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Abstract In the current context of emerging drug-resistant fungal pathogens such as Candida albicans and Candida parapsilosis, discovery of new antifungal agents is an urgent matter. This research aimed to evaluate the antifungal potential of 2-chloro-N-phenylacetamide against fluconazole-resistant clinical strains of C. albicans and C. parapsilosis. The antifungal activity of 2-chloro-N-phenylacetamide was evaluated in vitro by the determination of the minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC), inhibition of biofilm formation and its rupture, sorbitol and ergosterol assays, and association between this molecule and common antifungal drugs, amphotericin B and fluconazole. The test product inhibited all strains of C. albicans and C. parapsilosis, with a MIC ranging from 128 to 256 µg.mL-1, and a MFC of 512-1,024 µg.mL-1. It also inhibited up to 92% of biofilm formation and rupture of up to 87% of preformed biofilm. 2-chloro-N-phenylacetamide did not promote antifungal activity through binding to cellular membrane ergosterol nor it damages the fungal cell wall. Antagonism was observed when combining this substance with amphotericin B and fluconazole. The substance exhibited significant antifungal activity by inhibiting both planktonic cells and biofilm of fluconazole-resistant strains. Its combination with other antifungals should be avoided and its mechanism of action remains to be established.

Resumo No atual contexto de patógenos fúngicos resistentes emergentes tais como Candida albicans e Candida parapsilosis, a descoberta de novos agentes antifúngicos é uma questão urgente. Esta pesquisa teve como objetivo avaliar o potencial antifúngico da 2-cloro-N-fenilacetamida contra cepas clínicas de C. albicans e C. parapsilosis resistentes a fluconazol. A atividade antifúngica da substância foi avaliada in vitro através da determinação da concentração inibitória mínima (CIM), concentração fungicida mínima (CFM), ruptura e inibição da formação de biofilme, ensaios de sorbitol e ergosterol, e associação entre esta molécula e antifúngicos comuns, anfotericina B e fluconazol. O produto teste inibiu todas as cepas de C. albicans e C. parapsilosis, com uma CIM variando de 128 a 256 µg.mL-1, e uma CFM de 512-1,024 µg.mL-1. Também inibiu até 92% da formação de biofilme e causou a ruptura de até 87% de biofilme pré-formado. A 2-cloro-N-fenilacetamida não promoveu atividade antifúngica pela ligação ao ergosterol da membrana celular fúngica, tampouco danificou a parede celular. Antagonismo foi observado ao combinar esta substância com anfotericina B e fluconazol. A substância exibiu atividade antifúngica significativa ao inibir tanto as células planctônicas quanto o biofilme das cepas resistentes ao fluconazol. Sua combinação com outros antifúngicos deve ser evitada e seu mecanismo de ação deve ser estabelecido.

Resumo

In the current context of emerging drug-resistant fungal pathogens such as Candida albicans and Candida parapsilosis, discovery of new antifungal agents is an urgent matter. This research aimed to evaluate the antifungal potential of 2-chloro-N-phenylacetamide against fluconazole-resistant clinical strains of C. albicans and C. parapsilosis. The antifungal activity of 2-chloro-N-phenylacetamide was evaluated in vitro by the determination of the minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC), inhibition of biofilm formation and its rupture, sorbitol and ergosterol assays, and association between this molecule and common antifungal drugs, amphotericin B and fluconazole. The test product inhibited all strains of C. albicans and C. parapsilosis, with a MIC ranging from 128 to 256 µg.mL-1, and a MFC of 512-1,024 µg.mL-1. It also inhibited up to 92% of biofilm formation and rupture of up to 87% of preformed biofilm. 2-chloro-N-phenylacetamide did not promote antifungal activity through binding to cellular membrane ergosterol nor it damages the fungal cell wall. Antagonism was observed when combining this substance with amphotericin B and fluconazole. The substance exhibited significant antifungal activity by inhibiting both planktonic cells and biofilm of fluconazole-resistant strains. Its combination with other antifungals should be avoided and its mechanism of action remains to be established.

No atual contexto de patógenos fúngicos resistentes emergentes tais como Candida albicans e Candida parapsilosis, a descoberta de novos agentes antifúngicos é uma questão urgente. Esta pesquisa teve como objetivo avaliar o potencial antifúngico da 2-cloro-N-fenilacetamida contra cepas clínicas de C. albicans e C. parapsilosis resistentes a fluconazol. A atividade antifúngica da substância foi avaliada in vitro através da determinação da concentração inibitória mínima (CIM), concentração fungicida mínima (CFM), ruptura e inibição da formação de biofilme, ensaios de sorbitol e ergosterol, e associação entre esta molécula e antifúngicos comuns, anfotericina B e fluconazol. O produto teste inibiu todas as cepas de C. albicans e C. parapsilosis, com uma CIM variando de 128 a 256 µg.mL-1, e uma CFM de 512-1,024 µg.mL-1. Também inibiu até 92% da formação de biofilme e causou a ruptura de até 87% de biofilme pré-formado. A 2-cloro-N-fenilacetamida não promoveu atividade antifúngica pela ligação ao ergosterol da membrana celular fúngica, tampouco danificou a parede celular. Antagonismo foi observado ao combinar esta substância com anfotericina B e fluconazol. A substância exibiu atividade antifúngica significativa ao inibir tanto as células planctônicas quanto o biofilme das cepas resistentes ao fluconazol. Sua combinação com outros antifúngicos deve ser evitada e seu mecanismo de ação deve ser estabelecido.

