In vitro antifungal activity of anidulafungin

Anidulafungin is a new and very useful pharmacological tool for the treatment of invasive mycoses. The antifungal spectrum of anidulafungin reaches the most common pathogenic fungi. Anidulafungin is especially active against the genera Candida and Aspergillus. Its antifungal mechanism is based on the inhibition of the beta-1,3-D-glucan synthesis, an essential molecule for the cell wall architecture, with different consequences for Candida and Aspergillus, being anidulafungin fungicide for the former and fungistatic for the latter. This review describes the in vitro antifungal spectrum of anidulafungin based in the scientific and medical literature of recent years. We can underline that most than 99% of Candida isolates are susceptible to < or = 2 microg/ml of anidulafungin. MIC are very low (< or =0.125 microg/ml) for most clinical isolates of the species Candida albicans, Candida glabrata, Candida tropicalis and Candida krusei while Candida parapsilosis and Candida guilliermondii isolates are susceptible to anidulafungin concentrations < or = 2 microg/ml. An excellent activity of anidulafungin has been also described against Aspergillus, Pneumocystis and other fungi. However, its activity is very low against Cryptococcus and the Zygomycetes. The excellent activity of anidulafungin has made this antifungal a first line therapeutic indication for candidemia and invasive candidiasis in non-neutropenic patients.     

In vitro susceptibility of fungal and yeast clinical isolates to itraconazole and voriconazole

The interest on the in vitro susceptibility to itraconazole has recently increased due the availability of the intravenous formulation. In this study, comparative MICs of this antifungal with voriconazole were carried out in 62 clinical isolates of filamentous fungi and 100 yeasts isolates using the NCCLS microbroth methods described in M38-A and M27-A2 documents. A MIC90 of 0.125 micrograms per ml was observed for itraconazole and voriconazole against Aspergillus fumigatus. Higher susceptibility to itraconazole was found for the filamentous form of Sporotrhix schenckii (p = 0.001). Voriconazole was more effective against Scedosporium apiospermium while Scedosporium prolificans isolates were resistant to both azoles. Some isolates of Rhizopus stolonifer were susceptible to itraconazole and resistant to voriconazole, but without statistical significance. Susceptibility of nine species of Candida was similar for both triazoles used in this study. However, Candida glabrata was more susceptible to voriconazole. Some fluconazole-resistant Candida albicans isolates were susceptible to itraconazole and / or voriconazole. Cryptococcus neoformans was more susceptible to itraconazole than to voriconazole. Itraconazole and voriconazole showed very close in vitro activity against the tested fungal isolated, except against S. schenckii. In spite of this, there were some differences in susceptibility among isolates within the same fungal species.     

Design,synthesis, and structure-activity relationship studies of novel triazole agents with strong antifungal activity against Aspergillus fumigatus

The incidence of invasive fungal infections has dramatically increased for several decades. In order to discover novel antifungal agents with broad spectrum and anti-Aspergillus efficacy, a series of novel triazole derivatives containing 1,2,3-benzotriazin-4-one was designed and synthesized. Most of the compounds exhibited stronger in vitro antifungal activities against tested fungi than fluconazole. Moreover, 6m showed comparable antifungal activity against seven pathogenic strains as voriconazole and albaconazole, especially against Aspergillus fumigatus (MIC = 0.25 μg/ml), and displayed moderate antifungal activity against fluconazole-resistant strains of Candida albicans. A clear SAR study indicated that compounds with groups at the 7-position resulted in novel antifungal triazoles with more effectiveness and a broader-spectrum.     

