Rapid identification of the Candida species from direct blood cultures by CHROMagar Candida

We evaluated the ability of CHROMagar Candida to identify Candida species isolated directly from blood cultures. A total of 50 clinical isolates of Candida were incubated at 35 degrees C, and once growth was established, an aliquot of each was plated onto CHROMagar Candida medium. A control specimen was plated directly from Sabouraud's dextrose agar. Following incubation at 30 degrees C, all yeast isolates were identified by colony morphology and colour. We were able to identify all isolates of C. albicans (n = 20), C. tropicalis (n = 14), C. glabrata (n = 6), and C. krusei (n = 5), which were isolated from blood or from control cultures. This study demonstrated that CHROMagar Candida reliably isolated and identified yeast taken directly from blood cultures. We conclude that this rapid and easy method of identifying Candida species will enable clinicians to quickly choose the appropriate antifungal agent. This should decrease patient morbidity and mortality.     

泌尿系患者真菌感染的病原学和耐药性研究

目的研究住院患者泌尿系真菌感染病原学特点及其耐药性,为泌尿系真菌感染的防治提供参考数据。方法通过对尿液标本细菌学检验和分离鉴定以及药敏试验,对该医院临床科室泌尿系感染患者尿液标本进行了检测与分析。结果该医院2010年度从28 804例住院患者的尿液标本中检出真菌492株,真菌检出率为1.71%;其中从4 042例中段尿中检出真菌484株,检出率为11.97%。尿液标本中真菌检出率最高的患者来自重症医学科,其次是泌尿科和神经外科。尿液标本检出的真菌中,以热带假丝酵母菌居首位,构成比为74.4%;其次为白色假丝酵母菌和光滑假丝酵母菌。居前3位的真菌中,以光滑假丝酵母菌耐药性最强。结论泌尿系感染患者采集中段尿真菌检出率最高,以热带假丝酵母菌为主,主要患者分布在重症医学科,以光滑假丝酵母菌耐药性最强。     

Fungal sepsis: an increasing problem in major thermal injuries

In major thermal burns there has been an alarming emergence of fungal sepsis as defined by involvement of three or more organs and/or repeated positive blood cultures. During an 18-month period, we treated 72 patients (aged 18 +/- 2 years; TBSA burn, 57 +/- 3%; percent of third-degree burn, 45 +/- 3) with fungal sepsis. In all patients with documented three-organ involvement, treatment was with intravenous amphotericin (0.5 mg/kg body weight/day), immediate wound debridement, and early wound closure. The mortality was 32% (23 patients); 49 (68%) survived infection. Sixty-two variables were reviewed retrospectively using multiple regression analysis to ascertain specific factors associated with fungal sepsis and their relationship to survival. In burn patients, fungal sepsis is a strong determinant of survival, and its occurrence overshadows traditional factors presently utilized to predict clinical outcome.     

Detection of Candida spp. in blood cultures using nucleic acid sequence-based amplification (NASBA)

Candida spp. are the main causes of fungal infections in immunocompromised patients. It is known, that the routinely used automated blood culture systems may fail to detect yeasts. We therefore investigated, whether Nucleic Acid Sequence-Based Amplification (NASBA) can be used to improve the detection rate of Candida spp. in blood cultures. Culture-positive as well as negative blood cultures from patients with a proven candidaemia were analyzed, and the results of BacT/Alert monitoring were compared with the results of NASBA-based detection of yeast RNA. With the NASBA-assay, the number of positive blood cultures increased from 21% to 34%. The NASBA-assay may confirm the diagnosis and demonstrate the need for prolonged treatment. In addition it may shorten the time to detection. In summary, using NASBA for the detection of yeast RNA in blood cultures, we have shown for the first time that it is possible to improve the detection rate of yeasts in blood cultures by using amplification technology.     