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Inga cylindrica, a tropical fruit tree of the Fabaceae family (subfamily Mimosoideae), is native to South America. The seeds from this family are an essential source of trypsin inhibitors, which display promising bioactivity for increasing host defense against pathogens. The aim of the present study was to characterize the antimicrobial and antibiofilm activities of the trypsin inhibitor extracted from I. cylindrica seeds, ICTI. ICTI demonstrated antifungal activity with a minimum inhibitory concentration (MIC) of 32.11 µmol.L-1, and a minimum fungicidal concentration (MFC) of 32.1 µmol.L-1, against Cryptococcus gattii, Candida albicans, Candida glabrata and Candida guilliermondii. Combining ICTI with Amphotericin B had a significant synergistic effect, reducing the concentration of the antibiotic by 75% for C. albicans and 94% for C. gatti. The significant increase (16 x) in activity observed with ergosterol (1.01 mol.L-1) for C. albicans and C. gatti, and the lack of activity against bacterial strains, suggests that ICTI interferes with the integrity of the fungal cell membrane. Treatment with ICTI at 10 x MIC resulted in a 51% reduction in biofilm formation and a 56% decrease in mature biofilm colonies for C. albicans. Finally, ICTI displayed no toxicity in the in vivo Galleria mellonella model. Given its antifungal and antibiofilm properties, ICTI could be a promising candidate for the development of new antimicrobial drug prototypes.

Inga cylindrica, uma árvore frutífera tropical da família Fabaceae (subfamília Mimosoideae), é nativa da América do Sul. As sementes desta família são uma fonte essencial de inibidores de tripsina, que apresentam bioatividade promissora para aumentar a defesa do hospedeiro contra patógenos. O objetivo do presente estudo foi caracterizar as atividades antimicrobiana e antibiofilme do inibidor de tripsina extraído de sementes de I. cylindrica, ICTI. O ICTI demonstrou atividade antifúngica com concentração inibitória mínima (CIM) de 32,11 µmol.L-1 e concentração fungicida mínima (CFM) de 32,1 µmol.L-1, contra Cryptococcus gattii, Candida albicans, Candida glabrata e Candida guilliermondii. A combinação de ICTI com Anfotericina B teve um efeito sinérgico significativo, reduzindo a concentração do antibiótico em 75% para C. albicans e 94% para C. gatti. O aumento significativo (16 x) na atividade observado com ergosterol (1,01 mol.L-1) para C. albicans e C. gatti, e a falta de atividade contra cepas bacterianas, sugerem que o ICTI interfere na integridade da membrana celular fúngica . O tratamento com ICTI a 10 x MIC resultou numa redução de 51% na formação de biofilme e numa diminuição de 56% nas colónias maduras de biofilme para C. albicans. Finalmente, o ICTI não apresentou toxicidade no modelo in vivo de Galleria mellonella. Dadas as suas propriedades antifúngicas e antibiofilme, o ICTI pode ser um candidato promissor para o desenvolvimento de novos protótipos de medicamentos antimicrobianos.

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Pythium insidiosum é o agente etiológico da pitiose, sendo ele um microrganismo da classe dos oomicetos, patogênico e causador da pitiose em várias espécies animais, incluindo humanos. A doença é caracterizada por lesões no tecido cutâneo/subcutâneo, lesões gastrointestinais, vasculares, oculares, podendo se disseminar para órgãos internos. A forma cutânea gera granulomas exsudativos, sendo comum a formação de "kunkers", um material necrótico de coloração amarelada, apenas nos equídeos. O diagnóstico é feito através dos sinais clínicos e histórico do paciente, cultura e identificação do agente e alguns testes como os sorológicos (imunodifusão, ELISA, Western blot, hemaglutinação e teste imunocromatográfico), histopatológico, imunohistoquímica, testes moleculares e recentemente, análises proteômicas, como o MALDI ­TOF. O tratamento baseia-se na exérese cirúrgica da lesão com margens livre do patógeno, sendo apenas possível em regiões sem comprometimento de importantes estruturas anatômicas; uso de medicamentos como Anfotericina B e Iodeto de Potássio e a imunoterapia como forma de modular a resposta imune do hospedeiro. Não existe fator predisponente para a aquisição da doença, sendo o principal risco a permanência em águas estagnadas com presença de material vegetal. Dessa forma, a principal forma de infecção é evitar, quando possível, a manutenção por longos períodos de animais no interior de lagoas, rios/riachos ou açudes, bem como utilizar equipamentos de proteção individual em humanos quando necessitarem adentrar em tais ambientes.

Pythium insidiosumis the etiological agent of pythiosis. This microorganism is a pathogenic oomycete that causes disease in several animal species, such as horses, dogs, cattle, sheep, goats, cats, birds and also humans. It is a disease characterized by lesions in the cutaneous/subcutaneous tissues in the form of exudative granulomas, the formation of "kunkers" being common in horses. It can also cause damage in gastrointestinal tract, vascular, ocular, and disseminated to internal organs. The diagnostic methods are, in addition to clinical signs and patient history, culture and identification of the agent, serological tests (immunodiffusion, ELISA, Western blot, hemagglutination and immunochromatographic tests), histopathological, immunohistochemical, molecular tests and recently, proteomic analysis, like MALDI-TOF. The treatment is based on surgical exeresis of the lesion with margins free of the pathogen, and this treatment is only possible in regions without involviment of important anatomical structures; use of drugs such as Amphotericin B and Potassium Iodide and immunotherapy as a way to modulate the host immune response. There is no predisposing factor for acquiring the disease, the main risk being staying in stagnant water with the presence of plant material. Thus, the main form of infection is to avoid, when possible, keeping animals inside ponds, rivers/streams or dams for long periods, as well as using personal protective equipment on humans when they need to enter such environments.

