Chinese Journal of Mycology

Construction of MYC-tagged Sch9p in Candida albicans by using of long primers PCR amplification

XU Qiu-rong, LV Quan-zhen, SUI Xue, WANG Xiao-juan, YAN Lan, JIANG Yuan-ying

2015, 10(3): 129-133.

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Objective To construct the MYC-tagged Sch9p in Candida albicans.Methods Using a pair of the long primers to amplify sequences containing the MYC tag and the ARG4 selection markers from the plasmid pFA-ARG4-MYC.The amplified plasmid sequences were transformed into the C-terminus of SCH9 open reading frame (ORF) in C.albicans SN152 by homologous recombination.The positive colonies were selected in the SC-Leu^- selective solid culture medium.Afterwards,the positive colonies with correct integration were confirmed by PCR of genomic DNA.Finally,these positive transfectants were examined by time-growth curve testing,drug sensitivity spot assays,and mycelium inducing experiments.Results Strains with MYC-tagged Sch9p which had normal phenotypes were selected.Conclusions The MYC-tagged Sch9p of C.albicans strains can be constructed correctly by using of long primers to amplify the plasmid sequences containing the MYC tag and ARG4 selection markers which were integrated into the C-terminus of SCH9 ORF in C.albicans by homologous recombination.

Effect of targeted gold nanoparticles on viability of Cryptococcus neoformans in vitro research

XIAO Qin, CHEN Min, XU Nan

2015, 10(3): 134-138.

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Objective To design targeted gold nanoparticles (GNP) conjugated with anti-Cryptococcus neoformans antibody,to explore targeted gold nanoparticles-mediated photothermal therapy of Cryptococcus neoformans in vitro.Methods Gold nanoparticles were synthesized according to seed-mediated template-assisted protocol,conjugated with anti-Cryptococcus neoformans antibody(Ab).Cryptococcus neoformans were incubated with targeted gold nanoparticles,then exposed to near-infrared (NIR) light.Cell viability was evaluated.Results Targeted gold nanoparticles were synthesized successfully and can be used to selectively target Cryptococcus neoformans cells and following NIR laser exposure,can be used to significantly reduce cell viability.Conclusions These results demongstrated that Ab-mediated immunity gold nanoparticles can enhance NIR photothermal treatments effect on Cryptococcus neoformans.

Inhibitory effects of baicalin in combination with fluconazole against Candida albicans biofilms

YAN Gui-ming, Shi Gao-xiang, SHAO Jing, WANG Tian-ming, XIA Dan, WANG Chang-zhong

2015, 10(3): 139-145.

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Objective This study aimed to investigate the antifungal activity of baicalin alone or in combination with fluconazole against Candida albicans biofilms and to explore the related mechanism.Methods Checkerboard method was uased to study the interrelation of baicalin and fluconazole on C.albicans;Time-kill curve was used to evaluate the effect of baicalin alone and in combination with fluconazole on C.albicans;XTT reduction assay and measurement of biofilm biomass assay were performed to investigate the antibiofilm activity of baicalin alone and in combination with fluconazole;Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) were used to observe the three-dimensional structure of C.albicans biofilms;Cellular surface hydrophobicity (CSH) assay was performed to investigate the effect of baicalin alone and in combination with fluconazole on adherence in C.albicans;Quantitative real time PCR (qRT-PCR) was performed to measure the expression of adhesion and hypal related genes.Results Synergistic effect was showed in anti-biofilm by baicalin in combination with fluconazole,and FICI was between 0.28-0.5;SEM and CLSM showed that baicalin alone and in combination with fluconazole could destruct and resulted in scant or nonexistent biofilms;And expressions of ALS1,ALS3,EAP1,SUN41,CSH1 were significantly downregulated 6%,51%,24%,13%和39% by qRT-PCR.Conclusion Synergistic effect of baicalin in combination with fluconazole on C.albicans biofilm included by influenced the expression of adhesion,CSH controlling genes,and decreased the CSH.

Inhibitory effect of different durations of incubation of a photosensitizer on photodynamics of C.albicans

Xu Hui, CHENG Hong-xia, ZOU Xian-biao

2015, 10(3): 146-150.

