白念珠菌14-α脱甲基酶K143Q氨基酸置换与氟康唑耐药形成的相关性研究

目的研究临床耐氟康唑白念珠菌14-α脱甲基酶的K143Q氨基酸置换与氟康唑耐药形成的关系。方法运用体外定点突变技术构建ERG11基因A427C突变型重组质粒,并利用酶切、连接构建YES2/CT酵母表达质粒。通过构建成的表达质粒在酿酒酵母中的异源性表达及体外药物敏感性表型的检测,分析其致K143Q氨基酸置换与白念珠菌对氟康唑产生耐药的关系。结果真菌体外药物敏感性表型检测显示,含氨基酸置换的表达质粒转染于酿酒酵母INVSc1后,氟康唑最低抑菌浓度(MIC)增加16倍。结论 K143Q可增强白念珠菌对氟康唑的耐药性。     

Candida albicans outbreak associated with total parenteral nutrition in the neonatal unit

Background: The most frequently isolated fungi in patients using TPN belongs to the Candida genus. Various infections including venous catheter infections, fungemia, endocarditis and ophthalmitis may be encountered. Objective: Upon growth of Candida in the blood cultures from the pediatric (neonatal) unit of our hospital, a surveillance was performed in this unit and involving the health care workers. Clonal relationships of the isolates were investigated with molecular tests. Methods: Blood samples obtained from the patients in pediatric neonatal unit were studied with automatized blood culture [BacT/Alert (Bio Mérioux, France)]. Yeast isolates from environmental surveillance cultures (TPN solutions, hands of healthcare personnel, étagère, etc) and patients were identified as C. albicans with conventional methods and ID 32 C and ATB^TM Fungus 3 (Biomerieux, France) kits. Clonal similarity was determined by using AP-PCR as initial method and we have also typified all strains by the method of REP-PCR (diversilab system,bioMérieux). Finally; Pulsed Field Gel Electrophoresis (PFGE) was used for confirmation. Results: C. albicans was isolated in blood cultures of seven patients. Similar antifungal susceptibility patterns were observed in all isolates. AP-PCR and REP-PCR showed that the C. albicans isolates grown in the TPN solution and from the patients’ blood cultures were clonally same strains. PFGE analysis further confirmed this clonality. Conclusion: According to results of the molecular methods, we thought that a C. albicans outbreak had occurred in the neonatal pediatric unit, due to contamination of TPN solution.     

Effective reprocessing of reusable dispensers for surface disinfection tissues – the devil is in the details

Background: It has recently been reported that reusable dispensers for surface disinfection tissues may be contaminated, especially with adapted Achromobacter species 3, when products based on surface-active ingredients are used. Fresh solution may quickly become recontaminated if dispensers are not processed adequately. Methods: We evaluated the abilities of six manual and three automatic processes for processing contaminated dispensers to prevent recolonisation of a freshly-prepared disinfectant solution (Mikrobac forte 0.5%). Dispensers were left at room temperature for 28 days. Samples of the disinfectant solution were taken every 7 days and assessed quantitatively for bacterial contamination. Results: All automatic procedures prevented recolonisation of the disinfectant solution when a temperature of 60–70°C was ensured for at least 5 min, with or without the addition of chemical cleaning agents. Manual procedures prevented recontamination of the disinfectant solution when rinsing with hot water or a thorough cleaning step was performed before treating all surfaces with an alcohol-based disinfectant or an oxygen-releaser. Other cleaning and disinfection procedures, including the use of an alcohol-based disinfectant, did not prevent recolonisation.Conclusions: These results indicate that not all processes are effective for processing reusable dispensers for surface-disinfectant tissues, and that a high temperature during the cleaning step or use of a biofilm-active cleaning agent are essential.     