Pythium insidiosumes el agente etiológico de la pitiosis. Este microorganismo es un oomicetopatógeno que causa enfermedad en varias especies animales, como caballos, perros, bovinos, ovinos, caprinos, felinos, aves y también humanos. Es una enfermedad caracterizada por lesiones en el tejido cutáneo/subcutáneo en forma de granulomas exudativos, siendo frecuente la formación de kunkers en los caballos. También puede causar lesiones gastrointestinales, vasculares y oculares y diseminarse a los órganos internos. Los métodos diagnósticos son, además de los signos clínicos y la historia del paciente, el cultivo e identificación del agente, las pruebas serológicas (inmunodifusión, ELISA, Western blot, hemaglutinación y prueba inmunocromatográfica), la histopatología, la inmunohistoquímica, las pruebas moleculares y, recientemente, el análisis proteómico,como el MALDI-TOF. El tratamiento se basa en la escisión quirúrgica de la lesión con márgenes libres de patógenos, y este tratamiento solo es posible en regiones sin compromiso de estructuras anatómicas importantes; uso de fármacos como la anfotericina B y el yoduro de potasio y la inmunoterapia como forma de modular la respuesta inmunitaria del huésped. No existe un factor predisponente para adquirir la enfermedad, siendo el principal riesgo permanecer en aguas estancadas con presencia de material vegetal. Así, la principal forma de infección es evitar, en lo posible, mantener a los animales dentro de estanques, ríos/arroyos o represas por períodos prolongados, así como usar equipo de protección personal en humanos cuando necesitan ingresar a dichos ambientes.

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The aim of the current report is to describe a cutaneous candidiasis case affecting a canine individual treated at the University Veterinary Hospital of State University of Maranhão (UEMA), in São Luís City. The patient had three-month history of skin diseases; it had been previously subjected to several treatments based on antibiotics, corticosteroids and antifungal drugs that have failed to show clinical improvements. Dermatological assessment has indicated generalized moist dermatitis, intense skin desquamation, alopecia, pruritus and meliceric crusts along the animal's body, mainly in its dorsal region. Complementary tests, such as skin cytology and microscopy, trichogram, qualitative PCR and serology for canine visceral leishmaniasis, as well as fungal culture and antifungigram were requested based on this scenario. Serology recorded inconclusive results for leishmaniasis, whereas PCR recorded negative results in the presence of the agent's DNA. Cytology, microscopy and trichogram results have evidenced fungal infection in the assessed samples. Moreover, mycological culture and antifungigram resulted in the growth of Candida sp. specimens capable of resisting antifungal agents such as amphotericin B, fluconazole, itraconazole and nystatin. The therapy adopted after candidiasis diagnosis confirmation comprised oral doses of manipulated ketoconazole, in combination to topical therapy with shampoo based on moisturizing formulas associated with Miconazole and Chlorhexidine (at 2%), for four weeks. After 30 days, when the adopted therapy was over, the aforementioned animal presented remission of the previously observed lesions and fully improved condition.

The aim of the current report is to describe a cutaneous candidiasis case affecting a canine individual treated at the University Veterinary Hospital of State University of Maranhão (UEMA), in São Luís City. The patient had three-month history of skin diseases; it had been previously subjected to several treatments based on antibiotics, corticosteroids and antifungal drugs that have failed to show clinical improvements. Dermatological assessment has indicated generalized moist dermatitis, intense skin desquamation, alopecia, pruritus and meliceric crusts along the animal's body, mainly in its dorsal region. Complementary tests, such as skin cytology and microscopy, trichogram, qualitative PCR and serology for canine visceral leishmaniasis, as well as fungal culture and antifungigram were requested based on this scenario. Serology recorded inconclusive results for leishmaniasis, whereas PCR recorded negative results in the presence of the agent's DNA. Cytology, microscopy and trichogram results have evidenced fungal infection in the assessed samples. Moreover, mycological culture and antifungigram resulted in the growth of Candida sp. specimens capable of resisting antifungal agents such as amphotericin B, fluconazole, itraconazole and nystatin. The therapy adopted after candidiasis diagnosis confirmation comprised oral doses of manipulated ketoconazole, in combination to topical therapy with shampoo based on moisturizing formulas associated with Miconazole and Chlorhexidine (at 2%), for four weeks. After 30 days, when the adopted therapy was over, the aforementioned animal presented remission of the previously observed lesions and fully improved condition.

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Background: Cryptoccocal meningitis continues to present high incidence among AIDS patients. The treatment of choice is the synergistic combination of flucytosine (5-FC) with amphotericin B deoxycholate (AmBd) or its lipid formulations. However, 5-FC is unavailable in many countries and AmB demands hospitalization. The combination of AmB with the fungistatic fluconazole (FLC) or the use of high FLC daily doses alone became the choice. Nonetheless, sterilization of cerebrospinal fluid is delayed with FLC monotherapy, mainly with high fungal burden. These findings suggest the search for new antifungal compounds, such as liriodenine. Methods: Liriodenine antifungal activity was evaluated by three procedures: determining the minimum inhibitory concentration (MIC) on 30 strains of the Cryptococcus neoformans (C. neoformans) complex and 30 of the Cryptococcus gattii (C. gattii) complex, using EUCAST methodology and amphotericin B deoxycholate as control; performing the time-kill methodology in two strains of the C. neoformans complex and one of the C. gattii complex; and injury to cryptococcal cells, evaluated by transmission electron microscopy (TEM). Liriodenine absorption and safety at 0.75 and 1.50 mg.kg-1 doses were evaluated in BALB/c mice. Results: Liriodenine MICs ranged from 3.9 to 62.5 µg.mL-1 for both species complexes, with no differences between them. Time-kill methodology confirmed its concentration-dependent fungicidal effect, killing all the strains below the limit of detection (33 CFU.mL-1) at the highest liriodenine concentration (32-fold MIC), with predominant activity during the first 48 hours. Liriodenine induced severe Cryptococcus alterations ­ cytoplasm with intense rarefaction and/or degradation, injury of organelles, and presence of vacuoles. Liriodenine was better absorbed at lower doses, with no histopathological alterations on the digestive tract. Conclusion: The fungicidal activity confirmed by time-kill methodology, the intense Cryptococcus injury observed by TEM, the absorption after gavage administration, and the safety at the tested doses indicate that the liriodenine molecule is a promising drug lead for development of anticryptococcal agents.(AU)

