儿童真菌感染的病原学研究

目的探讨广州地区儿童真菌感染的病原分布特点及其耐药状况,为防治儿童真菌感染提供实验室依据。方法对患儿感染部位的真菌进行分离培养和鉴定：以ATB^TM FUNGUS3酵母样真菌药敏试验条进行常用抗真菌药物的敏感性分析。结果从患儿标本中分离出558株真菌,主要来自呼吸道有299株,占53．58％;其次是消化道、伤口（创口）、泌尿系统和血液等,分别占28．14％、6．27％、4．66％、3．76％。其中白色假丝酵母菌367株,占65．77％;其次为热带假丝酵母菌、光滑假丝酵母菌、近平滑假丝酵母菌、克柔假丝酵母菌、季也蒙假丝酵母菌等,分别占15．28％、5．02％、4．48％、3．41％、2．69％。从骨髓中检出5株马尔尼菲青霉,从脑脊液中检出3株新型隐球菌。真菌对两性霉素B、5-氟胞嘧啶、氟康唑、伊曲康唑、伏立康唑等总耐药率分别为8．78％、4．84％、10．54％、1．36％、0．85％。结论引起儿童真菌感染的主要病原菌是白色假丝酵母菌。对真菌感染应该有针对性地使用高效的抗真菌药物进行早期治疗。     

假丝酵母菌药敏实验及影响药物敏感性的因素分析

目的了解深圳地区育龄妇女外阴阴道假丝酵母菌的耐药情况及药物敏感性,探讨影响其药物敏感性的因素,为外阴阴道假丝酵母菌病（vulvovaginal candidiasis,VVC）的治疗提供参考依据。方法从体检妇女人群中阴道分泌物培养念珠菌感染阳性病例中选择103例为研究对象,对样本采用氟康唑、两性霉素B、伊曲康唑、酮康唑、益康唑进行药物敏感性实验;并回顾性调查检测对象病史、用药情况等。结果研究对象平均年龄（36.16±6.72）岁,文化程度以初中、高中和大专为主,分别占14.3%、29.7%和52.0%。对5种药物都敏感的样本有55例,占53.5%,对1种以上药物不敏感的样本有48例,占46.5%。药物敏感性依次是：两性霉素B（97.0%）、酮康唑（94.2%）、伊曲康唑（92.2%）、氟康唑（80.6%）、益康唑（72.8%）。近期使用抗生素更加容易使菌株对伊曲康唑和益康唑产生不敏感。使用抗生素的对象对两性霉素B、伊曲康唑、益康唑的耐药率分别为7.1%、14.3%和35.7%;而没有使用抗生素的对象对两性霉素B、伊曲康唑、益康唑的耐药率分别为2.3%、6.7%和25.8%。反复感染者的样本对两性霉素B（100.0%）和酮康唑（100.0%）非常敏感,但是会增加对伊曲康唑（8.7%）和益康唑（30.4%）的不敏感性。结论深圳地区外阴阴道假丝酵母菌整体耐药性较少,但近期使用抗生素和反复感染者对药物的敏感性有影响。临床治疗宜首选两性霉素B和酮康唑,反复感染者建议联合用药治疗。     

侵袭性真菌感染237例治疗分析

目的观察侵袭性真菌感染应用氟康唑、伊曲康唑和两性霉素B治疗的临床疗效及安全性。方法选择南方医科大学附属南方医院2002年2月至2007年2月收治的侵袭性真菌感染患者237例，单用常规剂量氟康唑107例、伊曲康唑84例、两性霉素B46例；氟康唑治疗无效者改用伊曲康唑39例和两性霉素B26例．伊曲康唑治疗无效改用两性霉素B和脂质体两性霉素B39例。结果237例侵袭性真菌感染总的治愈率和有效率分别为54．85％和72．57％。单用氟康唑治疗组的有效率显著低于伊曲康唑和两性霉素B治疗组，分别为39．25％、53．57％和56．52％（P〈0．05）；用氟康唑治疗无效改用伊曲康唑或两性霉素B治疗的有效率分别为76．93％和84．61％。结论伊曲康唑和两性霉素B抗真菌谱广且疗效相似，优于氟康唑；氟康唑和伊曲康唑安全性良好，而两性霉素B的不良反应发生率高。     

