应用基因扩增技术对三种临床常用酵母样真菌鉴定方法的评价
发布时间：2019-05-12 05:49
【摘要】：目的： 分子生物学技术的发展使真菌通过基因序列分析能够准确快速鉴定到种，但是临床实验室常使用商品化的生化鉴定方法来鉴定真菌。本实验以分子生物学方法鉴定酵母样真菌的结果为“金标准”，探讨API20C AUX和BD PhoenixTM酵母菌鉴定板鉴定酵母样真菌的临床应用价值。 方法： 收集吉林大学第一医院二部微生物室2012~2013年无菌体液标本分离出的酵母样真菌121株，，已剔除同一患者同一部位或不同部位分离出的相同菌株。将121株菌株传至沙氏培养基进行充分复苏、分纯培养，并传至科玛嘉显色培养基进行初步显色鉴定，同时对所有研究菌株平行进行API20CAUX及BD PhoenixTM的鉴定。煮沸法提取DNA，以真菌通用引物ITS1和ITS4扩增真菌rDNA，扩增产物的测序结果提交GenBank，通过基因序列分析法将真菌准确鉴定到种，不同鉴定方法的鉴定正确率行统计学分析，比较API20CAUX和BD PhoenixTM酵母菌鉴定板鉴定酵母样真菌的临床应用价值。 结果： 1.121株酵母样真菌经分子生物学方法准确鉴定到种，共包含8个菌种，均在API20C AUX的可鉴定菌种数据库内。除1株奥默毕赤酵母是BD PhoenixTM酵母菌鉴定板的库外菌种外，其余120株菌株均在BD PhoenixTM酵母菌鉴定板的可鉴定菌种数据库内，占临床分离菌株的99.2%（120/121）。 2.科玛嘉显色培养鉴定结果中有108株能够鉴定至种，13株无法鉴定；108株鉴定至种的结果中有8株鉴定错误，总体鉴定正确率为82.65%（100/121）。API20CAUX鉴定结果中有5株菌株鉴定错误，4株菌株无鉴定结果，其余112株菌株均鉴定正确，鉴定正确率为92.56%（112/121）。BD PhoenixTM酵母菌鉴定板对库内菌种120株酵母样真菌的鉴定结果中有1株葡萄牙念珠菌未能鉴定及1株季也蒙念珠菌鉴定错误，对库外菌种1株奥默毕赤酵母鉴定错误，其余118株菌株均鉴定正确，库内鉴定正确率为98.33%（118/120），对全部121株菌株的总体鉴定正确率为97.52%（118/121）。科玛嘉显色结合API20C AUX和科玛嘉显色结合BD PhoenixTM的总体鉴定正确率分别为90.08%（109/121）和91.74%（111/121）。BD PhoenixTM对库内菌种的鉴定正确率较API20C AUX的鉴定正确率高（2检验，P0.05）。科玛嘉显色培养的总体鉴定正确率与其结合BD PhoenixTM的鉴定正确率之间有显著差异（2检验，P0.05），与其结合API20CAUX的鉴定正确率之间无显著差异（2检验，P0.05）。 结论： 1.科玛嘉显色培养、API20C AUX、BD PhoenixTM酵母菌鉴定板在鉴定酵母样真菌中有较好的临床应用价值。 2. BD PhoenixTM酵母菌鉴定板较API20CAUX有更好的临床应用价值。 3. BD PhoenixTM酵母菌鉴定板和API20C AUX对少见菌种的鉴定效能有待进一步验证。
[Abstract]:Objective: with the development of molecular biology technology, fungi can be identified accurately and quickly by gene sequence analysis, but commercial biochemical identification methods are often used in clinical laboratories to identify fungi. In this experiment, the results of identification of yeast-like fungi by molecular biological method were "gold standard", and the clinical value of identification of yeast-like fungi by API20C AUX and BD PhoenixTM yeast identification plate was discussed. Methods: 121 strains of yeast-like fungi isolated from aseptic body fluid samples from the second microbiology department of the first Hospital of Jilin University in 2013 were collected and the same strains isolated from the same site or different parts of the same patient were eliminated. The strains were transferred to Shah's medium for full resuscitation, cultured in pure culture, and passed to Komaga color medium for preliminary color identification. At the same time, all the studied strains were identified by API20CAUX and BD PhoenixTM in parallel. DNA, was extracted by boiling method. The results of sequencing of fungal rDNA, products amplified by universal primers ITS1 and ITS4 were submitted to GenBank, for accurate identification of fungi to species by gene sequence analysis. The identification accuracy of different identification methods was statistically analyzed. The clinical value of API20CAUX and BD PhoenixTM yeast identification plate for identification of yeast-like fungi was compared. Results: 1.121 strains of yeast-like fungi were accurately identified by molecular biology, including 8 strains, all of which were in the identifiable strain database of API20C AUX. With the exception of one strain of Pichia omeptica which is outside the library of the BD PhoenixTM yeast identification board, all the other strains are in the identifiable strain database of the BD PhoenixTM yeast identification board, accounting for 99.2% (120 渭 121) of the clinical isolates. 2. 108 strains could be identified and 13 strains could not be identified. Of the 108 strains identified, 8 were wrong, the overall correct rate was 82.65% (100 脳 121). 