Evaluation of antifungal activity, mechanisms of action and toxicological profile of the synthetic amide 2-chloro-N-phenylacetamide.

Aspergillus niger causes infections such as otitis and pulmonary aspergillosis in immunocompromised individuals. Treatment involves voriconazole or amphotericin B, and due to the increase in fungal resistance, the search for new compounds with antifungal activity has intensified. In the development of new drugs, cytotoxicity and genotoxicity assays are important, as they allow predicting possible damage that a molecule can cause, and in silico studies predict the pharmacokinetic properties. The aim of this study was to verify the antifungal activity and the mechanism of action of the synthetic amide 2-chloro-N-phenylacetamide against Aspergillus niger strains and toxicity. 2-Chloro-N-phenylacetamide showed antifungal activity against different strains of Aspergillus niger with minimum inhibitory concentrations between 32 and 256 µg/mL and minimum fungicides between 64 and 1024 µg/mL. The minimum inhibitory concentration of 2-chloro-N-phenylacetamide also inhibited conidia germination. When associated with amphotericin B or voriconazole, 2-chloro-N-phenylacetamide had antagonistic effects. Interaction with ergosterol in the plasma membrane is the probable mechanism of action.2-Chloro-N-phenylacetamide has favorable physicochemical parameters, good oral bioavailability and absorption in the gastrointestinal tract, crosses the blood-brain barrier and inhibits CYP1A2. At concentrations of 50 to 500 µg/mL, it has little hemolytic effect and a protective effect for type A and O red blood cells, and in the cells of the oral mucosa it promotes little genotoxic change. It is concluded that 2-chloro-N-phenylacetamide has promising antifungal potential, favorable pharmacokinetic profile for oral administration and low cytotoxic and genotoxic potential, being a promising candidate for in vivo toxicity studies.

Cytotoxicity and Biological Activities of Geopopolis Extract from the Stingless bee (Melipona scutellaris) in Isolates of Staphylococcus aureus.

Geopropolis resins are produced by stingless bees (Meliponinae), developed from the collection of resinous materials, waxes and exudates, from the flora of the region where stingless bees are present, in addition to the addition of clay or earth in its composition. Several biological activities are attributed to Ethanol Extracts of Geopropolis (EEGP). The bioactive properties are associated with the complex chemical composition that the samples have. This work aims to evaluate the biological activities of the EEGP, in order to contribute with a natural therapeutic alternative, to face infections, mainly those caused by resistant strains of Staphylococcus aureus. The EEGP MIC tests showed antibacterial activity against two strains of S. aureus, both at concentrations of 550 µg/mL. The MBC performed with the inhibition values showed that the EEGP has bacteriostatic activity in both strains. Biofilm inhibition rates exhibited an average value greater than 65 % at the highest concentration. The EEGP antioxidant potential test showed good antioxidant activity (IC50) of 11.05±1.55 µg/mL. In the cytotoxicity test against HaCat cells, after 24 hours, EEGP induced cell viability at the three tested concentrations (550 µg/mL: 81.68±3.79 %; 1100 µg/mL: 67.10±3.76 %; 2200 µg/mL: 67.40±1.86 %). In view of the above, the safe use of EEGP from the brazilian northeast could be proven by the cytotoxicity test, and its use as an antioxidant and antibacterial agent has proven to be effective, as an alternative in combating oxidative stress and microorganisms such as S. aureus, which, through the spread and ongoing evolution of drug resistance, generates an active search for effective solutions.

Docking Prediction, Antifungal Activity, Anti-Biofilm Effects on Candida spp., and Toxicity against Human Cells of Cinnamaldehyde.