5356例血培养病原菌分布及耐药性分析

目的 了解血培养阳性标本中的病原菌分布和耐药性,以指导临床合理用药.方法患者血标本用BacT/Alert 3D全自动血培养仪进行培养,阳性标本分离出的病原菌使用VITEK-32全自动微生物分析系统进行菌株鉴定和药物敏感试验.采用WHONET 5.4软件对2009年1月至2011年12月5356例血培养标本所分离的病原菌及药敏结果进行统计分析.结果 5356份血培养标本中共分离出病原菌587株,阳性率10.96%.其中革兰阳性球菌233株(39.69%);革兰阴性杆菌201株(34.24%);真菌111株(18.91%);革兰阳性杆菌42株(7.16%).前10位分离菌种共389株,占总分离菌株66.27%,凝固酶阴性葡萄球菌(120株)居分离菌株首位,其后依次为白色念珠菌(61株)、金黄色葡萄球菌(56株)、大肠埃希菌(47株)、粪肠球菌(23株)、鲍曼不动杆菌(22株)、乙酸钙不动杆菌(17株)、土生拉乌尔菌(16株)、热带念珠菌(14株)和肺炎克雷伯菌(13株).革兰阳性菌对万古霉素、利奈唑胺敏感性较高,耐药率均为0%.革兰阴性杆菌对碳青霉烯类药物的耐药率低,对其他抗菌药物均存在不同程度耐药,耐药率在12.5%~87.23%之间.白色念珠菌、季也蒙念珠菌和热带念珠菌对两性霉素B、5-氟胞嘧啶、氟康唑和伊曲康唑等耐药率较低(1.64%~13.33%),未检出对伏立康唑耐药株.结论 血培养中革兰阳性球菌是最常见分离细菌,其次为革兰阴性杆菌,真菌分离率在增加.大部分菌株对多种抗菌药物耐药,提示临床应予以重视.     

The European Confederation of Medical Mycology (ECMM) survey of candidaemia in Italy: antifungal susceptibility patterns of 261 non-albicans Candida isolates from blood

OBJECTIVES: To analyse the in vitro antifungal susceptibility of 261 non-albicans Candida bloodstream strains isolated during the European Confederation of Medical Mycology survey of candidaemia performed in Lombardia, Italy (September 1997-December 1999). METHODS: In vitro susceptibility to flucytosine, fluconazole, itraconazole, posaconazole and voriconazole was determined using the broth microdilution method described in the NCCLS M27-A guidelines. Etest strips were used to assess susceptibility to amphotericin B. In vitro findings were correlated with the patient's underlying condition and previous antifungal treatment. RESULTS: MICs (mg/L) at which 90% of the strains were inhibited were, respectively, 2 for flucytosine, 8 for fluconazole, 0.5 for itraconazole, 0.25 for voriconazole and 0.25 for posaconazole. Amphotericin B MIC endpoints were <0.50 mg/L in all the isolates tested. Flucytosine resistance was detected in 19 isolates (7%), mainly among Candida tropicalis strains (30%). Innate or secondary fluconazole resistance was detected in 13 strains (5%). Among the 13 patients with fluconazole-resistant Candida bloodstream infection, three were HIV positive, including one treated with fluconazole for oral candidosis; the four who were HIV negative had received the azole during the 2 weeks preceding the candidaemia. Cross-resistance among fluconazole and other azoles was a rare event. CONCLUSIONS: Resistance is still uncommon in non-albicans Candida species recovered from blood cultures. However, in fungaemias caused by C. tropicalis, Candida glabrata and Candida krusei, there is a high prevalence of resistance to fluconazole and flucytosine. Fluconazole resistance should be suspected in patients treated previously with azoles, mainly those with advanced HIV infection.     

Lack of immediate effects of wound excision on the hyperdynamic circulation of burned patients

The high cardiac output (CO) observed during the chronic phase in burned patients has been ascribed, among other factors, to the elevated blood flow in the burn wound. The hemodynamic effects of wound excision to fascia have been studied in eight patients with second- and third-degree burns ranging from 42% to 70% total body surface area (TBSA) undergoing debridement and skin-grafting procedures. The study was performed on the 4th to the 51st postburn day when all patients were in a hyperdynamic state. Serial hemodynamic measurements, including arterial and mixed-venous blood O2 saturation and content, were made before induction of anesthesia, during surgery both before and after wound excision, and in the recovery room a few hours after surgery. During anesthesia, the elevated CO decreased probably as a consequence of decreased metabolic requirements, but no further hemodynamic change was observed following wound excision. The size of burn wound excised to fascia averaged 24% TBSA and ranged from one-third to more than one-half of the initial burn. With discontinuation of anesthesia, CO rose rapidly above preoperative values, apparently to meet a similar increase in oxygen consumption. The excision of large areas of burned tissue did not attenuate the hyperdynamic circulation in burned patients, at least during and immediately after surgery. The data suggest that the elevated blood flow in the burn wound does not play a significant role in the pathogenesis of the hyperdynamic state.     