5 of the 108 strains identified were wrong, 4 strains were not identified, and the other 112 strains were correctly identified, and the results of identification were as follows: (1) the total identification rate was 82.65% (100 脳 121), and that of the other 112 strains was correct. The correct rate of identification was 92.56% (112 / 121). BD PhoenixTM yeast identification board). One of the strains of Candida Portugal was not identified and one of the strains was wrong in the identification of Candida mongolicum in the library, and the results of the identification of yeast-like fungi in the library were as follows: (1) the identification rate was 92.56%. The identification error of one strain of Pichia pastoris outside the library was wrong, and the other 118 strains were correctly identified. The correct rate of identification in the library was 98.33% (118 鈮
本文编号：2475153
[Abstract]:Objective: with the development of molecular biology technology, fungi can be identified accurately and quickly by gene sequence analysis, but commercial biochemical identification methods are often used in clinical laboratories to identify fungi. In this experiment, the results of identification of yeast-like fungi by molecular biological method were "gold standard", and the clinical value of identification of yeast-like fungi by API20C AUX and BD PhoenixTM yeast identification plate was discussed. Methods: 121 strains of yeast-like fungi isolated from aseptic body fluid samples from the second microbiology department of the first Hospital of Jilin University in 2013 were collected and the same strains isolated from the same site or different parts of the same patient were eliminated. The strains were transferred to Shah's medium for full resuscitation, cultured in pure culture, and passed to Komaga color medium for preliminary color identification. At the same time, all the studied strains were identified by API20CAUX and BD PhoenixTM in parallel. DNA, was extracted by boiling method. The results of sequencing of fungal rDNA, products amplified by universal primers ITS1 and ITS4 were submitted to GenBank, for accurate identification of fungi to species by gene sequence analysis. The identification accuracy of different identification methods was statistically analyzed. The clinical value of API20CAUX and BD PhoenixTM yeast identification plate for identification of yeast-like fungi was compared. Results: 1.121 strains of yeast-like fungi were accurately identified by molecular biology, including 8 strains, all of which were in the identifiable strain database of API20C AUX. With the exception of one strain of Pichia omeptica which is outside the library of the BD PhoenixTM yeast identification board, all the other strains are in the identifiable strain database of the BD PhoenixTM yeast identification board, accounting for 99.2% (120 渭 121) of the clinical isolates. 2. 108 strains could be identified and 13 strains could not be identified. Of the 108 strains identified, 8 were wrong, the overall correct rate was 82.65% (100 脳 121). 5 of the 108 strains identified were wrong, 4 strains were not identified, and the other 112 strains were correctly identified, and the results of identification were as follows: (1) the total identification rate was 82.65% (100 脳 121), and that of the other 112 strains was correct. The correct rate of identification was 92.56% (112 / 121). BD PhoenixTM yeast identification board). One of the strains of Candida Portugal was not identified and one of the strains was wrong in the identification of Candida mongolicum in the library, and the results of the identification of yeast-like fungi in the library were as follows: (1) the identification rate was 92.56%. The identification error of one strain of Pichia pastoris outside the library was wrong, and the other 118 strains were correctly identified. The correct rate of identification in the library was 98.33% (118 鈮
本文编号：2475153
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