OBJECTIVE: This study evaluated the antifungal activity of cinnamaldehyde on Candida spp. In vitro and in situ assays were carried out to test cinnamaldehyde for its anti-Candida effects, antibiofilm activity, effects on fungal micromorphology, antioxidant activity, and toxicity on keratinocytes and human erythrocytes. Statistical analysis was performed considering α = 5%. RESULTS: The minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of cinnamaldehyde ranged from 18.91 µM to 37.83 µM. MIC values did not change in the presence of 0.8 M sorbitol, whereas an 8-fold increase was observed in the presence of ergosterol, suggesting that cinnamaldehyde may act on the cell membrane, which was subsequently confirmed by docking analysis. The action of cinnamaldehyde likely includes binding to enzymes involved in the formation of the cytoplasmic membrane in yeast cells. Cinnamaldehyde-treated microcultures showed impaired cellular development, with an expression of rare pseudo-hyphae and absence of chlamydoconidia. Cinnamaldehyde reduced biofilm adherence by 64.52% to 33.75% (p < 0.0001) at low concentrations (378.3-151.3 µM). Cinnamaldehyde did not show antioxidant properties. CONCLUSIONS: Cinnamaldehyde showed fungicidal activity through a mechanism of action likely related to ergosterol complexation; it was non-cytotoxic to keratinocytes and human erythrocytes and showed no antioxidant activity.

Antifungal Activity of Essential Oils against Candida albicans Strains Isolated from Users of Dental Prostheses.

OBJECTIVE: The objective of this study was to analyze the antifungal activity of citral, selected by screening natural products, against Candida albicans isolates from subjects who use dental prostheses. METHODOLOGY: Screening of essential oils, including those from Mentha piperita L. (Briq), Origanum vulgare, and Zingiber officinale L., and the phytoconstituents citral and limonene, to select an appropriate natural product. Citral, which mediated the best antifungal response, was selected for biological assays. The minimum inhibitory concentrations (MICs) and minimum fungicidal concentrations (MFCs) for citral and nystatin were determined by the microdilution method. Micromorphological analyses, time-kill curve, and modulation tests were performed. RESULTS: The MIC and MFC of citral were established as 32 µg/mL, consistent with fungicidal activity. The clinical strains were resistant to nystatin. Citral caused micromorphological alteration in the strains. In the time-kill curve, the growth of the clinical strain was reduction in growth equal to 3 log10 colony-forming units per milliliter after exposure to the MIC and MIC × 2 of citral for 2 h. Citral did not modulate the resistance of the studied strains to nystatin. CONCLUSION: This study revealed the potential of citral as a fungicidal agent and highlighted the resistance of clinical strains of C. albicans to nystatin.

Estudo do potencial antifúngico e do mecanismo de ação do timol contra cepas de Candida parapsilosis resistentes ao fluconazol e a anfotericina B / Study of the antifungal potential and mechanism of action of thymol against strains of Candida parapsilosis resistant to fluconazole and amphotericin B

Objetivo: investigar a suscetibilidade de cepas fúngicas de Candida parapsilosis isoladas de sangue humano frente ao timol, bem como seu mecanismo de ação. Metodologia: foram utilizadas técnicas de microdiluição em placas de 96 poços para determinar a concentração inibitória mínima (CIM) e concentração fungicida mínima (CFM). Além disso, foram realizados testes com o sorbitol e o ergosterol para investigar a ação do timol na parede e na membrana celular fúngica respectivamente. Resultados: nos testes de CIM e CFM, foi observado que as cepas de C. parapsilosis são resistentes ao fluconazol e a anfotericina B, no entanto, o timol desempenhou efeito fungicida com razão CFM/CIM entre 1 e 2. Além disso, a CIM do timol não aumentou quando o sorbitol ou o ergosterol foi adicionado no meio, sugerindo fortemente que este monoterpeno não age na parede celular fúngica ou por ligação ao ergosterol na membrana plasmática. Conclusão: portanto, esses resultados contribuem para a elucidação do mecanismo de ação do timol, sugerindo outros possíveis alvos de interação fármaco-receptor. No entanto, mais investigações de caráter enzimático e molecular em modelos in vitro são necessários para que se possa elucidar completamente o modo de ação desse promissor monoterpeno.

