Chinese Journal of Mycology

The study of differentially expressed proteins in yeast and mycelial phases of Malassiza furfur

XIANG Yun, RAN Yu-ping, TONG Ai-ping, GOU Lan-tu, WANG Wei, DAI Ya-ling

2014, 9(5): 257-263.

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Objectives To identify differential proteins in the yeast and mycelial phases of Malassezia furfur by using the two-dimensional electrophoresis (2-DE) protocol and tandem spectrum (MS/MS), and to understand the pathogenic mechanism of M.furfur and its morphological switching mechanism at protein level as well.Methods The type strain of M.furfur was inoculated in the yeast and mycelial phase media respectively. Vortexing with glass beads and ultrasonication were used to break up the cell wall into pieces, and trichloroacetate/acetone precipitation was applied to obtain the total proteins. These proteins were separated and visualized by 2-DE, analyzed by PDQuest soft to detect the differentially expressed protein spots. Electrospray-tandem spectrum analysis combined with homology search by Mascot and NCBI's Blast was used to identify proteins.Results The stable protein profiles from the mycelial and yeast phases were compared, more than 800 spots were detected respectively. Of these, 64 spots were regulated more than 3-fold, 11 were identified only in the yeast phases and 9 were identified only in the mycelial phases. Selected 40 differential spots, 22 were identified, corresponding to 16 unique proteins. In the identified proteins,9 proteins including actin, cofilin etc. were up-regulated and 5 proteins including glutathione transferase, cytoskeletal signaling protein etc, were down-regulated in mycelial phase.Conclusions Sixteen protein spots implicated in some cellular processes, such as metobalism, motility, oxidase stress etc. were positively identified by experiencements and important information would be provided for understanding the mechanisms underlying phenotype transformation of M.furfur and its pathogenesis.

MLST genotyping and mating type identification of C.neoformans in genotypes with virulent difference

KANG Ying-qian, CHEN Yu-ru, WANG Mei-zhu, ZHAO Liang, LI Xiao-ling, JIN Fang, LIU Tao-hua

2014, 9(5): 264-267.

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Objectives To study the Multilocus Sequence Typing (MLST) characteristics of C.neoformans var.grubii and to identify their mating types.Methods Primers of 7 housekeeping genes (CAP59,GPD1,LAC1,PLB1,SOD1,URA5 and IGS1) were designed for PCR amplification; the genotypes of 20 environment and clinical isolates were analyzed with MLST result and the matting types of these strains were identified. And both the results of MLST and Multilocus Microsatellite Typing (MLMT) were compared to each other for evaluating their stability and reliability in taxonomy.Results The multilocus microsatellite genotype of all the 10 environment isolates was MLMT-13 in previous experiment and the MLST type of there strains was ST-15 in this experiment. Similarly, all the MLMT-36 clinical isolates belong to ST-32. And all the strains were MAT-α.Conclusion The MLST Results are highly consistent with MLMT Results in previous experiment which reveals that both molecular typing Methods are high resoluted and stable on classification of different isolation background on species or varieties level.

Using LC-MS/MS for determination of Gliotoxin in vitro model of human bronchial epithelial infected by Aspergillus fumigatus

SHEN Liang-liang, Han yu, Zheng Jian-feng, Zhao Jing-jun

2014, 9(5): 268-271.

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Objectives To establish a LC-MS/MS method for the in vitro determination of Gliotoxin (GT) in co-culture model of human bronchial epithelial (HBE) infected by Aspergillus fumigatus (AF).Methods Developing co-cultured model of HBE infected by AF,the concentration of GT were detected by LC-MS/MS in different periods.Results The GT concentration of supernatant from co-culture increased over time (P<0.05) and the recovery of GT was 68.7%-72.6%.Conclusion This method is sensitive, simple and accurate, and suitable for the determination of GT in the co-culture model.

Comparative diagnostic values of anti-enolase assay and the (1,3)-β-D-glucan assay in early invasive candidiasis

LIAO Hong, LI Fang-qiu, ZHANG Guo-yong, LI Wei, HAN Dan-dan, LIU Qian

2014, 9(5): 272-274,271.

