侵袭性念珠菌病的实验室诊断研究进展

现代医学日益发展,广谱抗生素、免疫制剂及肿瘤放化疗等的广泛应用延长并改善了许多重症患者的生命,但同时也使得真菌感染日渐增多,特别是在免疫抑制疾病、恶性血液系统疾病、干细胞移植和器官移植中。念珠菌是侵袭性真菌病的主要病原体。据报道,在美国,念珠菌是引起院内血源性     

侵袭性念珠菌病的实验室诊断研究进展

现代医学日益发展,广谱抗生素、免疫制剂及肿瘤放化疗等的广泛应用延长并改善了许多重症患者的生命,但同时也使得真菌感染日渐增多,特别是在免疫抑制疾病、恶性血液系统疾病、干细胞移植和器官移植中。念珠菌是侵袭性真菌病的主要病原体。据报道,在美国,念珠菌是引起院内血源性感染（BSI）的第4位最常见病原体,使得BSI死亡率高达49％ 。在过去的40年中,白念珠菌引起的感染数量有所下降,而其他少见的念珠菌种如光滑念珠菌、热带念珠菌、克鲁念珠菌和近年新发现的都柏林念珠菌等感染显著增多。另外,不同种的念珠菌对抗真菌药物的敏感性和耐药性也有所不同。因此,提高念珠菌病诊断方法的敏感性,并对致病念珠菌鉴定到种的水平十分必要。目前,念珠菌的鉴定方法已由传统的培养法发展到非培养法,包括血清学方法和分子生物学技术等。     

用重组抗原诊断侵袭性念珠菌病

侵袭性念珠菌病(invasive candidiasis,IC)缺乏特异性临床症状和快速准确的诊断方法,迄今IC的诊断仍很困难。检测血清中的特异性念珠菌抗体有助于IC的诊断。但传统检测抗体的方法使用粗制真菌抗原,存在重复性差和交叉反应等问题。用分子克隆技术制备的重组抗原易于大量生产,且无翻译后修饰。将其应用于血清抗体检测,既消除了交叉反应,又易于方法的标准化,为IC早期诊断提供了新的诊断途径。     

卡泊芬净治疗中国成年人侵袭性念珠菌病和食管念珠菌病的安全性、耐受性及疗效的非对照、多中心、开放性研究

目的评价卡泊芬净注射剂治疗中国成年人侵袭性念珠菌病和食管念珠菌病的疗效及安全性。方法本研究为非对照、多中心、开放性临床研究,入选对象为年龄≥18岁,确诊为侵袭性念珠菌病或食管念珠菌病需要进行抗真菌治疗的中国成人患者。采用卡泊芬净静脉给药,每日50 mg,首日采用70 mg负荷剂量。在最后一次血液或其他正常无菌部位培养阳性后继续治疗至少14 d。食管念珠菌病患者每日50 mg,疗程至少7 d,症状缓解后继续治疗72 h。结果共入选63例患者,包括侵袭性念珠菌病60例,食管念珠菌病3例,其中安全性分析集（SS）63例,全分析集（FAS）63例,符合方案集（PPS）50例。SS 63例患者中,14例发生19例次严重不良事件,均与研究药物无关;发生与药物有关的非严重不良事件31例73例次,其中4例同时发生临床不良事件和实验室指标异常;8例发生12例次临床不良事件,主要为皮疹等,其中91.7%（11/12）属轻、中度。27例发生实验室指标异常,主要为ALT等肝酶升高、血钾降低等。与研究药物相关的不良事件总发生率为49.2%（31/63）,其中临床不良事件发生率为12.7%（8/63）,实验室指标异常发生率为42.9%（27/63）。1例因不良事件终止治疗,占1.6%（1/63）。FAS和PPS中的总有效率分别为58.1%（36/62）和70.0%（35/50）。FAS中,侵袭性念珠菌病的有效率为57.6%（34/59）,食管念珠菌病为2/3。PPS中,侵袭性念珠菌病的有效率为68.8%（33/48）,食管念珠菌病为3/3。结论卡泊芬净治疗中国成年人侵袭性念珠菌病和食管念珠菌病临床不良反应大多为轻、中度,患者可耐受。卡泊芬净可有效治疗中国成年人侵袭性念珠菌病和食管念珠菌病。     

