白念珠菌基因分型的相关研究

白色念珠菌定植于大多数人群的口腔中,在一定条件下可成为优势菌种而导致感染。基因分型是近年来白念分子生物学研究中的一个热点,随着医学科学技术的发展,由于深部真菌感染比例不断增加,分子生物学方法已经越来越广泛的应用于临床真菌病的研究中,从而为控制白念感染及为早期诊断、治疗提供基础。本文综述了限制性片段长度多态性、随机扩增多态性DNA分析、ITS区域序列分析等分子生物学技术在白念珠菌基因分型方面的相关研究,比较了它们的优缺点,并且讨论了将基因分型研究应用于临床诊断、治疗及开发新型抗真菌药物的发展趋势和广阔前景。认为目前更倾向于多种分型方法联合应用,并借助计算机软件进行分析,但是仍需进一步探索。     

Millimeter-Wave Effects on Candida Albicans Cells

In preliminary measurements, Candida albicans cultures exposed to 1 kHz square-wave-modulated microwaves at 72 GHz exhibited a significantly reduced number of colony-forming units in comparison with nonirradiated controls. To study whether the same effect could also be caused by continuous-wave (CW) irradiation, sedimented cells were exposed to either 1 kHz square-wave-modulated or CW microwaves at 72 GHz, with the same peak power. CW-exposed cells showed a higher growth rate (about 25%), whereas a reduction of about 15% was seen in cells exposed to square-wave-modulated microwaves in comparison with sham-exposed controls.     

Bacterial Endocarditis with Candida Albicans Superinfection

Clinical and pathological features of two fatal cases of bacterial endocarditis with Candida albicans superinfection are described. One patient presented with combined Streptococcus viridans and Candida endocarditis of the aortic valve. The second patient, an addict to paregoric injected intravenously, developed Staphylococcus aureus of the tricuspid valve with eventual Candida endocarditis. The responsible organisms were identified from blood cultures during the hospital course, and by culture or tissue section of postmortem material. Candida endocarditis has emerged as a disease entity in the past 20 years. The incidence is increasing and patients with bacterial endocarditis are among those at risk. Antibiotic therapy appeared to facilitate the development of Candida endocarditis in these two cases.     

Etiological Significance of Candida Albicans in Otitis Externa

A study covering 79 patients (42 males, 37 females) of different age groups clinically diagnosed as otomycosis were investigated mycologically to elucidate the role of Candia albicans, an opportunistic polymorphic yeast, in otitis externa. C. albicans was diagnosed as the sole pathogen in two patients (1 male and 1 female) aged 18 and 20 years, respectively. The organism was repeatedly demonstrated in the aural specimens both by direct microscopy as well as culture isolation. Both the patients had unilateral otomycosis and used antibiotic solution and removed wax with wooden stick. The topical application of one per cent clotrimazole lotion showed good response both clinically as well as mycologically. The growing significance of opportunistic fungi emphasizes on comprehensive studies to establish the etiologic role in various clinical disorders in human and animal medicine.     

Genetic Analysis of Prototrophic Natural Variants of Candida Albicans

To facilitate genetic analysis of Candida albicans natural variants, we have isolated a dominant mycophenolic acid-resistant mutant. Mycophenolic acid-resistant auxotrophs were used to analyze prototrophic natural variants by spheroplast fusion. The fusion products were shown to be heterozygous for many of the parental chromosomes by molecular and genetic criteria. Using this approach, we have found that one type of morphologic variation is due to a recessive change and identified three dominant 5-fluorocytosine-resistant mutants. Rare fusion products express recessive parental markers. These exceptional progeny should be useful for linkage analysis and strain construction.     