The discovery of enfumafungin, a novel antifungal compound produced by an endophytic Hormonema species biological activity and taxonomy of the producing organisms

In a screening of natural products with antifungal activity derived from endophytic fungi, we detected a potent activity in a culture belonging to the form-genus Hormonema, isolated from leaves of Juniperus communis. The compound is a new triterpene glycoside, showing an antifungal activity highly potent in vitro against Candida and Aspergillus and with moderate efficacy in an in vivo mouse model of disseminated candidiasis. The agent is especially interesting since its antifungal spectrum and its effect on morphology of Aspergillus fumigatus is comparable to that of the glucan synthase inhibitor pneumocandin B,,, the natural precursor of the clinical candidate MK-0991 (caspofungin acetate). An additional search for other Hormonema isolates producing improved titers or derivatives resulted in the isolation of two more strains recovered from the same plant host showing identical activity. The producing isolates were compared with other non-producing Hormonema strains by DNA fingerprinting and sequencing of the rDNA internal transcribed spacers. Comparison of rDNA sequences with other fungal species suggests that the producing fungus could be an undetermined Kabatina species. Kabatina is a coelomycetous genus whose members are known to produce Hormonema-like states in culture.     

Modern antifungals in therapy of nosocomial mycosis in oncologic patients

Rational position of voriconazole in the treatment of oncologic inpatients was shown and the criteria of its use in the algorithms of the therapy and prophylaxis of nosocomial fungal infections were developed. The clinical trial enrolled 50 patients with oncologic pathologies. The patients were divided into two groups of possible invasive candidiasis risk. The patients of one group were treated with fluconazole (Diflucan) and those of the other group were treated with voriconazole (Vifend). The spectrum of the hospital fungal flora was determined and susceptibility of 310 clinically important opportunistic fungi was investigated. All the isolates of Candida albicans and C.tropicalis were susceptible to amphotericin B, fluconazole and voriconazole and 79 and 50% of the isolates were susceptible to intraconazole respectively. As for the C.krusei isolates, 67% was susceptible to amphotericin B, 50% was susceptible to fluconazole, 100% was susceptible to voriconazole and none of the strains was susceptible to intraconazole. By the clinical efficacy voriconazole was superior to fluconazole and comparable with amphotericin B, while superior to it by the number of the side effects and by the cost of the treatment course. It was concluded that voriconazole should be considered as the main agent in the antifungal therapy of oncologic patients.     

In vitro antifungal activities of anidulafungin and micafungin, licensed agents and the investigational triazole posaconazole as determined by NCCLS methods for 12,052 fungal isolates: review of the literature

The echinocandins anidulafungin and micafungin and the triazole posaconazole are currently undergoing phase III clinical trials. Caspofungin and voriconazole have recently been licensed for the treatment of aspergillosis (both agents), other less common mould (voriconazole) and candidal (caspofungin) infections. This review summarizes the published in vitro data obtained by NCCLS or NCCLS modified methods on the in vitro fungistatic and fungicidal activities of these five agents for yeasts and moulds in comparison to the established agents, amphotericin B, fluconazole, itraconazole, and flucytosine. Among the yeasts, the echinocandins have less activity for Candida parapsilosis and Candida guilliermondii, no activity for Cryptococcus neoformans and Trichosporon spp., but good fungistatic and fungicidal activity in vivo and in vitro for most of the other Candida spp.; this fungicidal activity has been reported by minimum fungicidal concentrations (MFCs) or time kill curve results. The new triazoles exhibit good fungistatic activity (but not fungicidal) for most Candida spp., C. neoformans, and Trichosporon spp. For the Aspergillus spp. evaluated, the echinocandins have similar or better fungistatic activity than those of amphotericin B and the triazoles, but fungicidal activity has been demonstrated only with amphotericin B and the triazoles, with the exception of fluconazole. Most studies showed posaconazole and voriconazole minimum inhibitory concentrations (MICs) ranging from 0.25 to 8 microg/ml for non-solani Fusarium spp., while MIC and minimum effective concentration (MEC) endpoints of the echinocandins were >8 microg/ml. The fungistatic activity of the triazoles is also superior to that of the echinocandins for most of the dimorphic fungi and the Zygomycetes. However, micafungin has activity for the mould phase of most dimorphic fungi, but not for the parasitic or yeast phase of Paracoccidioides brasiliensis. The echinocandins appear to have variable and species dependent fungistatic activity for the dematiaceous fungi, but all agents have poor or no activity against most isolates of Scedosporium prolificans. Only amphotericin B exhibit good fungistatic activity against the Zygomycetes. The combination of caspofungin with some triazoles, amphotericin B or liposomal amphotericin B has been synergistic in vitro, in animal models and in patients. Breakpoints are not available for any mould and antifungal agent combination. In vitro/in vivo correlations should aid in the interpretation of these results, but standard testing conditions are needed for the echinocandins, especially for mould testing, to obtain reliable results.     

