e M. canis nos meios de cultura semeados com confirmação das características macro e micromorfológicas das duas diferentes espécies fúngicas. Assim, mediante processamento micológico estabeleceu-se, juntamente com o histórico do caso, o diagnóstico de microsporose mista canina. Desse modo, conclui-se a necessidade de exames laboratoriais para a confirmação do diagnóstico definitivo de dermatofitose, sendo a profilaxia, prevenção, controle e potencial zoonótico dessa micose dependente da espécie infectante, tendo importância o seu controle também por seu impacto em saúde pública.

Dermatophytosis mycoses are caused by fungi such as Epidermophyton, Microsporum and Trichophyton. M. canis and M. gypseum are the main responsible for dermatophytic infections in small animals and affect mainly young animals. These mycoses occur with lesions in the keratinized layers of the skin and the mixed infections and the intensity of the lesions can be increased because the causative agents act enhancing the clinical picture. We describe a case of mixed microsporose dog in a puppy dog breed Rottweiler. A canine breed Rottweiller served in a private clinic because he had alopecia, scaly lesions at different body sites. It was collected through skin scrapings and the material was sent for mycological examination. Direct examination of the structures revealed by the characteristics of dermatophytosis with of hyphae artrosporadas and arthroconidia clusters. The mycological cultivation resulted in growth of fungal colonies suggestive of M. canis and M. gypseum in culture media seeded with confirmation of macro and micromorphological characteristics of the two different fungal species. Thus, by processing mycological established together with the case history, the diagnosis of canine microsporose mixed. Through the described case is concluded the need for laboratory tests to confirm the definitive diagnosis of dermatophytosis and prophylaxis, prevention, control and zoonotic potential of this mycosis dependent on the infecting species, taking control of the same importance also for their impact on public health.

Dermatofitosis son micosis causadas por hongos como: Epidermophyton, Microsporum y Trichophyton. M.canis y M. gypseum que son los principales responsables en las infecciones por dermatofitos en animales pequeños y afectan principalmente animales jóvenes. Esas micosis se producen con lesiones en las capas queratinizadas de la piel, pelos y en infecciones mixtas, la intensidad de las lesiones puede ser aumentada, pues los agentes causantes actúan potencializando el cuadro clínico. Se describe un caso de microsporosis mixta canina en un cachorro de perro de la raza Rottweiller. Un canino de la raza Rottweiller atendido en una clínica privada que presentaba lesiones de alopecia y lesiones escamosas en diferentes partes del cuerpo. Se realizó recolección por medio de raspado cutáneo y el material fue enviado para examen micológico. El examen directo del pelo reveló estructuras características de dermatofitosis hifas artrosporadas y aglomerados de artroconidios. El cultivo micológico resultó en crecimiento de colonias fúngicas sugestivas de M. gypseum y M canis en los medios de cultura sembrados, con confirmación de características macro y micro morfológicas de dos especies fúngicas. Así, mediante procesamiento micológico establecido junto con el histórico del caso, el diagnóstico de microsporosis mixta canina. Se concluye la necesidad de exámenes de laboratorio para confirmación del diagnóstico definitivo de dermatofitosis, siendo que la profilaxis, prevención, control y potencial zoonótico de esa micosis depende de la especie infectante, siendo importante su control por el impacto que causa a la salud pública.

Pesquisa de trematódeos digenéticos em Heleobia spp. (Mollusca: Hydrobiidae) em área de ocorrência da Ehrlichiose monocítica equina, no Rio Grande do Sul, Brasil / Research of trematodes digenetics from Heleobia spp. (Mollusca: Hydrobiidae) in area of occurrence of equine monocytic ehrlichiosis, in Rio Grande do Sul, Brazil