Isogenic Strain Construction and Gene Mapping in Candida Albicans

Genetic manipulation of Candida albicans is constrained by its diploid genome and asexual life cycle. Recessive mutations are not expressed when heterozygous and undesired mutations introduced in the course of random mutagenesis cannot be removed by genetic back-crossing. To circumvent these problems, we developed a genotypic screen that permitted identification of a heterozygous recessive mutation at the URA3 locus. The mutation was introduced by targeted mutagenesis, homologous integration of transforming DNA, to avoid introduction of extraneous mutations. The ura3 mutation was rendered homozygous by a second round of transformation resulting in a Ura(-) strain otherwise isogenic with the parental clinical isolate. Subsequent mutation of the Ura(-) strain was achieved by targeted mutagenesis using the URA3 gene as a selectable marker. URA3 selection was used repeatedly for the sequential introduction of mutations by flanking the URA3 gene with direct repeats of the Salmonella typhimurium hisG gene. Spontaneous intrachromosomal recombination between the flanking repeats excised the URA3 gene restoring a Ura(-) phenotype. These Ura(-) segregants were selected on 5-fluoroorotic acid-containing medium and used in the next round of mutagenesis. To permit the physical mapping of disrupted genes, the 18-bp recognition sequence of the endonuclease I-SceI was incorporated into the hisG repeats. Site-specific cleavage of the chromosome with I-SceI revealed the position of the integrated sequences.     

小鼠念珠菌感染模型和抗感染免疫

目前,临床相关的小鼠念珠菌感染疾病模型种类日益增多.与早期小鼠念珠菌感染模型相比,近期感染模型与临床上免疫抑制机会性念珠菌感染患者的感染方式、感染的发展进程、临床表现、靶器官临床病理组织学特征更相近.建立了特定免疫缺陷小鼠(如转基因/基因敲除小鼠等);感染源从白色念珠菌转向非白念珠菌(如近平滑念珠菌、光滑念珠菌等);感染方式从局部口腔-消化道感染逐步转变为血源性深部多器官组织感染.特定模型的建立更深入地探讨了宿主-念珠菌感染源之间相互的免疫机制.细胞介导的免疫反应在宿主抗念珠菌感染免疫反应中占主导,吞噬细胞直接杀伤真菌;细胞分化为Th1和Th2型细胞,分泌相关细胞因子进行免疫调控.体液免疫中一些保护性抗体也具有一定的保护作用.     

Antifungal Activity of Schinifoline Against Candida Albicans in Caenorhabditis Elegans

Zanthoxylum schinifolium has been used as spices and traditional medicine in China for hundreds of years. A variety of active substances have been isolated from Zanthoxylum schinifolium using biological and chemical techniques. Among these substances, the effect of schinifoline has gradually attracted much attention. Candida albicans is one of the most common pathogens isolated from the gastrointestinal tract, vagina, and mouth in healthy individuals. In a healthy population, there are various mechanisms in host, such as the microbial flora, the epithelial barriers, and the innate immune system, that can control the presence of Candida albicans. However, when host immunity is compromised, an invasive fungal infection is more likely to occur. In this study, we explored the antifungal activity of schinifoline against Candida albicans in Caenorhabditis elegans. To determine the optimal concentration of schinifoline, we investigated the lifespan, defecation cycle and locomotion behavior of Caenorhabditis elegans after treatment with schinifoline. In addition, we examined colony formation in the intestine of Caenorhabditis elegans after Candida albicans infection. The results indicated that 100 and 200 mg/L of schinifoline could prolonged the lifespan, shorten the defecation cycle and increased the locomotion behavior of Caenorhabditis elegans, with 100 mg/L of schinifoline being the optimal concentration. Moreover, 100 mg/L of schinifoline increased the lifespan of Caenorhabditis elegans after infection and inhibited the colony formation of Candida albicans in Caenorhabditis elegans intestine. Therefore, we concluded that schinifoline exhibits anti-fungal effects and its potential use as natural drugs should be further explored in future studies.     

Identification of Candida albicans using the chromogenic medium Albicans ID

The correct identification of the microrganism is the base for epidemiological studies and treatment of infections. The aim of our study was to evaluate the efficacy of the chromogenic media Albicans ID (bioMerieux, France) in the identification of Candida albicans. A total of 190 yeasts strains were evaluated in the study. A rate of 100% of all C. albicans (80) and Candida dubliniensis (five) strains exhibited blue color. Nevertheless, the blue color was also observed with cultures of Candida rugosa (3/5) and Candida tropicalis (3/17). Albicans ID cromogenic media presented specificity rate of 90% and positive and negative predictive values of 88% and 100%, respectively, in the identification of C. albicans.     