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Objectives To compare the anti-enolase (anti-Eno) assay with traditional (1,3)-β-D-glucan (BG) assay for the early diagnosis of invasive candidiasis (IC).Methods Blood samples collected from patients of suspected IC were analyzed with both anti-Eno and BG detection. Diagnostic values were compared based upon the results of fungal culture.Results A total of 210 patients were enrolled into this study. Fifteen patients (7.1%) were diagnosed as definitive IC by fungal culture. Positive results were obtained in 54 (25.7%) and 33 (15.7%) patients by BG and anti-Eno assay, respectively. The sensitivity, specificity, positive predictive value and negative predictive value were 46.7%, 86.7%, 21.2% and 95.5% in anti-Eno assay while 33.3%, 74.9%, 9.3% and 93.6% in (1,3)-β-D-glucan assay. The sensitivity for IC diagnosis was enhanced to 66.7% upon the combination of anti-Eno and (1,3)-β-D-glucan assay.Conclusion Anti-Eno assay is superior to (1,3)-β-D-glucan assay in the diagnosis of early IC. Combining these two assays can improve the sensitivity of IC diagnosis.

Evaluation of domestic serum galactomannan detection kit for the diagnosis of invasive pulmonary aspergillosis

WANG Yuan-fang, KANG Mei, YUAN Yu, HUANG Yan-chun, MA Ying

2014, 9(5): 275-278.

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Objectives To evaluate the performance of domestic serum galactomannan (GM) test kit for the diagnosis of invasive pulmonary aspergillosis.Methods According to the reference standards set by the diagnostic criteria and treatment of invasive pulmonary fungal infection principle^[1],the serum of confirmed invasive pulmonary aspergillosis (IPA),clinical diagnosis of IPA,suspected IPA and without IPA were collected.Using Tianjin Bio-Enoche reagent kit of ELISA method, 86 specimens were analyzed to calculate the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV).Results All 86 clinical cases are consisted of 27 clinical proven IPA, 12 probable IPA, 47 without IPA. According to the three different kinds of positive judgment standard, the degree of sensitivity, specificity,PPV,NPV is respectively, sensitivity: 94.44%, 96.30%, 62.96%;Specificity:94.44%,96.30%, 62.96%;PPV:94.44%,96.30%,62.96%;NPV: 94.44%, 96.30%,62.96%. Statistical analysis confirmed that standard No.1 (The positive GM test was defined as GM value >0.95 μg/L and the negative defined as <0.75 μg/L) was the optimal, so standard No.1 was choosed as the final judgment standard.Conclusion Performance of the domestic serum GM detection kit is eligible and can be used for the auxiliary diagnosis of invasive pulmonary aspergillosis.

Morphological observation and genetic identification of a variant of granular type Trichophyton rubrum isolated from tinea corporis and cruris

YANG yan-ping, LI wen, OU ru-jun, CHEN he-lian, HUANG wen-ming

2014, 9(5): 279-282,278.

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Objectives To investigate the morphological features and sequences of ribosomal genes in a variant of granular type Trichophyton rubrum isolated from tinea corporis and cruris.Methods The morphological features of one T.rubrum strain recovered from a patient with tinea corporis and cruris were observed. The ribosomal internal transcribed spacer (ITS), D1/D2 and non-transcribed region were determined by PCR.Results The phenotype of the isolate produced antlers mycelium, macroconidium had hollow phenomenon, unilateral budding and it grew at 42℃. The sequences of ITS and D1/D2 were identical to T.rubrum in the GenBank, but sequences of TRS-1 and TRS-2 in the non-transcribed region were different from the reference strains by 2 and 1 base, respectively.Conclusion The strain might be a variant of granular type T.rubrum.

A case of cutaneous manifestations as an initial presentation of disseminated candidiasis

LIU Fang, ZHANG Min, KONG Qing-tao, YAN Wen-liang, HU Wen-xing, LIU Hai-bo, SANG Hong

2014, 9(5): 283-286.

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Objectives Disseminated candidiasis is a frequently fatal condition that is rising steadily in immunocompromised patients. We reported one case of disseminated candidiasis and reviewed currently available case reports on the cutaneous manifestations of disseminated candidiasis.Methods A 37-year-old woman with nodular on lower extremities as the initial presentation, involved of the lung and brain. She had focal segmental glomerulosclerosis (FSG) undergoing therapy of tacrolimus and prednisone for two years. Skin biopsies and culture of skin tissues, stools, urine and sputum were performed.Results With microbiological evidence of Candida albicans in skin tissue and kinds of excretions except cerebrospinal fluid and blood. Chest CT showed bilateral patchy infiltration. She was diagnosed as disseminated candidiasis and had anti-fungal therapy. A significant improvement of clinical symptoms and resolution of pulmonary were found in following days. But she had diffuse brain dissemination after interruption of antifungal treatment by herself.Conclusions Although widespread organ involvement is characteristic of disseminated candidiasis, reports of skin lesions are rare. Early cultures and biopsies resulted in early diagnosis, which prompted proper antifungal therapy and a positive outcome.

