Phytochemical Analysis and Evaluation of Antifungal and Antioxidant Activities of Essential Oil of Fruits from Juniperus oxycedrus L. Obtained from Morocco

Abstract The present study was aimed at conducting phytochemical analysis and evaluating the in vitro antifungal and antioxidant activities of the essential oil obtained from the fruits of J. oxycedrus L. Hydro-distillation was used to extract the essential oil from the fruits of Juniper oxycedrus. The essential oil was analyzed using gas chromatography with a flame ionization detector (GC-FID) and gas chromatography coupled with mass spectrometry (GC/MS). The antioxidant activity of the essential oil against 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals was determined in vitro using varying concentrations of the essential oil and vitamin C as a standard antioxidant compound. A disc diffusion test was employed to evaluate the antifungal activity of the essential oil against two test fungal strains, Penicillium citrinum, and Aspergillus niger. The results revealed that 49 constituents were identified in fruit oil, representing 91.56% of the total oil and the yield was 1.58%. Juniper fruit oil was characterized by having high contents of ß-pinene (42.04%), followed by limonene (15.45%), sabinene (9.52%), α-pinene (5.21%), (E)-caryophyllene (3.77%), ρ-cymene (1.56%), caryophyllene oxide (2.02%), and myrcene (1.02%). The radical scavenging activity (% inhibition) of the essential oil was highest (81.87± 2.83%) at a concentration of 200 µg/mL. The essential oil of J. oxycedrus exhibited antifungal activity against A. niger and P. citrinum with minimum inhibitory concentration values (MIC) ranging from 2.89 to 85.01 µl/mL. The findings of the study reveal that the antioxidant and antifungal properties of J. oxycedrus essential oil and their chemical composition are significantly correlated

Identification of the target site of antimicrobial peptide AMP-17 against Candida albicans / 生物工程学报

Candida albicans is one of the major causes of invasive fungal infections and a serious opportunistic pathogen in immunocompromised individuals. The antimicrobial peptide AMP-17 has prominent anti-Candida activity, and proteomic analysis revealed significant differences in the expression of cell wall (XOG1) and oxidative stress (SRR1) genes upon the action of AMP-17 on C. albicans, suggesting that AMP-17 may exert anti-C. albicans effects by affecting the expression of XOG1 and SRR1 genes. To further investigate whether XOG1 and SRR1 genes were the targets of AMP-17, C. albicans xog1Δ/Δ and srr1Δ/Δ mutants were constructed using the clustered regulatory interspaced short palindromic repeats-associated protein 9 (CRISPR/Cas9) system. Phenotypic observations revealed that deletion of two genes had no significant effect on C. albicans growth and biofilm formation, whereas XOG1 gene deletion affected in vitro stress response and mycelium formation of C. albicans. Drug sensitivity assay showed that the MIC80 values of AMP-17 against xog1Δ/Δ and srr1Δ/Δ mutants increased from 8 μg/mL (for the wild type C. albicans SC5314) to 16 μg/mL, while the MIC80 values against srr1Δ/Δ: : srr1 revertants decreased to the level of the wild type SC5314. In addition, the ability of AMP-17 to inhibit biofilm formation of both deletion strains was significantly reduced compared to that of wild type SC5314, indicating that the susceptibility of the deletion mutants to AMP-17 was reduced in both the yeast state and during biofilm formation. These results suggest that XOG1 and SRR1 genes are likely two of the potential targets for AMP-17 to exert anti-C. albicans effects, which may facilitate further exploration of the antibacterial mechanism of novel peptide antifungal drugs.

Artemisinins inhibit oral candidiasis caused by Candida albicans through the repression on its hyphal development / 国际口腔科学杂志·英文版

Candida albicans is the most abundant fungal species in oral cavity. As a smart opportunistic pathogen, it increases the virulence by switching its forms from yeasts to hyphae and becomes the major pathogenic agent for oral candidiasis. However, the overuse of current clinical antifungals and lack of new types of drugs highlight the challenges in the antifungal treatments because of the drug resistance and side effects. Anti-virulence strategy is proved as a practical way to develop new types of anti-infective drugs. Here, seven artemisinins, including artemisinin, dihydroartemisinin, artemisinic acid, dihydroartemisinic acid, artesunate, artemether and arteether, were employed to target at the hyphal development, the most important virulence factor of C. albicans. Artemisinins failed to affect the growth, but significantly inhibited the hyphal development of C. albicans, including the clinical azole resistant isolates, and reduced their damage to oral epithelial cells, while arteether showed the strongest activities. The transcriptome suggested that arteether could affect the energy metabolism of C. albicans. Seven artemisinins were then proved to significantly inhibit the productions of ATP and cAMP, while reduced the hyphal inhibition on RAS1 overexpression strain indicating that artemisinins regulated the Ras1-cAMP-Efg1 pathway to inhibit the hyphal development. Importantly, arteether significantly inhibited the fungal burden and infections with no systemic toxicity in the murine oropharyngeal candidiasis models in vivo caused by both fluconazole sensitive and resistant strains. Our results for the first time indicated that artemisinins can be potential antifungal compounds against C. albicans infections by targeting at its hyphal development.

