Unité de Recherche sur les Maladies Infectieuses et Tropicales Emergentes, UMR CNRS 6236 IRD 3R198, IFR 48 Faculté de Médecine, Université de la Méditerranée, Marseille, France.
Environ Microbiol Rep. 2010 Dec;2(6):752-60. doi: 10.1111/j.1758-2229.2010.00176.x.
We have developed a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based identification technique for Planctomycetes organisms, which are used here as bacteria of suitable diversity at genus and species level for testing resolution of the method. Planctomyces maris ATCC 29201, Planctomyces brasiliensis ATCC 49424(T) , P. brasiliensis ATCC 49425, Planctomyces limnophilus ATCC 43296(T) , Blastopirellula marina ATCC 49069(T) , Rhodopirellula baltica DSM 10527(T) and Gemmata obscuriglobus DSM 5831(T) were cultured on half-strength marine broth and agar, or alternatively on caulobacter broth and agar. The resulting pellets of organisms (liquid) or colonies (solid agar) were directly applied to a MALDI-TOF plate. This yielded a reproducible, unique protein profiles comprising 23-39 peaks ranging in size from 2403 to 12 091 Da. These peaks were unambiguously distinguished from any of the 3038 bacterial spectra in the Brüker database. Matrix-assisted laser desorption/ionization time-of-flight patterns were similar for isolates grown in solid and in liquid medium, albeit the patterns from solid growth were more easily interpretable. After the incorporation of the herein determined profiles into the Brüker database, Planctomycetes isolates were blindly identified within 10 min, with an identification score in the range of 1.8 to 2.3. Matrix-assisted laser desorption/ionization time-of-flight-based clustering of these Planctomycetes organisms was consistent with 16S rDNA-based phylogeny. However, the incorporation of additional non-Planctomycetes MALDI-TOF profiles in the analysis resulted in inconsequential clustering. In conclusion, MALDI-TOF protein profiling is a new approach for the rapid and accurate identification of cultured environmental organisms, as illustrated in this study through the analysis of Planctomycetes.
我们开发了一种基于基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）的浮霉菌鉴定技术，这里使用的浮霉菌是在属和种水平上具有适当多样性的细菌，用于测试该方法的分辨率。Planctomyces maris ATCC 29201、Planctomyces brasiliensis ATCC 49424(T)、P. brasiliensis ATCC 49425、Planctomyces limnophilus ATCC 43296(T)、Blastopirellula marina ATCC 49069(T)、Rhodopirellula baltica DSM 10527(T)和 Gemmata obscuriglobus DSM 5831(T)在半强度海洋肉汤和琼脂或 alternatively 在 caulobacter 肉汤和琼脂上培养。将生物体的沉淀物（液体）或菌落（固体琼脂）直接涂在 MALDI-TOF 板上。这产生了可重复的、独特的蛋白质谱，包含 23-39 个大小在 2403 到 12091 Da 之间的峰。这些峰与 Bruker 数据库中 3038 个细菌谱中的任何一个都可以明确区分。尽管来自固体生长的图案更容易解释，但在固体和液体培养基中生长的分离物的基质辅助激光解吸/电离飞行时间模式相似。将在此处确定的图谱纳入 Bruker 数据库后，在 10 分钟内即可对浮霉菌分离物进行盲目鉴定，鉴定得分在 1.8 到 2.3 之间。基于这些浮霉菌的基质辅助激光解吸/电离飞行时间聚类与基于 16S rDNA 的系统发育一致。然而，在分析中加入额外的非浮霉菌 MALDI-TOF 图谱不会导致聚类结果发生重大变化。总之，MALDI-TOF 蛋白谱分析是一种快速准确鉴定培养环境生物的新方法，正如本研究通过对浮霉菌的分析所表明的那样。
