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体外诱导耐药阿萨希毛孢子菌氟康唑耐药相关基因筛选
Screening of fluconazole resistance-related genes in
Trichosporon asahii with induced fluconazole resistancein vitro - 第三军医大学学报 2020年42卷第10期 页码：996-1001
- 作者机构：
1.400038 重庆，陆军军医大学(第三军医大学)研究生院
2.100700 北京，解放军总医院第七医学中心全军皮肤损伤修复研究所
3.100120 北京，军委政治工作部原机关门诊部
4.150001 哈尔滨，93199 部队门诊部
- 作者简介：
[ 杨蓉娅，E-mail: sumyry@163.com YANG Rongya-mail: sumyry@163.com
- 基金信息：国家自然科学基金面上项目(81271764);国家自然科学基金面上项目(81571972);北京市自然科学基金面上项目(7202201)
DOI：10.16016/j.1000-5404.202001156
中图分类号： R379.9;R394-33;R915收稿：2020-01-07，
修回：2020-03-05，
纸质出版：2020-05-30
- 稿件说明：
移动端阅览
夏志宽, 国晶, 张德全, 等. 体外诱导耐药阿萨希毛孢子菌氟康唑耐药相关基因筛选[J]. 第三军医大学学报, 2020,42(10):996-1001.
Zhikuan XIA, Jing GUO, Dequan ZHANG, et al. Screening of fluconazole resistance-related genes in
Trichosporon asahii with induced fluconazole resistancein vitro [J]. Journal of Third Military Medical University, 2020, 42(10): 996-1001.夏志宽, 国晶, 张德全, 等. 体外诱导耐药阿萨希毛孢子菌氟康唑耐药相关基因筛选[J]. 第三军医大学学报, 2020,42(10):996-1001. DOI： 10.16016/j.1000-5404.202001156.
Zhikuan XIA, Jing GUO, Dequan ZHANG, et al. Screening of fluconazole resistance-related genes in
Trichosporon asahii with induced fluconazole resistancein vitro [J]. Journal of Third Military Medical University, 2020, 42(10): 996-1001. DOI： 10.16016/j.1000-5404.202001156.
摘要
目的
2
筛选阿萨希毛孢子菌氟康唑耐药相关基因，为研究该菌氟康唑耐药机制及开发新型抗真菌剂提供理论依据。
方法
2
将临床分离的氟康唑敏感阿萨希毛孢子菌通过不断倍增氟康唑药物浓度的方法进行体外诱导培养，直至药物浓度≥128 μg/mL，再进行无药培养30代，测定其MIC值。分别提取敏感菌株和诱导耐药菌株的总RNA，构建RNA文库，转录组测序后进行生物信息学分析，寻找差异表达基因。相同方法诱导另外2株敏感菌株为耐药菌株，并对3对敏感与耐药菌株的基因组进行测序，比较ERG11基因序列。
结果
2
成功获得稳定的诱导耐药菌株，转录组发现耐药菌株中与药物转运相关基因、麦角固醇合成相关基因、抗氧化基因等上调表达，其中氟康唑靶酶编码基因ERG11基因表达上调了3.7倍。而基因组测序证实3株耐药菌株中ERG11基因的碱基序列未发生突变。
结论
2
阿萨希毛孢子菌耐药分子机制复杂，麦角固醇合成途径中的基因，特别是ERG11基因上调表达是其主要机制之一，同时还与外排转运基因、抗氧化基因等有关。
Abstract
Objective
2
To screen the genes associated with fluconazole resistance in
Trichosporon asahii
to facilitate mechanistic studies of fluconazole resistance and the development of new antifungal agents.
Methods
2
Three clinical isolates of fluconazole-sensitive
Trichosporon asahii
were cultured in the presence of increasing concentrations of fluconazole until the drug concentration reached 128 μg/mL. After further culture for 30 generations in the absence of fluconazole
the MIC value of the induced bacteria was measured. The total RNA was extracted from both the sensitive and resistant strains of the bacteria for construction of the stranded RNA-Seq libraries and transcriptome sequencing to identify the differentially expressed genes (DEGs) in the resistant strains. The genomes of the 3 sensitive and resistant strains were sequenced for comparison of the sequence of the differentially expressed ERG11 gene.
Results
2
We successfully obtained stable
Trichosporon asahii
strains with fluconazole resistance. Transcriptome sequencing data showed that the genes associated with drug transport
ergosterol synthesis and antioxidation were up-regulated in t
he resistant strains
and among them ERG11 gene
the target gene of fluconazole
was up-regulated by 3.7 folds. Genomic sequencing proved no mutations in the sequence of ERG11 gene in the 3 resistant strains.
Conclusion
2
The molecular mechanism for drug resistance in
Trichosporon asahii
is complex. The upregulation of ergosterol synthesis pathway
especially ERG11 gene
along with other genes associated with efflux transfer and oxidative stress
is one of the main mechanisms for drug resistance in
Trichosporon asahii
.
关键词
Keywords
references
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