A ehrlichiose monocítica equina (EME) na região Sul do Rio Grande do Sul tem demonstrado ser importante nas criações de cavalos Crioulos. A enfermidade foi relatada e diagnosticada como causa de diarreia, prejuízos com tratamentos e com a morte de equinos não estabulados, sendo apontada como um fator limitante na criação de equinos em algumas regiões. O modo de transmissão pela via oral, intermediada por trematódeos em ambientes aquáticos, tem sido sustentado. Caracóis dulciaquícolas estão envolvidos como hospedeiros intermediários de trematódeos albergadores de Neorickettsia risticii. Um total de 16.846 caracóis Heleobia foi coletado nos municípios de Arroio Grande, Rio Grande, Palmares do Sul e Santa Vitória do Palmar, 92,2% dos quais foram encontrados nas raízes de aguapés (Eichornea spp.). A frequência de trematódeos presentes nos caracóis variou de 2,3 a 12,8% nas propriedades coletadas. Foram encontrados três tipos de cercárias, morfotipo 1, morfotipo 2 e morfotipo 3, e dois morfotipos de metacercárias nos caracóis. Um total de 357 insetos da ordem Odonata foi coletado, fases de metacercárias foram encontradas no tegumento da subordem Anisoptera com frequência de 5,3%. Mais estudos são necessários para identificar as fases larvais encontradas, bem como para conhecer o hospedeiro definitivo, identificar o parasito adulto e a relação de seu ciclo de vida com a ocorrência da ehrlichiose monocítica equina.

The equine monocytic ehrlichiosis in the region South of the Rio Grande do Sul has demonstrated to be important in the creations of Crioulo horses. It has been reported as cause of diarrhea in equine not surround and as cause of losses with treatments and death of the animals, being pointed as a limited factor in the range breeding in some regions. The way of transmission for the oral way, intermediated for trematodes in aquatic environments, has been supported. Freshwater snails are involved as intermediate hosts of trematodes and harbor of the Neorickettsia risticii. A total of 16,846 Heleobia snails had been collected in the cities of Arroio Grande, Rio Grande, Palmares do Sul and Santa Vitoria do Palmar, 92.2% of which had been found in the roots of aquatics plants (Eichornea spp.). The frequency of trematodes present in the snails varied of 2.3 to 12.8% in the collected regions. Three types of cercariae were found, morphology type 1, morphology type 2 and morphology type 3, and two morphologic type of metacercariae of the snails. A total of 357 insects of the Odonata order were collected, stages of metacercariae had been found in the tissues of suborder Anisoptera with 5.3% of frequency. More studies are necessary for identification of the joined larval phases, as well as knowing the host definitive and identifying the adult parasite and the relation of its cycle of life with the occcurrence of equine monocytic ehrlichiosis.

Efeito antifúngico de agentes químicos sobre leveduras com potencial patogênico isoladas de ambiente hospitalar veterinário / Antifungal effect of chemical agents in yeasts with pathogenic potential isolated from veterinary hospital environment

Os ambientes hospitalares podem albergar micro-organismos patogênicos e oportunistas, capazes de infectar, particu- larmente, os indivíduos imunossuprimidos. O processo de limpeza e desinfecção assume importância capital para o controle de tais doenças. O presente trabalho avaliou a suscetibilidade de leveduras isoladas de superfícies de ambiente hospitalar veterinário frente a quatro desinfetantes/antissépticos comumente utilizados na rotina de desinfecção. O teste de microdiluição em caldo foi realizado com 24 isolados leveduriformes frente ao hipoclorito de sódio 4%, cloreto de benzalcônico 2%, clorexidina-cetrimida 6,6% e derivado de cloro-fenol 3%. A clorexidina-cetrimida, cloreto de ben- zalcônio e derivado de cloro-fenol foram eficazes em todos os isolados com concentração inibitória mínima e concen- tração fungicida inferiores à concentração recomendada pelo fabricante. Por outro lado, a ação fungicida do hipoclorito de sódio em 79,1% dos isolados testados foi obtida na concentração recomendada pelo fabricante, com desempenho inferior aos demais desinfetantes avaliados.

