Table of Content

28 February 2014, Volume 9 Issue 1

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Application of conditional promoter pCTR4 in Cryptococcal gene expression regulation

ZHAO Jing-yu, FA Zhen-zong, FANG Wei, MENG Yun-fang, ZHANG Chao, ZHOU Zhao-jing, PAN Wei-hua, LIAO Wan-qing

2014, 9(1):  1-6. 

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Objective To explore the application of conditional promoter pCTR4 in Cryptococcal gene expression regulation via construction of new vector pNEO/CTR4 and promoter replacement.Methods Constructed the new plasmid pNEO/CTR4 fusing report gene NEO and copper-repressive promoter pCTR4 via overlap PCR.On the basis, constructed its promoter replacement cassette, and then transformed them into Cryptococcal competent cells via gene gun to get its reconstitution strains, which were tested via regular and real-time PCR.Results Correct new vector pNEO/CTR4 and Cryptococcal reconstitution strains were obtained and confirmed.Conditional promoter pCTR4 played induced or repressed effect in UBI1 transcription under different media as expected.Conclusion The new vector of copper-repressed promoter was applicable in regulating Cryptococcal gene expression.And the ubiquitin encoding gene UBI1 was not essential for viability in Cryptococcus neoformans.Our study laid the groundwork for following functional studies of ubiquitin system in Cryptococcus neoformans.

The effect of CRK1 gene deletion on morphogenesis,adhesion and biofilm formation of Candida albicans

ZHAO Jing, LIU Ji-yong, ZHANG Hai, CAO Yong-bing, ZHEGN Qing-hu, WU Jian-hua

2014, 9(1):  7-11. 

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Objective To observe if the CRK1 gene deletion influence its virulence of C.albicans.Methods Cells were observed microscopically for formation of hyphae. The numbers of adherent C.albicans were counted by colony counting method after interaction at different time. Biofilm was quantified by MTT assay and CV assay.Results Compared with SC5314, the Δcrk1 had a weak ability to form hyphae at the condition of 10% Serum and RPMI-1640 medium. The Δcrk1 had a significant reduction in adhension with human intestinal epithelial cells in vitro at 30,60,90,120 minutes. It was weak for the Δcrk1 to form biofilm quantified by CV-assay and MTT-assay.Conclusions CRK1 gene deletion can weaken its morphogenesis,adhesion and biofilm formation.

Electroporation-mediate transformation of Penicillium marneffei and optimization of the transforming conditions

CHEN Yang-xia, GU An-jing, LIANG Yu-heng, MA Tuan, XI Li-yan

2014, 9(1):  12-15. 

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Objective To establish and optimize the electroporation-mediated transformation system for Penicillium marneffei.It would provide a good platform for the gene function research of P.marneffei.Method The selectable marker pyrG gene was transformed into the germinated spores of Penicillium marneffei strain SPM4 (pyrG^-,niaD^-) by electroporation method. It aimed at finding out the optimal protocol by changing some important effecting factors including spore stadium,germination time,electric field intensity, and so on.Results The appropriate electroporation conditions for SPM4 were as follows: spore stadium 6 d, germination time 4 h and electric field intensity 5 kV/cm. Under these conditions, 21 and 13 transformants were obtained using 1 μg circular and linearized plasmid DNA, separately.Conclusion Electroporation-mediate transformation of Penicillium marneffei can get high transformation efficiency and repeatability. It is implied that circular plasmid got higher transformation frequency than the linearized by electroporation for the germinated spores of SPM4.

The use of a method for determining the levels of cellular phosphatidic acid in detecting the Aspergillus fumigatus activating phospholipase D activity in pulmonary epithelial cells

WANG Shuo, HAN Xue-lin, GAO Huan, ZONG Hao, HAN Li

2014, 9(1):  16-19. 

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Objective To establish a method to detect Phospholipase D (PLD) activity in the A549 cells stimulated by A.fumigatus by measuring the levels of cellular phosphatidic acid (PA).Method Using the interaction model of epithelial cells stimulated by A.fumigatus ATCC13073;Chloroform-methanol method was used to extrat the intracellular lipid. A modified method was established to measure PA standard and the levels of cellular PA.Results The calibration curve of PA standard was linear between 5 and 250 μmol/L. Swollen conidia, but not resting conidia increased the level of PA in A549 cells.Conclusion This improved method for determining the levels of cellular PA was high sensitive, stable and convenient for measuring indirectly the intracellular PLD activity. Our data indicated that the PLD activity in the pulmonary epithelial cells could be significantly activated by A.fumigatus swollen conidia.

The efficacy of laser therapy, drug therapy in the treatment for mild or moderate onychomycosis

XU Jing, ZHUO Feng-lin, ZHANG Rui-na, ZHAO Jun-ying

2014, 9(1):  20-23. 