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ABSTRACT: Studies on the fungal microbiota of reptiles and amphibians are necessary to better understand of host-microbe interactions and the establishment of fungal disease in these animals. However, these studies are limited. The present researchidentified yeasts from free-ranging reptiles and amphibians from the Caatinga biome andevaluated the virulence factors production, the antifungal susceptibility in planktonic and biofilm growth and the pathogenicity of Candida famata isolates. Twenty-nine isolates of the genera Candida, Cryptococcus and Rhodotorula were identified by phenotypic and/or molecular methods and production of hydrolytic enzymes in vitro by these genera of fungi was evaluated. In addition, susceptibility of planktonic cells and biofilms to azoles and amphotericin B was evaluated. The pathogenicity of C. famata, the most prevalent yeast species isolated, was evaluated using Caenorhabditis elegans model. C. famata was the most prevalent yeast in amphibian and reptilian microbiota. Phospholipase and protease production was observed in 18/29 and 11/29 of the yeast isolates, respectively, while 100% formed biofilms. Itraconazole presented high minimal inhibitory concentrations against C. famata and C. tropicalis. Amphotericin B reduced the biomass and metabolic activity of biofilms. C. famata induced the mortality of C. elegans. In conclusion, reptiles and amphibians are colonized by yeasts capable of producing important virulence factors, especially by Candida spp. that present low susceptibility to azoles which may result from imbalances in ecosystem. Finally, C. famata isolated from these animals presented high pathogenicity, showing the importance of the study of reptile and amphibians fungal microbiota.

RESUMO: Estudos sobre a microbiota fúngica de répteis e anfíbios são necessários para melhor compreender as interações hospedeiro-microrganismo e o estabelecimento de doenças fúngicas nesses animais. No entanto, esses estudos são limitados. O objetivo da presente pesquisa foi identificar leveduras isoladas de répteis e anfíbios do bioma Caatinga e avaliar a produção de fatores de virulência, a sensibilidade a antifúngicos no crescimento planctônico e de biofilme e a patogenicidade de Candida famata. Vinte e nove isolados dos gêneros Candida, Cryptococcus e Rhodotorula foram identificados por métodos fenotípicos e/ou moleculares e a produção de enzimas hidrolíticas in vitro por esses gêneros de fungos foi avaliada. Além disso, foi avaliada a suscetibilidade de células planctônicas e biofilmes a azólicos e anfotericina B. A patogenicidade de C. famata, a espécie de levedura isolada mais prevalente, foi avaliada usando Caenorhabditis elegans. C. famata foi a levedura mais prevalente na microbiota de anfíbios e répteis. A produção de fosfolipase e protease foi observada em 18/29 e 11/29 dos isolados de levedura, respectivamente, enquanto 100% formaram biofilmes. O itraconazol apresentou altas concentrações inibitórias mínimas contra C. famata e C. tropicalis. A anfotericina B reduziu a biomassa e atividade metabólica dos biofilmes. C. famata induziu a mortalidade de C. elegans. Em conclusão, répteis e anfíbios são colonizados por leveduras capazes de produzir importantes fatores de virulência, especialmente por cepas de Candida spp. que apresentam baixa suscetibilidade a azólicos que podem resultar de desequilíbrio no ecossistema. Por fim, C. famata isolados desses animais apresentaram alta patogenicidade, mostrando a importância do estudo da microbiota fúngica de répteis e anfíbios.

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La leishmaniasis visceral canina (LVC) es una zoonosis de extrema importancia en la salud pública, en todo el mundo, con el 90% de los casos registrados en Brasil.  Es causada por el protozoario género Leishmania y su principal huésped es el perro doméstico y, para que se produzca la transmisión, es obligatoria la presencia del vector, el mosquito hematófago género Lutzomyia. El diagnóstico de la enfermedad es bastante difícil, ya que del 60 al 80% de los animales seropositivos son asintomáticos y las manifestaciones clínicas son muy inespecíficas.  Después de los cambios realizados por el Ministerio de Salud, actualmente está permitido llevar a cabo el tratamiento de animales positivos para LVC, en el que el medicamento más aceptado es Miltefosina, sin embargo, independientemente del tratamiento, el mejor método contra LVC es la prevención.  En el presente trabajo, se produjo un cuestionario a través de la plataforma Google Forms con 14 preguntas que cubren la leishmaniasis.  Se recogieron 65 respuestas de veterinarios de diferentes edades y estados de Brasil.  El 80% de los veterinarios dijeron que indicaban el tratamiento del animal en caso de leishmaniasis confirmada, mientras que el 3.08% todavía indica eutanasia.  Aunque no se recomienda, aún se observa el uso de anfotericina B y antimoniales pentavalentes en...

The canine visceral leishmaniasis (CVL) is a zoonosis extremely important for public health, of worldwide relevance, with 90% of the cases registered in Brazil. It´s caused by the protozoan from the genus Leishmania and its main host is the domestic dog. For the successful transmission, it must have the presence of the vector, a hematophagous mosquito from genus Lutyzomia. The diagnostic from the disease is extremely difficult, seen that 60% to 80% of the seropositive animals are asymptomatic and the clinical manifestations are very nonspecific. After changes made by the Health Ministry, it actually is permitted to realize the treatment of the positive animals to CVL, in which the most accepted medicament is Miltefosine. Yet, independent of the treatment, the best method against CVL is prevention. In the present job was produced a questionnaire through the platform Google Forms with 14 questions encompassing the Leishmaniasis. Were collected 65 answers veterinarians of different ages and states of Brazil.  80% of vets said to indicate the treatment of the animal in cases of leishmaniasis confirmed, while 3.08% still indicate euthanasia. Although not recommended, the use of amphotericin B and pentavalent antimonials is still seen in the treatment protocols, however, the majority (75.4%) opts for the use of Miltefosina, recommended. Vaccination, although accept...