Expression of BmSPI38 tandem multimers in Escherichia coli and its antifungal activity / 生物工程学报

The aim of this study was to prepare tandem multimeric proteins of BmSPI38, a silkworm protease inhibitor, with better structural homogeneity, higher activity and stronger antifungal ability by protein engineering. The tandem multimeric proteins of BmSPI38 were prepared by prokaryotic expression technology. The effects of tandem multimerization on the structural homogeneity, inhibitory activity and antifungal ability of BmSPI38 were explored by in-gel activity staining of protease inhibitor, protease inhibition assays and fungal growth inhibition experiments. Activity staining showed that the tandem expression based on the peptide flexible linker greatly improved the structural homogeneity of BmSPI38 protein. Protease inhibition experiments showed that the tandem trimerization and tetramerization based on the linker improved the inhibitory ability of BmSPI38 to microbial proteases. Conidial germination assays showed that His6-SPI38L-tetramer had stronger inhibition on conidial germination of Beauveria bassiana than that of His6-SPI38-monomer. Fungal growth inhibition assay showed that the inhibitory ability of BmSPI38 against Saccharomyces cerevisiae and Candida albicans could be enhanced by tandem multimerization. The present study successfully achieved the heterologous active expression of the silkworm protease inhibitor BmSPI38 in Escherichia coli, and confirmed that the structural homogeneity and antifungal ability of BmSPI38 could be enhanced by tandem multimerization. This study provides important theoretical basis and new strategies for cultivating antifungal transgenic silkworm. Moreover, it may promote the exogenous production of BmSPI38 and its application in the medical field.

Antibacterial and antifungal activity of methanolic extracts of Salix alba L. against various disease causing pathogens / Atividade antibacteriana e antifúngica de extratos metanólicos de Salix alba L. contra vários patógenos que causam doenças

Abstract The present study was aimed to manifest the antibacterial and antifungal activity of methanolic extracts of Salix alba L. against seven Gram-positive and Gram-negative bacterial pathogens e.g. Streptococcus pyogenes, Staphylococcus aureus (1), S. aureus (2), Shigella sonnei, Escherichia coli (1), E. coli (2) and Neisseria gonorrhoeae and three fungal isolates from the air such as Aspergillus terreus, A. ornatus, and Rhizopus stolonifer. Two different serotypes of S. aureus and E. coli were used. The agar well-diffusion method results showed the dose-dependent response of plant extracts against bacterial and fungal strains while some organisms were found resistant e.g. E. coli (1), S. sonnei, A. terreus and R. stolonifer. The highest antibacterial activity was recorded at 17.000±1.732 mm from 100 mg/mL of leaves methanolic extracts against S. pyogenes while the activity of most of the pathogens decreased after 24 h of incubation. The highest antifungal activity was reported at 11.833±1.0 mm against A. ornatus at 50 mg/mL after 48 h of the incubation period. These experimental findings endorse the use of S. alba in ethnopharmacological formulations and suggest the use of methanolic extracts of the said plant to develop drugs to control the proliferation of resistant disease causing pathogenic microbes.

Resumo O presente estudo teve como objetivo manifestar a atividade antibacteriana e antifúngica de extratos metanólicos de Salix alba L. contra sete patógenos bacterianos Gram-positivos e Gram-negativos. Streptococcus pyogenes, Staphylococcus aureus (1), S. aureus (2), Shigella sonnei, Escherichia coli (1), E. coli (2) e Neisseria gonorrhoeae e três isolados de fungos do ar, como Aspergillus terreus, A. ornatus, e Rhizopus stolonifer. Dois sorotipos diferentes de S. aureus e E. coli foram usados. Os resultados do método de difusão em ágar mostraram a resposta dependente da dose de extratos de plantas contra cepas de bactérias e fungos, enquanto alguns organismos foram considerados resistentes, e.g. E. coli (1), S. sonnei, A. terreus e R. stolonifer. A maior atividade antibacteriana foi registrada em 17.000 ± 1.732 de 100 mg/mL de extratos metanólicos de folhas contra S. pyogenes, enquanto a atividade da maioria dos patógenos diminuiu após 24 h de incubação. A maior atividade antifúngica foi relatada em 11,833 ± 1,0 contra A. ornatus a 50 mg/mL após 48 h do período de incubação. Esses achados experimentais endossam o uso de S. alba em formulações etnofarmacológicas e sugerem o uso de extratos metanólicos da referida planta para o desenvolvimento de fármacos que controlem a proliferação de doenças resistentes que causam micróbios patogênicos.