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Objective To observe the efficacy and safety of Nd:YAG 1064 nm laser therapy, itraconazole therapy in the treatment for onychomycosis.Method Sixty patients with 125 nails infected by fungi were randomly assigned to two groups. The nails in group A were treated by Nd:YAG 1064 nm laser. The nails in group B were treated by itraconazole.By following the patients of each group at 8 weeks, 16 weeks, and 24 weeks, both clinical and mycological efficacy was evaluated.Results In group A, the effective rates at 8 weeks, 16 weeks, and 24 weeks were 78.9%, 52.6% and 52.6% respectively. In group B the effective rates were 80.9%, 66.2% and 67.6% respectively. There was no significant difference in the treatment efficacy between the two groups (P<0.05).Conclusion Long pulse Nd:YAG 1064 nm laser therapy was a simple and effective method without significant complications or side effects. The difference in the treatment efficacy between the laser therapy and the drug therapy for patients with mild or moderate onychomycosis was not significant, so laser therapy could be considered in first-line therapy.

Trends of fungal spectrum and drug resistance changes in Intensive Care Unit

ZHANG Wei, LIN Zhao-fen, ZHAO Jin, XU Shi-qiong, SHAN Hong-wei

2014, 9(1):  24-27,33. 

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Objective To investigate the trends of fungal spectrum and drug resistance changes in Intensive Care Unit in six years.Methods One-thousand four hundred and thirty five stains of first-time isolated fungi were isolated during 2007-2012 in ChangZheng Hospital Intensive Care Unit. Cox-Stuart test was used to analysis trends in distribution of fungi and drug resistance changes in this retrospective study.Results During 2007-2012, the number of fungi inspected increased from 159 to 396 strains per year, among which the ratio of Candida albican,parapsilosis,aspergillus ranging 34.6%-50.3%,6.3%-9.6%,1.3%-3.2% with a trend of increase,P=0.02;Candida tropicalis,glabrata,and krusei ranging 13.6%-17.3%,9.4%-22.6%,1.4%-5.2%,P=0.18;The average drug resistant of Candida albican to itraconazole and fluconazole showed a trend of decrease P=0.02;The average drug resistant of Candida tropicalis to itraconazole and fluconazole showed a decreased trend as well,P=0.02 whereas the average drug resistant of Candida glabrata to itraconazoleshowed an increased trend,P=0.02.Conclusion The most common isolated strains of fungi in ICU were Candida albican,followed by Candida tropicalis and Candida glabrata.Candida albican,Candida parapsilosis and Aspergillus had increase ratio of distribution.There are decrease trends in the drug resistance of Candida albican and Candida tropicalis to fluconazole and itraconazole whereas Candida glabrata showed a increase trend to itraconazole.

Analysis of 297 cases of superficial mycosis and its pathogenic fungi strains

ZHANG Wei-zheng, CHEN Li, ZHAO Jin, CHEN Jiang-han

2014, 9(1):  28-30,35. 

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Objective To study pathogenic fungi strains in 297 cases of superficial mycosis.Methods The clinical specimens from 297 patients with superficial mycosis, which presented positive results of the direct microscopic examination, were examined with the culture and identification of isolates of causative fungi. Sometimes accurate identification needed BioMerieux API 20C AUX or the sequences of ITS region of ribosomal DNA (rDNA) identify. The results were analysed by SPSS 17.0 statistical software.Results All 13 kinds of pathogenic fungi strains were isolated, including Trichophyton rubrum 86 strains (29.0%),Trichophyton mentagrophytes 68 strains (22.9%),Candida 59 strains (19.9%),Dematiaceous hyphomycetes 13 strains (4.4%),Aspergillus 13 strains (4.4%),Rhodotorula 12 strains (4.0%),Penicillium 9 strains (3.0%),Mucor 9 strains (3.0%),Microsporum canis 5 strains (1.7%),Cryptococcus albidus 3 strains (1.0%),Trichosporon 2 strains (0.7%),Epidermophyton floccosum 1 strains (0.3%) and 17 mixed fungal infection (5.7%).Conclusion The predominant superficial mycosis is onychomycosis, and the predominant pathogenic fungi strains is Trichophyton rubrum, but Candida infection have grown significantly.

Clinical observation of two different operations in treatment of fungal keratitis

CHEN Jin-peng, WANG Rong, XU Hui-yong, ZHANG Peng, ZHANG Jian

2014, 9(1):  36-38,46. 

