Antifungal activity of diphenyl diselenide alone and in combination with itraconazole against Sporothrix brasiliensis.

Sporothrix brasiliensis is the main agent of feline and zoonotic sporotrichosis. Given the scarcity of options for sporotrichosis treatment and the promising activity of diphenyl diselenide (PhSe)2 against a diversity of fungal pathogens, this study aimed to evaluate the in vitro susceptibility of Sporothrix brasiliensis to (PhSe)2 alone and in combination with itraconazole (ITC). Forty clinical isolates of S. brasiliensis were subjected to broth microdilution method to evaluate their susceptibility to (PhSe)2 in concentrations ranging between 0.25 and 128 µg/ml, and to ITC (0.0313-16 µg/ml) following the protocol of the Clinical and Laboratory Standards Institute (CLSI, 2008). All of these isolates were also used to evaluate the interaction between these drugs by checkerboard technique. Geometric mean of (PhSe)2 MIC values was 12.12 µg/ml, ranging from 4 to 32 µg/ml, and for ITC was 0.37 µg/ml (0.125 to 1 µg/ml). Benefic effect (additive or synergistic) in the interaction test between (PhSe)2 and ITC occurs in 73% (29/40) of the isolates. The promising (PhSe)2in vitro activity against S. brasiliensis suggests that it may not only be a potential alternative therapy but also be used as a combination with ITC, which is the first choice of therapy to sporotrichosis.

Sporotrichosis in Southern Brazil, towards an epidemic?

Rio Grande do Sul (RS) is the second highest state with respect to sporotrichosis incidence in Brazil, with most cases occurring in the southern region. Given the importance of epidemiologic monitoring in hyperendemic areas, this study evaluated the spatial, geographical and annual sporotrichosis incidence over a period of 7 years in the southern region of RS, as well as the disease evolution over the last two decades. Data were collected from the Mycology Laboratory of the Federal University of Rio Grande (FAMED-FURG) and from the Centre for Zoonosis Control (CZC) of the Prefeitura Municipal of Pelotas city. All feline cases of sporotrichosis diagnosed between 2010 and 2016 were included and analysed. In addition, cases of human and canine sporotrichosis were accounted for. Over the 7-year period (2010-2016), 372 feline, 34 canine and 83 human cases of sporotrichosis were diagnosed, being the mean number of cases/year 18.33 in the first 3 years of the study and 116.33 in the last 3 years. Cases were distributed among 34 and 28 neighbourhoods in Rio Grande and Pelotas city, respectively. Socioeconomic features reinforced the result that the disease is already spreading across a major part of the southern region. In addition, the study demonstrated the intensification and expansion of the high endemicity areas. Therefore, given that the number of feline sporotrichosis cases in southern RS reached alarming proportions, we suggested that this region will promptly face an epidemic of sporotrichosis if no preventive or control measures are undertaken.

Use of mueller-iinton broth and agar in the germ tue test / Utilização de ágar e caldo Mueller-Hinton na prova do tubo germinativo

Candida albicans is often isolated from clinical samples, thus its presumptive differentiation from other species of the same genus can be based on its ability to form the germ tube in human serum. Nevertheless, there are two other species that share this characteristic: C. dubliniensis and C. africana. The aim of this study was to compare four different substrates to perform the germ tube (GT) test. The Candida spp. isolates were identified using a manual system (135 C. albicans, 24 C. tropicalis and one C. dubliniensis). The germ tube test was performed with fresh, previously frozen serum and Mueller-Hinton (MH) broth and agar. GT was observed in 96% (130/136) of the isolates through the fresh serum technique, 94% (128/136) through previously frozen serum, 92% (125/136) in MH agar, and 90% (122/136) in MH broth. The sensitivity of each test was higher than 90%, with 100% specificity. Both the MH agar and broth were able to identify the true positives, and false positives were not found. However, some C. albicans isolates were not identified. MH agar and broth may be used in laboratory for the rapid presumptive identification of C. albicans, as an alternative method for germ tube test.

Candida albicans é frequentemente isolada em amostras clínicas, assim a sua diferenciação presuntiva de outras espécies do gênero pode ser baseada na habilidade em formar o tubo germinativo em soro humano. Entretanto, existem outras duas espécies que também possuem essa característica, C. dubliniensis e C. africana. O objetivo foi comparar quatro diferentes substratos para a realização da prova do tubo germinativo (TG). Utilizou-se isolados de Candida spp. identificados através de meio manual (135 C. albicans, 24 C. tropicalis e um C. dubliniensis). A prova do tubo germinativo foi realizada utilizando soro previamente congelado e fresco, caldo e ágar Mueller-Hinton (MH). O TG através da técnica do soro a fresco foi observado em 96% (130/136), 94% (128/136) através do soro previamente congelado, 92% (125/136) no ágar e 90% (122/136) no caldo MH. A sensibilidade de cada teste foi maior que 90% e especificidade de 100%. Tanto o caldo quanto o ágar MH foram capazes de identificar apenas os verdadeiros positivos e não ocorrendo falsos positivos, porém deixaram de identificar alguns isolados de C. albicans. O ágar e o caldo MH podem ser utilizados na rápida e presuntiva identificação laboratorial de C. albicans, como uma alternativa para o teste do tubo germinativo.

