Emele Matthias F, Joppe Felix M, Riedel Thomas, Overmann Jörg, Rupnik Maja, Cooper Paul, Kusumawati R Lia, Berger Fabian K, Laukien Friederike, Zimmermann Ortrud, Bohne Wolfgang, Groß Uwe, Bader Oliver, Zautner Andreas E
Institut für Medizinische Mikrobiologie, Universitätsmedizin Göttingen, Göttingen, Germany.
Leibniz-Institut DSMZ-Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Braunschweig, Germany.
Front Microbiol. 2019 Sep 10;10:2087. doi: 10.3389/fmicb.2019.02087. eCollection 2019.
, a Gram-positive spore-forming bacterium, is the leading cause of nosocomial diarrhea worldwide and therefore a substantial burden to the healthcare system. During the past decade, hypervirulent PCR-ribotypes (RT) e.g., RT027 or RT176 emerged rapidly all over the world, associated with both, increased severity and mortality rates. It is thus of great importance to identify epidemic strains such as RT027 and RT176 as fast as possible. While commonly used diagnostic methods, e.g., multilocus sequence typing (MLST) or PCR-ribotyping, are time-consuming, proteotyping offers a fast, inexpensive, and reliable alternative solution. In this study, we established a MALDI-TOF-based typing scheme for A total of 109 ribotyped strains representative for five MLST clades were analyzed by MALDI-TOF. MLST, based on whole genome sequences, and PCR-ribotyping were used as reference methods. Isoforms of MS-detectable biomarkers, typically ribosomal proteins, were related with the deduced amino acid sequences and added to the proteotyping scheme. In total, we were able to associate nine biomarkers with their encoding genes and include them in our proteotyping scheme. The discriminatory capacity of the proteotyping scheme was mainly based on isoforms of L28-M (2 main isoforms), L35-M (4 main isoforms), and S20-M (2 main isoforms) giving rise to at least 16 proteotyping-derived types. In our test population, five of these 16 proteotyping-derived types were detected. These five proteotyping-derived types did not correspond exactly to the included five MLST-based clades, nevertheless the subtyping depth of both methods was equivalent. Most importantly, proteotyping-derived clade B contained only isolates of the hypervirulent RT027 and RT176. Proteotyping is a stable and easy-to-perform intraspecies typing method and a promising alternative to currently used molecular techniques. It is possible to distinguish the group of RT027 and RT176 isolates from non-RT027/non-RT176 isolates using proteotyping, providing a valuable diagnostic tool.
作为一种革兰氏阳性产芽孢细菌，是全球医院获得性腹泻的主要原因，因此给医疗系统带来了沉重负担。在过去十年中，高毒力PCR核糖体分型（RT），例如RT027或RT176在全球迅速出现，与病情严重程度和死亡率增加都有关联。因此，尽快识别诸如RT027和RT176等流行菌株至关重要。虽然常用的诊断方法，例如多位点序列分型（MLST）或PCR核糖体分型很耗时，但蛋白质分型提供了一种快速、廉价且可靠的替代解决方案。在本研究中，我们建立了一种基于基质辅助激光解吸电离飞行时间质谱（MALDI-TOF）的分型方案，用于对总共109株代表五个MLST进化枝的核糖体分型菌株进行MALDI-TOF分析。基于全基因组序列的MLST和PCR核糖体分型用作参考方法。MS可检测生物标志物的同工型，通常是核糖体蛋白，与推导的氨基酸序列相关联，并添加到蛋白质分型方案中。总共，我们能够将9种生物标志物与其编码基因相关联，并将它们纳入我们的蛋白质分型方案。该蛋白质分型方案的鉴别能力主要基于L28-M（2种主要同工型）、L35-M（4种主要同工型）和S20-M（2种主要同工型）的同工型，产生了至少16种蛋白质分型衍生类型。在我们的测试群体中，检测到了这16种蛋白质分型衍生类型中的5种。这5种蛋白质分型衍生类型与所纳入的5个基于MLST的进化枝并不完全对应，不过两种方法的亚型分型深度相当。最重要的是，蛋白质分型衍生的进化枝B仅包含高毒力RT027和RT176的分离株。蛋白质分型是一种稳定且易于实施的种内分型方法，是当前使用的分子技术的一种有前景的替代方法。使用蛋白质分型可以将RT027和RT176分离株组与非RT027/非RT176分离株区分开来，提供了一种有价值的诊断工具。