In Vitro Antifungal Susceptibility of Oral <Emphasis Type="Italic">Candida</Emphasis> Isolates from Patients Suffering from Caries and Chronic Periodontitis

Caries and chronic periodontitis are common oral diseases where a higher Candida colonization is reported. Antifungal agents could be adjuvant drugs for the therapy of both clinical conditions. The aim of the current study has been to evaluate the in vitro activities of conventional and new antifungal drugs against oral Candida isolates from patients suffering from caries and/or chronic periodontitis. In vitro activities of amphotericin B, fluconazole, itraconazole, miconazole, nystatin, posaconazole and voriconazole against 126 oral Candida isolates (75 Candida albicans, 18 Candida parapsilosis, 11 Candida dubliniensis, six Candida guilliermondii, five Candida lipolytica, five Candida glabrata, four Candida tropicalis and two Candida krusei) from 61 patients were tested by the CLSI M27-A3 method. Most antifungal drugs were highly active, and resistance was observed in less than 5% of tested isolates. Miconazole was the most active antifungal drug, being more than 98% of isolates susceptible. Fluconazole, itraconazole, and the new triazoles, posaconazole and voriconazole, were also very active. Miconazole, fluconazole and voriconazole have excellent in vitro activities against all Candida isolates and could represent suitable treatment for a hypothetically adjunctive therapy of caries and chronic periodontitis.     

Structure-based rational design,synthesis and antifungal activity of oxime-containing azole derivatives

In an attempt to find novel azole antifungal agents with improved activity and broader spectrum, computer modeling was used to design a series of new azoles with piperidin-4-one O-substituted oxime side chains. Molecular docking studies revealed that they formed hydrophobic and hydrogen-bonding interactions with lanosterol 14α-demethylase of Candida albicans (CACYP51). In vitro antifungal assay indicates that most of the synthesized compounds showed good activity against tested fungal pathogens. In comparison with fluconazole, itraconazole and voriconazole, several compounds (such as 10c, 10e, and 10i) show more potent antifungal activity and broader spectrum, suggesting that they are promising leads for the development of novel antifungal agents.     

Broad and complex antifungal activity among environmental isolates of lactic acid bacteria

More than 1200 isolates of lactic acid bacteria isolated from different environments were screened for antifungal activity in a dual-culture agar plate assay. Approximately 10% of the isolates showed inhibitory activity and 4% showed strong activity against the indicator mould Aspergillus fumigatus. The antifungal spectra for 37 isolates with strong activity and five isolates with low or no activity were determined. Several of the strains showed strong inhibitory activity against the moulds A. fumigatus, Aspergillus nidulans, Penicillium commune and Fusarium sporotrichioides, and also against the yeast Rhodotorula mucilaginosa. Penicillium roqueforti and the yeasts Pichia anomala and Kluyveromyces marxianus were not inhibited. Several isolates showed reduced antifungal activity after storage and handling. The majority of the fungal inhibitory isolates were identified by 16S rDNA sequencing as Lactobacillus coryniformis. Lactobacillus plantarum and Pediococcus pentosaceus were also frequently identified among the active isolates. The degree of fungal inhibition was not only related to production of lactic or acetic acid. In addition, antifungal cyclic dipeptides were identified after HPLC separation and several other active fractions were found suggesting a highly complex nature of the antifungal activity.     