Use of Mueller-Hinton broth and agar in the germ tube test.

Candida albicans is often isolated from clinical samples, thus its presumptive differentiation from other species of the same genus can be based on its ability to form the germ tube in human serum. Nevertheless, there are two other species that share this characteristic: C. dubliniensis and C. africana. The aim of this study was to compare four different substrates to perform the germ tube (GT) test. The Candida spp. isolates were identified using a manual system (135 C. albicans, 24 C. tropicalis and one C. dubliniensis). The germ tube test was performed with fresh, previously frozen serum and Mueller-Hinton (MH) broth and agar. GT was observed in 96% (130/136) of the isolates through the fresh serum technique, 94% (128/136) through previously frozen serum, 92% (125/136) in MH agar, and 90% (122/136) in MH broth. The sensitivity of each test was higher than 90%, with 100% specificity. Both the MH agar and broth were able to identify the true positives, and false positives were not found. However, some C. albicans isolates were not identified. MH agar and broth may be used in laboratory for the rapid presumptive identification of C. albicans, as an alternative method for germ tube test.

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.

Isolation of dermatophytes in wild felids from screening centers.

The aim of this study was detect the presence of dermatophyte fungi on wild felids from screening centers. Samples were taken from 30 animals, assembled in two groups: "free-ranging" and "transitory captivity". The dermatophytes (Trichophyton genus), isolated from two felids (6.6%), both of the group "free-ranging".

Determination of germ tube, phospholipase, and proteinase production by bloodstream isolates of Candida albicans.

INTRODUCTION: Candida albicans is a commensal and opportunistic agent that causes infection in immunocompromised individuals. Several attributes contribute to the virulence and pathogenicity of this yeast, including the production of germ tubes (GTs) and extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate GT production and phospholipase and proteinase activities in bloodstream isolates of C. albicans. METHODS: One hundred fifty-three C. albicans isolates were obtained from blood samples and analyzed for GT, phospholipase, and proteinase production. The assays were performed in duplicate in egg yolk medium containing bovine serum albumin and human serum. RESULTS: Detectable amounts of proteinase were produced by 97% of the isolates, and 78% of the isolates produced phospholipase. GTs were produced by 95% of the isolates. A majority of the isolates exhibited low levels of phospholipase production and high levels of proteinase production. CONCLUSIONS: Bloodstream isolates of C. albicans produce virulence factors such as GT and hydrolytic enzymes that enable them to cause infection under favorable conditions.

Determination of germ tube, phospholipase, and proteinase production by bloodstream isolates of Candida albicans

Introduction Candida albicans is a commensal and opportunistic agent that causes infection in immunocompromised individuals. Several attributes contribute to the virulence and pathogenicity of this yeast, including the production of germ tubes (GTs) and extracellular hydrolytic enzymes, particularly phospholipase and proteinase. This study aimed to investigate GT production and phospholipase and proteinase activities in bloodstream isolates of C. albicans. Methods One hundred fifty-three C. albicans isolates were obtained from blood samples and analyzed for GT, phospholipase, and proteinase production. The assays were performed in duplicate in egg yolk medium containing bovine serum albumin and human serum. Results Detectable amounts of proteinase were produced by 97% of the isolates, and 78% of the isolates produced phospholipase. GTs were produced by 95% of the isolates. A majority of the isolates exhibited low levels of phospholipase production and high levels of proteinase production. Conclusions Bloodstream isolates of C. albicans produce virulence factors such as GT and hydrolytic enzymes that enable them to cause infection under favorable conditions. .

[Susceptibility of Candida albicans blood isolates to 3 antifungal drugs: retrospective study in Rio Grande do Sul, Brazil, 1999-2009]. / Sensibilidad de aislamientos de Candida albicans de hemocultivos a 3 fármacos: estudio retrospectivo en Rio Grande do Sul, Brasil, 1999 a 2009.

BACKGROUND: Candidiasis is one of the most important among recurrent invasive yeast infections in patients, thus antifungal treatment becomes a challenge. AIMS: The aim of this study was to evaluate the in vitro activity of clinical Candida albicans isolates from blood cultures to fluconazole, amphotericin B and anidulafungin, in a hospital from Rio Grande do Sul, Brazil. METHODS: The susceptibility of 153 isolates to the 3 drugs mentioned was tested according to Clinical and Laboratory Standars Institute. Minimal inhibitory and fungicidal concentrations (MIC, MFC, respectively) of each drug were determined, as well as the epidemiological cutoff value (ECV). RESULTS: All of the isolates were susceptible to anidulafungin, MIC and MFC ≤ 1 µg/ml; however, when compared with ECV, 3% of the isolates exhibited higher values against fluconazole, 96% were susceptible, 3% susceptible dose-dependent, and 1% resistant. Also, it was observed that 21% of the isolates exhibited higher values than ECV. One isolate was resistant to amphotericin B; the other ones, susceptible, based on the MFC; furthermore, 1.5% of the isolates exhibited higher values. CONCLUSIONS: C. albicans isolates exhibited more susceptibility to anidulafungin, and 90% of them (MIC90) exhibited the lowest values against amphotericin B. Based on ECV and Pfaller classification, isolates could be resistant to fluconazole, demonstrating the importance of the combination of these parameters.