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Objective To evaluate the efficacy and safety of deep lamellar keratoplasty (DLKP) and penetrating keratoplasty (PKP) in the treatment of fungal keratitis.Methods Forty-four patients (44 eyes) were diagnosed as fungal keratitis and no obvious effect with drug treatment. DLKP (28 cases, 28 eyes) or PKP (16 cases, 16 eyes) was performed according to whether involved corneal endothelium by confocal laser microscopy. Patients were followed up for 6 to 24 months to observe recurrence postoperation, visual acuity and corneal topography astigmatism two weeks after suture removal.Results Corneal histologiacal examinations of all cases were detected of fungus. One eye by DLKP and three eyes by PKP appeared graft edema and improved after anti-rejection therapy. The best corrected visual acuity (BCVA) of DLKP group was 0.59±0.04, and BCVA of PKP group was 0.41±0.05,there was significant statistically difference (t=2.577,P=0.01). The corneal topographic astigmatism of DLKP group was (2.0±0.17)D, and that of PKP group was (2.9±0.30)D, there was statistically difference (t=0.088,P=0.016).Conclusions Laser confocal microscopy is helpful in the diagnosis of fungal keratitis and in judging the depth of the filament infiltration, providing a favourable objective evidence for choice of surgical methods. Both methods have good effects in treatment of fungal keratitis. The visual functional of DLKP is outshine than that of PKP.

Record of the articles on fungi published in late Qing journals and newspapers

2014, 9(1):  41-46. 

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The western mycological knowledge transmitted to China during late Qing dynasty has a seminal influence on the establishment and construction of modern Chinese mycology. Prior to the analysis of the influence, it is necessary to comprehensively investigate the relevant literature which is mainly distributed in various late Qing journals and newspapers. According to the statistics, it is found that there are 89 articles devoted to fungi and are distributed in 26 different journals and newspapers. 44 of them are on macrofungi, while 27 and 18 of them are on molds and yeasts. Only 28 articles are known to be translations, but those of the rest ones concerning the scientific knowledge of fungi are actually all introductions and translations. The western mycological knowledge in these articles is partly and indirectly introduced from Japanese books or journal articles, and partly translated from western scientific literature.

Progress on vaccine and antibodies against Candida albicans infections

ZHANG Yan-xia, YAN Lan, JIANG Yuan-ying

2014, 9(1):  47-51,64. 

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Candida albicans is a commensal opportunistic fungal pathogen in human that causes superficial as well as life-threatening systemic infections in immunocompromised patients.Since the lack of diagnosis methods of candidiasis in early stage, in addition to the narrow range and significant toxicity of available antifungal agents, and the research and development of new drugs are rather difficult, the systemic candidiasis becomes the main causes resulting in death in immunocompromised patients. Therefore,antifungal treatment is still faced with severe challenges.Recently, most research has been devoted to clarify the host immune response mechanisms against C.albicans infections and try to develop the immunotherapy for C.albicans infections; so the vaccine and antibody are expected to be the effective way for prevention and treatment of fungal infections. In this article, progress in research in vaccine and antibody against C.albicans is reviewed, so as to provide a reference for the study of novel vaccine and antibody against C.albicans infections.

Innate immunity response in Anti-systemic Candida albicans infection

CHEN Xia, CUI Fan, LIN Zhao-chun

2014, 9(1):  52-55,59. 

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Candida albicans is a common pathogen which may induce superficial and deep infection of human.It is believed that innate immune response plays a leading role in the host anti-systemic infection of Candida albicans.Innate immune response mediating anti-Candida infection comprises a range of fungal recognition receptors and immune effector cells. Host immune response to systemic Candida albicans infection is critical for the prognosis of patients.In this paper,the mechanism of innate immune response in host anti-systemic Candida albicans infection was summarized.

The effect of UV on Candida albicans

ZHANG Lei, YANG Wei-ling

2014, 9(1):  56-59. 

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Ultraviolet as one of the important environmental factors is significant to the growth and the physiological reactivity of many organisms including Candida albicans.It was found that the progression of hyphal growth was inhibited, formation of spore was increased and no phototropic effects were seen; PL-irradiation inactivated Candida albicans strains through a multi-targeted process, What's more, null strains of rad51 was more susceptible to UV damage than the rad52 null and loss of heterozygosity was induced in Candida albicans by ultraviolet irradiation.In addition, statistical analysis demonstrated that UVC treatment significantly reduced the fungal microbial infection after burns; growth inhibition zone was significantly greater after treatment with riboflavin/UVA.So the influence of UV is remarkable to Candida albicans.

Advances in the method of fungi diversity

SHI Li-na, LIU Xiao-feng

2014, 9(1):  60-64. 

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Fungi widely exists in nature, playing an important role in the ecosystem. The pervasive application of molecular biology technology in microbial diversity and the emergence of metagenomic breaked the limitations of traditional microbial culture method, so that improve the human's recognition of fungal diversity. This paper reviewed the main development course of studying fungal diversity approaches, introduced several commonly used methods of molecular biology on fungal diversity research, mainly including clone library PCR based, Denaturing gradient gel electrophoresis, Terminal restriction fragment length polymorphism, Real-time fluorescent quantitative PCR, Fluorescence in situ hybridization and barcoded pyrosequencing, and expounded the huge potential of metagenomics in this field.