Objective: to investigate the susceptibility of fungal strains of Candida parapsilosis isolated from human blood against thymol, as well as its mechanism of action. Methodology: microdilution techniques were used in 96-well plates to determine minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC). In addition, tests were performed with sorbitol and ergosterol to investigate the action of thymol on the wall and on the fungal cell membrane respectively. Results: in the CIM and CFM tests, it was observed that C. parapsilosis strains are resistant to fluconazole and amphotericin B, however, thymol had a fungicidal effect with MFC/MIC ratio between 1 and 2. In addition, thymol MIC did not increase when sorbitol or ergosterol was added in the medium, strongly suggesting that this monoterpene does not act on the fungal cell wall or by binding to ergosterol on the plasma membrane. Conclusion: therefore, these results contribute to the elucidation of the mechanism of action of thymol, suggesting other possible targets of drug-receptor interaction. However, further investigations of enzymatic and molecular character in in vitro models are necessary to fully elucidate the mode of action of this promising monoterpene.

Evaluation of Antifungal Activity and Mode of Action of New Coumarin Derivative, 7-Hydroxy-6-nitro-2H-1-benzopyran-2-one, against Aspergillus spp.

Aspergillus spp. produce a wide variety of diseases. For the treatment of such infections, the azoles and Amphotericin B are used in various formulations. The treatment of fungal diseases is often ineffective, because of increases in azole resistance and their several associated adverse effects. To overcome these problems, natural products and their derivatives are interesting alternatives. The aim of this study was to examine the effects of coumarin derivative, 7-hydroxy-6-nitro-2H-1-benzopyran-2-one (Cou-NO2), both alone and with antifungal drugs. Its mode of action against Aspergillus spp. Cou-NO2 was tested to evaluate its effects on mycelia growth and germination of fungal conidia of Aspergillus spp. We also investigated possible Cou-NO2 action on cell walls (0.8 M sorbitol) and on Cou-NO2 to ergosterol binding in the cell membrane. The study shows that Cou-NO2 is capable of inhibiting both the mycelia growth and germination of conidia for the species tested, and that its action affects the structure of the fungal cell wall. At subinhibitory concentration, Cou-NO2 enhanced the in vitro effects of azoles. Moreover, in combination with azoles (voriconazole and itraconazole) Cou-NO2 displays an additive effect. Thus, our study supports the use of coumarin derivative 7-hydroxy-6-nitro-2H-1-benzopyran-2-one as an antifungal agent against Aspergillus species.

Evaluation of Antifungal Activity and Mechanism of Action of Citral against Candida albicans.

Candida albicans is a yeast that commensally inhabits the human body and can cause opportunistic or pathogenic infections. Objective. To investigate the antifungal activity of citral against C. albicans. Methodology. The minimum inhibitory concentration (MIC) and the minimum fungicidal concentration (MFC) were determined by the broth microdilution techniques. We also investigated possible citral action on cell walls (0.8 M sorbitol), cell membranes (citral to ergosterol binding), the time-kill curve, and biological activity on the yeast's morphology. Results. The MIC and MFC of citral were, respectively, 64 µg/mL and 256 µg/mL. Involvement with the cell wall and ergosterol binding were excluded as possible mechanisms of action. In the morphological interference assay, it was observed that the product inhibited pseudohyphae and chlamydoconidia formation. The MIC and the MFC of citral required only 4 hours of exposure to effectively kill 99.9% of the inoculum. Conclusion. Citral showed in vitro antifungal potential against strains of C. albicans. Citral's mechanism of action does not involve the cell wall or ergosterol, and further study is needed to completely describe its effects before being used in the future as a component of new antifungals.

Distribution of dermatophytes from soils of urban and rural areas of cities of Paraiba State, Brazil.

