Matrix-assisted laser desorption/ionisation-time of flight mass spectrometry azole susceptibility assessment in Candida and Aspergillus species

BACKGROUND Matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF MS) allows rapid pathogen identification and potentially can be used for antifungal susceptibility testing (AFST). OBJECTIVES We evaluated the performance of the MALDI-TOF MS in assessing azole susceptibility, with reduced incubation time, by comparing the results with the reference method Broth Microdilution. METHODS Resistant and susceptible strains of Candida (n = 15) were evaluated against fluconazole and Aspergillus (n = 15) against itraconazole and voriconazole. Strains were exposed to serial dilutions of the antifungals for 15 h. Microorganisms' protein spectra against all drug concentrations were acquired and used to generate a composite correlation index (CCI) matrix. The comparison of autocorrelations and cross-correlations between spectra facilitated by CCI was used as a similarity parameter between them, enabling the inference of a minimum profile change concentration breakpoint. Results obtained with the different AFST methods were then compared. FINDINGS The overall agreement between methods was 91.11%. Full agreement (100%) was reached for Aspergillus against voriconazole and Candida against fluconazole, and 73.33% of agreement was obtained for Aspergillus against itraconazole. MAIN CONCLUSIONS This study demonstrates MALDI-TOF MS' potential as a reliable and faster alternative for AFST. More studies are necessary for method optimisation and standardisation for clinical routine application.

Perfiles de expresión de los genes ERG11, MDR1 y AFR1 en Cryptococcus neoformans var. grubii aislados de pacientes con HIV / Gene expression profiles of ERG11, MDR1 y AFR1 in Cryptococcus neoformans var. grubbi isolated from HIV patients

Introducción. El fluconazol es el antifúngico más utilizado para la prevención y el tratamiento de infecciones causadas por el género Cryptococcus, agente etiológico de la criptococosis. La resistencia al fluconazol en los aislamientos de Cryptoccocus neoformans puede hacer fracasar el tratamiento y generar recaídas de la infección. Objetivo. Evaluar los perfiles de expresión de los genes AFR1, MDR1 y ERG11 en aislamientos clínicos de C. neoformans var. grubii, durante la respuesta in vitro a la inducción con fluconazol. Materiales y métodos. Se estudiaron 14 aislamientos de C. neoformans var. grubii provenientes de pacientes con HIV, de los cuales 6 eran sensibles al fluconaol y 8 presentaban sensibilidad disminuida. Los niveles de expresión de los genes ERG11, AFR1 y MDR1 se determinaron mediante PCR en tiempo real. Resultados. Los aislamientos resistentes al fluconazol mostraron sobreexpresión de los genes AFR1 y MDR1, mientras que la expresión de los fenotipos de resistencia evaluados se mantuvo homogénea en ERG11, en todos los aislamientos de C. neoformans var. grubii. Conclusiones. La sobreexpresión de los genes AFR1 y MDR1 que codifican las bombas de eflujo, contribuye a la resistencia al fluconazol en los aislamientos estudiados. Sin embargo, los patrones de resistencia que se registran en este hongo, sumado a los casos de recaídas en pacientes con HIV, no pueden atribuirse únicamente a los casos de resistencia por exposición al fármaco. Otros mecanismos podrían también estar involucrados en este fenómeno, como la resistencia emergente (resistencia mediante otros genes ERG) y la heterorresistencia, los cuales deben ser estudiados en estos aislamientos.

Introduction: Fluconazole is the most used antifungal drug for prevention and treatment of Cryptococcus spp. infections, the etiological agent of cryptococcosis. Resistance to fluconazole among Cryptococcus neoformans isolates can lead to treatment failure and generate relapses. Objective: To evaluate the expression profles of the AFR1, MDR1 and ERG11 genes in C. neoformans var. grubii clinical isolates during the in vitro response to fluconazole induction. Materials and methods: Fourteen C. neoformans var. grubii isolates recovered from HIV patients were studied, in which 6 showed sensitivities to fluconazole and 8 decreased sensitivity. The expression levels of ERG11, AFR1 and MDR1 genes were determined by real-time PCR from extracted mRNA. Results: AFR1 and MDR1 genes from C. neoformans var. grubii were overexpressed in fluconazole resistant isolates, whereas ERG11 maintains homogeneous expression in all the evaluated resistance phenotypes of C. neoformans var. grubii isolates. Conclusions: The overexpression of AFR1 and MDR1 genes, which codify for efflux pumps, contributes to fluconazole resistance in the studied isolates. However, the resistance patterns in this fungus and the relapse cases in HIV patients cannot be attributed solely to the exposure to the drug. Heteroresistance and the emerging resistance (resistance through other ERG genes), might be other mechanisms involved in this phenomenon, which must be studied in these isolations.