Hospital environments can have pathogens and opportunistic pathogens, important for immunocompromising individuals. The process of cleaning and disinfection constitutes an important measure for the control of these diseases. This study evaluated the sensitivity of yeasts isolated from surfaces of nosocomial veterinary environment to four disinfectants/antiseptics used in the routine of disinfection. The test of broth microdilution was carried in 24isolates of yeasts against 4% sodium hypochlorite, 2% benzalkonium chloride, 6.6% chlorexidine-cetrimide and 3% chloro-phenol derivate. Chlorexidine-cetrimide, benzalkonium chloride and chloro-phenol derivate were efficient in all isolates with minimal inhibitory concentration and minimal fungicide concentration lower than the concentra- tion recommended by manufacturer. By other hand, fungicide action of the sodium hypochlorite in the concentration recommended by the manufacturer was verified in 79.1% of the isolated tested, with the lower performance of than the others evaluated disinfectants.

Clinical and mycological analysis of dog's oral cavity

The oral microbiota of humans and animals is made up of a wide variety of yeasts and bacteria, but microbiota of dogs is not totally described. Although such identification is an important step to establish the etiopathogenesis and adequate therapy for the periodontal disease The aim of this study was to evaluate and correlate oral alterations with the presence of yeasts in oral cavity of female dogs. After clinical evaluation samples from healthy and from dogs with oral diseases were obtained from three different oral sites by swabs, curettes, millimeter periodontal probes and HA membrane tip in cellulose ester. Yeast identification was performed through macroscopic and microscopic colony features and biochemical tests. Dental calculus was the most prevalent occurrence in the oral cavity of 59 females. However, the isolation of yeasts was significantly higher (p < 0.05) in animals suffering from halitosis. Eleven yeast species were identified, namely: Malassezia pachydermatis, Rhodotorula spp., Candida albicans, C. catenulata, C. famata, C. guilliermondii, C. parapsilosis, C. intermedia, Trichosporon asahii, T. mucoides and Cryptococcus albidus. It could be concluded that the yeasts are part of the microbiota from the different sites of the oral cavity of the female canines studied without causing any significant alterations except halitosis.

Alterações hematológicas em felinos com esporotricose cutânea / Hematological alterations of cats with cutaneous sporotrichosis / Alterações hematológicas en felinos con esporotricosis cutânea

A esporotricose é a micose subcutânea de maior ocorrência em pequenos animais, especialmente felinos e caninos. Esta doença causa lesões cutâneas ulceradas de caráter subagudo ou crônico que podem resultar no comprometimento do estado geral do animal. Este estudo avaliou alterações hematológicas em gatos com esporotricose relacionando-as com a forma clínica da doença. As alterações hematológicas foram observadas em 73,3% dos animais, sendo caracterizadas principalmente por leucocitose por neutrofilia e anemia. Os resultados demonstram que animais com a forma cutânea fixa da micose não apresentam alterações no hemograma, enquanto animais com a forma cutânea disseminada desenvolvem importantes alterações que indicam o agravamento do quadro clínico com possibilidade de evolução da micose para uma forma sistêmica.

Sporotrichosis is the most frequent subcutaneous mycosis of small animals, specially cats and dogs. This disease leads toof the mycosis do not present alterations in the blood count. However, animals with the cutaneous disseminated form develop important alterations that can aggravate the disease and evolve the mycosis to a systemic form.

La esporotricosis es la micosis subcutánea de mayor ocurrencia en pequeños animales, específicamente felinos y caninos. Esta enfermedad produce lesiones cutáneas ulceradas de carácter sub agudo o crónico que pueden resultar en el comprometimiento del estado general del animal. Este estudio evalúo alteraciones hematológicas en gatos con esporotricosis, relacionándolas con la forma clínica de la enfermedad. Se ha observado alteraciones hematológicas en 73,3% de los animales, siendo caracterizadas principalmente por leucocitosis por neutrofilia y anemia. Los resultados demuestran que animales con la forma cutánea fija de la micosis no presentan alteraciones en el hemograma, mientras que los animales con la forma cutánea diseminada desarrollan importantes alteraciones que indican el comprometimiento del cuadro clínico con posibilidad de evolución de la micosis para una forma sistémica.