白念珠菌临床分离调查及基因分型研究

目的 了解白念珠菌临床分离情况,并探讨其药敏结果与基因分型的相关性.方法 回顾性分析本院2011年3～11月间临床分离白念珠菌分布及耐药性;随机选取232株,采用PCR方法扩增白念珠菌25S rDNA基因内含子区进行基因分型研究;采用ATB真菌药敏试剂条进行药敏分析;统计分析药敏结果与基因分型的相关性.结果 期间共检出酵母样真菌973例,占病原菌阳性样本数比率为15.7％ (973/6196);其中分离白念珠菌562株,占58％ (562/973),主要分布科室为呼吸科(39.1％)、老年科(13.2％)、ICU(7.7％)、神经内科(7.5％)、免疫科(6.0％)以及其他科室(26.5％);标本类型以下呼吸道为主(81.7％),其次为尿路(9.4％)、血液(1.8％)等.对氟胞嘧啶、两性霉素B、氟康唑、伊曲康唑及伏立康唑的耐药率分别为0.9％、0％、1.4％、1.6％和1.1％.随机选取的232株白念珠菌经PCR方法可分为3型:A型125株,B型96株,C型11株.各型在5种药物的耐药性上并无差异.结论 临床分离酵母样真菌以白念珠菌为主,感染部位以下呼吸道为主;临床分离株对5种抗真菌药物敏感度较高,主要基因型为A和B型,不同基因分型间药敏结果并无统计学差异.     

Antifungal activity of coumarins

The antifungal activity of 40 coumarins was tested against the fungal strains: Candida albicans (ATCC 14053), Aspergillus fumigatus (ATCC 16913) and Fusarium solani (ATCC 36031), using the broth microdilution method. Osthenol showed the most effective antifungal activity among all the compounds tested, with a MIC value of 125 microg/ml for Fusarium solani and 250 micro/ml for Candida albicans and Aspergillus fumigatus. The antifungal potential of this prenylated coumarin can be related to the presence of an alkyl group at C-8 position.     

Correlation between broth microdilution and disk diffusion methods for antifungal susceptibility testing of caspofungin, voriconazole, amphotericin B, itraconazole and fluconazole against Candida glabrata

Candida glabrata is one of the most frequent organisms isolated from superficial and invasive fungal infections, after Candida albicans. This organism also exhibits intrinsically low susceptibility to azole antifungals and treatment often fails. The microdilution method is not very practical for use in routine susceptibility testing in the clinical laboratory, thus necessitating the use of other methods. In this study, we compared the in vitro activity of five antifungal agents in three different groups (echinocandin, polyene and azole) against 50 C. glabrata isolates by broth microdilution and disk diffusion methods recommended by Clinical Laboratory Standards Institute CLSI M27-A3 and CLSI M44-A, respectively. All the isolates were susceptible to amphotericin B (100%) and 98% of the isolates were susceptible to caspofungin by the broth microdilution method. Within the azole group drugs, voriconazole was the most active followed by fluconazole and itraconazole in vitro. The highest rate of resistance was obtained against itraconazole with a high number of isolates defined as susceptible-dose dependent or resistant. Although the disk diffusion method is easy to use in clinical laboratories, it shows very poor agreement with the reference method for fluconazole and itraconazole against C. glabrata (8% and 14%, respectively).     

The activity of echinocandins, amphotericin B and voriconazole against fluconazole-susceptible and fluconazole-resistant Brazilian Candida glabrata isolates

The extensive use of azole antifungal agents has promoted the resistance of Candida spp to these drugs. Candida glabrata is a problematic yeast because it presents a high degree of primary or secondary resistance to fluconazole. In Brazil, C. glabrata has been less studied than other species. In this paper, we compared the activity of three major classes of antifungal agents (azoles, echinocandins and polyenes) against fluconazole-susceptible (FS) and fluconazole-resistant (FR) C. glabrata strains. Cross-resistance between fluconazole and voriconazole was remarkable. Among the antifungal agents, the echinocandins were the most effective against FS and FR C. glabrata and micafungin showed the lowest minimal inhibitory concentrations.     