A Leishmaniose visceral canina (LVC) é uma zoonose de extrema importância na saúde pública, de distribuição mundial, com 90% dos casos registrados no Brasil. É causada pelo protozoário gênero Leishmania e tem como principal hospedeiro o cão doméstico. Para que ocorra a transmissão, é obrigatória a presença do vetor, mosquito hematófago Lutzomyia sp. O diagnóstico da doença é bastante difícil, já que 60 a 80% dos animais soropositivos são assintomáticos e as manifestações clínicas são muito inespecíficas. Após mudanças realizadas pelo Ministério da Saúde, atualmente é permitido realizar o tratamento dos animais positivos para LVC, no qual o medicamento mais aceito é a miltefosina, porém, independente do tratamento, o melhor método contra a LVC é a prevenção. No presente trabalho foi produzido um questionário através da plataforma Google Forms com 14 perguntas englobando a Leishmaniose. Foram coletadas 65 respostas de médicos veterinários de diferentes idades e estados do Brasil.  80% dos veterinários disseram indicar o tratamento do animal em caso de leishmaniose confirmada, enquanto 3,08% ainda indicam a eutanásia. Embora não recomendados, ainda se nota o uso de anfotericina B e antimoniais pentavalentes nos protocolos de tratamento, porém a maioria (75,4%) opta pelo uso da miltefosina, recomendada. A vacinação, embora aceita pela maioria dos veterinários (95,38%)...

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Susceptibility testing is essential to inform the correct management of Aspergillus infections. In this study we present antifungal susceptibility profile of A. fumigatus isolates recovered from lungs of birds with and without aspergillosis. Fifty three isolates were tested for their antifungal susceptibility to voriconazole (VRC), itraconazole (ITZ), amphotericin (AMB) and caspofungin (CSP) using the M38-A2 broth microdilution reference method. Five isolates were resistant to more than one antifungal drug (CSP + AMB, VRC + ITZ and AMB + ITZ). Fifteen (28%) isolates with susceptible increased exposure (I) to ITZ were sensible to VRC. Resistance to AMB (>2µg/mL) was observed in only four isolates. Eleven (21%) A. fumigatus present resistance to ITZ (13%) and VRC (8%). Fungal isolation from respiratory samples has been regarded as being of limited usefulness in the ante mortem diagnosis of aspergillosis in birds. However, the results suggest that the detection and antifungal susceptibility profile may be helpful for monitoring of therapy for avian species and where antifungal resistance might be emerging and what conditions are associated to the event.(AU)

Os testes de suscetibilidade são essenciais para informar o correto manejo das infecções por Aspergillus. Neste estudo apresentamos o perfil antifúngico de isolados de A. fumigatus provenientes de pulmões de aves com e sem aspergilose. Cinqüenta e três isolados foram testados quanto à susceptibilidade antifúngica ao voriconazol (VRC), itraconazol (ITZ), anfotericina B (AMB) e caspofungina (CSP) pelo método de referência de microdiluição do caldo M38-A2. Cinco isolados foram resistentes a mais de um antifúngico (CSP + AMB, VRC + ITZ e AMB + ITZ). Quinze (28%) isolados suscetíveis - com exposição aumentada (I) ao ITZ foram sensíveis ao VRC. A resistência ao AMB (>2µg/mL) foi observada em apenas quatro isolados. Onze (21%) A. fumigatus apresentaram resistência a ITZ (13%) e VRC (8%). O isolamento de fungos de amostras respiratórias tem sido considerado de utilidade limitada no diagnóstico ante mortem de aspergilose em aves. No entanto, os resultados sugerem que a detecção e o perfil de suscetibilidade a antifúngicos podem ser úteis para o monitoramento da terapia de espécies aviárias, assim como a emergência da resistência antifúngica e quais condições podem estar associadas ao evento.(AU)

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A analgesia com opioides é limitada ou indefinida em ruminantes. Neste trabalho, objetivou-se a realização de um estudo comparativo entre dois analgésicos opioides: a morfina e o tramadol, com base nas avaliações clínica e pedométrica de animais submetidos a artrite e sinovite experimental transitórias, desenvolvidas na articulação interfalângica distal, após administração intra-articular de anfotericina B. Utilizou-se seis animais, em dois tratamentos distintos, com morfina, na dose de 0,5 mg/kg e 20 dias depois com tramadol, na dose de 1,8 mg/kg, ambos via intramuscular. Os animais foram avaliados em intervalos de três horas, num total de 27 horas, observando-se parâmetros fisiológicos, deambulação e atividade pedométrica. Claudicação e alterações pedométricas foram observadas para ambos os fármacos. Por meio das variáveis da atividade pedométrica, observou-se um padrão de inquietação compatível com nocicepção podal, não havendo a interferência dos fármacos sobre a claudicação. Concluiu-se que a morfina e o tramadol, nas doses testadas, foram incapazes de interferir na atenuação do grau de claudicação no momento de máxima estimulação dolorosa, frente ao modelo experimental de dor ortopédica.

The use of opioids for analgesia is limited or undefined in ruminants. The aim of this study was to compare two analgesics opioids, morphine and tramadol, based on the clinical and pedometric evaluations of animals submitted to a temporary experimental arthritis and synovitis, developed at the distal interphalangeal joint after an intra-articular administration of amphotericin B. Six animals were used in two different treatments, using morphine, dose of 0,5 mg/kg, and twenty days later, tramadol, dose of 1,8 mg/kg, both intramuscularly. The animals were evaluated at 3-hour intervals for a total of 27 hours, observing the physiological parameters, ambulation and pedometer activity. Lameness and pedometer variations were observed in both treatments. Based on the variables of pedometric activity, a pattern of restlessness compatible with podal nociception was observed, and there was no interference of any of the drugs on lameness. It was concluded that morphine and tramadol, at the tested doses, were incapable of interfering in the lameness attenuation during the maximum painful stimulation moment in this experimental protocol of orthopedic pain.