Caracterização da microbiota por fungos filamentosos no tegumento hígido de bovinos de corte / Characterization of filamentous fungal flora from the integument of healthy cattle

O tegumento de bovinos pode albergar uma grande diversidade de fungos filamentosos, potencialmente capazes de causar dermatoses, incluindo fungos do grupo dos dermatófitos. O presente estudo teve por objetivos caracterizar a microbiota de fungos filamentosos do tegumento hígido de bovinos de corte, verificar a presença de dermatófitos como parte integrante da microbiota, assim como avaliar a ocorrência de dermatófitos no solo dos estabelecimentos estudados. Durante o período de um ano, 56 bovinos sadios de quatro propriedades da fronteira Oeste do RS foram submetidos a coletas mensais de pelos e raspado cutâneo. No mesmo período, foram obtidas amostras de solo para pesquisa de dermatófitos. As análises dos resultados indicaram uma microbiota de fungos filamentosos classificada em 30 gêneros, dos quais os mais frequentes foram (nesta ordem): Nigrospora, Fusarium, Curvularia, Alternaria, Epicoccum, Paecilomyces e Trichoderma. Nas amostras de pelame, obteve-se o isolamento de Trichophyton mentagrophytes (0,4 por cento), Microsporum gypseum (0,3 por cento) e Trichophyton verrucosum (0,1 por cento). M. gypseum foi o dermatófito mais frequentemente isolado do solo. Os resultados obtidos evidenciaram que a microbiota fúngica do tegumento hígido de bovinos de corte comporta-se de maneira transitória e que bovinos sadios não atuam como portadores assintomáticos de T. verrucosum, sugerindo que esta espécie de dermatófito é não residente no tegumento dos animais.

The tegument of cattle may harbor a great diversity of filamentous fungi that can potentially cause skin diseases, including fungi belonging to the dermatophyte group. This study aimed to characterize the population of filamentous fungi from a healthy coat of cattle, checking for dermatophytes as part of the microbiota, as well as evaluating the presence of dermatophytes in the soil occupied by the animals. During the period of one year, 56 healthy cattle in four properties in the western boundary of the state of Rio Grande do Sul, Southern Brazil, were subjected to monthly samples of hair and skin scrapings. In the same period soil samples were obtained for the detection of dermatophytes. The results indicated a fungal biota classified into 30 different genera, among which the most frequent were (in order of frequency): Nigrospora, Fusarium, Curvularia, Alternaria, Epicoccum, Paecilomyces and Trichoderma. Isolates of Trichophyton mentagrophytes (0.4 percent), Microsporun gypseum (0.3 percent), and T. verrucosum (0.1 percent) were obtained in the samples of skin scrapings. M. gypseum was the dermatophyte most frequently isolated from soil. The results showed that the fungal biota of the external tegument of healthy cattle is transient. Moreover, it was shown that healthy cattle do not act as asymptomatic carriers of T. verrucosum, suggesting that this species of dermatophytes is not resident in the tegument of animals.

Ultrastructural study of the mycelial phase of clinical isolates of Sporothrix schenckii obtained from feline, canine and human cases of sporotrichosis

Using transmission electron microscopy, we studied the presence of melanin and cell wall thickness of clinical isolates of Sporothrix schenckii obtained from cats, dogs and humans as compared to reference strains. We detected differences regarding presence of the melanin among the clinical isolates of S. schenckii and a correlation between presence of melanin and cell wall thickness.