Detection of helper T cells and regulatory T cells of 21 cases with invasive pulmonary aspergillosis patients

WANG Run-chao, WAN Zhe, LI Ruo-yu

2014, 9(5): 287-292.

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Objectives To analysis CD4⁺T cells in peripheral blood mononuclear cell and clinical data of invasive pulmonary aspergillosis(IPA) patients and explore function of cellular immunity mediated by helper T cells and regulatory T cells in IPA.Methods A total of 21 IPA patients were evaluated. 19 healthy volunteers as the normal control group. We detected Th1,Th2,Th17 and Treg expression in CD4⁺T cells, related cytokine secretion and key transcription factors of IPA patients and controls.Results Patients with IPA exhibited decreased Th1 and Treg expression, related cytokines (IFN-γ, IL-17A, TGF-β) and T-bet, RORγt and Foxp3 mRNA levels when compared with the normal control group. while there was no significant difference between patients with IPA and the normal control group about peripheral Th2 frequency, Th2 related cytokines IL-4, and GATA-3 mRNA levels.Conclusions Th1,Th17and Treg-mediate cellular immunity were inhibited in IPA patients.

A couple suffering from tinea corporis transmitted from cat caused by Microsporum canis

LI Zong-hui, WU Fan, DENG Lin, HU Zhi-li, SANG Hong

2014, 9(5): 296-298.

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To report two cases of tinea corporis caused by Microsporum canis.The two patients were husband and wife, with a pet cat at home. Multiple annular erythemas were presented on their chest, neck and forearm for 2 weeks.Tinea corporis caused by Microsporum canis was diagnosed by fungal examination.The lesions were cured by oral and topical antifungal therapy.

The research developments in the therapy of allergic fungal asthma

YIN Qing-xin, SHI Dong-mei, LIAN Cui-hong, LIU Wei-da

2014, 9(5): 299-302,292.

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Allergic fungal asthma is a chronic airway inflammatory disease induced by environmental exposure to airborne fungal allergens, which are ubiquitous in many indoor and outdoor environments. As a result of complex interactions among inflammatory cells, structural cells, and the intercellular matrix of the allergic lung, patients with allergic fungal asthma may experience a greater degree of airway wall remodeling and progressive, accumulated pulmonary dysfunction, along with the symptoms of wheeze, chest stufly and dyspnea, and have a serious impact on the job as well as personal life of the patients. However, the current standard asthma treatment options, such as glucocorticoids and antifungal agents, can hardly meet the requirements of this disease. Researches on the mechanism and therapeutics of allergic fungal asthma are gradually increased in the recent decades, and have made some progress, such as immunological therapy. In this review, we analyzed and summarized the Results of these researches.

The research progress of Candida albicans wall-breaking method

WANG Xiao-fei, ZHANG Lu-lu, CAO Yong-bing, JIANG Yuan-ying

2014, 9(5): 310-315.

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Cell wall as a special and necessary cell structure of fungus.it's hard to make it fully broken in a simple way. Therefore, to achieve the ideal wall-breaking effect is one of the key steps in the study of Candida albicans. In extraction such as RNA, DNA and protein components of Candida albicans, in order to obtain sufficient and stable experimental samples, the researchers shoulduse different methods breaking the cell wall according to different research purposes. This paper summarizes a variety of Candida albicans wall-breaking methods, so as to provide applicable and efficient solution of wall-breaking for related research in Candida albicans.

Mechanism of Candida albicans adhering to the host

LI Dan, CUI Fan, XIONG Jun-hao

2014, 9(5): 316-320.

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Adherence of Candida albicans to the host is the critical first step in the infection. Therefore, clarifying the mechanism of adhesion is very important for exploring new methods for the prevention and treatment of Candida albicans infection. In recent years, a lot of research have been done on the adhesion mechanism of Candida albicans, including the surface structure, adhesins and adhesion related genes, et al. In this paper, the mechanism of Candida albicans adhering to the host was summarized.

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