Antifungal activity of Lactobacillus against Microsporum canis, Microsporum gypseum and Epidermophyton floccosum

A total of 220 lactic acid bacteria isolates were screened for antifungal activity using Aspergillus fumigatus and Aspergillus niger as the target strains. Four Lactobacillus strains exhibited strong inhibitory activity on agar surfaces. All four were also identified as having strong inhibitory activity against the human pathogenic fungi Microsporum canis, Microsporum gypseum and Epidermophyton floccosum. One of the four lactobacilli, namely Lb. reuteri ee1p exhibited the most inhibition against dermatophytes. Cell-free culture supernatants of Lb. reuteri ee1p and of the non-antifungal Lb. reuteri M13 were freeze-dried and used to access and compare antifungal activity in agar plate assays and microtiter plate assays. Addition of the Lb. reuteri ee1p freeze-dried cell-free supernatant powder into the agar medium at concentrations greater than 2% inhibited all fungal colony growth. Addition of the powder at 5% to liquid cultures caused complete inhibition of fungal growth on the basis of turbidity. Freeze-dried supernatant of the non-antifungal Lb. reuteri M13 at the same concentrations had a much lesser effect. As Lb. reuteri M13 is very similar to the antifungal strain ee1p in terms of growth rate and final pH in liquid culture, and as it has little antifungal activity, it is clear that other antifungal compounds must be specifically produced (or produced at higher levels) by the anti-dermatophyte strain Lb. reuteri ee1p. Reuterin was undetectable in all four antifungal strains. The cell free supernatant of Lb. reuteri ee1p was analyzed by LC-FTMS using an Accela LC coupled to an LTQ Orbitrap XL mass spectrometer. The high mass accuracy spectrum produced by compounds in the Lb. reuteri ee1p strain was compared with both a multianalyte chromatogram and individual spectra of standard anti-fungal compounds, which are known to be produced by lactic acid bacteria. Ten antifungal metabolites were detected.     

Novel fluconazole derivatives with promising antifungal activity

The fungistatic nature and toxicity concern associated with the azole drugs currently on the market have resulted in an increased demand for new azole antifungal agents for which these problematic characteristics do not exist. The extensive use of azoles has resulted in fungal strains capable of resisting the action of these drugs. Herein, we report the synthesis and antifungal activity of novel fluconazole (FLC) analogues with alkyl-, aryl-, cycloalkyl-, and dialkyl-amino substituents. We evaluated their antifungal activity by MIC determination and time-kill assay as well as their safety profile by hemolytic activity against murine erythrocytes as well as cytotoxicity against mammalian cells. The best compounds from our study exhibited broad-spectrum activity against most of the fungal strains tested, with excellent MIC values against a number of clinical isolates. The most promising compounds were found to be less hemolytic than the least hemolytic FDA-approved azole antifungal agent voriconazole (VOR). Finally, we demonstrated that the synthetic alkyl-amino FLC analogues displayed chain-dependent fungal membrane disruption as well as inhibition of ergosterol biosynthesis as possible mechanisms of action.     

Molecular Identification and Antifungal Susceptibilities of Black Aspergillus Isolates from Otomycosis Cases in Hungary

Otomycosis, also known as fungal otitis externa, has been used to describe a fungal infection of the external auditory canal, but sometimes involving the middle ear. Many fungal species have been identified as infectious agents in otomycosis, with Aspergillus and Candida species being the most common. Among aspergilli, Aspergillus niger is the most commonly described species in the literature. In this study, 14 black Aspergillus strains were analyzed, which were isolated from otomycosis cases in Hungary between 2010 and 2011. These strains were identified as A. niger according to conventional morphological methods. Species identification was based on sequencing of part of the calmodulin gene. Our results indicate that instead of A. niger, A. awamori and A. tubingensis are the predominant species that cause ear infections in Southern Hungary. Antifungal susceptibility tests were carried out against four antifungal drugs: amphotericin B, itraconazole, ketoconazole and terbinafine. All isolates were found to exhibit low in vitro MIC values to amphotericin B, terbinafine and itraconazole. However, the examined isolates exhibited high in vitro MIC values to ketoconazole.     