Molecular heterogeneity of Malassezia pachydermatis through RAPDPCR / Heterogeneidade molecular da Malassezia pachydermatis através de RAPD-PCR

Several methodologies in molecular biology have been used in theinvestigation of Malassezia pachydermatis and its differentiation into subtypes. Recentmolecular research of this species includes the use of samples isolated from canine otitisexterna and dermatitis, as well as from healthy animals, having in view an epidemiologicalstudy of the yeast. The aim of this study was to identify molecular differences in M.pachydermatis samples isolated from dogs with otitis externa. The M. pachydermatis strainswere analyzed by means of the Random Amplification Primer DNA - Polimerase ChainReaction (RAPD–PCR) for molecular heterogeneity research. DNA extraction was carriedout with phenol-chloroform and the RAPD technique using the AGAATCCGCC primer.A variation was observed in the number and arrangement of the bands among the 49studied isolates, grouped into nine patterns. Isolate groupings were not found to be relatedto animal breed, age or sex. It was concluded that M. pachydermatis has differences in itsmolecular profile, as shown by the molecular technique (RAPD – PCR), which allowsisolates to be classified into nine subtypes.

Várias metodologias em biologia molecular têm sido aplicadas para estudar aM. pachydermatis diferenciando-a em subgrupos. Recentemente utiliza-se a investigaçãomolecular desta espécie isolada de otite externa e dermatite, e também de isolados da mesmade animais hígidos, para um estudo epidemiológico da levedura. O objetivo deste trabalhofoi identificar diferenças moleculares entre isolados de M. pachydermatis obtidos de casos deotite externa canina. Para isto, amostras da levedura provenientes de cães com estaenfermidade foram estudadas através da técnica de Polimorfismo de DNA Amplificado aoAcaso - Reação da Polimerase em Cadeia (RAPD–PCR) para pesquisa de heterogeneidademolecular. A extração de DNA foi realizada no processo fenol-cloroformio e a técnica deRAPD foi estudada com o primer AGAATCCGCC. Pode-se observar com estametodologia, variação no número e posição das bandas entre os 49 isolados estudados,podendo-se agrupá-los em nove padrões. Os agrupamentos formados pelos isolados nãoapresentaram relação com raça, idade ou sexo do animal. Concluindo-se que a M.pachydermatis apresenta diferenças em seu perfil molecular, observado pela técnica molecular(RAPD-PCR) que permite classificar os isolados desta espécie em até nove subtipos.

Ocorrência de Cryptococcus neoformans em excretas de pombos na cidade de Pelotas, Estado do Rio Grande do Sul / Occurrence of Cryptococcus neoformans in pigeon excrement in the city of Pelotas, State of Rio Grande do Sul

INTRODUÇÃO: O Cryptococcus neoformans é uma levedura capsulada, agente etiológico da criptococose em humanos e animais, encontrado em fontes ambientais, incluindo excretas de pombos, é uma importante causa de mortalidade em indivíduos imunodeprimidos em todo o mundo. MÉTODOS: Com o objetivo de verificar a ocorrência do Cryptococcus neoformans, em excretas de pombos, na Cidade de Pelotas, pesquisou-se 70 ambientes, incluindo prédios, praças e locais ao ar livre, da Cidade de Pelotas, RS. Após a coleta, os excrementos foram adicionados de salina com cloranfenicol, homogeneizados, semeados em ágar Sabouraud com cloranfenicol e ágar Níger e incubados a 32ºC. Para identificação, realizou-se exame direto, prova da fenoloxidase, urease, assimilação de carboidratos e cultura em meio CGB. RESULTADOS: Dos locais estudados (nº =70), em 26 (37,1 por cento) havia excretas de pombos. Estes lugares foram representados por prédios históricos (nº =8), torre de igreja (nº =1), engenhos e armazéns de arroz (nº =7), praça (nº =1) e locais ao ar livre (nº =9), o isolamento de Cryptococcus neoformans ocorreu em 26,9 por cento (nº =7/26), destes locais. CONCLUSÕES: O estudo chama a atenção, para o isolamento do fungo em áreas urbanas, que apresentavam grande acúmulo de excretas, indicando um risco para a saúde pública, especialmente para indivíduos imunocomprometidos.