Resumo

Abstract Background: Three drugs - pentavalent antimonials, amphotericin B and pentamidine - are currently used for leishmaniasis treatment. They are administered for long periods, only parenterally, and have high cardiac, renal and hepatic toxicities. Therefore, the investigation of new compounds is required. Nitro-heterocyclic derivatives have been used as possible drug candidates to treat diseases caused by trypanosomatids. Methods: Leishmania (L.) amazonensis promastigotes (MHO/BR/73/M2269), maintained in the Laboratório de Soroepidemiologia - Instituto de Medicina Tropical- USP, were exposed to five nitroheterocyclic derivatives, with differences at phenyl-ring position 4: BSF-C4H9, BSF-H, BSF-NO2, BSF-CH3 and BSF-Cl, for 48 hours. After analyzing viability (MTT assay), we evaluated cellular-morphology activity of compounds by transmission electron microscopy (TEM) and measurement of apoptosis (phosphatidylserine expression) by flow cytometry. Results: EC50 of amphotericin B and BSF-CH3 were 0.50 (M and 0.39 (M respective. Other nitro-heterocyclic compounds presented EC50 higher than amphotericin B. All compounds showed greater AV- and PI-positive expression than amphotericin B at 100 (M, except BSF-NO2. TEM showed complete nuclear disfigurement with 100 (M of BSF-NO2, 25 and 6.25 (M of BSF-H, and 6.25 (M BSF-Cl; presence of vesicles within the flagellar pocket with 25 (M BSF-H; alteration of the kinetoplast with 25 (M BSF-C4H9, 25 (M of BSF-H, 6.25 (M BSF-CH3 and 6.25 (M of BSF-Cl. Conclusions: Nitro-heterocyclic compounds have shown activity against promastigotes of L. amazonensis, at lower concentrations. However, improvement of compound scaffolds are needed to assist the elucidation of the mechanism of action and to achieve greater activity.

Resumo

Background: Deep fungal infections of the orbit and nasal passages causing rhinitis and ulcerative keratomycosis areuncommonly reported in cats. Hyalohyphomycetes and phaeohyphomycetes have rarely been associated with this disorder.Sino-orbital fungal diseases are emerging and more invasive than sino-nasal fungal diseases with poor response to therapyand a worse prognosis. Brachycephalic feline breeds seem to be at increased risk for development of upper respiratoryfungal diseases. Diagnosis is based on the demonstration of fungal hyphae by cytology or histology and definitive confirmation by fungal culture and molecular methods. This is the first case report of a cat with clinical mixed fungal ball withAspergillus and Scopulariopsis in Brazil.Case: A 3-year-old male Persian cat, in São José city, Santa Catarina, Brazil, was presented with exophthalmos and corneal ulcer of the left eye and protrusion, hyperemia, quemosis and fibroses of the left third eyelid. The retropulsion of theglobe was negative in this eyeball and a presumptive diagnosis of a retrobulbar mass was made. The patient underwenta surgical procedure for inspection and collection of samples for bacterial and mycological culture. Culture revealed nobacterial growth, however, unique and abundant growth of Aspergillus spp. was present. A subconjunctival enucleation ofthe left eye was made and the mass was sent for histopathology examination. Histology showed inflammatory proliferativenecrotizing pyogranulomatous reaction; with the presence of severe fungal infection evidenced by large number of hyalineseptated regular and irregular mold hyphae. Molecular identification was performed using panfungal primers (ITS3-F /ITS4-R). Patient was treated with systemic itraconazole associated with amphotericin B and topical clotrimazole. A massstarted to grow rapidly in the left pterygopalatine fossa and was surgically removed, but recurrence occurred seven daysafter...

Resumo

Abstract This study compares patients with and without non-viral microbial keratitis in relation to sociodemographic variables, clinical aspects, and involved causative agent. Clinical aspects, etiology and therapeutic procedures were assessed in patients with and without keratitis that were diagnosed in an Eye Care Center in Campo Grande, MS, Brazil. Patients were divided into two groups: (a) cases: 64 patients with non-viral microbial keratitis diagnosed at biomicroscopy; and (b) controls: 47 patients with other eye disorders that were not keratitis. Labor activity related to agriculture, cattle raising, and contact lens use were all linked to keratitis occurrence (p 0.005). In patients with keratitis, the most common symptoms were pain and photophobia, and the most frequently used medicines were fourth-generation fluoroquinolones (34.4%), amphotericin B (31.3%), and natamycin (28.1%). Microbial keratitis evolved to corneal perforation in 15.6% of cases; transplant was indicated in 10.9% of cases. Regarding the etiology of this condition, 23 (42.2%) keratitis cases were caused by bacteria (Pseudomonas aeruginosa, 12.5%), 17 (39.1%) by fungi (Fusarium spp., 14.1% and Aspergillus spp., 4.7%), and 4 (6.3%) by Acanthamoeba. Patients with keratitis present with a poorer prognosis. Rapid identification of the etiologic agent is indispensable and depends on appropriate ophthalmological collection and microbiological techniques.

Resumo

Abstract Amphotericin B is a fungicidal substance that is treatment of choice for most systemic fungal infections affecting immunocompromised patients. However, severe side effects have limited the utility of this drug. The aim of this study was to evaluate the antifungal effect of the combination of amphotericin B with black tea or white tea and protective of citotoxic effect. The present study shows that white and black teas have additive effects with amphotericin B against some species Candida. In addition, the combination of white and black tea with amphotericin B may reduce the toxicity of amphotericin B to red blood cells. Our results suggest that white and black tea is a potential agent to combine with amphotericin for antifungal efficacy and to reduce the amphotericin dose to lessen side effects.

Resumo A anfotericina B é o tratamento de escolha para a maioria das infecções fúngicas sistémicas que afetam os doentes imunocomprometidos. No entanto, efeitos secundários graves têm limitado a utilidade desta droga. O objetivo deste estudo foi avaliar o efeito antifúngico da combinação de anfotericina B com chá preto ou chá branco, bem como o efeito citotóxico desta combinação sobre hemáceas. O presente estudo demonstra que o chá branco e preto de Camellia sinensis têm efeitos aditivos com anfotericina B contra algumas espécies de Candida sp. Além disso, a combinação de chá branco e preto com anfotericina B pode reduzir a toxicidade da anfotericina B em hemáceas. Nossos resultados sugerem que o chá branco e preto são agentes potenciais para associação com anfotericina B contribuindo para eficácia antifúngica, bem como redução de toxicidade.