Activities and ultrastructural effects of antifungal combinations against simulated Candida endocardial vegetations

In vitro pharmacodynamic model (PDM) simulation of serum antifungal concentrations may predict the value of combination antifungal regimens against Candida sp. endocarditis. We investigated the effects of combinations of flucytosine (5FC), micafungin (Mica), and voriconazole (Vor) against Candida-infected human platelet-fibrin clots, used as simulated endocardial vegetations (SEVs). Single clinical bloodstream isolates of Candida albicans, Candida glabrata, Candida parapsilosis, and Candida tropicalis were used. All four isolates were susceptible to 5FC, while C. glabrata was resistant to Vor and C. tropicalis had a paradoxical resistance phenotype to Mica. The SEVs were prepared with an initial inoculum of 1 x 10(6) CFU/g of SEV and added to a PDM, which utilized yeast nitrogen broth-2% glucose and incubation at 35 degrees C and simulated antifungal pharmacokinetic profiles. Fungal densities in the SEVs were determined in quadruplicate over 72 h. Scanning electron microscopy (SEM) was used to evaluate treatment and control SEVs. Vor was the least active single agent against all Candida spp. except for C. parapsilosis, where it was comparable to Mica. In contrast, 5FC was the most active against all Candida spp. except for C. tropicalis, where it was comparable to Mica. The combination of 5FC plus Vor was superior to either agent alone against C. parapsilosis. The combination of Vor plus Mica was inferior to the use of Mica alone against C. tropicalis. The triple combination of 5FC plus Vor plus Mica was no better than single or dual agents against any of the Candida spp. The ultrastructural features of infected SEVs were unique for each Candida sp., with C. parapsilosis in particular manifesting friable biofilm clusters. In general, 5FC and Mica were superior in their rates and extents of fungal burden reduction compared to Vor against Candida-infected SEVs. Evaluation of 5FC and Mica in animal models of Candida endocarditis is warranted.     

In vivo activity of micafungin in a persistently neutropenic murine model of disseminated infection caused by Candida tropicalis

Micafungin is a new echinocandin with broad-spectrum in vitro and in vivo antifungal activity against both Aspergillus and Candida species. We compared the activity of micafungin with that of amphotericin B and fluconazole in a persistently immunocompromised murine model of disseminated candidiasis against a strain of Candida tropicalis that was resistant to amphotericin B and fluconazole in vitro. Mice were rendered persistently neutropenic with multiple doses of cyclophosphamide and infected intravenously with C. tropicalis. Mice were treated with either intraperitoneal amphotericin B (0.5-5 mg/kg per dose), oral fluconazole (50 mg/kg twice a day), intravenous micafungin (1-10 mg/kg per dose) or solvent control for 7 days. Mice were killed at 11 days post-infection and kidneys, lungs, brain and liver removed for quantitative culture. Overall mortality in the model was low, with rates varying between 10% and 25% in treatment groups. Micafungin at doses between 2 and 10 mg/kg were the only regimes able to reduce cfu below the level of detection of tissues infected with C. tropicalis. Micafungin was well tolerated by the mice and was much more effective than amphotericin B or fluconazole against an amphotericin B- and fluconazole-resistant C. tropicalis.     

The comparative efficacy of cilofungin, fluconazole and amphotericin B in disseminated Candida tropicalis infection in neutropenic mice.