Perfil de resistencia antifúngica en el tratamiento de candidiasis vaginal: Un diagnóstico de agentes etiológicos / Antifungal resistance profile in the treatment of vaginal candidiasis: A diagnosis of etiologic agents

Introducción: En las micosis, la resistencia a los antifúngicos aumenta debido a los diagnósticos y tratamientos incorrectos. Por tanto, se ha sugerido la vigilancia y el seguimiento de las cepas resistentes para garantizar una terapia adecuada. Objetivo: Determinar las especies de Candida y el perfil de resistencia a fluconazol y voriconazol, que presentan los aislados obtenidos de muestras almacenadas durante los meses diciembre 2017 y marzo 2018 de pacientes provenientes del Laboratorio del Hospital Regional "Miguel Ángel Mariscal Llerena" del departamento de Ayacucho, Perú. Material y Métodos: Estudio descriptivo transversal, en el cual se aislaron 110 cepas de Candida sp almacenadas por un período de cuatro meses, y procesadas por métodos estandarizados y aprobados por el Ministerio de Salud (MINSA) y el Instituto Nacional de Salud (INS) del Perú relacionados con el diagnóstico de agentes etiológicos de micosis humanas y la sensibilidad antifúngica con los métodos estandarizados de Clinical Laboratory Standard Institute (CLSI). Resultados: Se observó C. albicans en 86,4 % de los aislados seguida por C. glabrata con 9,1 %, C. parapsilosis 2,7 % y 0,9 % de C. tropicalis y C. krusei. El 10,5 % de C. albicans fue resistente al fluconazol y voriconazol con CMI ≥ 128 μg/mL y ≥ 16 μg/mL respectivamente, mientras que 20 % de C. glabrata mostró sensibilidad dosis dependiente y 10 % de resistencia al fluconazol. Conclusiones: Existe una gran variedad de especies de Candida, siendo la C. albicans la más común, seguida por C. glabrata, con un mayor porcentaje de aislamiento en comparación con otras especies. Se puede observar que estas especies poseen grados de vulnerabilidad considerables a la aplicación de fármacos como el fluconazol y voriconazo(AU)

Introduction: In mycoses, resistance to antifungals increases due to incorrect diagnosis and treatment. Therefore, surveillance and monitoring of resistant strains has been suggested to ensure adequate therapy. Objective: To determine the Candida species and the resistance profile to fluconazole and voriconazole presented by the isolates obtained from samples stored during the months of December 2017 and March 2018 from patients coming from the Laboratory of the Regional Hospital "Miguel Ángel Mariscal Llerena" in the department of Ayacucho, Peru. Material and Methods: Cross-sectional descriptive study in which 110 strains of Candida sp were isolated and stored for a period of four (04) months, and processed by standardized methods approved by the Ministry of Health (MINSA) and the National Institute of Health (INS) of Peru related to the diagnosis of etiological agents of human mycosis and antifungal sensitivity with the standardized methods of the Clinical Laboratory Standard Institute (CLSI). Results: C. albicans was observed in 86,4 % of isolates, followed by C. glabrata (9,1 %), C. parapsilosis (2,7 %), and 0,9 % of C. tropicalis and C. krusei; 10,5 % of C. albicans was resistant to fluconazole and voriconazole with MIC ≥ 128 μg/mL and ≥ 16 μg/mL respectively, while 20 % of C. glabrata showed dose dependent sensitivity and 10 % resistance to fluconazole. Conclusions: There is a wide variety of Candida species, with C. albicans being the most common, followed by C. glabrata, with a higher percentage of isolation compared to other species. It can be seen that these species have considerable degrees of vulnerability to the application of drugs such as fluconazole and voriconazole(AU)

Distribution and antifungal susceptibility of Candida species isolated from clinical samples in southern Brazil

Abstract Invasive infections caused by Candida species have been strongly associated with poor prognosis and high resistance rates to some antifungals. This study aimed to identify Candida species isolated from different anatomical sites and to describe their susceptibility profile to antifungals. Ninety-four clinical isolates of Candida were obtained from a Medical Laboratory of Santa Catarina/Brazil. Species identification was performed by MALDI-TOF MS. Susceptibility assays were performed as described by Clinical Laboratory Standard Institute (CLSI) microboth method. Among the analyzed samples, C. albicans was the pathogen most incident (59.9%) followed by C. parapsilosis complex (14.9%), C. glabrata complex (8.5%), and C. tropicalis (6.3%). 37 Candida strains were isolated from vaginal content (39.3%), 21 from the nail (22.4%), 8 from tracheal aspirates (8.5%), and 7 from urine (7.4%). Together, the Candida isolates presented decreased susceptibility to azole drugs, mainly to fluconazole and itraconazole. Amphotericin B showed sensibility in 95.7% of samples analyzed. Previous knowledge about etiology and antifungal susceptibility becomes indispensable to conduct an efficient treatment.

