棘白菌素对氟康唑耐药的念珠菌体外药物敏感性的研究

目的评价3种棘白菌素类药物（卡泊芬净、米卡芬净、阿尼多芬净）体外对氟康唑耐药念珠菌的药物敏感性。方法采用微量液体稀释法和琼脂稀释法测定最小抑制浓度（MIC）。结果微量液体稀释法：59株耐药白念珠菌3种药物MIC50均为0.06μg/mL,米卡芬净、阿尼多芬净的MIC范围均为0.015～0.125μg/mL,卡泊芬净为0.015～0.25μg/mL;8株耐药光滑念珠菌MIC值均为0.063μg/mL。琼脂稀释法：59株耐药白念珠菌和8株耐药光滑念珠菌3种药物MIC值均为0.063μg/mL。结论3种棘白菌素类药物可能具有治疗氟康唑耐药的念珠菌感染的临床价值。     

白念珠菌耐药机制新进展

近年来对于深部真菌感染的研究报道越来越多,其已日益成为一些重要疾病临床治疗过程中的常见并发症,其中,白念珠菌病的发病率仍居高不下.虽然目前有多种抗真菌药物应用于临床,但其耐药现象愈来愈严重,给临床治疗带来了极大的挑战.近来有关白念珠菌耐药机制的研究有了较新的进展.该文就新发现的白念珠菌的耐药机制,作一概述.     

棘白菌素B提取液脱色工艺

研究碱性阴离子交换树脂WD-6对发酵棘白菌素B(ECB)提取液的脱色效果。通过对ECB提取液和ECB标准溶液的全波长扫描,确定ECB提取液中色素的最佳吸收波长为315nm;考察了树脂添加量、脱色温度、摇床转速和脱色时间对脱色率和ECB损失率的影响,获得最佳脱色工艺:树脂添加量4%(质量分数),脱色温度30℃,摇床转速150r/min,脱色时间100min。在此条件下,ECB提取液的脱色率可达88.3%,ECB损失率仅为4.8%。     

白念珠菌耐药的分子机制研究进展

近年来,免疫受损人群不断增多,该人群念珠菌病发病率呈上升趋势。随着抗真菌药物的广泛应用,临床分离到的白念珠菌耐药株增多,有关白念珠菌对抗真菌药物的耐药机制的研究又有了进一步的进展。就白念珠菌对唑类、多烯类、5-氟胞嘧啶、棘白菌素类等抗真菌药物的耐药机制方面的研究进展,作了介绍。     

光滑念珠菌致病性和耐药机制研究进展

<正>光滑念珠菌(Candida glabrata)是一种黏膜表面定植的机会致病菌,可引起侵袭性念珠菌病。光滑念珠菌是仅次于白念珠菌的念珠菌病相关第二大病原体^[1]。其高发病率与HIV、肿瘤、糖尿病患者数量的增加有关,人口老龄化和侵入性治疗也会增加感染风险。光滑念珠菌与致病性相关的毒力因子包括对宿主细胞及医疗器械黏附性、生物膜形成、逃避宿主防御、复制性老化、分泌水解酶(包括蛋白酶、磷脂酶、酯酶和溶血素等)等,在感染过程中发挥重要作用^[2]。此外,光滑念珠菌对抗真菌药物具有普遍耐受性,常常导致治疗的失败^[3]。由于致病性与耐药性的完美组合,使光滑念珠菌感染呈现出“发病率高、病死率高、耐药率高”的现象,需引起足够重视^[4]。本文就光滑念珠菌致病相关毒力因子和耐药机制研究进展进行综述。     

白念珠菌耐药机制研究进展CSCD

白念珠菌是侵袭性念珠菌病最常见的致病菌,其耐药问题使临床治疗面临着严峻的挑战。白念珠菌常见的耐药机制包括药物靶点突变或上调、药物外排增加、生物被膜形成等,近年来代谢调节、线粒体功能改变、选择性剪切等机制也受到了广泛关注。了解白念珠菌耐药机制有助于探索研究全新结构的抗真菌药物和开发更多有效的抗耐药真菌策略。该文就白念珠菌耐药机制研究进展进行综述。     