念珠菌病诊断与治疗:专家共识

中华医学会第二届感染与抗微生物治疗论坛:念珠菌病诊治策略高峰论坛于2010年9月26日在上海举行.50余位来自血液病、重症医学、感染、呼吸、肾病、普外、烧伤、眼科、皮肤、妇科、儿科等各临床学科以及从事K学真菌实验研究的资深专家就如何规范念珠菌病的诊断和合理进行抗真菌治疗进行了深人的讨论,拟定了"念珠菌病诊断与治疗:专家共识"的内容纲要,并通过了由全体专家起草"念珠菌病诊治专家共识"总则,由各学科专家分别负责撰写本学科涉及的念珠菌病诊治细则的建议.经数月的稿件交流、意见反馈和相互审阅形成了本文,供各科医师临床实践中参照.     

念珠菌血症的诊断和抗真菌治疗

近20年来侵袭性真菌感染呈明显增多趋势,研究显示自1979年至2000年真菌所致的脓毒症增加了207%。念珠菌血症为住院患者最常见的侵袭性真菌病。念珠菌血流感染的发病率在不同国家和不同医学中心存在差异。发病率按人口统计为1.5/     

1,3-β-D-葡聚糖和抗白念珠菌芽管抗体检测对侵袭性念珠菌病诊断的临床价值

目的探讨1,3-β-D-葡聚糖(BG)和抗白念珠菌芽管抗体(CAGTA)对侵袭性念珠菌病(IC)的诊断和治疗监测的临床价值。方法收集IC患者26例、念珠菌定植者100例。动态监测患者血浆BG和血清抗CAGTA含量,分析2项指标单用和联合应用诊断IC的敏感性、特异性、阳性预测值及阴性预测值,分析其与抗真菌治疗疗效间的关系。结果单用BG和抗CAGTA诊断IC的敏感性、特异性、阳性预测值、阴性预测值分别为61.5%、90.0%、61.5%、90.0%和57.7%、83.0%、46.9%、88.3%;BG和抗CAGTA联合检测诊断IC的特异性和阳性预测值均达100.0%,明显高于单项检测。抗真菌治疗有效组随着抗真菌治疗时间的延长,BG和抗CAGTA含量逐渐下降;治疗无效组BG和抗CAGTA含量呈持续上升趋势。结论联合监测BG和抗CAGTA不仅能提高IC诊断的特异性和阳性预测值,而且对IC的早期诊断和治疗监测具有潜在应用价值。     

念珠菌IgG抗体与1-3-β-D葡聚糖检测对侵袭性念珠菌病的诊断价值

目的 探讨念珠菌IgG抗体检测与1-3-β-D葡聚糖检测对侵袭性念珠菌病(IC)的诊断价值.方法 收集56例血清,其中34例为IC确诊病例(确诊组),22例为排除深部真菌感染的住院患者血清(对照组).56例血清标本均采用酶联免疫吸附试验(ELISA)检测念珠菌IgG抗体浓度,用动态显色法检测1-3-β-D葡聚糖含量(真...     

卡泊芬净治疗耐药念珠菌血症伴肾衰2例报告

入住ICU的患者由于各种慢性基础疾病合并急性并发症致免疫抑制,以及高危的腹部外科手术、高龄、各种导管的留置、广谱抗菌药和糖皮质激素的广泛应用等危险因素的存在,侵袭性真菌感染（IFI）患病率逐年增加并严重威胁患者的生命。以念珠菌为主的酵母样真菌和以曲霉菌为主的丝状真菌是IFI最常见的病原菌,分别占91．4％和5．9％。     

Treatment of murine Candida krusei or Candida glabrata infection with L-743,872.

L-743,872 is a broad-spectrum pneumocandin antifungal drug developed by Merck Research Co., and in the present work it was evaluated in vivo in murine models of Candida krusei and Candida glabrata infection. Mice were infected intravenously with two isolates of C. krusei and treated with fluconazole or L-743,872. Fluconazole was beneficial only in immune-competent mice infected with isolate 94-2696. At > 0.5 mg/kg of body weight/day, L-743,872 was effective against both infecting isolates in immune-competent and immune-suppressed mice. Against C. glabrata, L-743,872 was effective, at doses > or = 0.5 mg/kg, in reducing fungal cell counts in the kidneys but not in the spleen. L-743,872 has significant potential for clinical development.     