Resumo

Background: Histoplasmosis is a systemic mycosis whose etiologic agent is the fungus Histoplasma capsulatum. This fungal infection, which is the second most frequent systemic mycotic fungal disease in felines in the United States, has rarely been found in cats in Brazil. This paper reports on a case of acute pulmonary histoplasmosis in a domestic cat treated with oral itraconazole associated with amphotericin B administered subcutaneously. This treatment resulted in clinical remission of the patient’s symptoms, as evidenced by radiographic follow-ups.Case: A domestic cat suffering from acute dyspnea was taken to a veterinary clinic. The animal was subjected to emergency oxygen therapy, and kept at rest through sedation with midazolam. A physical examination revealed normally colored mucosa, 8% dehydration, bristly fur, body condition score 2/9, tachypnea with respiratory rate of 100 breaths per minute and expiratory dyspnea. The radiographic examination showed marked opacification of all the pulmonary fields, with a mixed pattern (interstitial and alveolar) of heterogeneous appearance and diffuse distribution, which are changes consistent with an inflammatory infectious process (pneumonia). A cytological analysis of the pleural fluid revealed round to oval-shaped intracytoplasmic structures, varying in size from 2 to 4 μm, inside foamy macrophages, consistent with Histoplasma capsulatum. Based on the diagnosis of pulmonary histoplasmosis, and in view of the patient’s acute respiratory distress, it was decided to treat the cat using itraconazole associated with amphotericin B. Itraconazole was administered orally at a dose of 100 mg/cat every 24 h, while amphotericin B was administered subcutaneously at a dose of 0.5 mg/kg, combined with 100 mL of sodium chloride 0.9% and 100 mL of 5% glycated serum, with monitoring of serum concentrations of symmetric dimethylarginine (SDMA).[...]

Resumo

Abstract Yeast infections have acquired great importance due to increasing frequency in immunocompromised patients or patients undergoing invasive diagnostic and therapeutic techniques, and also because of its high morbidity and mortality. At the same time, it has been seen an increase in the emergence of new pathogenic species difficult to diagnose and treat. The aim of this study was to determine the in vitro susceptibility of 89 yeasts from different sources against the antifungals amphotericin B, voriconazole, fluconazole and flucytosine, using the VITEK® 2 Compact system. The antifungal susceptibility was performed automatically by the Vitek® 2 Compact system. The origin of the yeasts was: Group 1 - microbiota of wild animals (W) (26/89), 2 - cow's milk with subclinical mastitis (M) (27/89) and 3 - hospital enviorment (H) (36/89). Of the 89 yeasts submitted to the Vitek® 2 test, 25 (20.9%) were resistant to fluconazole, 11 (12.36%) to amphotericin B, 3 (3.37%) to voriconazole, and no sample was resistant to flucytosine. Regarding the minimum inhibitory concentration (MIC), fluconazole showed an MIC between 1 and 64 mg/mL for the three groups, voriconazole had an MIC between 0.12 and 8 mg/mL, amphotericin B had an MIC between 0.25 and 4 mg/mL for group H and group W respectively, between 0.25 and 16 mg/mL for group M and flucytosine had an MIC equal to 1μg/mL for all groups. The yeasts isolated from the H group showed the highest resistance to fluconazole 12/89 (13.49%), followed by group W (7.87%) and group M (5.62%). The more resistant group to voriconazole was followed by the M and H groups, the W group showed no resistance to this antifungal. Group H was the least resistant (2.25%) to amphotericin.

Resumo As infecções por leveduras têm adquirido grande importância, devido ao aumento da sua frequência em pacientes imunocomprometidos ou pacientes submetidos a técnicas diagnosticas e terapêuticas agressivas, e devido sua alta morbidade e mortalidade. Paralelamente tem-se observado um incremento na aparição de novas espécies patógenas difíceis de diagnosticar e tratar. O objetivo desse estudo foi avaliar a suscetibilidade in vitro de 89 leveduras de diferentes origens frente aos antifúngicos Anfotericina B, Voriconazol, Fluconazol e Fluocitocina pelo Sistema Vitek® 2. O antifungigrama foi realizado automaticamente pelo Vitek® 2 Compact. A origem das leveduras foi: Grupo 1- Microbiota de Animais Silvestres (S) (26/89), 2- Leite com mastite bovina subclínica (L) (27/89) e 3- Ambiente Hospitalar (H) (36/89). Das 89 leveduras submetidas à carta Vitek®, 25 (20.09%) foram resistentes ao fluconazol, oito (8.99%) à anfotericina B, três (3.37%) ao voriconazol, e nenhuma amostra mostrou-se resistente a fluocitosina. O grupo três (H) foi mais resistente ao fluconazol que os demais, já o dois (L) foi mais resistente ao voriconazol e a anfotericina B que os outros dois. O fluconazol pode ter apresentado maior número de resistências devido ser um fármaco comumente usado principalmente em humanos. As leveduras isoladas de humanos apresentaram maior número de resistências aos fármacos testados do que as leveduras isoladas de animais silvestres. O que pode ocorrer devido a uma maior exposição dos humanos aos fármacos em relação aos animais que vivem isolados em ambientes selvagens e na maioria dos casos nunca teve contato com fármacos de qualquer origem.