白念珠菌生物膜与侵袭及耐药

为了解近年来白念珠菌(Candida a lbicans)表面生长及生物膜形成研究的动向和进展,本文从几个方面阐述了白念珠菌表面生长和生物膜形成的生理病理学及其抗真菌耐药性的特征。包括白念珠菌宿主表面黏附、菌丝转换基因、微菌落定量感触调节、外分泌酶降解作用、菌丝地形趋向性和抗真菌耐药形成机制。结论提示,白念珠菌生物膜形成是其表面生长的重要结构,是临床上医疗植入物发生血源性白念珠菌感染传播的主要诱因,也是抗真菌耐药形成的重要因素,具有重要的病理学和生物学意义。     

光滑念珠菌的生物学、流行病学以及耐药机制研究进展

随着HIV感染患者的增多、器官移植、放疗化疗及抗真菌药物的广泛使用,近年来全世界范围内念珠菌感染趋势发生了明显变化,除白念珠菌外,光滑念珠菌在临床上的检出率逐年增加,在部分国家和地区已成为第二常见的侵袭性念珠菌。光滑念珠菌临床分离株通常对一线抗真菌药物高度耐药,由于目前治疗策略匮乏,其造成的系统感染死亡率可高达50%。为了进一步加深人们对光滑念珠菌的认识,研发遏制其感染的诊疗策略,本文综述了近年来光滑念珠菌的流行病学、毒力因子以及耐药机制等方面的进展,为国内同行深入探究其耐药特性和致病机理提供参考。     

棘白菌素B脱酰基酶的研究

棘白菌素B脱酰基酶是合成阿尼芬净重要中间产物棘白菌素B母核的关键酶。本文综述了近年来国内外学者对该酶的成熟机制、酶学性质、重组表达及生物催化应用等方面的相关研究进展。     

微粒添加对棘白菌素B发酵过程的影响

阿尼芬净是一种新型的抗真菌药物,能够抑制各种致病念珠菌在活体内外的活性。棘白菌素B(Echinocandin B,ECB)是合成阿尼芬净的关键前体,其发酵单位的高低直接关系到阿尼芬净的价格及市场前景。文中考察了构巢曲霉在摇瓶发酵生产ECB的过程中,添加滑石粉、Al2O3、玻璃珠等微粒对其发酵单位的影响。发现微粒的粒径和添加浓度是菌体形态和ECB产量的关键影响因素,添加20 g/L滑石粉(d50=14.2μm)和7颗玻璃珠(d=6 mm)可使ECB发酵产量分别比对照提高33.2%和41.7%,达到1 262.9 mg/L和1 344.1mg/L。结果表明微粒的添加可以显著地改善丝状微生物发酵过程中的菌丝形态,提高其产物的发酵产量,为丝状微生物发酵过程的优化提供了一种重要手段。     

Green synthesis of selenium nanoparticles and evaluate their effect on the expression of ERG3, ERG11 and FKS1 antifungal resistance genes in Candida albicans and Candida glabrata

Drug resistance in Candida species has been considerably increased in the last decades. Given the opposition to antifungal agents, toxicity and interactions of the antimicrobial drugs, identifying new antifungal agents seems essential. This study assessed the antifungal effects of nanoparticles (NPs) on the standard strains of Candida albicans and Candida glabrata and determined the expression genes, including ERG3, ERG11 and FKS1. Selenium nanoparticles (Se-NPs) were biosynthesized with a standard strain of C. albicans and approved by several methods including, ultraviolet-visible spectrophotometer, X-ray diffraction technique, Fourier-transform infrared analysis, field-emission scanning electron microscopy and EDX diagram. The antifungal susceptibility testing performed the minimum inhibitory concentrations (MICs) using the CLSI M27-A3 and M27-S4 broth microdilution method. The expression of the desired genes was examined by the real-time PCR assay between untreated and treated by antifungal drugs and Se-NPs. The MICs of itraconazole, amphotericin B and anidulafungin against C. albicans and C. glabrata were 64, 16 and 4 µg ml⁻¹. In comparison, reduced the MIC values for samples treated with Se-NPs to 1 and 0·5 µg ml⁻¹. The results obtained from real-time PCR and analysis of the ∆∆C_q values showed that the expression of ERG3, ERG11 and FKS1 genes was significantly down-regulated in Se-NPs concentrations (P < 0·05). This study's evidence implies biosafety Se-NPs have favourable effects on the reducing expression of ERG3, ERG11 and FKS1 antifungal resistance genes in C. albicans and C. glabrata.     