Caspofungin dose escalation for invasive candidiasis due to resistant Candida albicans

Previous in vivo studies have reported caspofungin dose escalation to be effective against Candida glabrata with reduced susceptibility. We hypothesized that higher doses of caspofungin would be effective against invasive candidiasis caused by the more virulent species Candida albicans, including isolates resistant to this echinocandin. Immunocompetent mice were inoculated with one of three C. albicans isolates, including one susceptible and two resistant isolates with different FKS1 hot spot 1 point mutations. Mice received daily caspofungin treatment for 7 days and were then followed off therapy for 2 weeks to assess survival. Kidney tissue and blood were collected, and fungal burden and serum (1 → 3)-β-D-glucan were measured. Significant differences in virulence were observed among the three C. albicans isolates, which translated into differences in responses to caspofungin. The most virulent of the resistant isolates studied (isolate 43001; Fks1p F641S) did not respond to caspofungin doses of up to 10 mg/kg of body weight, as there were no differences in survival (survival range, 0 to 12% with treatment), tissue burden, or (1 → 3)-β-D-glucan concentration compared to those for untreated controls. Higher doses of caspofungin did improve survival against the second resistant isolate (53264; Fks1p S645P) that demonstrated reduced virulence (5 and 10 mg/kg; 80% survival). In contrast, caspofungin doses as low as 1 mg/kg improved survival (85 to 95%) and reduced tissue burden and (1 → 3)-β-D-glucan concentration against the susceptible isolate (ATCC 90028). These data suggest that caspofungin dose escalation for invasive candidiasis may not be consistently effective against resistant C. albicans isolates, and this may be associated with the virulence of the strain.     

Development of caspofungin resistance following prolonged therapy for invasive candidiasis secondary to Candida glabrata infection

We report a case of Candida glabrata invasive candidiasis that developed reduced susceptibility to caspofungin during prolonged therapy. Pre- and posttreatment isolates were confirmed to be isogenic, and sequencing of hot spots known to confer echinocandin resistance revealed an F659V substitution within the FKS2 region of the glucan synthase complex.     

Use of pharmacokinetic-pharmacodynamic analyses to optimize therapy with the systemic antifungal micafungin for invasive candidiasis or candidemia

Echinocandins have become a first-line therapy for invasive candidiasis (IC). Using phase 3 trial data for patients with IC, pharmacokinetic-pharmacodynamic (PK-PD) relationships for efficacy for micafungin were examined. Micafungin exposures were estimated using a population pharmacokinetic model, and univariable and multivariable logistic regressions were used to identify factors associated with outcome, including the micafungin area under the concentration-time curve (AUC)/MIC ratio. Monte Carlo simulation was used to evaluate the probability of achieving AUC/MIC ratios associated with efficacy. Mycological and clinical success rates for evaluable cases were 89.4 and 90.9, respectively. MIC50s and MIC90s for Candida species inhibition were 0.008 and 0.5 mg/liter, respectively. The median AUC/MIC ratio was 15,511 (range, 41.28 to 98,716). Univariable analyses revealed a significant relationship between the AUC/MIC ratio and mycological response, with the worst response being among patients with lower (≤3,000) AUC/MIC ratios (P=0.005). For patients with Candida parapsilosis, AUC/MIC ratios of ≥285 were predictive of a higher mycological response (P=0.11). Multivariable logistic regression demonstrated the AUC/MIC ratio, APACHE II score, and history of corticosteroid use to be significant independent predictors of a favorable response. PK-PD target attainment analyses suggested that 76.7% and 100% of patients would achieve an AUC/MIC ratio of ≥3,000 for an MIC of 0.03 mg/liter and an AUC/MIC ratio of ≥285 for an MIC of <0.5 mg/liter, respectively. The identification of a lower AUC/MIC ratio target for C. parapsilosis than other Candida species suggests consideration of species-specific echinocandin susceptibility breakpoints and values that are lower than those currently approved by regulatory agencies.     