HIV Infection as Risk Factor for Death among Hospitalized Persons with Candidemia,South Africa, 2012–2017

We determined the effect of HIV infection on deaths among persons >18 months of age with culture-confirmed candidemia at 29 sentinel hospitals in South Africa during 2012–2017. Of 1,040 case-patients with documented HIV status and in-hospital survival data, 426 (41%) were HIV-seropositive. The in-hospital case-fatality rate was 54% (228/426) for HIV-seropositive participants and 37% (230/614) for HIV-seronegative participants (crude odds ratio [OR] 1.92, 95% CI 1.50–2.47; p<0.001). After adjusting for relevant confounders (n = 907), mortality rates were 1.89 (95% CI 1.38–2.60) times higher among HIV-seropositive participants than HIV-seronegative participants (p<0.001). Compared with HIV-seronegative persons, the stratum-specific adjusted mortality OR was higher among HIV-seropositive persons not managed in intensive care units (OR 2.27, 95% CI 1.47–3.52; p<0.001) than among persons who were (OR 1.56, 95% CI 1.00–2.43; p = 0.05). Outcomes among HIV-seropositive persons with candidemia might be improved with intensive care.     

Association of Bartonella spp bacteremia with Chagas cardiomyopathy,endocarditis and arrythmias in patients from South America

Infection with Bartonella spp may cause cardiac arrhythmias, myocarditis and endocarditis in humans. The aim of the present study was to evaluate a possible association between Bartonella spp bacteremia and endocarditis, arrhythmia and Chagas cardiomyopathy in patients from Brazil and Argentina. We screened for the presence of bacterial 16S rRNA in human blood by PCR using oligonucleotides to amplify a 185-bp bacterial DNA fragment. Blood samples were taken from four groups of subjects in Brazil and Argentina: i) control patients without clinical disease, ii) patients with negative blood-culture endocarditis, iii) patients with arrhythmias, and iv) patients with chronic Chagas cardiomyopathy. PCR products were analyzed on 1.5% agarose gel to visualize the 185-bp fragment and then sequenced to confirm the identity of DNA. Sixty of 148 patients (40.5%) with cardiac disease and 1 of 56 subjects (1.8%) from the control group presented positive PCR amplification for Bartonella spp, suggesting a positive association of the bacteria with these diseases. Separate analysis of the four groups showed that the risk of a Brazilian patient with endocarditis being infected with Bartonella was 22 times higher than in the controls. In arrhythmic patients, the prevalence of infection was 45 times higher when compared to the same controls and 40 times higher for patients with Chagas cardiomyopathy. To the best of our knowledge this is the first report of the association between Bartonella spp bacteremia and Chagas disease. The present data may be useful for epidemiological and prevention studies in Brazil and Argentina.     

Effect of serine‐type protease of Candida spp. isolated from linear gingival erythema of HIV‐positive children: critical factors in the colonization

J Oral Pathol Med (2010) 39 : 753–760 Background: There are several kinds of oral soft tissue lesions that are common manifestations observed in human immunodeficiency virus (HIV)‐infected children; for example, linear gingival erythema (LGE) that is a distinctive fiery red band along the margin of the gingivae. The etiology and pathogenesis of LGE are questionable, but a candidal origin has been suggested. Proteases are key virulence attributes produced by a variety of pathogenic fungi, including Candida. The objective of the present study is to identify the protease production in Candida species including, C. albicans (n = 5), C. dubliniensis (n = 1) and C. tropicalis (n = 1), isolated directly from typical LGE lesions observed in six HIV‐positive children, and also to test the effect of a serine protease inhibitor on the interaction of Candida spp. and epithelial cells in vitro. Methods: The ability of Candida strains to release proteases in the culture supernatant fluids was visualized by gelatin‐SDS–PAGE. Gel strips containing 30‐fold concentrated supernatant (1.5 × 10⁸ yeasts) were incubated at 37°C for 48 h in 50 mM sodium phosphate buffer, pH 5.5. The concentrated supernatants were also incubated with fibronectin, laminin, immunoglobulin G, bovine serum albumin and human serum albumin. The effect of serine protease inhibitor on the interaction of Candida spp. and epithelial cells (MA 104) was measured after pre‐treatment of fungi with the inhibitor (phenylmethylsulphonyl fluoride, PMSF). Results: All the extracellular proteases were completely inhibited by PMSF, identifying these activities as serine‐type proteases. Interestingly, a common 62‐kDa serine protease was observed in all Candida strains. The culture supernatants, rich in serine protease activities, cleaved several soluble proteinaceous substrates. Additionally, we demonstrated that pre‐treatment of C. albicans, C. dubliniensis and C. tropicalis with PMSF diminished the interaction with epithelial cells. Conclusions: Collectively, our results show that Candida spp. isolated from LGE lesions produced and secreted serine proteases and these enzymes may be involved in the initial colonization events.     