Synthesis of New Thiazole Derivatives Bearing Thiazolidin-4(5H)-One Structure and Evaluation of Their Antimicrobial Activity

The first report about antimicrobial resistance was published in the 1940s. And today, the antimicrobial resistance has become a worldwide problem. Because of this problem, there is a need to develop new drugs. That's why we synthesized some novel thiazolidine-4-one derivatives and evaluated their antimicrobial activity. The final compounds were obtained by reacting 2-[(4,5-diphenylthiazol-2-yl)imino]thiazolidin-4-one with some aryl aldehydes. The synthesized compounds were investigated for their antimicrobial activity against four Candida species, five gram-negative and four gram-positive bacterial species. The lead compounds (4a- h) were obtained with a yield of at least 70%. All compounds showed antimicrobial activity. Compound 4f (MIC: 31.25 µg/ml) exhibited more efficacy than the other compounds against C. glabrata (ATCC 24433). Compound 4b (MIC: 62.5 µg/ml) was the most active compound against all bacterial species, particularly K. pneumoniae (NCTC 9633). Whereas, compound 4c (MIC: <31.25 µg/ml) was observed as the most active compound against E. coli (ATCC 25922). In general, all compounds (4a-4h) showed antimicrobial activity against all fungi and bacterial species. Compounds 4b (2,6-dichlorobenzylidene), 4c (2,6-dihydroxybenzylidene), 4f (1H-pyrrol-2- yl)methylene), 4g (4-triflouromethylbenzylidene) and 4h (2,3,4-trimethoxybenzylidene) were determined as the most active compounds

Épidémiologie des candidoses systémiques dans les services à haut risque au CHU et au centre anti cancer de Batna ­Algérie / Epidemiology of systemic candidiasis in high-risk departments at the university hospital and cancer center of Batna -Algeria

Introduction : Les candidoses systémiques sont des affections graves responsables d'une mortalité élevée. L'objectif de ce travail est de décrire l'épidémiologie des candidoses systémiques dans les services à haut risque au CHU et au CAC de BATNA. Patients et méthodes : Il s'agit d'une étude prospective descriptive durant une période de trois ans (1er janvier 2016 au 31 décembre 2018). Les patients inclus sont ceux ayant au moins un prélèvement profond positif á Candida spp. Résultats : Un total de 69 cas de candidoses systémiques correspondant à 75 isolats et concernant 63 patients a pu être analysé. L'incidence globale était de 2,62 cas pour 1000 admissions. Les principaux motifs d'hospitalisation étaient les hémopathies malignes et le choc septique. La présence d'une colonisation ( 2 sites), une antibiothérapie á large spectre, d'un cathéter intra vasculaire, une corticothérapie, une chimiothérapie, une neutropénie étaient les facteurs de risque les plus retrouvés. L'analyse des souches isolées a montré la prédominance des espèces non albicans. L'index de colonisation ≥ 0,5 a été significativement associé au risque de candidose systémique. L'utilisation des Azolés a été associée á un taux de mortalité le plus élevé (19%). Le taux de mortalité est significativement élevé 51%. Conclusion : Les facteurs de risque et un index de colonisation ≥ 0,5 dans les services á haut risque constituent un facteur prédictif de candidose systémique. La prise en charge thérapeutique doit être instaurée pour réduire le taux de mortalité et éviter les complications liées á ces infections.

Background: Systemic candidiasis are serious conditions responsible for high mortality. The objective of this work is to describe the epidemiology of systemic candidiasis in high-risk departments at the UHC and the ACC of BATNA. Patients and methods: This is a descriptive prospective study over a period of three years (January 1, 2016 to December 31, 2018). The patients included are those with at least one positive deep sample for Candida spp. Results: 69 cases of systemic candidiasis corresponding to 75 isolates and concerning 63 patients could be analyzed. The overall incidence was 2.62 cases per 1,000 admissions. The main reasons for hospitalization were hematologic malignancies and septic shock. The presence of colonization ( 2 sites), broad-spectrum antibiotic therapy, an intravascular catheter, corticosteroid therapy, chemotherapy, neutropenia were the most common risk factors. Analysis of the isolated strains showed the predominance of nonalbicans species. Colonization index ≥ 0.5 was significantly associated with the risk of systemic candidiasis. Azole's use was associated with the highest mortality rate (19%). The mortality rate is significantly high 51%. Conclusion. Risk factors and a colonization index ≥ 0.5 in high-risk wards are a predictor of systemic candidiasis. Therapeutic care must be instituted to reduce the mortality rate and avoid complications linked to these infections