In vivo pharmacodynamic target investigation for micafungin against Candida albicans and C. glabrata in a neutropenic murine candidiasis model

Previous studies using in vivo candidiasis models have demonstrated that the concentration-associated pharmacodynamic indices, the maximum concentration of a drug in serum/MIC and 24-h area under the curve (AUC)/MIC, are associated with echinocandin treatment efficacy. The current investigations used a neutropenic murine model of disseminated Candida albicans and C. glabrata infection to identify the 24-h AUC/MIC index target associated with a stasis and killing endpoint for the echinocandin, micafungin. The kinetics after intraperitoneal micafungin dosing were determined in neutropenic infected mice. Peak levels and AUC values were linear over the 16-fold dose range studied. The serum drug elimination half-life ranged from 7.5 to 16 h. Treatment studies were conducted with 4 C. albicans and 10 C. glabrata isolates with micafungin MICs varying from 0.008 to 0.25 μg/ml to determine whether similar 24-h AUC/MIC ratios were associated with efficacy. The free drug AUC/MICs associated with stasis and killing (1-log) endpoints were near 10 and 20, respectively. The micafungin exposures associated with efficacy were similar for the two Candida species. Furthermore, the free drug micafungin exposures required to produce stasis and killing endpoints were similar to those recently reported for another echinocandin, anidulafungin, against the identical Candida isolates in this model.     

Anidulafungin for the treatment of candidaemia/invasive candidiasis in selected critically ill patients

A prospective, multicentre, phase IIIb study with an exploratory, open-label design was conducted to evaluate efficacy and safety of anidulafungin for the treatment of candidaemia/invasive candidiasis (C/IC) in specific ICU patient populations. Adult ICU patients with confirmed C/IC meeting ≥ 1 of the following criteria were enrolled: post-abdominal surgery, solid tumour, renal/hepatic insufficiency, solid organ transplant, neutropaenia, and age ≥ 65 years. Patients received anidulafungin (200 mg on day 1, 100 mg/day thereafter) for 10-42 days, optionally followed by oral voriconazole/fluconazole. The primary efficacy endpoint was global (clinical and microbiological) response at the end of all therapy (EOT). Secondary endpoints included global response at the end of intravenous therapy (EOIVT) and at 2 and 6 weeks post-EOT, survival at day 90, and incidence of adverse events (AEs). The primary efficacy analysis was performed in the modified intent-to-treat (MITT) population, excluding unknown/missing responses. The safety and MITT populations consisted of 216 and 170 patients, respectively. The most common pathogens were Candida albicans (55.9%), C. glabrata (14.7%) and C. parapsilosis (10.0%). Global success was 69.5% (107/154; 95% CI, 61.6-76.6) at EOT, 70.7% (111/157) at EOIVT, 60.2% (77/128) at 2 weeks post-EOT, and 50.5% (55/109) at 6 weeks post-EOT. When unknown/missing responses were included as failures, the respective success rates were 62.9%, 65.3%, 45.3% and 32.4%. Survival at day 90 was 53.8%. Treatment-related AEs occurred in 33/216 (15.3%) patients, four (1.9%) of whom had serious AEs. Anidulafungin was effective, safe and well tolerated for the treatment of C/IC in selected groups of ICU patients.     

Ergosterol biosynthesis inhibitors become fungicidal when combined with calcineurin inhibitors against Candida albicans,Candida glabrata,and Candida krusei

Azoles target the ergosterol biosynthetic enzyme lanosterol 14alpha-demethylase and are a widely applied class of antifungal agents because of their broad therapeutic window, wide spectrum of activity, and low toxicity. Unfortunately, azoles are generally fungistatic and resistance to fluconazole is emerging in several fungal pathogens. We recently established that the protein phosphatase calcineurin allows survival of Candida albicans during the membrane stress exerted by azoles. The calcineurin inhibitors cyclosporine A (CsA) and tacrolimus (FK506) are dramatically synergistic with azoles, resulting in potent fungicidal activity, and mutant strains lacking calcineurin are markedly hypersensitive to azoles. Here we establish that drugs targeting other enzymes in the ergosterol biosynthetic pathway (terbinafine and fenpropimorph) also exhibit dramatic synergistic antifungal activity against wild-type C. albicans when used in conjunction with CsA and FK506. Similarly, C. albicans mutant strains lacking calcineurin B are markedly hypersensitive to terbinafine and fenpropimorph. The FK506 binding protein FKBP12 is required for FK506 synergism with ergosterol biosynthesis inhibitors, and a calcineurin mutation that confers FK506 resistance abolishes drug synergism. Additionally, we provide evidence of drug synergy between the nonimmunosuppressive FK506 analog L-685,818 and fenpropimorph or terbinafine against wild-type C. albicans. These drug combinations also exert synergistic effects against two other Candida species, C. glabrata and C. krusei, which are known for intrinsic or rapidly acquired resistance to azoles. These studies demonstrate that the activity of non-azole antifungal agents that target ergosterol biosynthesis can be enhanced by inhibition of the calcineurin signaling pathway, extending their spectrum of action and providing an alternative approach by which to overcome antifungal drug resistance.     