The dermatophytes, keratinophilic fungi, represent important microorganisms of the soil microbiota, where there are cosmopolitan species and others with restricted geographic distribution. The aim of this study was to broaden the knowledge about the presence of dermatophytes in soils of urban (empty lots, schools, slums, squares, beaches and homes) and rural areas and about the evolution of their prevalence in soils of varying pH in cities of the four mesoregions of Paraiba State, Brazil. Soil samples were collected from 31 cities of Paraiba State. Of 212 samples, 62% showed fungal growth, particularly those from the Mata Paraibana mesoregion (43.5%), which has a tropical climate, hot and humid. Soil pH varied from 4.65 to 9.06, with 71% of the growth of dermatophytes occurring at alkaline pH (7.02 - 9.06) (ρ = 0.000). Of 131 strains isolated, 57.3% were geophilic species, particularly Trichophyton terrestre (31.3%) and Mycrosporum gypseum (21.4%). M. nanum and T. ajelloi were isolated for the first time in Paraiba State. The zoophilic species identified were T. mentagrophytes var. mentagrophytes (31.3 %) and T. verrucosum (7.6 %), and T. tonsurans was isolated as an anthropophilic species. The soils of urban areas including empty lots, schools, slums and squares of cities in the mesoregions of Paraiba State were found to be the most suitable reservoirs for almost all dermatophytes; their growth may have been influenced by environmental factors, soils with residues of human and/or animal keratin and alkaline pH.

Increasing antibiotic activity against a multidrug-resistant Acinetobacter spp by essential oils of Citrus limon and Cinnamomum zeylanicum.

The genus Acinetobacter has gained importance in recent years due to involvement in serious infections and antimicrobial resistance. Many plants have been evaluated not only for direct antimicrobial activity, but also as resistance modifying agents. The Essential oil of Citrus limon (EOCL) addition at 156.25 µgmL(-1) (MIC/8) sub-inhibitory concentration in the growth medium led to MIC decrease for amikacin, imipenem and meropenem. The Essential oil of Cinnamomum zeylanicum (EOCZ) addition at 78.125 µg mL(-1) (MIC/8) sub-inhibitory concentrations in the growth medium caused drastic MIC reduction of amikacin. Results of combining antibiotics and essential oils had shown us a synergistic effect with both essential oils/amikacin combinations. An additive effect was observed with the combinations of both essential oils and gentamicin. The results of this study suggest that essential oil of C. limon and C. zeylanicum may suppress the growth of Acinetobacter species and could be a source of metabolites with antibacterial modifying activity.

Distribution ofddermatophytes from soils of urbn and rural areas of cities of paraiba state, Brazil / Distribuicao de dermatofitos isolados de solos de cidades do Estado da Paraiba, Brasil

SUMMARY The dermatophytes, keratinophilic fungi, represent important microorganisms of the soil microbiota, where there are cosmopolitan species and others with restricted geographic distribution. The aim of this study was to broaden the knowledge about the presence of dermatophytes in soils of urban (empty lots, schools, slums, squares, beaches and homes) and rural areas and about the evolution of their prevalence in soils of varying pH in cities of the four mesoregions of Paraiba State, Brazil. Soil samples were collected from 31 cities of Paraiba State. Of 212 samples, 62% showed fungal growth, particularly those from the Mata Paraibana mesoregion (43.5%), which has a tropical climate, hot and humid. Soil pH varied from 4.65 to 9.06, with 71% of the growth of dermatophytes occurring at alkaline pH (7.02 - 9.06) (ρ = 0.000). Of 131 strains isolated, 57.3% were geophilic species, particularly Trichophyton terrestre (31.3%) and Mycrosporum gypseum (21.4%). M. nanum and T. ajelloi were isolated for the first time in Paraiba State. The zoophilic species identified were T. mentagrophytes var. mentagrophytes (31.3 %) and T. verrucosum (7.6 %), and T. tonsurans was isolated as an anthropophilic species. The soils of urban areas including empty lots, schools, slums and squares of cities in the mesoregions of Paraiba State were found to be the most suitable reservoirs for almost all dermatophytes; their growth may have been influenced by environmental factors, soils with residues of human and/or animal keratin and alkaline pH. .