重症监护病房白念珠菌耐药性8年变化趋势

目的调查上海长征医院重症监护室(ICU)近8 a中临床分离白念珠菌的耐药性变化,为临床治疗提供参考。方法上海长征医院ICU 2002～2009年从414例患者中首次分离出414株白念珠菌,对其中277株进行药敏试验。采用Cox-Stuart趋势检验回顾性分析临床分离真菌中白念珠菌所占比例变化趋势和白念珠菌对常用抗真菌药物耐药率的变化趋势。结果 2002～2009年间,上海长征医院ICU白念珠菌分离株数从2002年的34株增加至2009年的92株,但白念珠菌占总真菌分离株数的百分比维持在34.6%～55.7%,P=0.03。白念珠菌对于5-氟胞嘧啶和两性霉素B平均耐药率分别为4.0%和0.7%,对其他常用抗真菌药的耐药率依次为咪康唑47.0%、酮康唑10.8%、伊曲康唑19.9%、特比萘芬42.6%、氟康唑14.6%及伏立康唑13.0%。白念珠菌对5-氟胞嘧啶和两性霉素B和伊曲康唑耐药率的8年变化无统计学差异。结论上海长征医院ICU近8 a来白念珠菌仍然为临床较为常见的真菌分离株,但白念珠菌占总分离株数的百分比有逐渐减少的趋势。白念珠菌对常用抗真菌药物耐药性均无明显变化趋势。     

Anidulafungin: a new therapeutic approach in antifungal therapy. Pharmacology of anidulafungin

Anidulafungin is a new echinocandin antifungal agent which inhibits beta-1,3-D-glucan synthase and disrupts fungal cell-wall synthesis. It has marked antifungal activity against Candida spp. and Aspergillus spp., including amphotericin B and triazole resistant strains. Due to the limited oral availability, anidulafungin in clinical use is available for parenteral administration only. Elimination of anidulafungin takes place via slow non-enzymatic degradation to inactive metabolites. Less than 10% and 1% of the initially administered drug is excreted unchanged into feces and urine, respectively. It does not require dosage adjustment in subjects with hepatic or renal impairment established. Anidulafungin is generally well tolerated. Adverse events appear not to be dose or infusion related. The most common treatment related adverse events are phlebitis, headache, nausea, vomiting and pyrexia. The lack of interactions with tacrolimus, cyclosporine and corticosteroids and its limited toxicity profile places anidulafungin as an attractive new option for the treatment of invasive fungal infections especially in transplant patients.     

Antifungal activity of synthetic di(hetero)arylamines based on the benzo[b]thiophene moiety

The antifungal activity of several di(hetero)arylamine derivatives of the benzo[b]thiophene system was evaluated against clinically relevant Candida, Aspergillus, and dermatophyte species by a broth macrodilution test based on CLSI (formerly NCCLS) guidelines. The most active compound showed a broad spectrum of activity (against all tested fungal strains, including fluconazole-resistant fungi), with particularly low MICs for dermatophytes. Results from the inhibition of the dimorphic transition in Candida albicans and flow cytometry studies further confirmed their biological activity. With this study it was possible to establish some structure-activity relationships (SARs). The hydroxy groups proved to be essential for the activity in the aryl derivatives. Furthermore, the spectrum of activity in the pyridine derivatives was broadened by the absence of the ester group on position 2 of the benzo[b]thiophene system.     

Clinical efficacy of new antifungal agents

Several new options are now available for treating serious fungal infections. All three echinocandin agents currently available have been shown in randomized, blinded clinical trials to be efficacious in treating candidemia and invasive candidiasis. By contrast, the demonstrated efficacy of the echinocandins for the treatment of invasive aspergillosis has been based on historically controlled salvage treatment trials in patients failing or intolerant of other therapies. The new triazole agents, voriconazole and posaconazole, have a broad spectrum of antifungal activity. Voriconazole has become the agent of choice for invasive aspergillosis. On the basis of compassionate treatment data, posaconazole appears to be effective for treatment of zygomycosis. These agents have also been shown to be effective in the treatment of non-Aspergillus mould infections, several of the endemic mycoses and serious Candida infections.