Background

The prevalence of opportunistic yeast infections has increased in recent decades as the result of an increasing immunocompromised patient population.

Aims

To evaluate ribosomal RNA (rRNA) gene sequence to identify medically important yeast species, to investigate the performance of both the rRNA gene internal transcribed spacer (ITS) and D1/D2 region in identifying clinically relevant yeasts, and to compare these results with those of a standard phenotypic method.

Methods

Both regions from 50 yeast strains, comprising 45 clinical isolates and 5 reference strains, were amplified using PCR and then sequenced. The sequences were compared to reference data available from the GenBank database of the National Center for Biotechnology Information using the BLASTn tool.

Results

Using ID32C, 88% (44/50) of all strains were identified accurately at the species level, although 6% were misidentified; two Candida eremophila isolates were identified as Candida glabrata and Candida tropicalis, and one Saprochaete clavata isolate was identified as Saprochaete capitata. Two of the four isolates identified by phenotypic methods as Trichosporon asahii were defined so by analyzing the ITS region, but the remaining two were not distinguishable from closely related species. Based on the D1/D2 region, these four isolates had 100% sequence identity with T. asahii, Trichosporon japonicum, and Trichosporon asteroides. The isolate identified as Trichosporon inkin using ID32C could not be distinguished from Trichosporon ovoides by analyzing the ITS and D1/D2 regions.

Conclusions

Identifying medically important yeasts by sequencing the ITS and D1/D2 region is a rapid and reliable alternative to conventional identification methods. For a diagnostic algorithm, we suggest a two-step procedure integrating conventional methods (e.g. microscopic morphology on corn meal agar with Tween® 80 and API ID32C®) and sequence analysis of the ITS and D1/D2 region.

中文翻译：

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通过对内部转录间隔区和大核糖体亚基的D1 / D2区进行序列分析，鉴定具有医学意义的酵母。

背景

近几十年来，由于免疫功能低下的患者人数增加，机会性酵母菌感染的患病率增加了。

目的

为了评估核糖体RNA（rRNA）基因序列以鉴定医学上重要的酵母菌种，研究rRNA基因内部转录间隔区（ITS）和D1 / D2区域在鉴定临床相关酵母中的性能，并将这些结果与标准表型方法。

方法

使用PCR扩增来自50个酵母菌株的两个区域，包括45个临床分离株和5个参考菌株，然后测序。使用BLASTn工具将序列与可从国家生物技术信息中心的GenBank数据库获得的参考数据进行比较。

结果

使用ID32C，尽管在物种级别上有6％的错误识别，但在物种级别上可以准确地识别出所有菌株的88％（44/50）。鉴定出2株念珠菌分离物为光滑念珠菌和热带念珠菌，1株腐腐菌菌鉴定为腐霉菌。通过分析ITS区域，对通过表型方法鉴定为Trichosporon asahii的四个分离株中的两个进行了定义，但其余两个与紧密相关的物种没有区别。基于D1 / D2区域，这四个分离株与T. asahii，Trichosporon japonicum和滴虫小行星。确定为分离丝孢inkin使用ID32C不能区别于丝孢ovoides通过分析ITS和D1 / D2的区域。

结论

通过对ITS和D1 / D2区域进行测序来鉴定医学上重要的酵母菌是常规鉴定方法的快速而可靠的替代方法。对于诊断算法，我们建议采用两步程序，将常规方法（例如，玉米粉琼脂的显微形态与Tween®80和APIID32C®）进行整合，并对ITS和D1 / D2区进行序列分析。