A purine permease in Candida glabrata

Abstract Competition experiments revealed that adenine and guanine were transported by a purine permease in both Candida glabrata 4 and a C. glabrata 4 cytosine permease negative mutant. The C. glabrata 4 cytosine permease negative mutant was isolated using 5-fluorocytosine selection. This mutant no longer transported cytosine, but transported adenine and guanine. A transport system for hypoxanthine was not detected. Hence, in addition to the cytosine permease, a purine permease exists in C. glabrata . This differs from the purine cytosine permeases in Saccharomyces cereuisiae and Candida albicans which transport adenine, cytosine, guanine and hypoxanthine.     

Silver nanoparticles: influence of stabilizing agent and diameter on antifungal activity against Candida albicans and Candida glabrata biofilms

Aim: The purpose of this work was to evaluate the size‐dependent antifungal activity of different silver nanoparticles (SN) colloidal suspensions against Candida albicans and Candida glabrata mature biofilms. Methods and Results: The research presented herein used SN of three different average sizes (5, 10 and 60 nm), which were synthesized by the reduction of silver nitrate through sodium citrate and which were stabilized with ammonia or polyvinylpyrrolidone. Minimal inhibitory concentration (MIC) assays were performed using the microdilution methodology. The antibiofilm activity of SN was determined by total biomass quantification (by crystal violet staining) and colony forming units enumeration. MIC results showed that all SN colloidal suspensions were fungicidal against the tested strains at very low concentrations (0·4–3·3 μg ml^?1). With regard to biomass quantification, SN colloidal suspensions were very effective only against C. glabrata biofilms, achieving biomass reductions around 90% at a silver concentration of 108 μg ml^?1. In general, all SN suspensions promoted significant log₁₀ reduction of the mean number of cultivable biofilm cells after exposure to silver concentrations at or higher than 108 μg ml^?1. Moreover, the results showed that the particle size and the type of stabilizing agent used did not interfere in the antifungal activity of SN against Candida biofilms. Conclusions: This study suggests that SN have antifungal therapeutic potential, but further studies are still required namely regarding formulation and delivery means. Significance and Impact of the Study: SN may contribute to the development of new strategies for the improvement of oral health and quality of life particularly of the complete denture wearers.     

住院患者光滑念珠菌检出的回顾性分析

目的调查住院患者光滑念珠菌检出的特征。方法回顾性调查分析白求恩国际和平医院2008年1月～2009年4月住院患者中光滑念珠菌检出阳性者的临床资料,以同期白念珠菌检出患者为对照。结果其间共有52例详细病史资料记录的光滑念珠菌检出患者,以60岁以上老年人为主,占65.4%;主要分离自痰标本,占76.9%。患者患有多种基础疾病,以肺部感染(28例,53.8%)、恶性肿瘤(20例,38.5%)、脑梗死(15例,28.8%)常见。使用抗生素(52例,100%)、留置导尿管(15例,28.8%)是光滑念珠菌检出者的主要实施医疗措施。氟康唑是临床最常用的治疗光滑念珠菌感染药物(23例,44.2%)。光滑念珠菌检出患者死亡率高(14例,26.9%),高于同期白念珠菌检出对照组(6.2%,P=0.004)。结论光滑念珠菌检出患者与白念珠菌检出具有相似的临床流行病学特征。     

Silver colloidal nanoparticles: antifungal effect against adhered cells and biofilms of Candida albicans and Candida glabrata