耐氟康唑的白假丝酵母菌主动外排机制初探

目的:了解对氟康唑耐药的白假丝酵母菌主动外排系统及主动外排基因CDR1的表达水平。方法:检测氟康唑敏感性和耐药性白假丝酵母菌对罗丹明6G主动外排情况,筛选出主动外排系统功能增强的菌株;采用Northern blot技术检测主动外排系统功能增强的菌株的CDR1基因的表达。结果:在由葡萄糖提供能量的体系中,5株耐药菌株外排罗丹明6G较敏感菌株明显增加,Northern blot发现其中4株CDR1基因表达水平升高。结论:耐氟康唑白假丝酵母菌主动外排基因CDR1表达升高。     

Repeated Use of Gal1 for Gene Disruption in Candida Albicans

A technique which has the potential to allow repeated use of the same selectable marker to create gene disruptions in Candida albicans has been developed. In this approach, originally described for Saccharomyces cerevisiae, the selectable marker is flanked by direct repeats. Mitotic recombination between these repeats leads to elimination of the selectable marker. A module in which the GALq1 gene is flanked by direct repeats of the bacterial CAT gene was constructed and used to disrupt one copy of the URA3 gene in a gal1 mutant. Gal- revertants were selected by plating on 2-deoxy-D-galactose (2DOG). The frequency of 2DOG-resistant colonies recovered was 20 times higher than that obtained with a similar construct not flanked by direct repeats. Of these, 20% had lost the GAL1 gene by recombination between the direct repeats. The GAL1 gene was used again to disrupt the remaining wild-type copy of the URA3 gene of one of these gal1 isolates, resulting in a stable ura3 mutant. This technique should be generally applicable to derive homozygous gene disruptions in this diploid organism.     

The occurrence of Candida Albicans in Lake Ontario bathing beaches.

There are inherent weaknesses associated with currently used bacterial fecal pollution indicator systems. Fecal pollution indicator data would be more meaningful if supplemented with information relating to the occurrence of pathogens in recreational water. Through surveys of four bathing beaches on Lake Ontario, it was established that the opportunistically pathogenic yeast Candida albicans occurs in near shore waters. The beaches surveyed could be differentiated on the basis of bacterial fecal pollution indicator levels and numbers of the pathogens C. albicans and Pseudomonas aeruginosa. The occurrence of C. albicans and P. aeruginosa appeared to be related to elevated fecal pollution indicator levels. Maximum numbers of all parameters were observed in July and August in association with peak bather loads at the beaches. In only one instance does the data suggest that a beach was subjected to human fecal contamination.     

Relevance of Antigenicity of Candida Albicans Growth Phases to Diagnosis of Systemic Candidiasis

The antigenicity of the two growth phases of Candida albicans has been compared by using crossed immunoelectrophoresis and double-diffusion techniques. Qualitative and quantitative differences in antigenic composition between the phases have been revealed and moreover a specific mycelial antigenic component demonstrated.It is postulated that the use of specific mycelial antigens for routine diagnostic precipitin testing in patients with suspected systemic candidiasis would give more reliable results than at present obtained with yeast cell antigens.     

Candida Albicans: a molecular revolution built on lessons from budding yeast

Candida albicans is an opportunistic fungal pathogen that is found in the normal gastrointestinal flora of most healthy humans. However, in immunocompromised patients, blood-stream infections often cause death, despite the use of anti-fungal therapies. The recent completion of the C. albicans genome sequence, the availability of whole-genome microarrays and the development of tools for rapid molecular-genetic manipulations of the C. albicans genome are generating an explosion of information about the intriguing biology of this pathogen and about its mechanisms of virulence. They also reveal the extent of similarities and differences between C. albicans and its benign relative, Saccharomyces cerevisiae.     

白色念珠菌对人口腔黏膜上皮角质细胞凋亡和增殖的影响

目的:探索白色念珠菌对人口腔黏膜上皮角质细胞(KB细胞)凋亡和增殖的影响。方法:分别培养孢子型白色念珠菌和菌丝型白色念珠菌,取健康志愿者的颊粘膜制备KB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),另外取单纯KB细胞作为对照组。对比分析各组细胞凋亡率及各个周期的细胞数,统计分析各组细胞增殖指数(PI)。结果:菌丝组凋亡率显著高于对照组及孢子组(P0.05);菌丝组在G0/G1期的细胞比例均显著低于对照组及孢子组(P0.05);菌丝组在S期和G2/M期的细胞比例均显著高于对照组及孢子组(P0.05);菌丝组的PI显著高于孢子组和对照组(P0.05)。结论:菌丝型白色念珠菌可诱导KB细胞凋亡率的上升及改变KB细胞周期变化,并引起KB细胞的PI升高,对于临床上治疗口腔念珠菌病具有重要的指导意义。