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.

Inhibitory effect of sodium hypochlorite and chlorhexidine digluconate in clinical isolates of Sporothrix schenckii.

The susceptibility of Sporothrix schenckii isolates from clinical cases of canine, feline and human sporotrichosis, and from the environment, was evaluated with 4% sodium hypochlorite and 6.6% chlorhexidine digluconate using the broth microdilution, agar diffusion and direct exposure techniques. The minimal inhibitory concentration was smaller than 0.8% for chlorhexidine digluconate and between 8% and 4% for sodium hypochlorite. Inhibition zones were not found in agar diffusion for sodium hypochlorite, and zones averaging 1.9 mm were found for chlorhexidine digluconate. In the direct exposure test, sodium hypochlorite demonstrated best performance at 20 min of contact, as chlorhexidine digluconate presented little antimicrobial activity.

Epidemiological findings and laboratory evaluation of sporotrichosis: a description of 103 cases in cats and dogs in southern Brazil.

Sporotrichosis is a subcutaneous mycosis, which affects mainly small animals, and is considered an important public health disease. This paper describes the epidemiological and laboratory characteristics of 103 clinical cases of sporotrichosis diagnosed over a 10-year period in southern Brazil. The 92 cats and 11 dogs from eight municipalities in Rio Grande do Sul State developed especially the disseminated cutaneous and fixed cutaneous forms of the disease. Respiratory signs such as sneezing, serous nasal discharge and dyspnea were found in about 57% of the animals. The detection of Sporothrix schenckii in different clinical samples showed highest isolation in testicles (46.6%), oral cavity (45.2%) and conjunctival mucosa (38.1%). A differentiated histological pattern was found between the fixed cutaneous and disseminated cutaneous (DC) manifestations of the disease; well-organized granulomas of nodular distribution and various fungal structures prevailed in the DC form in cats. Melanin detection in S. schenckii cells by the Fontana-Masson technique was positive in 45.4% of the samples. The study revealed that the State of Rio Grande do Sul is an endemic sporotrichosis area and demonstrated the possibility of involvement of other pathways in the infection and spread of the disease. In addition, it emphasized the importance of laboratory tests for mycosis confirmation, especially in dogs that develop clinical manifestations without the presence of cutaneous lesions.

Viabilidade de Candida albicans estocada em água destilada estéril a 20C / Viability of Candida albicans stored in sterile distilled water at 20 degrees C

Esse trabalho teve como objetivo avaliar o método de conservação de leveduras do gênero Candida estocadas em água destilada estéril a baixas temperaturas. Utilizaram-se 240 cepas de Candida albicans estocadas em água destilada estéril a - 20 graus C, durante o período de seis meses a oito anos. As cepas foram subcultivadas em duplicata em ágar de Sabouraud e incubadas a 35 graus C por 96 horas. Houve recuperação de 68,4% das 240 leveduras estocadas, sendo que a maior porcentagem de viabilidade foi referente ao período de seis meses e a menor foi de três anos. O método utilizado para o armazenamento das leveduras possui vantagens quando comparado a outros, além de ocupar pouco espaço, é facilmente executado e possui baixo custo.

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.

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.

Esporotricose óssea e cutânea em canino / Cutaneos and osseous sporotrichosis in dog

A esporotricose é uma micose subcutânea causada pelo fungo dimórfico Sporothrix schenckii. Este artigo descreve o primeiro caso de esporotricose óssea e cutânea, em canino, na cidade de Pelotas, Rio Grande do Sul. O animal apresentava lesões ulceradas e crostosas, há aproximadamente três anos no plano nasal e membro torácico direito, dispnéia e apatia. Para confirmação do diagnóstico, foram realizados exames micológico, histopatológico, radiológico e hematológico. O animal foi tratado durante três meses com 10mg/kg de itraconazol,por via oral, obtendo-se a cura das lesões. Este estudo alerta clínicos de pequenos animais para a ocorrência desta micose em caninos na região de Pelotas, RS.

The sporotrichosis is a subcutaneous mycosis caused by dimorphic fungus Sporothrix schenckii. This article describes the first cutaneous and osseous sporotrichosis case in canine in the Pelotas city, Rio Grande do Sul. The animal presented crusts and ulcerated lesions,approximately the three years in the nasal plan and right hind-foot,dispneic and apathy. Mycological, histopathological, radiological and hematological diagnosis was realized. The animal was treated by three months with itraconazole, administered orally at a dosage of 10mg/kg, until lesions disappeared. This study alert small animals clinicians for the occurrence of this mycosis in dogs in the region of Pelotas, RS.