RESUMO Os dermatófitos, fungos queratinofílicos, representam importantes microrganismos da microbiota do solo, onde existem espécies cosmopolitas e outras de distribuição geográfica restrita. Este estudo teve como objetivo ampliar o conhecimento da distribuição de dermatófitos do solo proveniente de áreas urbanas (terrenos baldios, escolas, favelas, praças, praias e residências) e rurais de quatro mesorregiões paraibanas e da influência do pH na adaptação desse grupo de fungos. Amostras de solos urbanos e rurais foram coletadas de 31 cidades do estado da Paraíba, Brasil. De 212 amostras 62% apresentaram crescimento fúngico, destacando-se a Mesorregião da Mata Paraibana (43.5%), a qual apresenta clima tropical, quente e úmido. O pH das amostras de solo variou de 4.65 a 9.06, com crescimento de 71% dos dermatófitos em pH alcalino (7.02 - 9.06) (ρ = 0.000). Das 131 cepas isoladas 57.3% eram espécies geofílicas, destacando-se Trichophyton terrestre (31.3%) e Microsporum gypseum (21.4%). M. nanum e T. ajelloi foram isolados pela primeira vez no estado da Paraíba. Entre as espécies zoofílicas foram identificadas T. mentagrophytes var. mentagrophytes (31.3%) e T. verrucosum (7.6%) e como espécie antropofílica foi isolada T. tonsurans. Os solos de terrenos baldios, escolas, favelas e praças de cidades paraibanas são os reservatórios mais adequados dos dermatófitos, cujo crescimento pode ter sido influenciado por fatores ambientais, solos com resíduos de queratina humana e ou animal e pH alcalino. .

Actividad antifúngica del aceite esencial de Eugenia caryophyllata sobre cepas de Candida tropicalis de aislados clínicos / Antifungal activity of the essential oil of Eugenia caryophyllata on Candida tropicalis strains from clinical isolates

Candidiasis is an opportunistic fungal infection caused by Candida yeasts. In Brazil, C. tropicalis is the second most frequently isolated microorganism after C. albicans. The arising of strains resistant to conventional antifungal agents has increased the search for new alternatives from natural products, especially essential oils. This research investigated essential oil activity against strains of C. tropicalis by disk diffusion method. The minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) were also determinate. In the disk diffusion, the essential oils of Cinnamomum zeylanicum, Eugenia caryophyllata and Origanum vulgare had the highest inhibition zones values. MIC and MFC values of E. caryophyllata essential oil were 512 and 1024 ug/mL, respectively. MIC and MFC amphotericin B values were identical (2 ug/mL). Therefore, it was concluded that E. caryophyllata essential oil has strong antifungal activity and may be subject to further studies.

La candidiasis es una infección fúngica oportunista causada por levaduras del género Candida. En Brasil, la especie C. tropicalis esta siendo aislada frecuentemente, es el segundo microorganismo más aislado después de C. albicans. La aparición de cepas resistentes a los antifúngicos convencionales ha aumentado la búsqueda de nuevas alternativas provenientes de productos naturales, especialmente los aceites esenciales. En este estudio se investigó la actividad de los aceites esenciales contra las cepas de C. tropicalis, utilizando el método de difusión en disco, la concentración inhibitoria mínima (CIM) y la concentración fungicida mínima (CFM). En el método de difusión en disco, con los aceites esenciales de Cinnamomum zeylanicum, Eugenia caryophyllata y Origanum vulgare se obtuvieron mayores valores de inhibición. La CIM y CFM del aceite esencial de Eugenia caryophyllata fueron 512 y 1024 ug/mL, mientras que los de la anfotericina B fueron idénticos, 2 ug/mL. Por lo tanto, se puede concluir que el aceite esencial de E. caryophyllata tiene potente actividad antifúngica y puede ser objeto de nuevos estudios sobre esta actividad.