利血平对耐药结核分枝杆菌临床株的作用

目的观察泵抑制剂（利血平）对耐药结核分枝杆菌临床分离株的作用。方法应用3组BacT．ALERT3D培养仪系统的培养基。分别命名为A、B、C组培养基。3组培养基均接种42株耐药结核分枝杆菌临床分离株，其中A、B组同时加入利福平1μg／mL；环丙沙星2μg／mL；利福平1μg／mL和环丙沙星2μg／mL；A、C组同时加入利血平20mg／L：C组未加入任何抗结核药物。结果在A组培养基中，耐利福平1μg／mL的14株菌株中有2株恢复对利福平的敏感性，12株仍然耐药；耐环丙沙星2μg／mL的6株菌株和耐利福平1μg／mL及环丙沙星2μg／mL的22株菌株全部恢复药物敏感性。在B组培养基中，42株耐药结核分枝杆菌临床分离株．药物耐药性无变化。在C组培养基中，42株耐药结核分枝杆菌临床分离株明显生长，说明利血平对所研究菌株无抑制作用。A组与B组培养基细菌的药物敏感性差异有统计学意义（P〈0．01）。结论结核分枝杆菌存在耐药机制之一的外排系统；泵抑制剂之一利血平对耐药的结核分枝杆菌临床分离株有抑制其外排作用，使结核分枝杆菌恢复敏感性。     

811例酵母样真菌感染的临床分布及药敏分析

目的分析临床酵母样真菌感染的分布情况,结合真菌的鉴定及药敏,指导临床合理用药。方法回顾性分析真菌感染者的临床资料、检出的酵母样真菌的分布及对抗真菌药物的耐药特点。结果真菌感染以60岁以上老年人居多,占69.4%,感染部位主要为呼吸道、泌尿道及肠道;基本都有抗生素使用史,多有创伤性介入治疗。检出真菌主要为白色假丝酵母菌,共526例,另有热带假丝酵母菌117例,光滑假丝酵母菌86例,近平滑假丝酵母菌57例,其他酵母菌25例。10种抗真菌药物中,耐药性较高的为特比奈芬、制霉菌素和两性霉素B,比较敏感的为5-氟胞嘧啶和酮康唑。两性霉素B33.9%和氟康唑22.6%的耐药率,大大高于以往熟知的程度,这与临床大量常规使用其预防真菌感染有密切联系。结论应加强真菌检测,指导临床合理使用抗生素,减少多重耐药出现。     

重症监护病房感染病原菌分布及耐药性分析

调查医院重症监护病房（ICU）病原菌感染的分布及耐药趋势,为临床合理选用抗菌药物提供依据。本文对ICU患者病原菌采用自动微生物检定仪鉴定,K-B纸片扩散法进行药敏试验。结果表明,255株菌中,革兰氏阴性菌120株（占47.1%）,革兰氏阳性菌75株（占29.4%）,真菌60株（占23.5%）;革兰氏阴性菌中非发酵菌比例较高（占G-66.7%）,其中铜绿假单胞菌居首位（占43.0%）,其次是鲍曼不动杆菌（17.0%）;大肠埃希氏菌、肺炎克雷伯菌及阴沟肠杆菌共22株,占18.0%,其中产超广谱内酰胺酶（ESBLs）比率大于70.0%;革兰氏阳性菌以金黄色葡萄球菌为主,有29株,耐甲氧西林金黄色葡萄球菌（MRSA）26株,占89.7%;真菌60株,对5-氟胞嘧啶、两性霉素、氟康唑、伊曲康唑及氟利康唑均敏感。这说明ICU患者感染多耐药严重,加强耐药性监测、合理使用抗菌药物十分重要。     