Análisis fitoquímico preliminar y actividad antifúngica In vitro del extracto etanólico de las hojas de Solanum hispidum pers. Colectadas en la localidad Obraje - Perú / Preliminary phytochemical analysis and in vitro antifungal activity of the ethanolic extract of the leaves of Solanum hispidum pers. collected in the locality in Obraje - Peru

RESUMEN Objetivo . Analizar y determinar la actividad antifúngica in vitro del extracto etanólico de las hojas de Solanum hispidum Pers. Materiales y métodos . Se realizó el análisis fitoquímico preliminar cualitativo mediante reacciones de color y precipitación. Se investigó la actividad antifúngica in vitro frente a Candida albicans, Aspergillus brasilensis y Trichophyton mentagrophytes usando el método de difusión en pozo de agar y el ensayo de la concentración mínima inhibitoria (CMI). Resultados . El análisis fitoquímico preliminar cualitativo mostró la presencia de compuestos fenólicos, taninos, flavonoides, esteroides, alcaloides y saponinas. La actividad antifúngica in vitro fue demostrada para todos cultivos fúngicos con halos de inhibición entre 23 a 26 mm. Los valores de la CMI fueron de 125, 250 y 125 µg/mL para C. albicans, A. brasilensis y T. mentagrophytes, respectivamente. Conclusiones. El extracto etanólico de las hojas de Solanum hispidum Pers contiene importantes metabolitos secundarios y tiene moderada actividad antifúngica.

ABSTRACT Objective. To analyze and determine the in vitro antifungical activity of the ethanolic extract of the leaves of Solanum hispidum Pers. Materials and methods. We carried out a preliminary qualitative phytochemical analysis by color and precipitation reactions. We evaluated the in vitro antifungical activity against Candida albicans, Aspergillus brasilensis and Trichophyton mentagrophytes by using the agar well diffusion method and the minimum inhibitory concentration (MIC) assay. Results. Preliminary qualitative phytochemical analysis showed the presence of phenolic compounds, tannins, flavonoids, steroids, alkaloids and saponins. In vitro antifungal activity was demonstrated for all fungal cultures with inhibition halos between 23 to 26 mm. The MIC values were 125, 250, and 125 μg/mL for C. albicans, A. brasilensis, and T. mentagrophytes, respectively. Conclusions. The ethanolic extract of the leaves of Solanum hispidum Pers. contains important secondary metabolites and has moderate antifungical activity.

Estudio comparativo de la actividad anti-fúngica de extractos secuenciales de Fuchsia lycioides contra Candida sp / comparative study of the antifungal activity of sequencial extracts of Fuchsia lycioides against Candida sp

The genus Fuchsia is generally used in herbal preparations to treat conditions caused by microorganisms. Based on the popular use of this type of plants, the objective of this study was to obtain sequential extracts of increasing polarity from the branches of Fuchsia lycioides by maceration at room temperature and by the Soxhlet method at 60ºC, to later evaluate the antifungal capacity of the extracts against different clinical isolates of the Candida genus. The ethyl acetate extract exhibited strong anti-fungal activity, selectively inhibiting C. albicans strains with MIC and CMF values of 10 and 15 µg/mL, respectively; comparable with the drug itraconazole®. The analysis of the extract by GC-MS showed a high concentration of terpenoids (mainly phytol) and phenylpropanoids (mainly cinnamic acid), possibly responsible for the antifungal activity of the ethyl acetate extract of F. lycioides.

El género Fuchsia se usa generalmente en preparaciones de hierbas para tratar afecciones provocadas por microorganismos. En base al uso popular de este tipo de plantas, el objetivo de este estudio fue obtener los extractos secuenciales de polaridad creciente de las ramas de Fuchsia lycioides por maceración a temperatura ambiente y por el método Soxhlet a 60ºC, para luego evaluar la capacidad antifúngica de los extractos frente a diferentes aislados clínicos del genero Candida. El extracto de acetato de etilo exhibió una fuerte actividad antifúngica inhibiendo en forma selectiva las cepas de C. albicans con valores de CMI y de CMF de 10 y 15 µg/mL, respectivamente; comparables con el fármaco itraconazol®. El análisis del extracto por CG-EM mostró una alta concentración de terpenoides (principalmente fitol) y fenilpropanoides (principalmente ácido cinámico), posibles responsables de la actividad antifúngica del extracto de acetato de etilo de F. lycioides.

Phytochemical Analysis, Antioxidant and Antifungal Activity of Essential oil and Extracts of Alpinia malaccensis (Burm. f. ) Roscoe flowers

Abstract The present study describes chemical composition, phytochemicals, antifungal activities, antioxidant assays and total phenolic content of essential oil and varied polarity solvent extract from flowers of Alpinia malaccensis (Burm.f.). Total 27 components were identified in essential oil by GC-MS with terpinen-4-ol (28.6%) and α- terpineol (12.8%) as the main constituent. The essential oil was found to have maximal levels of phenolic content (64.60 µg/mL) as compared to the other extracts. The antioxidant assay evaluated in extracts and essential oil by different methods revealed good-to-moderate antioxidant potential with different IC50 values viz. (188.02 -250.25 µg/mL) in Fe3+ reducing power, (153.15-201.59 µg/mL) in Fe2+ metal-chelating ability, (130.39-181.12 µg/mL) in DPPH, (88.29-187.32 µg/mL) in OH radical, (79.04-156.79 µg/mL), in NO radical and (138.72-233.00 µg/mL) in superoxide anion scavenging activities, respectively. The methanolic extract display remarkable fungicidal activity against the tested pathogens followed by dichloromethane extract, essential oil, hexane extract and petroleum ether extract respectively, with MIC values ranging from 31.25 to 500 µg/mL. Based on results, it can be inferred that the flower of A. malaccensis if explored further for its medicinal properties, might be a good source to develop a safe and sustainable natural food preservative