Azoles de antes y ahora: una revisión / Azoles of then and now: a review

Resumen Los azoles son fármacos que inhiben la enzima 14α-esteroldemetilasa, impidiendo la unión de ergosterol; esto altera la estructura y función de la pared celular fúngica. Especialmente el grupo de los triazoles: fluconazol, itraconazol, voriconazol, posaconazol e isavuconazol, son una alternativa farmacológica para el tratamiento de la enfermedad fúngica invasora causada por Aspergillus spp, Candida spp, Cryptococcus spp, patógenos emergentes como los Mucorales, y de micosis endémicas como las ocasionadas por Histoplasma spp y Coccidioides spp. Los efectos adversos de los triazoles son menos frecuentes comparados con los ocasionados por anfotericina B, un antifúngico de uso común para estas micosis. Los principales efectos adversos de los triazoles son hepáticos, gastrointestinales y cardiovasculares como la prolongación del intervalo QT. Las interacciones farmacológicas son usuales y se presentan con moléculas que usan sustratos del citocromo CYP3A4, lo que incluye anti-retrovirales, anti-tuberculosos e inmunomoduladores. En este trabajo se revisan la historia, características farmacológicas y los ensayos clínicos que evidencian su eficacia clínica en los diferentes escenarios clínicos.

Abstract The azoles are drugs that inhibit the 14α-sterol-demethylase enzyme preventing the binding of ergosterol, altering the functionality and structure of the fungal cell wall. Especially the group of triazoles: fluconazole, itraconazole, voriconazole, posaconazole and isavuconazole, are a pharmacological alternative for the treatment of the invasive fungal disease, caused by Aspergillus spp, Candida spp, Cryptococcus spp, by emerging pathogens for example, the Mucoral and finally of endemic mycosis as those caused by Histoplasma spp. and Coccidioides spp. The adverse effects of the triazoles are less frequent compared to those caused by amphotericin B, the main ones being hepatics, gastrointestinals and cardiovasculars, such as the prolongation of the QT interval. The pharmacological interactions are common and occur with molecules that use the substrates of the CYP3A4 cytochrome, for example: antiretroviral, anti-tuberculous and immunomodulators. The history, pharmacological characteristics and clinical trials are reviewed.

Mecanismos de resistencia a fluconazol expresados por Candida glabrata: una situación para considerar en la terapéutica / Resistance mechanisms to fluconazole expressed by Candida glabrata: a situation to consider in therapy / Mecanismos de resistência a fluconazol expressos por Candida glabrata: uma situação a considerar na terapêutica

Introducción: Los esfuerzos terapéuticos orientados a atender las micosis por Candidaspp. se han enfocado en el empleo de azoles; sin embargo, en la literatura científica se discute su beneficio, por los amplios y descritos mecanismos de resistencia. Objetivo: Describir los mecanismos de resistencia al fluconazol expresados por la especie Candida glabrata, con la intención de que sean considerados dentro de las variables de elegibilidad para la intervención. Método: Se realizó una revisión integrativa utilizando la pregunta orientadora: ¿cuáles son los mecanismos de resistencia al fluconazol expresados por la especie Candida glabrata? Veintinueve estudios obtenidos de la base de datos PubMed cumplieron los criterios del análisis crítico propuesto por el instrumento PRISMA, utilizado para la selección de los artículos incluidos para su revisión en este manuscrito. Las categorías bajo las cuales se organizaron los elementos de análisis fueron: sobrexpresión de bombas de eflujo y modificaciones en la enzima lanosterol 14-alfa-desmetilasa. Resultados: Los mecanismos de resistencia al fluconazol expresados por Candida glabrata están determinados principalmente por la regulación a la alza de bombas de adenosina-trifosfato Binding Cassette (ABC) y por la modificación del punto de unión con su blanco farmacológico: la enzima lanosterol 14-alfa-desmetilasa. Conclusión: Los mecanismos de resistencia expresados por Candida glabrata se asocian con la modificación estructural de la diana farmacológica y la sobreexpresión de bombas de eflujo de manera diferencial a otras especies. Se sugiere que Candida glabrata es intrínsecamente menos susceptible al fluconazol.

Introduction: Therapeutic efforts aimed at treating mycosis caused by Candida spp. have focused on the use of azoles; however, their benefits have been subject to discussion in scientific literature, due to the extensive and well-described resistance mechanisms. Objective: To describe the resistance mechanisms to fluconazole expressed by the Candida glabrata species, so they are considered within the variables of eligibility for intervention. Method: An integrative review was carried out using the guiding question: what are the fluconazole resistance mechanisms expressed by the Candida glabrata species? Twenty-nine studies obtained from the PubMed database met the criteria for the critical analysis proposed by the PRISMA instrument, which was used for the selection of articles for review included in this paper. The analysis elements were organized in the following categories: overexpression of efflux pumps and modifications in the enzyme lanosterol 14-alpha-demethylase. Results: The resistance mechanisms to fluconazole expressed by Candida glabrata are mainly determined by the upregulation of Adenosine triphosphate Binding Cassette (ABC) pumps and by the modification of the point of attachment with its pharmacological target: the enzyme lanosterol 14-alpha-demethylase. Conclusion: The resistance mechanisms expressed by Candida glabrata are associated with the structural modification of the pharmacological target and the overexpression of efflux pumps, in a way different to other species. It is suggested that Candida glabrata is intrinsically less susceptible to fluconazole.