艾滋病合并皮肤马尔尼菲青霉感染4例

目的探讨艾滋病（AIDS）合并皮肤马尔尼菲青霉感染的临床及实验室特征。方法分析2005年1月至2006年6月本院收治的明确诊断为AIDS合并皮肤马尔尼菲青霉感染的4例患者临床特征；取皮损、血和骨髓分别在25℃和37℃进行真菌培养。观察菌落形态、显微镜下特征；对皮肤活检组织行HE及六胺银染色。观察镜下皮损组织学及马尔尼菲青霉的特征。结果AIDS合并皮肤马尔尼菲青霉感染伴多系统损害。皮损特征：早期表现为淡红色丘疱疹、糜烂性丘疹，继而为坏死性丘疹、传染性软疣样丘疹、皮肤溃疡及血痂。37℃培养呈酵母相，25℃呈菌丝相。皮肤病理活检六胺银染色（＋）。使用二性霉素B、伊曲康唑治疗。3例临床症状缓解、皮疹消退出院。1例死亡。结论AIDS合并皮肤马尔尼菲青霉感染皮损特征：坏死性丘疹、传染性软疣样丘疹。皮损25℃、37℃真菌培养结合皮肤病理活检是确诊的关键。二性霉素B、伊曲康唑是目前治疗AIDS合并皮肤马尔尼菲青霉感染的首选药物。     

临床常见真菌的分布和药物敏感性分析

目的:了解真菌感染的临床分布、真菌种类及耐药情况,为临床诊治提供依据。方法:采用沙保弱平板或CHROMa-gar平板进行菌株分离、API Candida、API20CAUX菌株鉴定,用Rosco纸片扩散法测定药敏。结果:690份标本,分离出700株真菌,其中假丝酵母菌属占53.5%,白假丝酵母菌占25.6%;痰标本占24.3%;住院患者中以ICU和肾内科分离率高且以白假丝酵母菌为主,分别占59.3%和45.2%;门诊患者以皮肤科分离率高,以近平滑假丝酵母菌为主,占23.4%;酵母样真菌对伊曲康唑、氟康唑、两性霉素B、制霉菌素、酮康唑的敏感率分别为89.4%、83.9%、98.2%、95.9%、96.0%。结论:假丝酵母菌是真菌培养中分离率最高的真菌,分离的主要真菌种类因科室和患者而异。真菌的分离、鉴定和药敏试验对临床使用抗真菌药物具有指导意义,特别是对氟康唑天然耐药菌株的鉴定,有利于抗真菌药物选择。     

816株真菌感染分布及药敏分析

目的了解本院医院感染真菌分布的特点及对常用抗真菌药物的耐药情况。方法CHROMagar显色培养基及ATB真菌试剂盒进行鉴定;药敏试验采用纸片扩散K—B法。结果2008年1月至2009年6月我院临床共分离真菌816株,其中白色念珠菌（占53．2％）是引起真菌感染的最常见菌种,其次是热带念珠菌（21．6％）、光滑念珠菌（15．6％）、近平滑念珠菌（5．0％）和克柔念珠菌（2．3％）。药敏试验结果显示各种真菌对两性霉素B、制霉菌素的敏感性最高,分别达98％和99％,其次是氟康唑、伊曲康唑。结论真菌感染率呈逐年上升趋势,耐药率也逐渐增高。因此应及时对送检标本进行真菌培养和药敏试验,合理使用抗真菌药物,减少医院感染多重耐药和深部真菌感染的发生。     

