Whole-Cell MALDI-TOF MS Versus 16S rRNA Gene Analysis for Identification and Dereplication of Recurrent Bacterial Isolates.
Whole-Cell MALDI-TOF MS Versus 16S rRNA Gene Analysis for Identification and Dereplication of Recurrent Bacterial Isolates.
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DOI：
10.3389/fmicb.2018.01294
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发表时间：
2018
影响因子：
5.2
通讯作者：
Uhlik O
中科院分区：
文献类型：
--
作者：
Strejcek M;Smrhova T;Junkova P;Uhlik O
许多生态学实验是基于从不同环境样品中提取和下游分析微生物。由于其高通量，成本效益和快速的性能，基质辅助激光解吸/电离质谱与飞行时间检测器（MALDI-TOF MS），这已被提出作为一个有前途的工具，用于细菌鉴定和分类，可以有利地用于去复制复发细菌分离株。在这项研究中，我们将49种细菌培养物的全细胞MALDI-TOF MS分析与两种基于几乎完整的16 S rRNA基因序列分析的成熟细菌鉴定和分类方法进行了比较：一种是使用最接近型菌株分配的基于突变型的方法，另一种是涉及98.65%序列相似性阈值的基于序列相似性的方法，已发现其最能描述细菌种类。使用基于参考的MALDI-TOF MS的文化分类是可比的，所产生的细菌型分配到属的水平。在物种水平上，发现16 S rRNA基因分析和MALDI-TOF MS之间的一致性有限，这可能表明光谱参考数据库需要改进。我们还评估了质谱相似性技术的物种水平划定，可以独立使用的参考数据库。我们建立了最佳的质谱相似性阈值组MALDI-TOF质谱的常见的环境分离物类似的，以避免类型和序列相似性为基础的方法。当使用质谱相似性方法时，我们建议分析的质量范围为4-10 kDa，其中包含稳定的质量信号，并包含大多数的血型决定峰。我们发现，余弦相似性（CS）阈值为0.79区分质谱类似于98.65%的物种级划定序列相似性阈值，相应的精度和召回值分别为0.70和0.73。当匹配到物种级别的分类器类型分配时，计算出最佳CS阈值为0.92，相关的精确度和召回率分别为0.83和0.64。总的来说，我们的研究表明，基于相似性的MALDI-TOF MS方法可常规用于下游分析的分离株的有效去复制，并且独特生物体的损失最小。此外，MALDI-TOF MS分析具有进一步改进的潜力，不像16 S rRNA基因分析，其方法学限制已达到平台期。
Many ecological experiments are based on the extraction and downstream analyses of microorganisms from different environmental samples. Due to its high throughput, cost-effectiveness and rapid performance, Matrix Assisted Laser Desorption/Ionization Mass Spectrometry with Time-of-Flight detector (MALDI-TOF MS), which has been proposed as a promising tool for bacterial identification and classification, could be advantageously used for dereplication of recurrent bacterial isolates. In this study, we compared whole-cell MALDI-TOF MS-based analyses of 49 bacterial cultures to two well-established bacterial identification and classification methods based on nearly complete 16S rRNA gene sequence analyses: a phylotype-based approach, using a closest type strain assignment, and a sequence similarity-based approach involving a 98.65% sequence similarity threshold, which has been found to best delineate bacterial species. Culture classification using reference-based MALDI-TOF MS was comparable to that yielded by phylotype assignment up to the genus level. At the species level, agreement between 16S rRNA gene analysis and MALDI-TOF MS was found to be limited, potentially indicating that spectral reference databases need to be improved. We also evaluated the mass spectral similarity technique for species-level delineation which can be used independently of reference databases. We established optimal mass spectral similarity thresholds which group MALDI-TOF mass spectra of common environmental isolates analogically to phylotype- and sequence similarity-based approaches. When using a mass spectrum similarity approach, we recommend a mass range of 4–10 kDa for analysis, which is populated with stable mass signals and contains the majority of phylotype-determining peaks. We show that a cosine similarity (CS) threshold of 0.79 differentiate mass spectra analogously to 98.65% species-level delineation sequence similarity threshold, with corresponding precision and recall values of 0.70 and 0.73, respectively. When matched to species-level phylotype assignment, an optimal CS threshold of 0.92 was calculated, with associated precision and recall values of 0.83 and 0.64, respectively. Overall, our research indicates that a similarity-based MALDI-TOF MS approach can be routinely used for efficient dereplication of isolates for downstream analyses, with minimal loss of unique organisms. In addition, MALDI-TOF MS analysis has further improvement potential unlike 16S rRNA gene analysis, whose methodological limits have reached a plateau.
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