INTRODUCTION: Cryptococcus neoformans is an encapsulated yeast and is the etiological agent for human and animal cryptococcosis. It is found in sources within the environment, including pigeon excrement, and is an important cause of mortality among immunocompromised individuals worldwide. METHODS: Seventy different environments in the city of Pelotas, Rio Grande do Sul, were surveyed for the purpose of investigating Cryptococcus neoformans occurrences in pigeon excreta. The environments included buildings, public squares and outdoor locations in the city. After collection, chloramphenicol saline solution was added to the excreta, which were then homogenized and seeded onto Sabouraud agar with chloramphenicol and onto Niger agar, and incubated at 32°C. Identification was performed by direct examination and by means of the phenoloxidase and urease tests, carbohydrate assimilation and culturing in CGB medium. RESULTS: Out of the sites investigated (n = 70), 26 (37.1 percent) of them contained pigeon excreta. These included historical buildings (n = 8), a church tower (n = 1), rice mills and warehouses (n = 7), a public square (n = 1) and outdoor locations (n = 9). Cryptococcus neoformans was isolated from 26.9 percent (n = 7/26) of these locations. CONCLUSIONS: This study draws attention to isolation of this fungus in urban areas that present large accumulations of pigeon excrement. This represents a risk to public health, especially for immunocompromised individuals.

In vitro activity of Origanum vulgare essential oil against Candida species

The aim of this study was to evaluate the in vitro activity of the essential oil extracted from Origanum vulgare against sixteen Candida species isolates. Standard strains tested comprised C. albicans (ATCC strains 44858, 4053, 18804 and 3691), C. parapsilosis (ATCC 22019), C. krusei (ATCC 34135), C. lusitaniae (ATCC 34449) and C. dubliniensis (ATCC MY646). Six Candida albicans isolates from the vaginal mucous membrane of female dogs, one isolate from the cutaneous tegument of a dog and one isolate of a capuchin monkey were tested in parallel. A broth microdilution technique (CLSI) was used, and the inoculum concentration was adjusted to 5 x 10(6) CFU mL-1. The essential oil was obtained by hydrodistillation in a Clevenger apparatus and analyzed by gas chromatography. Susceptibility was expressed as Minimal Inhibitory Concentration (MIC) and Minimal Fungicidal Concentration (MFC). All isolates tested in vitro were sensitive to O. vulgare essential oil. The chromatographic analysis revealed that the main compounds present in the essential oil were 4-terpineol (47.95 percent), carvacrol (9.42 percent), thymol (8.42 percent) and □-terpineol (7.57 percent). C. albicans isolates obtained from animal mucous membranes exhibited MIC and MFC values of 2.72 µL mL-1 and 5 µL mL-1, respectively. MIC and MFC values for C. albicans standard strains were 2.97 µL mL-1 and 3.54 µL mL-1, respectively. The MIC and MFC for non-albicans species were 2.10 µL mL-1 and 2.97 µL mL-1, respectively. The antifungal activity of O. vulgare essential oil against Candida spp. observed in vitro suggests its administration may represent an alternative treatment for candidiasis.

Esporotricose cutânea experimental: Avaliação in vivo do itraconazol e terbinafina / Experimental cutaneous sporotrichosis: in vivo evaluation of itraconazole and terbinafine

O estudo objetivou avaliar a atividade in vivo do itraconazol e terbinafina no tratamento da esporotricose cutânea experimental. Foram utilizados 80 ratos Wistar divididos em quatro grupos (TERB20, TERB30, ITRA e CONT) inoculados no coxim plantar esquerdo com 0,2ml de solução contendo 2x103 células de Sporothrix schenckii/ml e tratados com terbinafina (20 e 30mg/kg), itraconazol (10mg/kg) e placebo durante 13 semanas. As lesões do sítio de inoculação foram avaliadas e mensuradas semanalmente, assim como a disseminação das mesmas. Após foi realizada análise micológica e histopatológica. Os resultados demonstraram que os animais do grupo ITRA diferiram estatisticamente em todos os parâmetros avaliados em relação ao CONT. Em relação à terbinafina, não houve diferenças estatísticas entre os grupos tratados e controle. Pode-se confirmar a boa atividade do itraconazol no tratamento da esporotricose e a pouca eficácia da terbinafina nas doses utilizadas, sendo necessários mais estudos com este antifúngico.