Introdução: Os esforços terapêuticos voltados ao tratamento de micose por Candida spp. se focaram no uso de azóis; no entanto, na literatura científica discute-se seu benefício devido aos extensos e descritos mecanismos de resistência. Objetivo: Descrever os mecanismos de resistência ao fluconazol expressos pela espécie Candida glabrata, com a intenção de serem considerados dentro das variáveis de elegibilidade para a intervenção. Método: Uma revisão integrativa foi realizada utilizando a questão norteadora: quais os mecanismos de resistência ao fluconazol expressos pela espécie Candida glabrata? Vinte e nove estudos obtidos da base de dados PubMed atenderam os critérios de análise crítica proposta pelo instrumento PRISMA, utilizado para a seleção dos artigos incluídos para revisão neste manuscrito. As categorias sob as quais se organizaram os elementos de análise foram: superexpressão de bombas de efluxo e modificações na enzima lanosterol 14-alfa-desmetilase. Resultados: Os mecanismos de resistência ao fluconazol expressos por Candida glabrata são determinados principalmente pela regulação positiva das bombas de adenosina-trifosfato Binding Cassette (ABC) e pela modificação do ponto de fixação com seu alvo farmacológico: a enzima lanosterol 14-alfa-desmetilasa. Conclusão: Os mecanismos de resistência expressos por Candida glabrata são associados à modificação estrutural da Diana farmacológica e a superexpressão de bombas de efluxo de maneira diferencial a outras espécies. Sugere-se que Candida glabrata é intrinsecamente menos susceptível ao fluconazol.

In Silico Identification of Novel Tuberculosis Drug Targets in Mycobacterium tuberculosis P450 Enzymes by Interaction Study with Azole Drugs

@#Introduction: Tuberculosis (TB) is one of the utmost serious infectious diseases worldwide. The emergence of multidrug resistance demands the development of better or new putative drug targets for tuberculosis. Recent studies suggest Mycobacterium tuberculosis cytochrome P450 enzymes as promising drug targets and azole drugs as potential inhibitors. Methods: Various computational tools, like Expasy Protparam, Swiss model, RaptorX and Phyre2 were used to analyze 12 Mycobacterium tuberculosis P450 enzymes and determine their three-dimensional structure. The structural validation was done through a Ramachandran plot using RAMPAGE server. The docking of P450 enzymes with azole drugs was done with autodock ver 4.2.6. Results: Based on sub-cellular localization prediction using CELLO tool, P450 enzymes CYP123A1, CYP132A1, CYP135A1, CYP136A1, CYP140A1, and CYP143A1 were predicted to be in the cytoplasm. Through structure assessment by Ramachandran plot, the best homology modelled proteins were docked with azole drugs like clotrimazole, croconazole, econazole, fluconazole, itraconazole, itraconazole, ketaconazole and micronazole by using autodock. By docking method it is identified that ketaconazole drug has a high affinity towards most of the mycobacterium P450 enzymes followed by the itrconazole drug. CYP123A1 enzyme is preferable as a drug target due to high binding affinity towards ketoconazole followed by CYP135A1, CYP140A1 enzymes. Conclusion: This study would help in identifying putative novel drug targets in Mycobacterium tuberculosis, which can lead to promising candidates for the optimization and development of novel anti-mycobacterial agents.

Paracoccidioidomicosis. Una enfermedad multisistémica / Paracoccidioidomycosis. A multisystemic disease

Resumen La paracoccidioidomicosis es una enfermedad crónica, sistémica y progresiva, sólo descrita en América Latina. Su presentación clínica crónica multifocal es la más prevalente, afectando mayormente a hombres adultos y comprometiendo principalmente a pulmones, sin embargo, puede diseminarse a cualquier órgano generando múltiples complicaciones en el paciente. Presentamos el caso de un paciente masculino, inmunocompetente, caficultor, quien debuta con compromiso de la glándula suprarrenal y en quien posteriormente se documenta compromiso pulmonar. El diagnóstico se confirmó mediante biopsia de lesiones en glándula suprarrenal, inmunodifusión en gel de agar y reacción en cadena de la polimerasa, la cual mostró compromiso por Paracoccidioides brasiliensis. (Acta Med Colomb 2018; 43: 111-114).