52株生殖道念珠菌病病原菌的鉴定及药敏分析

目的分析女性生殖道念珠菌感染状况以及耐药性情况。方法临床标本接种到沙保弱培养基,分离培养出的真菌经革兰染色、芽管形成试验、CHROMagar显色培养进行鉴定,并进行氟康唑、两性霉素B药敏分析。结果120份临床标本分离培养出念珠菌52株。白色念珠菌占69.2%,热带念珠菌占7.7%,克柔念珠菌占5.8%,其他念珠菌占17.3%;分离出的52株念珠菌对氟康唑、两性霉素B敏感率分别为86.5%、94.2%。结论女性生殖道念珠菌感染仍以白色念珠菌为主。念珠菌对氟康唑和两性霉素B的敏感性有差异,应重视念珠菌的培养鉴定和药敏试验,以指导临床合理选择抗真菌药物。     

In vitro antifungal-drug susceptibilities of mycelial and yeast forms of Penicillium marneffei isolates in Cambodia

Penicillium marneffei infection is an important disease among human immunodeficiency virus patients in Southeast Asia. The in vitro antifungal-drug susceptibilities of 29 clinical isolates and 5 isolates from bamboo rats collected from 2002 to 2004 were determined. The P. marneffei yeast form is more susceptible than the mycelial form to amphotericin B and ketoconazole, while the mycelial and yeast forms displayed similar susceptibilities to flucytosine and itraconazole. The MICs of fluconazole were higher for both mycelial and yeast forms.     

In-vitro susceptibility testing by agar dilution method to determine the minimum inhibitory concentrations of amphotericin B, fluconazole and ketoconazole against ocular fungal isolates

Purpose: To standardize in-vitro antifungal susceptibility testing by agar dilution method to find out the minimum inhibitory concentration (MIC) of amphotericin B, fluconazole and ketoconazole on ocular fungal isolates. Methods: A total of 180 ocular fungal isolates (130 filamentous fungi and 50 yeasts) were included. The antifungal drugs such as amphotericin B (0.0625-8 μg/mL), fluconazole (0.2-819.6 μg/mL) and ketoconazole (0.025-6.4 μg/mL) were incorporated in doubling dilutions in the yeast nitrogen base medium. The MIC was determined as the lowest concentration of the antifungal drug preventing growth of macroscopically visible colonies on drug containing plates when there was visible growth on the drug - free control plates. Results: All 50 ocular isolates of yeast were susceptible to amphotericin B, while two (4%) and five (10%) strains were resistant to fluconazole and ketoconazole respectively. Of the 130 filamentous fungi tested, six (4.6%) were resistant to amphotericin B, 49 (37.7%) and 10 (7.6%) were resistant to fluconazole and ketoconazole respectively. Percentile 50 (MIC 50) and Percentile 90 (MIC 90) for all the three antifungal agents were calculated. Aspergillus niger, Aspergillus terreus and Candida krusei were found to be resistant to fluconazole and ketoconazole. Conclusion: This technique was found to be reliable, cost effective and easy to perform with consistent results.     

Synergic effect of 5-fluorocytosine and imidazole derivatives against yeast strains resistant to 5-fluorocytosine

Antifungal agents associations are widely used in therapy of deep mycotic diseases, particularly amphotericin B-5-fluorocytosine association. Synergistic effect has also been described between 5-fluorocytosine and imidazole derivates. The authors have tested here eventual synergy between 5-fluorocytosine and imidazole derivatives (miconazole, ketoconazole, fluconazole, itraconazole) against 57 years isolates resistant to 5-fluorocytosine by a semi-automated methods in liquid medium (Yeast Nitrogen Base and Brain Heart Infusion). The synergistic effect between 5-fluorocytosine and antifungal imidazoles varies widely with the drug tested. It's more frequent with ketoconazole. Itraconazole and fluconazole present very little synergistic effects in vitro.     

In vitro antifungal susceptibility testing of Candida blood isolates and evaluation of the E-test method.