The aim of this study was to evaluate the in vivo activity of itraconazole and terbinafine for treating experimental cutaneous sporotrichosis. Eighty Wistar rats were used, divided into four groups (TERB20, TERB30, ITRA and CONT). They were inoculated in the left plantar pad with 0.2 ml of solution containing 2x10³ cell/ml of Sporothrix schenckii and treated with terbinafine (20 and 30 mg/kg), itraconazole (10 mg/kg) or placebo for 13 weeks. The lesions at the inoculation site were evaluated and measured weekly, along with their dissemination. Mycological and histopathological analyses were performed subsequently. The results showed that the animals in the ITRA group differed statistically in all parameters evaluated, in relation to CONT. For terbinafine, there were no statistical differences between the treated and control groups. It could be confirmed that itraconazole presented good activity for treating sporotrichosis, while terbinafine was ineffective for this disease at the doses used. However, more studies on the latter antifungal agent are needed.

CLSI broth microdilution method for testing susceptibility of Malasseziapachydermatis to thiabendazole / CLSI método de Microdiluição em Caldo para teste de suscetibilidade da Malassezia pachydermatis frente ao tiabendazol

Thiabendazole, classified as antiparasitic and also used as an antifungal drug, can be found as otological solution indicated for treatment of parasitic and fungal external otitis in small animals. Malassezia pachydermatis is a yeast recognized as a normal inhabitant on the skin and mucous membranes of dogs andcats. However, it is considered an opportunistic agent that causes external otitis and dermatitis in these animals. The aim of this study was to evaluate the in vitro effect of thiabendazole against 51 isolates of M. pachydermatis using the CLSI Broth Microdilution method that has been adapted for this yeast species (NCCLS, 2002). Based on this test, the Minimum Inhibitory Concentrations (MIC) of thiabendazol was calculated. Subsequently, the susceptibility of each isolate against this antifungal was determined. It wasobserved that the MIC of thiabendazole against M. pachydermatis ranged from 0.03 to > 4 μg/mL. A total of 13.7% of the isolates were found to be resistant, 47.1% were intermediate and 39.2% were sensitive to the drug. The rate of resistance of the yeasts against thiabendazole was similar to the results previously obtained with other antifungals, while the adapted broth microdilution technique used in this study proved to be efficient.

Tiabendazol, um fármaco classificado como antiparasitário e também usado como antifúngico, pode ser encontrado como solução otologica indicada no tratamento da otite externa parasitária e fungica em pequenos animais. Malassezia pachydermatis é uma levedura considerada habitante normalda pele e das mucosas de cães e gatos. Entretanto, considerase um agente do oportunista causador de otite externa edermatite nestes animais. A finalidade deste estudo foi avaliar o efeito in vitro do tiabendazol frente a 51 amostras de M. pachydermatis através do método CLSI de Microdiluição em Caldo adaptado para esta espécie de levedura (NCCLS, 2002). Baseado neste teste calculou-se as Concentrações InibitóriasMínimas (CIM) do tiabendazol e, subseqüentemente, foi detectada a suscetibilidade de cada amostra frente a este antifungico. Observou-se que a CIM do tiabendazol frente a M. pachydermatis variou de 0,03 a > 4 μg/mL. Estas amostras foram classificadas em resistente (13,7%), sensibilidade intermediária (47,1%) e sensível (39,2%). A resistência dalevedura frente ao tiabendazol mostrou similaridade com resultados anteriormente observados com outros antifúngicos e a adaptação da técnica de Microdiluição em Caldo utilizada mostrou-se eficiente.