Chemical composition and antifungal properties of commercial essential oils against the maize phytopathogenic fungus Fusarium verticillioides / Composición química y propiedades antifúngicas de aceites esenciales comerciales contra el hongo fitopatógeno de maíz Fusarium verticillioides

ABSTRACT The aim of the present study was to analyze the chemical composition ofCurcuma longa,Pimenta dioica,Rosmarinus officinalis, andSyzygiumaromaticumessential oils (EOs) and their antifungal and anti-conidiogenic activity againstFusarium verticillioides. The chemical profile of the EOs was determined by GC/MS. The antifungal and anti-conidiogenic activities were evaluated by the agar dilution method. The tested concentrations were 1000ppm, 500ppm, 250ppm and 125ppm.S. aromaticumEO exhibited the highest antifungal effect, followed byP. dioicaand to a lesser extentC. longa. The major compounds of these EOs were eugenol (88.70% inS. aromaticumand 16.70% inP. dioica), methyl eugenol (53.09% inP. dioica), and α-turmerone (44.70%), β-turmerone (20.67%), and Ar-turmerone (17.27%) inC. longa.Rosmarinus officinalispoorly inhibited fungal growth; however, it was the only EO that inhibited conidial production, with its major components being 1,8-cineole (53.48%), α-pinene (15.65%), and (−)-camphor (9.57%). Our results showed that some compounds are capable of decreasing mycelial growth without affecting sporulation, and vice versa. However, not all the compounds of an EO are responsible for its bioactivity. In the present work, we were able to identify different major compounds or mixtures of major compounds that were responsible for antifungal and anti-conidiogenic effects. Further experiments combining these pure components are necessary in order to achieve a highly bioactive natural formulation against the phytopathogenic fungusF. verticillioides.

RESUMEN El objetivo del presente estudio fue analizar la composición química de los aceites esenciales (AE) deCurcuma longa,Pimenta dioica,RosmarinusofficinalisySyzygiumaromaticum, y su actividad antifúngica y anti-esporuladora contraFusarium verticillioides. La composición de los AE se analizó por CG-EM. La actividad antifúngica y anti-esporuladora se evaluó a través del método de dilución en agar usando las siguientes concentraciones: 1.000, 500, 250 y 125ppm. El AE deS. aromaticumdemostró el mayor efecto antifúngico, seguido del deP. dioicay, en menor medida, delC. longa. Los compuestos principales de estos AE fueron eugenol (88,70% enS. aromaticumy 16,70% enP. dioica), metileugenol (53,09% enP. dioica) y α-turmerona (44,70%), β-turmerona (20,67%) y Ar-turmerona (17,27%) enC. longa. El AE deR. officinalisfue el que menor efecto inhibitorio presentó sobre el crecimiento fúngico, pero fue el único que inhibió la producción de conidias; sus principales componentes fueron 1,8-cineol (53,48%), α-pineno (15,65%) y (−)-alcanfor (9,57%). Nuestros resultados demostraron que algunos compuestos son capaces de disminuir el crecimiento del micelio deF. verticillioidessin afectar la esporulación, y vice versa. Sin embargo, no todos los compuestos de un AE son responsables de su bioactividad. En el presente trabajo, pudimos identificar diferentes compuestos o mezclas de compuestos que fueron responsables de los efectos antifúngicos y anti-esporuladores. Se necesitan nuevos experimentos que evalúen la combinación de estos compuestos puros para lograr una formulación bioactiva y de origen natural para el control deF. verticillioides.

Antifungal efficacy of Moringa oleifera leaf and seed extracts against Botrytis cinerea causing gray mold disease of tomato (Solanum lycopersicum L. ) / Eficácia antifúngica de extratos de folha e de semente de Moringa oleifera contra Botrytis cinerea causador da doença do molde cinza de tomate (Solanum lycopersicum L. )

Abstract Drawbacks associated with the use of chemical fungicides to control plant pathogenic fungi such as Botrytis cinerea stimulate the need for alternatives. Therefore, the present study was carried out to determine the antifungal potentials of Moringa oleifera extracts against B. cinerea. Phytochemical analysis using qualitative chemical tests revealed the presence of huge amount of crucial phytochemicals compounds like phenolic compounds, alkaloids and saponins in the M. oleifera leaf extract. Antifungal bioassay of the crude extracts indicated better mycelial growth inhibition by methanol leaf extract (99%). The minimum inhibitory concentration (MIC) was 5 mg/ml with 100% spore germination inhibition and minimum fungicidal concentration (MFC) was 10 mg/ml with 98.10% mycelial growth inhibition using broth micro dilution and poisoned food techniques. Gas chromatography-mass spectrometry (GC-MS) analysis led to the identification of 67 volatile chemical compounds in the leaf extract with 6-decenoic acid (Z)- (19.87%) was the predominant compound. Further chemical elucidation of the crude extracts performed by liquid chromatography with tandem mass spectrometry (LC-MS/MS) showed the presence of non-volatile chemical compounds, mostly flavones, flavonoids and phenolic acids (i.e. quercetin and kaempferol). Scanning electron microscopy and transmission electron microscopy analysis showed positive effect of M. oleifera leaf extract on the treated conidia and mycelium of B. cinerea. Findings revealed that irreversible surface and ultra-structural changes with severe detrimental effects on conidia and mycelium morphology compared to control treatment. Overall findings suggested that M. oleifera leaf extract is a promising candidate for biological control of fungal pathogens, thus limiting overdependence on chemical fungicides.