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Objective To facilitate the selection of the best lighting time by determining the level at which C.albicans and 5-ALA PDT generate protoporphyrin IX (PpIX) at different time points and by measuring the bacteriostatic rate of 5-amino ketones pentanoic acid photodynamic therapy (5-ALA PDT).Method C.albicans suspension was prepared and incubated with ALA in the dark.The consequent mixture was divided into the control group and experimental group.The generation of PpIX was observed under a laser confocal microscope.The inhibitory rate of 5-ALA PDT on C.albicans was determined by MTT method.Results After dark incubation,C.albicans and ALA produced fluorescent PpIX that reached the peak amount between 30 min and 90 min,but began to decrease significantly at 120 min.There was no fluorescent material in control group.Conclusion The inhibitory effect of ALA PDT on C.albicans was closely related to the concentration of the photosensitizer,which provides data for treatment of C.albicans disease.

The sensitivity testing in vitro of major Candida to terconazole

ZHU Jun-hao, HAN De-ming, LI Li, LIU Jian-hui, ZHU Xiao-chan, ZHANG Qiang-qiang

2015, 10(3): 151-153.

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Objective To research the sensitivity of major Candida,isolated from severe vulvovaginal candidiasis to terconazole.Methods Isolates were collected from the patients with vulvovaginal candidiasis,who were from the Gynecology clinic of Shanghai Renji Hospital,Shanghai Sixth People's Hospital and Shanghai First Maternity and Infant Hospital.CHROM agar candida medium and API 20C AUX were used to identify Candida species.The sensitivity of 213 Candida isolates to terconazole,clotrimazole and miconazole were tested by microdilution broth method according to the CLSI M27-A3 microdilution method.Results 192 isolates were identified to C.albicans (90.14%),21 isolates were identified to C.glabrata (9.86%).The sensitivity of C.albicans isolates to terconazole were as follows:MIC range:0.024-12.5 μg/mL,MIC₅₀:0.39 μg/mL,MIC₉₀:1.56 μg/mL,GM:0.248 μg/mL;The sensitivity of C.glabrata isolates to terconazole were as follows:MIC range:0.024-1.56 μg/mL,MIC₅₀:0.049 μg/mL,MIC₉₀:1.56 μg/mL,GM:0.107 μg/mL.Conclusions Currently vaginal candidiasis infection is still based on C.albicans.The research reprompted terconazole with clotrimazole,miconazole three kinds of antifungal drugs on the 213 Candida have good sensitivity.Terconazole has strong inhibition to C.albicans and C.glabrata and did not find resistance phenomenon.

Clinical and etiologic features in 52 oral candidiasis patients with HIV/AIDS

LI Ze-hui, LI Gang, XUE Rui

2015, 10(3): 154-157.

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Objective Oral candidiasis is considered one of the most common oral diseases of Human Immunodeficiency Virus /Acquired Immune Deficiency Syndrome patients.Understanding the clinical and etiologic features of the disease plays a vital role in its diagnosis and use of medication.Methods Mucosal swab samples were collected from 70 HIV/AIDS patients with oral candidiasis.And yeast colonies were developed and identified by CHROMagar Candida.Results There were 52 cases of oral candidiasis with 70 HIV/AIDS patients,mainly manifested in Pseudomembranous and erythema types,of which Pseudomembranous is the most common.The detection rate of Candida albicans was the highest (60.32%) in patients with oral candidiasis HIV/AIDS.Other identified Candida species were C.tropicalis (19.05%),C.glabrata (12.70%) and C.crusei (7.94%).There were 9 cases of oral candidiasisin of HIV/AIDS patients with mixed infection and 2 cases with three kinds of pathogenic bacterias.Conclusion The Clinical manifestation of HIV/AIDS patients with oral candidiasis is complex and concurrent.However,diagnosing based merely on its clinical appearance of the disease is insufficient.Therefore,accurate classification of pathogenic bacterias can guide the usage of clinical medication and reduce the production of drug resistant bacterias.

Efficacy and safety of Meifute^® antifungal solution in the treatment of tinea pedis cases from troops

ZHU Hong-mei, WEN Hai, XU Jing-feng, ZHANG Guo-en, CHENG Peng

2015, 10(3): 158-162.