Abstract Paracoccidioidomycosis is a chronic, systemic and progressive disease which is described only in Latin America. Its chronic and multifocal clinical presentation is the most prevalent, affecting mainly adult men and compromising mainly lungs; however, it can spread to any organ generating multiple complications in the patient. The case of an immunocompetent male patient, coffee grower, who debuted with compromise of the adrenal gland and in who subsequently pulmonary involvement was documented, is presented. The diagnosis was confirmed by biopsy of lesions in the adrenal gland, agar gel immunodiffusion and polymerase chain reaction, which showed compromise by Paracoccidioides brasilensis. (Acta Med Colomb 2018; 43: 111-114).

In vitro antifungal susceptibility of clinical and environmental isolates of Aspergillus fumigatus and Aspergillus flavus in Brazil

ABSTRACT The in vitro susceptibility of 105 clinical and environmental strains of Aspergillus fumigatus and Aspergillus flavus to antifungal drugs, such as amphotericin B, azoles, and echinocandins was evaluated by the broth microdilution method proposed by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). Following the EUCAST-proposed breakpoints, 20% and 25% of the clinical and environmental isolates of A. fumigatus, respectively, were found to be resistant to itraconazole (Minimal Inhibitory Concentration, MIC > 2.0 mg/L). Voriconazole showed good activity against A. fumigatus and A. flavus strains, except for one clinical strain of A. fumigatus whose MIC was 4.0 mg/L. Posaconazole (≤0.25 mg/L) also showed appreciable activity against both species of Aspergillus, except for six A. fumigatus strains with relatively higher MICs (0.5 mg/L). The MICs for Amphotericin B ranged from 0.06 to 1.0 mg/L for A. fumigatus, but were much higher (0.5-8.0 mg/L) for A. flavus. Among the echinocandins, caspofungin showed a geometric mean of 0.078 and 0.113 against the clinical and environmental strains of A. flavus, respectively, but had elevated minimal effective concentrations (MECs) for seven of the A. fumigatus strains. Anidulafungin and micafungin exhibited considerable activity against both A. fumigatus and A. flavus isolates, except for one environmental isolate of A. fumigatus that showed an MEC of 1 mg/L to micafungin. Our study proposes that a detailed investigation of the antifungal susceptibility of the genus Aspergillus from different regions of Brazil is necessary for establishing a response profile against the different classes of antifungal agents used in the treatment of aspergillosis.

Efficacy of voriconazole on leishmaniasis by Leishmania major: An in vitro and in vivo study

Objective:To appraise the activity of voriconazole against Leishmania major (L. major) in vitro and its effectiveness on wound regeneration in cutaneous leishmaniasis in BALB/c mice.Methods:The ICResults:The ICConclusions:Our results demonstrate that voriconazole can be an option in treating the cutaneous leishmaniasis by L. major.

Efficacy of voriconazole on leishmaniasis by Leishmania major: An in vitro and in vivo study

Objective: To appraise the activity of voriconazole against Leishmania major (L. major) in vitro and its effectiveness on wound regeneration in cutaneous leishmaniasis in BALB/c mice. Methods: The IC

In Vitro Antifungal Activities against Moulds Isolated from Dermatological Specimens

Background: This study aimed to determine the minimum inhibitory concentrations (MICs) of various antifungal agents against moulds isolated from dermatological specimens. Methods: We identified 29 moulds from dermatological specimens between October 2012 and March 2013 by conventional methods. We performed antifungal susceptibility testing on six antifungal agents, amphotericin B, clotrimazole, itraconazole, ketoconazole, miconazole and terbinafine, according to the Clinical and Laboratory Standards Institute guidelines contained in the M38-A2 document. Results: Most antifungal agents were active against the dermatophytes, except for terbinafine against Trichophyton rubrum (geometric mean MIC, MICGM 3.17 µg/mL). The dematiaceous moulds were relatively susceptible to amphotericin B and azoles (MICGM 0.17-0.34 µg/mL), but not to terbinafine (MICGM 3.62 µg/mL). Septate hyaline moulds showed variable results between the relatively more susceptible Aspergillus spp. (MICGM 0.25-4 µg/mL) and the more resistant Fusarium spp. (MICGM 5.66-32 µg/mL). The zygomycetes were susceptible to amphotericin B (MICGM 0.5 µg/mL) and clotrimazole (MICGM 0.08 µg/mL), but not to other azoles (MICGM 2.52-4 µg/mL). Conclusion: Amphotericin B and clotrimazole were the most effective antifungal agents against all moulds excepting Fusarium spp., while terbinafine was useful against dermatophytes (except T. rubrum) and Aspergillus spp. However, a larger study is required to draw more solid conclusions.