Resumo As desvantagens associadas ao uso de fungicidas químicos para controlar fungos fitopatogênicos, como Botrytis cinerea, estimulam a necessidade de alternativas. Portanto, o presente estudo foi realizado para determinar o potencial antifúngico de extratos de Moringa oleifera contra B. cinerea. A análise fitoquímica usando testes químicos qualitativos revelou a presença de uma grande quantidade de compostos fitoquímicos cruciais, como compostos fenólicos, alcaloides e saponinas no extrato da folha de M. oleifera. O bioensaio antifúngico dos extratos brutos indicou melhor inibição do crescimento micelial pelo extrato de folhas de metanol (99%). A concentração inibitória mínima (MIC) foi de 5 mg/mL com 100% de inibição da germinação de esporos e a concentração fungicida mínima (MFC) foi de 10 mg/mL com 98,10% de inibição do crescimento micelial usando microdiluição em caldo e técnicas de alimentos envenenados. A análise por cromatografia gasosa-espectrometria de massa (GC-MS) levou à identificação de 67 compostos químicos voláteis no extrato da folha, sendo o ácido 6-decenoico (Z) (19,87%) o composto predominante. Elucidação química adicional dos extratos brutos realizada por cromatografia líquida com espectrometria de massa em tandem (LC-MS/MS) mostrou a presença de compostos químicos não voláteis, principalmente flavonas, flavonoides e ácidos fenólicos (ou seja, quercetina e kaempferol). As análises de microscopia eletrônica de varredura e microscopia eletrônica de transmissão mostraram efeito positivo do extrato de folhas de M. oleifera sobre os conídios e micélios tratados de B. cinerea. Os resultados revelaram a superfície irreversível e alterações ultraestruturais com graves efeitos prejudiciais sobre os conídios e a morfologia micelial, em comparação com o tratamento de controle. Os resultados gerais sugeriram que o extrato da folha de M. oleifera é um candidato promissor para o controle biológico de patógenos fúngicos, limitando assim a dependência excessiva de fungicidas químicos.

Phytochemical composition, antifungal activity, in vitro and in vivo toxicity of Syzygium cumini (L) Skeels leaves extract / Composición fitoquímica, actividad antifúngica, toxicidad in vitro e in vivo del extracto de hojas de Syzygium cumini (L) Skeels

This study determined phytochemical composition, antifungal activity and toxicity in vitro and in vivo of Syzygium cumini leaves extract (Sc). Thus, was characterized by gas chromatography coupled to mass spectrometry and submitted to determination of Minimum Inhibitory (MIC) and Fungicidal concentrations (MFC) on reference and clinical strains of Candida spp. and by growth kinetics assays. Toxicity was verified using in vitro assays of hemolysis, osmotic fragility, oxidant and antioxidant activity in human erythrocytes and by in vivo acute systemic toxicity in Galleria mellonella larvae. Fourteen different compounds were identified in Sc, which showed antifungal activity (MIC between 31.25-125µg/mL) with fungistatic effect on Candida. At antifungal concentrations, it demonstrated low cytotoxicity, antioxidant activity and neglible in vivotoxicity. Thus, Sc demonstrated a promising antifungal potential, with low toxicity, indicating that this extract can be a safe and effective alternative antifungal agent.

Este estudio determinó la composición fitoquímica, la actividad antifúngica y la toxicidad in vitro e in vivo del extracto de hojas de Syzygium cumini (Sc). Así, se caracterizó mediante cromatografía de gases acoplada a espectrometría de masas y se sometió a determinación de Concentraciones Mínimas Inhibitorias (CMI) y Fungicidas (MFC) sobre cepas de referencia y clínicas de Candida spp. y mediante ensayos de cinética de crecimiento. La toxicidad se verificó mediante ensayos in vitro de hemólisis, fragilidad osmótica, actividad oxidante y antioxidante en eritrocitos humanos y por toxicidad sistémica aguda in vivo en larvas de Galleria mellonella. Se identificaron catorce compuestos diferentes en Sc, que mostraron actividad antifúngica (CMI entre 31.25-125 µg/mL) con efecto fungistático sobre Candida. En concentraciones antifúngicas, demostró baja citotoxicidad, actividad antioxidante y toxicidad in vivo insignificante. Por lo tanto, Sc demostró un potencial antifúngico prometedor, con baja toxicidad, lo que indica que este extracto puede ser un agente antifúngico alternativo seguro y eficaz.