There is insufficient in vivo data on the efficacy of new antifungal agents against invasive Candida tropicalis infection. Disseminated infection with Candida tropicalis in neutropenic mice was treated with cilofungin, fluconazole, or amphotericin B intraperitoneally, and compared to untreated controls. Early survival rates at the end of treatment (day 10) were similar for amphotericin B (97.5%) and fluconazole (100%), and superior to cilofungin (62.6%) which was better than no treatment (0%). Late survival rates (day 31) were highest for amphotericin B (95%), and significantly lower for cilofungin (48.7%) and fluconazole (43.9%), p = 0.0001. Rates of sterilization of the lung, liver, and spleen were high in survivors for all regimens (85.1-100%) but lower for the kidneys: fluconazole, 21.3%; amphotericin B, 39.3%; and cilofungin, 65.5%. Amphotericin B was the most effective agent in this study of disseminated Candida tropicalis (C. tropicalis) infection.     

Toxic epidermal necrolysis complicated by severe wound sepsis: a case study

The case of a 74-year-old female who developed toxic epidermal necrolysis (30% TBSA) is presented. Despite early, aggressive treatment, the patient developed severe wound sepsis and multiple-system organ failure. The fulminant wound sepsis was preceded by early endotoxemia. Patient serum suppressed T lymphocyte blastogenesis in the mixed lymphocyte reaction test and neutrophil chemotaxis in vitro. Endotoxin removal (affinity chromatography) resulted in a reverse of serum suppressive activity against T lymphocytes but did not reverse inhibition of PMN chemotaxis. Circulating endotoxin levels remained detectable during the entire course while blood cultures remained negative. It is our belief that early endotoxemia (detected before clinical signs of wound sepsis becomes apparent) leads to immunosuppression and other systemic complications.     

2018年陕西省人民医院全自动血培养仪及配套鉴定系统分离病原菌的实验室特征及耐药分析

目的　分析2018 年陕西省人民医院全自动血培养仪及配套鉴定系统分离病原菌的实验室特征及耐药率分析, 为临床合理用药提供参考。方法　对2018 年陕西省人民医院临床送检的血培养标本中病原菌的分布及耐药率进行回顾 分析。结果　血标本培养的阳性率为9.92%,共分离病原菌962 株,革兰阴性菌598 株、革兰阳性菌354 株、真菌10 株; 其中革兰阴性菌主要为大肠埃希菌占23.0% (221/962) 和肺炎克雷伯菌占18.3% (176/962);革兰阳性菌主要为表皮葡萄 球菌占10.2% (98/962) 和人葡萄球菌占8.5% (82/962);真菌主要为白假丝酵母菌占0.6%（6/962）和近平滑假丝酵母菌 占0.4%（4/962）。血培养阳性率较高的科室分别为肝胆外科、急诊外科、血液内科、重症医学科和呼吸内科。大肠埃 希菌和肺炎克雷伯菌对亚胺培南和美罗培南的敏感率为98.2%;葡萄球菌属对万古霉素、利奈唑胺和替加环素的敏感率 为100%。结论　血流感染病原菌主要为革兰阴性菌,其对常用抗生素耐药率较高。临床应重视血培养标本的送检情况 和细菌耐药监测,合理使用抗生素,早期控制血流感染,降低患者的病死率。     

Polymicrobial bloodstream infections involving Candida species: analysis of patients and review of the literature

Candida species are the 4th most common cause of nosocomial bloodstream infections in North America. It is not widely appreciated, however, that many of these infections are polymicrobial, that is, that bacteria and occasionally more than 1 species of Candida are present in the same blood culture bottle. Analysis of 2 groups of candidemic patients and a review of the literature were performed. Review of 141 candidemic patients from 8 Veterans Affairs hospitals and 231 patients from a tertiary care hospital with transplant services was performed. Of the 372 patients with candidemia, 100 (27%) had polymicrobial blood cultures: 88 patients (24%) had synchronous bacteremia and 12 patients (3%) had more than 1 species of Candida. One hundred bacteria were isolated from these patients, 69 were Gram positive, and 31 were Gram negative. Candidemia was shown to occur in a setting of polymicrobial bacteremia extending over days, whereas Staphylococcus aureus and coagulase-negative Staphylococcus were less frequently associated with polymicrobial bloodstream infections. Review of more than 8000 reported episodes of candidemia revealed high rates of polymicrobial infection occurring with candidemia. Of blood cultures isolating Candida, 23% were polymicrobial and 4% had more than 1 species of Candida. Thus, almost 1 in 4 patients with candidemia will have a polymicrobial bloodstream infection. As detection of bloodborne infections evolves toward nonculture methodologies, documentation of the frequency of polymicrobial bloodstream infections involving Candida is important. This finding may have treatment implications for clinicians.     

