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Histoplasma capsulatum (H. capsulatum) is a thermal-dimorphic fungus, the causal agent of histoplasmosis. Its presence in the environment is related with chicken manure due to their high nitrogen and phosphorus content. In Colombia, chicken manure is the most used raw material in the composting process; however, there is no information about the capacity of H. capsulatum to survive and remain viable in a composted organic fertilizer. To address this question, this study shows three assays based on microbiological culture and the Hc100 nested PCR. First, a composting reactor system was designed to transform organic material under laboratory conditions, and the raw material was inoculated with the fungus. From these reactors, the fungus was not isolated, but its DNA was detected. In the second assay, samples from factories where the DNA of the fungus was previously detected by PCR were analyzed. In the raw material samples, 3 colonies of H. capsulatum were isolated and its DNA was detected. However, after the composting process, neither the fungus was recovered by culture nor DNA was detected. In the third assay, sterilized and nonsterilized organic composted samples were inoculated with H. capsulatum and then evaluated monthly during a year. In both types of samples, the fungus DNA was detected by Hc100 nested PCR during the whole year, but the fungus only grew from sterile samples during the first two months evaluated. In general, the results of the assays showed that H. capsulatum is not able to survive a well-done composting process.

In order to assess the antifungal activity of methanolic extracts from neem tree (Azadirachta indica A. Juss.), several bioassays were conducted following M38-A2 broth microdilution method on 14 isolates of the dermatophytes Trichophyton mentagrophytes, Trichophyton rubrum, Microsporum canis and Epidermophyton floccosum. Neem extracts were obtained through methanol-hexane partitioning of mature green leaves and seed oil. Furthermore, high performance liquid chromatography (HPLC) analyses were carried out to relate the chemical profile with their content of terpenoids, of widely known antifungal activity. The antimycotic Terbinafine served as a positive control. Results showed that there was total growth inhibition of the dermatophytes isolates at minimal inhibitory concentrations (MIC) between 50 μg/mL and 200 μg/mL for leaves extract, and between 625 μg/mL and 2500 μg/mL for seed oil extract. The MIC of positive control (Terbinafine) ranged between 0.0019 μg/mL and 0.0313 μg/mL. Both neem leaves and seed oil methanol extracts exhibited different chromatographic profiles by HPLC, which could explain the differences observed in their antifungal activity. This analysis revealed the possible presence of terpenoids in both extracts, which are known to have biological activity. The results of this research are a new report on the therapeutic potential of neem to the control of dermatophytosis.Keywords: HPLC, neem, microbial sensitivity tests, minimum inhibitory concentration, terpenoids. RESUMENSe determinó la actividad antifúngica de extractos metanólicos de la especie Azadirachta indica A. Juss. (Meliaceae), conocida comúnmente como neem, empleando el método de microdilución en caldo M38-A2 de referencia para hongos filamentosos y dermatofitos. Se evaluaron 14 aislamientos de los dermatofitos Trichophyton mentagrophytes, Trichophyton rubrum, Microsporum canis y Epidermophyton floccosum. Los extractos de neem fueron obtenidos mediante partición metanol-hexano a partir de aceite de semillas y hojas verdes maduras. Adicionalmente, se analizaron por cromatografía líquida de alta resolución (CLAR) con el fin de relacionar su perfil químico con el contenido de terpenoides, de conocida actividad antifúngica. Se empleó como control positivo el antimicótico Terbinafina. Los resultados mostraron inhibición total del crecimiento de los aislamientos de dermatofitos a concentraciones mínimas inhibitorias (CMI) entre 50 μg/mL y 200 μg/mL para el extracto de hojas y entre 625 μg/mL y 2500 μg/mL para el extracto de aceite de semillas. La CMI encontrada para el control positivo (Terbinafina) fluctuó entre 0,0078 μg/mL y 0,0313 μg/mL. Los extractos metanólicos de hojas y aceite de semillas de neem exhibieron diferentes perfiles cromatográficos en CLAR, lo cual podría explicar las diferencias observadas en su actividad antifúngica. Éste análisis químico reveló la posible presencia de compuestos INTRODUCTIONThe research about neem has been focused not only on its known antifeedant activity on insects, but also on its antifung...

ResumenLas líneas celulares de neem (Azadirachta indica A. Juss.) cultivadas en suspensión líquida han demostrado producir metabolitos secundarios bioactivos, particularmente triterpenoides. En consecuencia, se han realizado estudios para el control de microorganismos de importancia médica, como los hongos dermatofitos. El objetivo principal de este trabajo fue evaluar a través de un método de referencia in vitro la actividad antifúngica de diferentes extractos de cultivos celulares de neem sobre varios aislamientos de Trichophyton mentagrophytes, Trichophyton rubrum y Epidermophyton floccosum. Se realizó un escalado de cultivos de suspensiones celulares de neem, a partir de los cuales se obtuvo un extracto crudo metanólico. Éste extracto fue fraccionado posteriormente por cromatografía en columna de silica gel. Con los extractos obtenidos se determinó la Concentración Mínima Inhibitoria (CMI), siguiendo el método de microdilución en caldo M38-A2, con cinco aislamientos de T. mentagrophytes, cinco de T. rubrum y tres de E. floccosum. Se usó como control positivo el antimicótico Terbinafina. Los resultados mostraron que el extracto crudo de biomasa celular de neem inhibe el crecimiento hasta en 100 % de T. mentagrophytes, T. rubrum y E. floccosum. Al evaluar las fracciones por separado, se observó que las de menor polaridad exhibieron en general mayor actividad antifúngica (CMI=109 µg/mL) que el extracto crudo per se (CMI=2500 µg/ mL) y las fracciones más polares (CMI=7000 µg/mL). Lo anterior indica que las células de neem cultivadas en suspensión producen compuestos con actividad antifúngica, siendo más bioactivos los presentes en las fracciones de menor polaridad.Palabras clave: método de microdilución en caldo, cromatografía, cultivo de células en suspensión, concentración mínima inhibitoria, compuestos bioactivos. AbstractCell lines of neem (Azadirachta indica A. Juss.) grown in liquid suspension have shown to produce bioactive secondary metabolites, particularly triterpenoids. In consequence, its use as a control of medical microorganisms (like dermatophytes) is proposed. The main goal of this study was to assess the antifungal activity of methanolic extracts from neem cultured cell suspensions on several isolates of Trichophyton mentagrophytes (five isolates), Trichophyton rubrum (five isolates) and Epidermophyton floccosum (three isolates). Neem cell suspension cultures were scaled up, from which a raw methanolic extract was obtained. This extract was fractionated by silica gel column chromatography. The raw methanolic extract and its fractions were used in order to determine the Minimal Inhibitory Concentration (MIC) on the dermatophytes isolates by following M38-A2 broth microdilution method. Antimycotic Terbinafine was used as positive control. The results shown that neem raw cellular biomass extract inhibits the growth of T. mentagrophytes, T. rubrum and E. floccosum in at least 100%. In the evaluations of the separated fractions, it was observed that the low polarity fractions had higher antifungal ac...