Evaluation of susceptibility and response in the surface of agents of surface mycoses (Trichophyton mentagrophytes; T. tonsurans) to antifungal drugs of interest in a medical clinic / Avaliação da susceptibilidade e resposta in vitro de agentes de micoses superficiais (Trichophyton mentagrophytes; T. tonsurans) à drogas antifúngicas de interesse em clínica médica

INTRODUCTION: The resistance of fungal species to drugs usually used in clinics is of great interest in the medical field. OBJECTIVE: To evaluate susceptibility and in vitro response of species of Trichophyton spp. to antifungal drugs of interest in clinical medicine. METHODS: 12 samples of clinical isolates from humans were used, nine of T. mentagrophytes and three of T. tonsurans. Susceptibility tests were performed according to the agar diffusion (AD) and broth microdilution (BM) methods. RESULTS: In the AD method, the species T. tonsurans presented a percentage of sensitivity of 33% in relation to amphotericin B and 66% to itraconazole, with 100% resistance to ketoconazole and fluconazole. T. mentagrophytes also showed 100% resistance to ketoconazole in this technique, with 11% sensitivity to ketoconazole, 22% to itraconazole and 22% of samples classified as sensitive dose dependent. In the MC method, the species T. tonsurans presented a sensitivity percentage of 66%, 55% and 33% in relation to ketoconazole, fluconazole and itraconazole, respectively. The T. mentagrophytes species presented sensitivity percentages of 11%, 11%, 33% and 55% for amphotericin B, itraconazole, ketoconazole and fluconazole, respectively. CONCLUSION: There was resistance in vitro of the species of T. mentagrophytes and T. tonsurans against the antifungal fluconazole and relative resistance against ketoconazole in the AD method. In BM, however, important percentages of sensitivity were observed for the two species analyzed in relation to the antifungals fluconazole and ketoconazole when compared to itraconazole and amphotericin B.

INTRODUÇÃO: A resistência de espécies fúngicas às drogas usualmente empregadas no meio clínico é motivo de grande interesse na área médica. OBJETIVO: Avaliar susceptibilidade e resposta in vitro de espécies de Trichophyton spp. a drogas antifúngicas de interesse em clínica médica. MÉTODOS: Foram utilizadas 12 amostras de isolados clínicos de humanos, sendo nove de T. mentagrophytes e três de T. tonsurans. Foram realizados testes de susceptibilidade segundo os métodos de difusão em ágar (DA) e microdiluição em caldo (MC). RESULTADOS: No método de DA, a espécie T. tonsurans apresentou percentual de sensibilidade de 33% em relação à anfotericina B e de 66% ao itraconazol, com 100% de resistência frente ao cetoconazol e ao fluconazol. A espécie T. mentagrophytes também apresentou 100% de resistência frente ao cetoconazol nesta técnica, com 11% de sensibilidade ao cetoconazol, 22% ao itraconazol e 22% das amostras classificadas como sensível dose dependente. No método de MC, a espécie T. tonsurans apresentou percentual de sensibilidade de 66%, 55% e 33% em relação ao cetoconazol, fluconazol e itraconazol, respectivamente. A espécie T. mentagrophytes apresentou percentuais de sensibilidade de 11%, 11%, 33% e 55% para anfotericina B, itraconazol, cetoconazol e fluconazol, respectivamente. CONCLUSÃO: Houve resistência in vitro das espécies do T. mentagrophytes e T. tonsurans frente ao antifúngico fluconazol e resistência relativa frente ao cetoconazol no método de DA. Na MC, no entanto, foram observados importantes percentuais de sensibilidade das duas espécies analisadas frente aos antifúngicos fluconazol e cetoconazol quando comparadas ao itraconazol e à anfotericina B.

Nanopartículas de prata em bases de próteses de PMMA para controle de atividade microbiana / Silver nanoparticles in PMMA prosthesis bases to control microbial activity

Objetivo:Avaliar in vitro a atividade de Staphylococcus aureus e Candida albicans em bases de próteses convencionais à base de polimetilmetacrilato de metila com nanopartículas de prata incorporadas a sua composição. Métodos: Foi realizado um estudo experimental laboratorial com resinas acrílicas autopolimerizáveis comercialmente disponíveis, Vipi Flash/VIPI e JET/Clássico. Foram confeccionados 80 corpos de prova, divididos em 16 grupos (n = 5), referentes ao tipo de resina, tratamento (incorporação e imersão na solução de nanopartículas de prata) e microrganismo inoculado. As nanopartículas foram sintetizadas com ácido polimetacrílico, nitrato de prata e irradiadas com luz ultravioleta de baixa potência (~8W) por 6 horas, e as suas concentrações idealizadas pelo método de microdiluição em placas para determinação da concentração mínima inibitória frente aos microrganismos selecionados. Verificou-se ação bactericida e fungicida com concentração inicial de 25% e após fator de diluição 12,5%. Resultados: Houve dificuldade de incorporação das nanopartículas na resina acrílica, que pode decorrer da alteração da proporção 3:1 recomendada pelo fabricante ou pela redução ou inativação da ação da nanopartícula de prata pela interação com o polimetilmetacrilato. VIPI com inclusão de nanopartícula obteve menor aderência de biofilme de Candida albicans. Conclusão:A nanopartícula de prata mostrou-se eficaz na sua ação de controle de Candida albicans e Staphylococcus aureus no método de imersão, entretanto, a sua ação antimicrobiana foi comprometida após inclusão nas resinas acrílicas.