Fungal infections have dramatically increased in recent years, along with the increase of drug-resistant isolates in immunocompromised patients. Ninety eight Candida species obtained from blood cultures at the Tri-Service General Hospital, Taiwan, from 1998 to 2000 were studied. These included 50 Candida albicans, 13 Candida glabrata, 24 Candida tropicalis and 11 Candida parapsilosis isolates. To investigate their susceptibility to commonly used antifungal drugs, minimum inhibitory concentrations (MIC) of amphotericin B, fluconazole, flucytosine, and ketoconazole were determined. Both the National Committee for Clinical Laboratory Standards reference broth macrodilution method and E-test were used in parallel. Ninety five isolates (95/98, 96.94%) were susceptible to amphotericin B at a concentration < or = 1 microg/mL. All isolates (100%, 98/98) were susceptible to flucytosine. Approximately 30% of these Candida isolates were resistant to fluconazole. The MIC for 90% of isolates (MIC90) values for both methods for these isolates were 0.5 microg/mL for amphotericin B, 32 microg/mL for fluconazole, 0.25 microg/mL for flucytosine (0.125 microg/mL by E-test method), and 4 microg/mL for ketoconazole. MIC for 50% of isolates (MIC50) values for these agents were 0.25, 2, 0.06, and 0.06 microg/mL, respectively. The essential agreement of MIC values within 2 dilutions for the 2 methods was 99.0% for amphotericin B, 90.8% for ketoconazole, 92.9% for fluconazole, and 91.8% for flucytosine. This study showed that E-test has equivalent performance to the broth macrodilution method and can be used as an alternative MIC technique for antifungal susceptibility testing.     

Antifungal susceptibility testing of yeast isolates from blood cultures by microbroth dilution and the E test

The results of microbroth dilution were compared with those of the E test for 169 yeast isolates tested for their susceptibility to antifungal agents. All isolates were tested by both methods against amphotericin B, ketoconazole, fluconazole, and itraconazole. The E test results generally correlated well with those obtained by the reference method. There was at least 80% agreement of minimum inhibitory concentration results within two dilutions for all yeast species and agents tested, except forCryptococcus neoformans tested with fluconazole (8% agreement). The E test appears to be a suitable alternative antifungal susceptibility test method for yeasts, although improvements are required for testingCryptococcus neoformans against fluconazole.     

Pathogenicity and antifungal susceptibility ofChaetomium species

Several reports have been published implicatingChaetomium spp. as opportunistic pathogens. A critical review of these cases was made, and the majority of the responsible strains were studied.Chaetomium globosum was the most common species, being isolated in at least nine clinical cases of infection. Some of these clinical isolates and others from environmental sources were tested against six antifungal agents (5-fluorocytosine, fluconazole, amphotericin B, itraconazole, ketoconazole and miconazole). The 23 strains tested were totally resistant to the first two drugs, and none of the other antifungal agents demonstrated fungicidal activity. There were no significant differences between the susceptibility of the clinical strains and the other strains.     

In vitro susceptibility of chromoblastomycosis and phaeohyphomycosis agents to antifungal drugs.

The in vitro susceptibility of chromoblastomycosis and phaeohyphomycosis agents to antifungal drugs was appraised using the reference macrodilution method proposed by the National Committee for Clinical Laboratory Standards (NCCLS) for yeasts modified for filamentous fungi. The antifungal drugs amphotericin B, 5-fluorocytosine, itraconazole and fluconazole were tested against one environmental and 18 clinical isolates. This work amended the macrodilution methods proposed by NCCLS and suggests that a conidial suspension free of hyphae leads to a more reliable assay and provides for better reproducibility. The macrodilution method was performed with 10(4) conidia ml-1. The MIC values ranged from 1.0 to 16.0 micrograms ml-1 for amphotericin B and 3.12 to 25.0 micrograms ml-1 for 5-fluorocytosine. A MIC range of 0.06 to 1.95 micrograms ml-1 was determined for itraconazole while 2.0 to 64.0 micrograms ml-1 was detected for fluconazole.     