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Objective To evaluate efficacy and safety of Meifute^® antifungal solution in treating tinea pedis cases from troops.Methods Generally,a single-session soak in Meifute^® antifungal solution for up to120-minute long was the suggested treatment of tinea pedis.As for some tinea pedis cases with hyperkeratosis or blisters,repeated treatment of one or two sessions at 14 day intervals might be recommended.Effect and safety evaluation was performed at week one (V1),week two (V2),and week four (V3) after initial treatment.Results Finally there were 206 cases of tinea pedis being enrolled into this study,including 104 cases of group Honour Guard (i.e.group H.G.)and 102 cases of group Armed Police (i.e.group A.P.).At week four (V3),overall effective rate was 95.6% (197/206),the cure rate was 78.2% (161/206),overall fungal clearance rate was 95.1%.Group H.G.had an effective rate of 96.1% (100/104) and a cure rate of 81.7% (85/104).Group A.P.got an effective rate of 95.1% (97/102) and a cure rate of 74.5% (76/102).There were only several cases of local side effects including mild pain (51/212) that could relieved once soak finished,and local mild edema (12/212)that self-subsided during 2 hours after soak.Follow-up of 6 months later revealed a recurrence rate of 8.97% (14/156).The recurrence rates of Group H.G.and Group A.P.were 3.06% (3/98) and 18.97% (11/58),respectively.The comprehensive satisfactory rate of patient reached up to 95.32%.Conclusion Meifute^® antifungal solution has showed good efficacy and is well tolerated in treating tinea pedis.It is especially recommended to be used in the patients who requiring a fastly and rapidly responsed therapeutic method to relieve their symptons and signs of tinea pedis within a short duration.

An investigation on RNA-interference method applicable for the filamentous fungi

ZHANG Zhen-ying, HOU Bin-bin, LIU Xia

2015, 10(3): 163-167.

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Objective To obtain a RNA-interference method mediated by Agrobacterium tumefaciens applicable for gene research on filamentous fungi.Methods Gene recombination and interference system selection were used.Results The recombinant vector PCB309-pfgrt was obtained and used in interfering the expression level of two-component DRK1 gene successfully.Conclusions This method optimized the transformation system,solving the problem of high efficient RNA interference vector construction,which has wide application prospect in genetic research in filamentous fungi.

The apprehension of British association of dermatologists' guidelines for the management of onychomycosis 2014

QIU Meng, ZOU Xian-biao

2015, 10(3): 171-174.

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British association of dermatologists' guidelines for the management of onychomycosis 2014 states epidemiology,aetiology,classification,treatment,and special individuals' administration of onychomycosis in detail.Above all,Candida and mould nail infection is interpreted more in-depth.The guideline plays an important role to clinical work.The paper summarized the essence of guideline.

The research progress on new targets of antifungal candidates

HUANG Xin, LIU Ying, CHEN Si-min, AN Mao-mao, JIANG Yuan-ying

2015, 10(3): 175-181.

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Invasive fungal infections are devastating with increasing morbidity and substantial mortality.Unfortunately,classic antifungal agents are limited by their antifungal spectrums,adverse effects,drug-drug interactions,variable pharmacokinetics,and limit types of formulation.Therefore,novel antifungal agents are urgently needed for the treatment of serious invasive fungal infections.It is possible that antifungal agents with novel targets will provide favorable options to treat fungal infections.It has made great development in antifungal agents research recent years,and the exciting advances are represented by four clusters of antifungal candidates.The first is the novel approach of candidates that destroy fungal cell walls,like E1210 and D11-2040.The second important finding is the candidates that block protein kinases signaling pathways or protein phosphatase signaling pathways,such as KP-372-1,17-AAG,and Mycograb;The third area pertains to candidates that target fungal virulence factors,like C7,²¹³Bi-18B7,¹⁸⁸Re-18B7 and so on.The fourth is the candidates that activate the host immune system,including PEV7 and β-(Man)₃-Fba-TT.Over all,these four areas demonstrate the exciting research progress on new targets of antifungal agents.

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