Aim:To perform an in vitro evaluation of the activity of Staphylococcus aureus and Candida albicansin conventional prosthesis bases, based on methyl polymethylmethacrylate with silver nanoparticles incorporated into the composition. Methods: An experimental laboratory study was carried out using commercially available self-curing acrylic resins, Vipi Flash/VIPI and JET/Clássico. Eighty specimens were manufactured and divided into 16 groups (n = 5), referent to the resin brand, treatment (incorporation and immersion in the silver nanoparticle solution), and inoculated microorganism. The nanoparticles were synthesized with polymethacrylic acid and silver nitrate, and were irradiated with a low power (~ 8W) ultraviolet light for 6 hours. Their concentrations were idealized by the method of microplate dilution to determine the minimum inhibitory concentration when compared to the selected microorganisms. Bactericidal and fungicidal activities were identified with an initial concentration of 25% and a subsequent dilution factor of 12.5%. Results:It was difficult to incorporate the AgNPs into the acrylic resin, which may well have resulted from the change from the 3:1 proportion recommended by the manufacturer or by reducing or inactivating the action of the silver nanoparticle by interaction with polymethylmethacrylate. VIPI with the inclusion of nanoparticles obtained a lesser Candida albicans biofilm adherence. Conclusion: Silver nanoparticles were effective in controlling Candida albicans and Staphylococcus aureus in the immersion method; however, the antimicrobial activity was compromised after inclusion in acrylic resins.

Antifungal susceptibility profile of Candida clinical isolates from 22 hospitals of São Paulo State, Brazil

This study aimed to evaluate the frequency of cryptic Candida species from candidemia cases in 22 public hospitals in São Paulo State, Brazil, and their antifungal susceptibility profiles. During 2017 and 2018, 144 isolates were molecularly identified as 14 species; C. parapsilosis (32.6%), C. albicans (27.7%), C. tropicalis (14.6%), C. glabrata (9.7%), C. krusei (2.8%), C. orthopsilosis (2.8%), C. haemulonii var. vulnera (2.1%), C. haemulonii (1.4%), C. metapsilosis (1.4%), C. dubliniensis (1.4%), C. guilliermondii (1.4%), C. duobushaemulonii (0.7%), C. kefyr (0.7%), and C. pelliculosa (0.7%). Poor susceptibility to fluconazole was identified in 6.4% of C. parapsilosis isolates (0.12 to >64 µg/mL), 50% of C. guilliermondii (64 µg/mL), 66.6% of C. haemulonii var. vulnera (16-32 µg/mL), and C. duobushaemulonii strain (MIC 64 µg/mL). Our results corroborated the emergence of C. glabrata in Brazilian cases of candidemia as previously reported. Importantly, we observed a large proportion of non-wild type C. glabrata isolates to voriconazole (28.6%; <0.015 to 4 µg/mL) all of which were also resistant to fluconazole (28.6%). Of note, C. haemulonii, a multidrug resistant species, has emerged in the Southeast region of Brazil. Our findings suggested a possible epidemiologic change in the region with an increase in fluconazole-resistant species causing candidemia. We stress the relevance of routine accurate identification to properly manage therapy and monitor epidemiologic trends.

Medicines for Malaria Venture COVID Box: a source for repurposing drugs with antifungal activity against human pathogenic fungi

BACKGROUND Treatment of mycoses is often ineffective, usually prolonged, and has some side effects. These facts highlight the importance of discovering new molecules to treat fungal infections. OBJECTIVES To search the Medicines for Malaria Venture COVID Box for drugs with antifungal activity. METHODS Fourteen human pathogenic fungi were tested against the 160 drugs of this collection at 1.0 µM concentration. We evaluated the ability of the drugs to impair fungal growth, their fungicidal nature, and morphological changes caused to cells. FINDINGS Thirty-four molecules (21.25%) presented antifungal activity. Seven are antifungal drugs and one is the agricultural fungicide cycloheximide. The other drugs with antifungal activity included antibiotics (n = 3), antimalarials (n = 4), antivirals (n = 2), antiparasitcs (n = 3), antitumor agents (n = 5), nervous system agents (n = 3), immunosuppressants (n = 3), antivomiting (n = 1), antiasthmatic (n = 1), and a genetic disorder agent (n = 1). Several of these drugs inhibited Histoplasma capsulatum and Paracoccidioides brasiliensis growth (15 and 20, respectively), while Fusarium solani was not affected by the drugs tested. Most drugs were fungistatic, but niclosamide presented fungicidal activity against the three dimorphic fungi tested. Cyclosporine affected morphology of Cryptococcus neoformans. MAIN CONCLUSIONS These drugs represent new alternatives to the development of more accessible and effective therapies to treat human fungal infections.