The aim of this study was to evaluate the effect of silver nanoparticles (SN) against Candida albicans and Candida glabrata adhered cells and biofilms. SN (average diameter 5 nm) were synthesized by silver nitrate reduction with sodium citrate and stabilized with ammonia. Minimal inhibitory concentration (MIC) tests were performed for C. albicans (n = 2) and C. glabrata (n = 2) grown in suspension following the Clinical Laboratory Standards Institute microbroth dilution method. SN were applied to adhered cells (2 h) or biofilms (48 h) and after 24 h of contact their effect was assessed by enumeration of colony forming units (CFUs) and quantification of total biomass (by crystal violet staining). The MIC results showed that SN were fungicidal against all strains tested at very low concentrations (0.4–3.3 μg ml^?1). Furthermore, SN were more effective in reducing biofilm biomass when applied to adhered cells (2 h) than to pre-formed biofilms (48 h), with the exception of C. glabrata ATCC, which in both cases showed a reduction ～90%. Regarding cell viability, SN were highly effective on adhered C. glabrata and respective biofilms. On C. albicans the effect was not so evident but there was also a reduction in the number of viable biofilm cells. In summary, SN may have the potential to be an effective alternative to conventional antifungal agents for future therapies in Candida-associated denture stomatitis.     

Susceptibility of Candida glabrata biofilms to echinocandins: alterations in the matrix composition

Candidiases are the most recurrent fungal infections, especially among immunosuppressed patients. Although Candida albicans is still the most widespread isolated species, non-Candida albicans Candida species have been increasing. The goal of this work was to determine the susceptibility of C. glabrata biofilms to echinocandins and to evaluate their effect on the biofilm matrix composition, comparing the results with other Candida species. Drug susceptibilities were assessed through the determination of minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC) and minimum biofilm eradication concentration (MBEC) of caspofungin (Csf) and micafugin (Mcf). The β-1,3 glucans content of the matrices was assessed after contact with the drugs. The data suggest that, generally, after contact with echinocandins, the concentration of β-1,3 glucans increased. These adjustments in the matrix composition of C. glabrata biofilms and the chemical differences between Csf and Mcf, seem responsible and may determine the effectivity of the drug responses.     

Pdr1蛋白N端携带Flag标签的光滑假丝酵母的构建

近年来,光滑假丝酵母已成为第二位引起侵袭性真菌感染的病原体。光滑假丝酵母对唑类药物(临床一线抗真菌药物)的敏感性低且易发生耐药,一直是研究的热点。介导光滑假丝酵母对唑类药物耐药的关键基因是转录因子pdr1,其功能性突变会使Pdr1蛋白功能过度活跃,导致下游唑类药物外排泵基因高表达,从而对唑类药物耐药。本研究利用同源重组技术,构建在基因pdr1的5′端定点插入3×Flag标签的重组菌株2a2和2b2,为后续利用免疫染色质共沉淀技术寻找Pdr1直接调控基因奠定基础。结果表明,3×Flag标签添加到Pdr1蛋白N端可成功表达Flag-Pdr1蛋白;与野生型菌株相比,表达Flag-Pdr1的菌株对氟康唑的耐药性增强。此外,与野生型菌株相比,表达Flag-Pdr1的菌株中cdr1和pup1基因表达水平显著上升,提示在Pdr1蛋白N端加Flag标签能使其功能活跃,表明N端对Pdr1蛋白功能具有重要意义。     

Effects of fluconazole on Candida glabrata biofilms and its relationship with ABC transporter gene expression

Candida glabrata has emerged as the second most prevalent fungal pathogen and its ability to form biofilms has been considered one of the most important virulence factors, since biofilms present a high tolerance to antifungal agents used in fungal infection treatment. The mechanisms of biofilm tolerance to antifungal agents remain poorly understood. Thus, the aim of this study was to evaluate the effects of fluconazole (FLU) on the formation and control of C. glabrata biofilms and its relation with the expression of genes encoding for ABC transporters, CDR1, SNQ2, and PDR1. For that, minimal inhibitory concentration values for seven C. glabrata strains were determined and the effect of FLU against C. glabrata biofilms was evaluated by total biomass quantification and viable cell enumeration. Matrices from biofilms were analyzed in terms of protein, carbohydrate and DNA content. ABC transporter gene expression was analyzed for quantitative real-time PCR. In addition to the high amounts of proteins and carbohydrates detected in the extracellular matrices in the presence of FLU, this work showed that the overexpression of efflux pumps is a possible mechanism of biofilm tolerance to FLU and this phenomenon alters the structure of C. glabrata biofilms by creating cell clusters.