Treatment of Candida glabrata infection in immunosuppressed mice by using a combination of liposomal amphotericin B with caspofungin or micafungin

While Candida albicans remains the most common Candida isolate, Candida glabrata accounts for approximately 15 to 20% of all Candida infections in the United States. In this study we used immunosuppressed mice infected with C. glabrata to investigate the efficacy of liposomal amphotericin B alone or in combination with the echinocandin caspofungin or micafungin. For monotherapy, mice were given six daily doses of liposomal amphotericin B (3 to 20 mg/kg of body weight), caspofungin (1 to 5 mg/kg), or micafungin (2.5 to 10 mg/kg). With concomitant therapy, mice received liposomal amphotericin B (7.5 mg/kg) in addition to caspofungin (2.5 mg/kg) or micafungin (2.5 mg/kg) for 6 days. For sequential therapy, liposomal amphotericin B was administered on days 1 to 3 and caspofungin or micafungin was given on days 4 to 6; conversely, caspofungin or micafungin was administered on days 1 to 3 and liposomal amphotericin B was given on days 4 to 6. Efficacy was based on the number of CFU per gram of kidney 21 days postchallenge. Monotherapy with liposomal amphotericin B (7.5 to 20 mg/kg) was significantly more effective than no drug treatment (control group) (P < 0.05) and demonstrated a dose-dependent response, with 20 mg/kg lowering the CFU/g from 6.3 to 4.2 (significantly different from the value for the control group [P < 0.001]). Monotherapy with all echinocandin doses lowered the CFU/g from 6.0 to 6.4 to 2.7 to 3.3 (significantly different from the value for the control group [P < 0.001]) with no dose-dependent response. Complete clearance of infection could be achieved only when liposomal amphotericin B was given either concomitantly with caspofungin or micafungin or if liposomal amphotericin B was given sequentially with caspofungin. In conclusion, the combination of liposomal amphotericin B with an echinocandin markedly improved the therapeutic outcome in murine C. glabrata systemic infection.     

Prior antimicrobial therapy and risk for hospital-acquired Candida glabrata and Candida krusei fungemia: a case-case-control study

The incidence of infections caused by Candida glabrata and Candida krusei, which are generally more resistant to fluconazole than Candida albicans, is increasing in hospitalized patients. However, the extent to which prior exposure to specific antimicrobial agents increases the risk of subsequent C. glabrata or C. krusei candidemia has not been closely studied. A retrospective case-case-control study was performed at a university hospital. From 1998 to 2003, 60 patients were identified with hospital-acquired non-C. albicans candidemia (C. glabrata or C. krusei; case group 1). For comparison, 68 patients with C. albicans candidemia (case group 2) and a common control group of 121 patients without candidemia were studied. Models were adjusted for demographic and clinical risk factors, and the risk for candidemia associated with exposure to specific antimicrobial agents was assessed. After adjusting for both nonantimicrobial risk factors and receipt of other antimicrobial agents, piperacillin-tazobactam (odds ratio [OR], 4.15; 95% confidence interval [CI], 1.04 to 16.50) and vancomycin (OR, 6.48; CI, 2.20 to 19.13) were significant risk factors for C. glabrata or C. krusei candidemia. For C. albicans candidemia, no specific antibiotics remained a significant risk after adjusted analysis. Prior fluconazole use was not significantly associated with either C. albicans or non-C. albicans (C. glabrata or C. krusei) candidemia. In this single-center study, exposure to antibacterial agents, specifically vancomycin or piperacillin-tazobactam, but not fluconazole, was associated with subsequent hospital-acquired C. glabrata or C. krusei candidemia. Further studies are needed to prospectively analyze specific antimicrobial risks for nosocomial candidemia across multiple hospital centers.