Synthesis and anti-breast cancer activity of azole derivatives / 第二军医大学学报

Objective To design and synthesize a new series of efficient, low toxicity azole derivatives using antifungal drug ketoconazole as the lead compound and to explore their anti-breast cancer activity. Methods Based on the docking mode of ketoconazole with estrogen receptor. We designed and synthesized eleven derivatives, whose 2, 4-dichlorophenyl and triazole ring were retained and the side chains were modified. Then the in vitro anticancer activities against breast cancer cells MDA- MB-231 and MCF-7 were determined by MTT using tamoxifene as the positive control drug. Results and Conclusion The synthesized compounds have been reported for the first time and they have been confirmed by1HNMR and13CNMR. The synthesized azole derivatives have greater inhibitory effects than tamoxifene against breast cancer MDA MB-231 cells.

Antifungal susceptibility of clinical and environmental Cryptococcus neoformans and Cryptococcus gattii isolates in Jabalpur, a city of Madhya Pradesh in Central India

In this study, we present antifungal susceptibility data of clinical and environmental isolates of Central Indian Cryptococcus neoformans (Serotype A, n = 8 and n = 50 respectively) and Cryptococcus gattii (Serotype B, n = 01 and n = 04 respectively). Susceptibilities to fluconazole, itraconazole and ketoconazole were determined by using NCCLS broth micro-dilution methodology. The total number of resistant strains for fluconazole in case of C. neoformans and C. gattii showed a significant difference by using chi-square test (p < 0.05*), while considering fisher's exact p value was nonsignificant (p > 0.05). However, the total number of resistant strains for itraconazole and ketoconazole was not found statistically significant. A comparison of geometric means of clinical and environmental strains of C. gattii and C. neoformans was not found statistically significant using student ‘t’ test (p value > 0.05 NS). Though less, the antifungal data obtained in this study suggests that primary resistance among environmental and clinical isolates of C. neoformans and C. gattii against tested antifungal was present and C. gattii comparatively was less susceptible than C. neoformans var. grubii isolates to fluconazole than to itraconazole and ketoconazole. A continuous surveillance of antifungal susceptibility of clinical and environmental isolates of C. neoformans and C. gattii is desirable to monitor the emergence of any resistant strains for better management of cryptococcosis patients.

Acute invasive pulmonary aspergillosis, shortly after occupational exposure to polluted muddy water, in a previously healthy subject / Aspergilose pulmonar invasiva aguda, logo após exposição ocupacional a água poluída barrenta, em indivíduo previamente saudável

Invasive pulmonary aspergillosis (IPA) predominantly occurs in severely neutropenic immunocompromised subjects. The occurrence of acute IPA after brief but massive exposure to Aspergillus conidia in previously healthy subjects has been documented, although only six such cases have been reported. The diagnosis was delayed in all six of the affected patients, five of whom died. We report the case of a 50-year-old HIV-negative male, a water pipeline maintenance worker, who presented with acute-onset dyspnea and fever one day after working for 2 h in a deep pit containing polluted, muddy water. Over a one-month period, his general condition deteriorated markedly, despite antibiotic therapy. Imaging showed bilateral diffuse nodules with cavitation, some of which were surrounded by ground-glass opacity suggestive of a halo sign (a hallmark of IPA). Cultures (of sputum/bronchial aspirate samples) and serology were positive for Aspergillus fumigatus. After being started on itraconazole, the patient improved. We conclude that massive exposure to Aspergillus conidia can lead to acute IPA in immunocompetent subjects.

A aspergilose pulmonar invasiva (API) ocorre predominantemente em indivíduos imunocomprometidos com neutropenia grave. A ocorrência de API aguda após exposição breve, mas maciça, a conídios de Aspergillus sp. em indivíduos previamente saudáveis já foi documentada, embora apenas seis casos tenham sido relatados. O diagnóstico foi tardio em todos os seis pacientes afetados, dos quais cinco foram a óbito. Relatamos o caso de um homem de 50 anos de idade, HIV negativo, trabalhador de manutenção de tubulação de água, que apresentou dispneia e febre de início agudo um dia após trabalhar 2 h em uma vala funda contendo água poluída e barrenta. Num período de um mês, seu estado geral se deteriorou acentuadamente, apesar da antibioticoterapia. Exames de imagem mostraram nódulos bilaterais difusos com cavitação, alguns dos quais circundados por opacidade em vidro fosco sugestiva de sinal do halo (uma característica da API). As culturas (de amostras de escarro/aspirado brônquico) e a sorologia foram positivas para Aspergillus fumigatus. Após iniciado o tratamento com itraconazol, o paciente melhorou. Concluímos que a exposição maciça a conídios de Aspergillus pode levar a API em indivíduos imunocompetentes.

Efficacy of moxifloxacin & econazole against multidrug resistant (MDR) Mycobacterium tuberculosis in murine model.