Antifungal resistance: an emerging problem in Mexico

BACKGROUND: An increase in mycosis associated with therapeutic failure has been observed worldwide. The dearth of data in Mexico led us to study antifungal resistance. MATERIAL AND METHODS: Seventy six isolates of patients from the Hospital de Especialidades, Centro Médico Nacional Siglo XXI, Instituto Mexicano del Seguro Social were included: 36 with dermatophytoses and 40 with candidiasis. Dermatophytes were assesed using the E-test method and Candida spp. using the broth microdilution method. Antifungal drugs included itraconazole, ketoconazole and fluconazole for dermatophytes; in addition, voriconazole and amphotericin B were used to treat yeasts. RESULTS: From the 36 dermatophytes, seven isolates (19.4%) showed resistance to one or more antifungal drugs: three to Trichophyton rubrum, three to T. mentagrophytes and one to T. tonsurans. One T. rubrum isolate was resistant to the three azoles; the other six isolates were resistant to fluconazole only. From the 40 Candida isolates, 11 (27.5%) showed resistance: seven to ketoconazole and itraconazole; three only to itraconazole and one to ketoconazole. One C. glabrata isolate showed resistance to the four azoles. None of the yeasts showed resistance to amphotericin B. CONCLUSION: Therapeutic failure could be caused by drug resistance. In our study we found an antifungal resistance of 20% and 27.5% in dermatophytes and in yeasts respectively.     

Multisite reproducibility of the Etest MIC method for antifungal susceptibility testing of yeast isolates.

A multicenter study was performed to establish the interlaboratory reproducibility of Etest, to provide an additional comparison of Etest MICs with reference broth macrodilution MICs, and to develop some tentative quality control (QC) guidelines for using Etest for antifungal susceptibility testing of Candida spp. Two QC strains, Candida parapsilosis ATCC 22019 and Candida krusei ATCC 6258, were tested by Etest against amphotericin B, fluconazole, flucytosine, itraconazole, and ketoconazole in each of four laboratories. The QC strains were tested 20 times each against the five antifungal agents by using a common lot of RPMI agar. A total of 80 MICs per drug per strain were generated during the study. Overall, 98 to 100% of the MICs fell within a 3 log2 dilution range for the respective yeast-antifungal agent combinations. The level of agreement of Etest MICs with broth macrodilution MICs was 86 to 100% with amphotericin B (C. krusei and C. parapsilosis), itraconazole (C. krusei and C. parapsilosis), flucytosine (C. parapsilosis), and fluconazole (C. parapsilosis). A lower level of agreement was observed with ketoconazole (C. krusei and C. parapsilosis). Although all participants reported identical Etest MICs, the MICs of flucytosine and fluconazole when tested against C. krusei fell well above the upper limits of the reference range for this strain. The tentative QC limits for the two QC strains and five antifungal agents when tested by the Etest methodology are the same as the QC limits when tested by the reference broth macrodilution method for amphotericin B and C. krusei, itraconazole and C. krusei, flucytosine and C. parapsilosis, fluconazole and C. parapsilosis, and itraconazole and C. parapsilosis. The Etest QC ranges are 1 dilution broader (4-dilution range) than the reference macrodilution method QC ranges for ketoconazole and C. krusei, amphotericin B and C. parapsilosis, and ketoconazole and C. parapsilosis.     

A flucytosine-resistant Cryptococcus neoformans (serotype D) strain isolated in turkey from cutaneous lesions.

A Cryptococcus neoformans strain from cutaneous lesions of a patient with thrombotic thrombocytopenia purpura was tested for in vitro susceptibility against seven conventional antifungal agents. The strain was susceptible to fluconazole, itraconazole, ketoconazole and miconazole but was resistant to 5-fluorocytosine (5-FC). Minimal inhibitory concentration (MIC) values obtained against amphotericin B and terbinafine were 1 and 4 microg ml(-1), respectively. The isolate belonged to serotype D. Few human cases of cryptococcosis have been reported over the last 50 years in Turkey. This is the first C. neoformans isolate in Turkey shown to have primary resistance to 5-FC. Primary resistance to flucytosine is rarely reported in C. neoformans and may be associated with treatment failure in some cases.