Recent progress on anti-Candida natural products / 中国天然药物

Candida is an intractable life-threatening pathogen. Candida infection is extremely difficult to eradicate, and thus is the major cause of morbidity and mortality in immunocompromised individuals. Morevover, the rapid spread of drug-resistant fungi has led to significant decreases in the therapeutic effects of clinical drugs. New anti-Candida agents are urgently needed to solve the complicated medical problem. Natural products with intricate structures have attracted great attention of researchers who make every endeavor to discover leading compounds for antifungal agents. Their novel mechanisms and diverse modes of action expand the variety of fungistatic agents and reduce the emergence of drug resistance. In recent decades, considerable effort has been devoted to finding unique antifungal agents from nature and revealing their unusual mechanisms, which results in important progress on the development of new antifungals, such as the novel cell wall inhibitors YW3548 and SCY-078 which are being tested in clinical trials. This review will present a brief summary on the landscape of anti-Candida natural products within the last decade. We will also discuss in-depth the research progress on diverse natural fungistatic agents along with their novel mechanisms.

Chemical constituents of Psidium guajava and their antitumor and antifungal activities / 中国中药杂志

Twenty-six compounds, including sixteen meroterpenoids(1-16), a triterpenoid(17), four terpenoid derivatives(18-21), and five aromatic compounds(22-26), were isolated from the leaves of Psidium guajava. Their structures were identified by spectroscopic analyses including NMR and MS. Compounds 21-26 were obtained from plants of Psidium for the first time. Based on the structure,(R)-2-ethylhexyl 2H-1,2,3-triazole-4-carboxylate(24 a), an α-glucosidase inhibitor recently isolated from Paramignya trimera, should be revised as compound 24. Meroterpenoids 1-16 were evaluated for their antitumor and antifungal activities. Meroterpenoids psiguajadial D(4), guapsidial A(5), 4,5-diepipsidial A(7), guadial A(14), and guadial B(15) showed cytotoxicities against five human tumor cell lines(HL-60, A-549, SMMC-7721, MCF-7, and SW-480), among which 5 was the most effective with an IC_(50) of 3.21-9.94 μmol·L~(-1).

A Ten-year Retrospective Study of Invasive Candidiasis in a Tertiary Hospital in Beijing / 生物医学与环境科学（英文）

Objective@#This study aimed to evaluate the epidemiological, clinical and mycological characteristics of invasive candidiasis (IC) in China.@*Methods@#A ten-year retrospective study including 183 IC episodes was conducted in a tertiary hospital in Beijing, China.@*Results@#The overall incidence of IC from 2010-2019 was 0.261 episodes per 1,000 discharges. Candidemia (71.0%) was the major infective pattern; 70.3% of the patients tested positive for @*Conclusion@#The incidence of IC has declined in the recent five years.

Effect of berberine hydrochloride on cell wall integrity of Candida albicans hypha / 中国中药杂志

The aim of this paper was to investigate the effect of berberine hydrochloride on the cell wall integrity of Candida albicans hypha. The minimal inhibitory concentration(MIC) of berberine hydrochloride against clinical and standard C. albicans strains was detected by micro liquid-based dilution method; the effect of berberine hydrochloride on the colony formation of C. albicans SC5314 was investigated by spot assay; the effect of berberine hydrochloride on the metabolism of C. albicans SC5314 hypha was checked by XTT reduction assay, and the viability of C. albicans SC5314 hypha was tested by fluorescent staining assay. The effect of berberine hydrochloride on the morphology of C. albicans SC5314 hypha was examined by scanning electron microscope. The changes in the cell wall of C. albicans SC5314 hypha after berberine hydrochloride treatment were detected by transmission electron microscopy. The effect of berberine hydrochloride on β-glucan from C. albicans SC5314 was detected by flow cytometry. The effect of berberine hydrochloride on hypha-specific gene ECE1 and β-glucan synthase genes FKS1 and FKS2 in C. albicans was examined by qRT-PCR. The results showed that berberine hydrochloride showed a strong inhibitory effect on both clinical and standard strains of C. albicans, and the MIC was 64-128 μg·mL~(-1). Spot assay, XTT redunction assay and fluorescent staining assay showed that with the increase of berberine hydrochloride concentration, the viability of C. albicans SC5314 gradually decreased. The transmission electron microscopy scanning assay showed that this compound could cause cell wall damage of C. albicans. The flow cytometry analysis showed the exposure degree of C. albicans β-glucan. The qRT-PCR further showed that berberine hydrochloride could significantly down-regulate hypha-specific gene ECE1 and β-glucan synthase-related gene FKS1 and FKS2. In conclusion, this compound can down-regulate C. albicans and β-glucan synthase-related gene expressions, so as to destroy the cell wall structure of C. albicans, expose β-glucan and damage the integrity of the wall.

Advances in the biosynthesis of natamycin and its regulatory mechanisms / 生物工程学报

Natamycin is a polyene macrolide antibiotics with strong and broad spectrum antifungal activity. It not only effectively inhibits the growth and reproduction of fungi, but also prevents the formation of some mycotoxins. Consequently, it has been approved for use as an antifungal food preservative in most countries, and is also widely used in agriculture and healthcare. Streptomyces natalensis and Streptomyces chatanoogensis are the main producers of natamycin. This review summarizes the biosynthesis and regulatory mechanism of natamycin, as well as the strategies for improving natamycin production. Moreover, the future perspectives on natamycin research are discussed.