Background & objectives: studies have shown the bactericidal potential of econazole and clotrimazole against Mycobacterium tuberculosis under in vitro and ex vivo conditions along with their synergism with conventional antituberculosis drugs. these molecules were also found to be effective against different multidrug resistant (MDR) M. tuberculosis isolates in vitro. Hence the present study was designed to evaluate the in vivo antimycobacterial potential of moxifloxacin and econazole alone and in combination against multidrug resistant tuberculosis (MDR-TB) in a mice model. Methods: Mice were infected with 2.5×107 bacilli of MDR strain of M. tuberculosis by aerosol route of infection. After four weeks of infection, chemotherapy was started orally by moxifloxacin 8.0 mg/kg body wt and econazole 3.3 mg/kg alone and in combination, as well as with four first line anti-tuberculosis drugs as a positive control. The animals were sacrificed and the lungs and spleen were excised under aspetic conditions. The tissues were homogenized with sterile normal saline, an aliquot of the homogenate was plated on Middlebrook 7H11 agar supplemented with oleate albumin dextrose catalase (OADC) and incubated at 37°C for four weeks. The number of visible and individual colonies were counted. Results: The first line anti-tuberculosis drugs (RIF+INH+EMB+PZA) after eight weeks of therapy had no impact as the bacillary load in lungs and spleens remained unchanged. However, econazole, moxifloxacin alone as well as in combination significantly reduced the bacillary load in lungs as well as in spleens of MDR-TB bacilli infected mice. Interpretation & conclusions: Co-administration of the two drugs (econazole and moxifloxacin) to MDR-TB strain JAL-7782 infected mice exhibited additive effect, the efficacy of the drugs in combination being higher as compared with ECZ or MOX alone. These results were substantiated by histopathological studies. This study suggests the utility of econazole for the treatment of MDR tuberculosis and warrants further work in this direction.

ERG11 mutations associated with azole resistance in Candida albicans isolates from vulvovaginal candidosis patients

Objective: To investigate the azole susceptibility of Candida albicans ( C. albicans) from vulvovaginal candidosis patients and to analyze the relationship between ERG11 gene mutations in these isolates and azole resistance. Methods: Three hundred and two clinical isolates of Candida species were collected. Azole susceptibility was tested in vitro in microdilution studies. The ERG11 genes of 17 isolates of C. albicans (2 susceptibles, 5 dose-dependent resistants and 10 resistants) were amplified and sequenced. Results: Of the 302 isolates collected, 70.2% were C. albicans, of which 8.5%, 3.8% and 4.2% were resistant to fluconazole, itraconazole and voriconazole, respectively. In total, 27 missense mutations were detected in ERG11 genes from resistant/susceptible dose-dependent isolates. Among them, Y132H, A114S, and Y257H substitutions were most prevalent and were known to cause fluconazole resistance. G464S and F72S also have been proved to cause fluconazole resistance. Two novel substitutions (T285A, S457P) in hotspot regions were identified. Conclusions: Twenty seven mutations in the ERG11 gene were identified in azole-resistant C. albicans isolates, which indicated a possible relation with the increase in resistance to azole drugs and the recurrence of vulvovaginal candidosis. The relationship of two novel substitutions (T285A, S457P) with fluconazole resistance needs to be further verified by site-directed mutagenesis.

ERG11 mutations associated with azole resistance in Candida albicans isolates from vulvovaginal candidosis patients

Objective:To investigate the azole susceptibility of Candida albicans (C. albicans) from vulvovaginal candidosis patients and to analyze the relationship between ERG11 gene mutations in these isolates and azole resistance. Methods:Three hundred and two clinical isolates of Candida species were collected. Azole susceptibility was tested in vitro in microdilution studies. The ERG11 genes of 17 isolates of C. albicans (2 susceptibles, 5 dose-dependent resistants and 10 resistants) were amplified and sequenced. Results:Of the 302 isolates collected, 70.2%were C. albicans, of which 8.5%, 3.8%and 4.2%were resistant to fluconazole, itraconazole and voriconazole, respectively. In total, 27 missense mutations were detected in ERG11 genes from resistant/susceptible dose-dependent isolates. Among them, Y132H, A114S, and Y257H substitutions were most prevalent and were known to cause fluconazole resistance. G464S and F72S also has been proved to cause fluconazole resistance. Two novel substitutions (T285A, S457P) in hotspot regions were identified. Conclusions:Twenty seven mutations in the ERG11 gene were identified in azole-resistant C. albicans isolates, which indicated a possible relation with the increase in resistance to azole drugs and the recurrence of vulvovaginal candidosis. The relationship of two novel substitutions (T285A, S457P) with fluconazole resistance needs to be further verified by site-directed mutagenesis.