Susceptibility patterns of Candida species recovered from Canadian intensive care units

OBJECTIVE: The objective of this study was to determine the speciation and susceptibility patterns of Candida species recovered from Canadian intensive care units (ICUs) during a 1-day point-prevalence study on fungal colonization/infection in Canadian ICUs. METHODS AND SETTING: Blood, urine, respiratory tract, rectal, and wound fungal cultures were performed for 357 patients present at any time during a single-day 24-hour period in 35 Canadian ICUs. Comparative in vitro activities of amphotericin B, fluconazole, itraconazole, voriconazole, posaconazole, micafungin, anidulafungin, and aminocandin were determined. RESULTS: Four hundred fifteen yeasts (409 Candida species and 6 non-Candida yeasts) were recovered. Almost 50% of the patients were found to have positive respiratory tract or rectal cultures. Candida albicans accounted for 72% of the Candida species isolated, followed by Candida glabrata (16%), Candida tropicalis (5%), Candida parapsilosis (3%), Candida krusei (2%), and other Candida species or nonspeciated isolates (2%). Minimum inhibitory concentrations (milligrams per liter) at which 90% of the strains were inhibited were 0.06 for micafungin as well as anidulafungin, 0.12 for voriconazole, 0.25 for itraconazole, posaconazole, as well as aminocandin, 1 for amphotericin B, and 4 for fluconazole. Only 4% of the isolates were resistant to fluconazole and/or itraconazole. CONCLUSIONS: Candida albicans is the predominant species colonizing Canadian ICU patients. Overall, the triazoles, both older and new compounds, and the echinocandins have excellent in vitro antifungal activities against Candida species recovered from Canadian ICUs patients.     

Facilitating residual wound closure after partial graft loss with vacuum assisted closure therapy.

Third degree burns require skin grafting. In most instances, if the graft becomes infected, it requires debridement of the site and re-grafting. The purpose of this report is to illustrate the successful healing of a skin graft using negative pressure wound therapy with silver impregnated foam and soft silicone wound contact layer in a 4% total body surface area burn of a lower extremity skin graft infected with Pseudomonas aerugenosa without regrafting. A 27-year-old Hispanic male sustained a gasoline flame burn and presented 72 hours postincident with right lower extremity cellulitis. After intravenous antibiotics, the area was grafted with a partial thickness sheet graft. At 9 days postoperatively, the patient developed a wound infection, with an eventual 40% graft loss and was started on a course of antibiotics. With continued graft loss, on the 22nd postoperative day, negative pressure wound therapy V.A.C. (Vacuum Assisted Closure-KCI, San Antonio, TX) with silver impregnated foam and soft silicone wound contact layer (Mepitel, Molnlycke Health Care, Gothenburg, Sweden) were applied. The wound was completely re-epithelialized by 9 days. In combination with antibiotics, it was possible to treat a residual open wound and prevent the need for regrafting.     

Two-year surveillance on fluconazole susceptibility of Candida spp isolates in a general and university hospital in Rome.

Fluconazole susceptibility was tested in 385 clinical yeast isolates (285 Candida albicans, 38 C. glabrata, 31 C. tropicalis, 31 other Candida subsp.) using the agar disk diffusion test. Yeasts were collected from specimens obtained from outpatients (69) and inpatients (intensive care unit: 79 isolates, major burn unit: 31 isolates, hematology ward: 45 isolates, gynecology ward: 67 isolates, other wards: 94 isolates). Three hundred and fifty-six (92%) yeast isolates showed to be susceptible, 18 (5%) were susceptible dose-dependent, and 10 (3%) were resistant to fluconazole. Of the resistant group, 3 isolates were C.albicans, while seven were Candida non-albicans (2 C. rugosa, 2 C. humicola, 1 C. tropicalis, 1 C. ciferrii, 1 C. glabrata). The disk-diffusion method was easy to perform and there were no difficulties in the interpretation of inhibition zone diameters. Fluconazole maintained a good activity against Candida spp despite its extensive use for the prophylaxis and treatment of fungal infections.     