Resumo

The therapeutic arsenal for the treatment of Leishmaniasis is limited and includes toxic compounds (antimonials, amphotericin B, pentamidine and miltefosine). Given these aspects, the search for new compounds based on floristic biodiversity is crucial. In the present work, we report the isolation, characterization and antileishmanial activity of six related neolignans (1­6) of bioactive extract from Nectandra leucantha (Lauraceae) twigs. Methods: Dried and powdered twigs of N. leucantha were exhaustively extracted using n-hexane. The crude extract was dereplicated by HPLC/HRESIMS and subjected to column chromatography to yield pure compounds 1­6. Their chemical structures were identified via NMR and comparison of obtained data with those previously published in the literature. Biological assays of compounds 1­6 and their respective monomers (eugenol and methyleugenol) were performed using promastigote and amastigote forms of Leishmania (L.) infantum. Results: Dereplication procedures followed by chemical characterization of isolated compounds by NMR enabled the identification of related neolignans 1­6. Neolignans 2, 4 and 6 showed potential against amastigote forms of L. (L.) infantum (EC50 values of 57.9, 67.7 and 13.7 µM, respectively), while compounds 1 and 3 were inactive. As neolignans 2­4 are chemically related, it may be suggested that the presence of the methoxyl group at C4 constitutes an important structural aspect to increase antileishmanial potential against amastigote forms. Compound 6, which consists of a methylated derivative of compound 5 (inactive) showed antileishmanial activity similar to that of the standard drug miltefosine (EC50 =16.9 µM) but with reduced toxicity (SI = 14.6 and 7.2, respectively). Finally, two related monomers, eugenol and methyleugenol, were also tested and did not display activity, suggesting that the formation of dimeric compounds by oxidative coupling is crucial for antiparasitic activity of dimeric compounds 2, 4 and 6. Conclusion: This study highlights compound 6 against L. (L.) infantum amastigotes as a scaffold for future design of new compounds for drug treatment of visceral leishmaniasis.(AU)

Resumo

Background: Sporotrichosis is caused by pathogenic fungi of the genus Sporothrix. The clinically relevant species are S. schenckii, S. globosa and S. brasiliensis. In Brazil, S. brasiliensis is the most prevalent etiological agent among humans and cats. In cats with sporotrichosis, skin lesions are mainly characterized by nodules and ulcers, usually located in the head, nasal region and limbs. The presence of respiratory signs concomitantly with cutaneous lesions is frequent, especially sneezing, and may be associated with lesions located in the nasal mucosa. Ketoconazole (KTZ), itraconazole (ITZ), potassium iodide (KI), sodium iodide (NaI), terbinafine (TRB), fluconazole (FLZ) and amphotericin B (AMB) are the drugs currently available for treating feline sporotrichosis. ITZ remains the drug of choice. ITZ combined with KI has been successfully used in the treatment of naïve cats (especially cases with lesions in the nasal region), cases of recurrence and refractory to ITZ. Clinical cure with NaI has been described in some cases, but its use has been limited by adverse reactions. The conventional formulation is the saturated solution and the recommended dose in the treatment of feline sporotrichosis is 10 mg/kg every 12 h. Cats are sensitive to iodide preparations and should be carefully monitored for clinical evidence of iodism, such as apathy, anorexia, vomiting, diarrhea, hypothermia, hyperthermia, cardiomyopathy, hyperexcitability, muscular spasms and ptyalism. The purpose of this study was to evaluate the therapeutic response of NaI capsules in feline sporotrichosis.Materials, Methods & Results: An observational cohort study was conducted in cats with sporotrichosis at the Laboratory of Clinical Research in Dermatozoonoses in Domestic Animals (Lapclin-Dermzoo), Evandro Chagas National Institute of Infectious Diseases (INI)/Oswaldo Cruz Foundation (Fiocruz), Rio de Janeiro, Brazil.[...]

Resumo

Abstract This study compares patients with and without non-viral microbial keratitis in relation to sociodemographic variables, clinical aspects, and involved causative agent. Clinical aspects, etiology and therapeutic procedures were assessed in patients with and without keratitis that were diagnosed in an Eye Care Center in Campo Grande, MS, Brazil. Patients were divided into two groups: (a) cases: 64 patients with non-viral microbial keratitis diagnosed at biomicroscopy; and (b) controls: 47 patients with other eye disorders that were not keratitis. Labor activity related to agriculture, cattle raising, and contact lens use were all linked to keratitis occurrence (p < 0.005). In patients with keratitis, the most common symptoms were pain and photophobia, and the most frequently used medicines were fourth-generation fluoroquinolones (34.4%), amphotericin B (31.3%), and natamycin (28.1%). Microbial keratitis evolved to corneal perforation in 15.6% of cases; transplant was indicated in 10.9% of cases. Regarding the etiology of this condition, 23 (42.2%) keratitis cases were caused by bacteria (Pseudomonas aeruginosa, 12.5%), 17 (39.1%) by fungi (Fusarium spp., 14.1% and Aspergillus spp., 4.7%), and 4 (6.3%) by Acanthamoeba. Patients with keratitis present with a poorer prognosis. Rapid identification of the etiologic agent is indispensable and depends on appropriate ophthalmological collection and microbiological techniques.

Resumo

Abstract Amphotericin B is a fungicidal substance that is treatment of choice for most systemic fungal infections affecting immunocompromised patients. However, severe side effects have limited the utility of this drug. The aim of this study was to evaluate the antifungal effect of the combination of amphotericin B with black tea or white tea and protective of citotoxic effect. The present study shows that white and black teas have additive effects with amphotericin B against some species Candida. In addition, the combination of white and black tea with amphotericin B may reduce the toxicity of amphotericin B to red blood cells. Our results suggest that white and black tea is a potential agent to combine with amphotericin for antifungal efficacy and to reduce the amphotericin dose to lessen side effects.

Resumo A anfotericina B é o tratamento de escolha para a maioria das infecções fúngicas sistémicas que afetam os doentes imunocomprometidos. No entanto, efeitos secundários graves têm limitado a utilidade desta droga. O objetivo deste estudo foi avaliar o efeito antifúngico da combinação de anfotericina B com chá preto ou chá branco, bem como o efeito citotóxico desta combinação sobre hemáceas. O presente estudo demonstra que o chá branco e preto de Camellia sinensis têm efeitos aditivos com anfotericina B contra algumas espécies de Candida sp. Além disso, a combinação de chá branco e preto com anfotericina B pode reduzir a toxicidade da anfotericina B em hemáceas. Nossos resultados sugerem que o chá branco e preto são agentes potenciais para associação com anfotericina B contribuindo para eficácia antifúngica, bem como redução de toxicidade.