Reduced fluconazole susceptibility of Candida albicans isolates in women with recurrent vulvovaginal candidiasis: effects of long-term fluconazole therapy

Fluconazole resistance and resultant failure to control Candida vaginitis (vulvovaginal candidiasis) remains extremely uncommon; however, long-term clinically relevant decrease in fluconazole susceptibility undoubtedly occurs and accompanies prolonged fluconazole chemoprophylaxis. Accordingly, in patients with refractory vaginitis or breakthrough infections due to Candida albicans, in vitro susceptibility testing is essential to optimally manage vaginitis.     

71株白念珠菌临床分离株微卫星基因分型

目的 建立高效、快速、简便的白念珠菌临床分离株三重保守基因微卫星分型技术。方法 以煮沸法处理的71株白念珠菌野生株为模板，三色荧光分别标记保守基因CDC3、EF3和HIS3微卫星序列引物，PCR扩增，产物行聚丙烯酰胺凝胶电泳。由基因扫描分析软件获取片段的精确长度，Genotyper软件行基因分型。结果 71株白念珠菌野生株行CDC3、EF3和HIS3基因分型分别获得5、10、8种等位基因，7、8、9种基因型，三位点联合分析共获得14种菌株基因型。结论 白念珠菌多重保守基因微卫星分型能从基因水平快速、高效的对临床分离株进行分型，对白念珠菌的流行病学研究有重要价值。     

白色念珠菌对人口腔黏膜上皮角质细胞的作用研究

目的研究白色念珠菌对人口腔黏膜上皮角质细胞的作用机制。方法将培养的口腔黏膜上皮角质细胞分别加入孢子型白色念珠菌(孢子组)和菌丝型白色念珠菌(菌丝组),另将单独培养的口腔黏膜上皮角质细胞作为阴性对照组。比较各组对口腔黏膜上皮角质细胞的黏附作用,并观察白色念珠菌对口腔黏膜上皮角质细胞的凋亡和增殖作用。结果菌丝组黏附指数(7.63±1.39)显著高于孢子组(3.47±1.04),差异有统计学意义(P<0.05);菌丝组凋亡率[(3.49±0.4)%]显著高于孢子组[(2.07±0.15)%]和阴性对照组[(1.98±0.07)%],差异有统计学意义(P<0.05),而孢子组和阴性对照组凋亡率相比差异无统计学意义(P>0.05)。菌丝组G0/G1期细胞比例[(38.17±7.83)%]显著低于孢子组[(49.47±11.41)%]和阴性对照组[(57.71±9.39)%],差异有统计学意义(P<0.05);S期[(8.54±4.03)%]、G2/M期[(20.18±9.59)%]细胞比例上升,且显著高于孢子组[(6.47±2.73)%、(10.71±3.19)%]和阴性对照组[(5.35±2.11)%、(12.38±4.35)%],差异有统计学意义(P<0.05);PI[(42.79±18.73)%]明显高于孢子组[(28.67±8.79)%]和阴性对照组[(26.87±7.64)%],差异有统计学意义(P<0.05),而孢子组和阴性对照组PI相比差异无统计学意义(P>0.05)。结论白色念珠菌能够导致口腔黏膜上皮角质细胞发生凋亡和增殖周期改变。