Comparative in vitro activities of caspofungin and micafungin, determined using the method of the European Committee on Antimicrobial Susceptibility Testing, against yeast isolates obtained in France in 2005-2006

The in vitro activities of caspofungin and micafungin against 1,038 yeast isolates have been determined. The caspofungin and micafungin MICs were lower for Candida albicans, Candida glabrata, and Candida tropicalis than for Candida parapsilosis, Candida guilliermondii, and Candida krusei. A clear correlation was seen between the MICs for the two drugs.     

纳米载银抗菌剂与四针状氧化锌抗白色念珠菌的性能

背景：口腔修复材料室温固化甲基丙烯酸甲酯的表面结构疏松多孔,极易附着各种细菌、微生物导致患者义齿性口炎的发生,所以在甲基丙烯酸甲酯中加入抗菌剂成为国内外学者研究的热点。目的：比较加入纳米载银抗菌剂与四针状氧化锌抗菌剂后,室温固化甲基丙烯酸甲酯对白色念珠菌的抗菌活性。方法：采用对倍稀释法测定纳米载银抗菌剂与四针状氧化锌抗菌剂对白色念珠菌的最小杀菌浓度。将纳米磷酸锆载银抗茵粉体与四针状氧化锌抗菌剂分别以0％,1％,2％,3％的质量比加入到室温固化甲基丙烯酸甲酯粉剂中,检测各组试件对白色念珠菌的抗菌率。结果与结论：纳米载银抗菌剂对白色念珠菌的最小杀菌浓度为40g／L,四针状氧化锌抗菌剂对白色念珠菌的最小杀菌浓度为25g／L。未加入纳米载银抗菌剂或四针状氧化锌抗菌剂的甲基丙烯酸甲酯几乎无抗菌活性,加入两种抗菌剂后其抗菌活性明显增加,随着加入抗菌剂质量比的增加,抗菌活性逐渐增强;当抗菌剂质量比增加到3％时,加入纳米载银抗菌剂的甲基丙烯酸甲酯抗菌率为92．23％,加入四针状氧化锌抗菌剂的甲基丙烯酸甲酯抗菌率为98．23％。表明纳米载银抗菌剂与四针状氧化锌抗菌剂对白色念珠菌均有抗茵效果,且四针状氧化锌抗菌剂比纳米载银系抗菌剂抗菌效果好。     

Pseudooutbreak of Candida versatilitis fungemia in a microbiology laboratory

Candida versatilitis was isolated from 10 blood cultures that had been supplemented with olive oil to promote the growth of Malassezia spp., and from the stock olive oil bottle in the laboratory. This unusual non-pathogenic yeast isolate was readily identified by DNA sequencing methodology. This report also points out that care must be taken to ensure the sterility of supplements added to blood culture media.     

In vitro susceptibility of 245 yeast isolates to amphotericin B, 5-fluorocytosine, ketoconazole, fluconazole and itraconazole.

The in vitro activity of amphotericin B, 5-fluorocytosine, ketoconazole, fluconazole and itraconazole was tested against 245 yeast strains isolated from clinical specimens (68 Candida albicans, 74 Candida tropicalis, 43 Candida krusei, 28 Candida glabrata, 19 Candida parapsilosis, 8 Candida lusitaniae and 5 Candida guilliermondii). An agar dilution method was employed to carry out testing. Minimal inhibitory concentrations to restrain 90% of isolate growth (MIC90) ranged from 0.12 to 2 mg/l for amphotericin B and for 5-fluorocytosine, from 0.03 to 8 mg/l for ketoconazole, from 0.05 to 50 mg/l for itraconazole and from 0.1 to > 100 mg/l for fluconazole. Among the azole derivatives, the most active was ketoconazole, followed by itraconazole. Only 1 strain of C. albicans was resistant to amphotericin B (MIC > 4 mg/l). Both C. tropicalis and C. krusei responded poorly to fluconazole and the former to itraconazole as well. The species most susceptible to the antifungal agents tested was C. glabrata and the most resistant were C. tropicalis and C. krusei.