IHU Méditerranée Infection, Marseille, France.
Aix-Marseille Université, IRD, APHM, MEPHI, Marseille, France.
Microbiol Spectr. 2021 Dec 22;9(3):e0106921. doi: 10.1128/spectrum.01069-21.
The growing application of metagenomics to different ecological and microbiome niches in recent years has enhanced our knowledge of global microbial biodiversity. Among these abundant and widespread microbes, the candidate phyla radiation (CPR) group has been recognized as representing a large proportion of the microbial kingdom (>26%). CPR are characterized by their obligate symbiotic or exoparasitic activity with other microbial hosts, mainly bacteria. Currently, isolating CPR is still considered challenging for microbiologists. The idea of this study was to develop an adapted protocol for the coculture of CPR with a suitable bacterial host. Based on various sputum samples, we tried to enrich CPR ( members) and to cocultivate them with pure hosts (Schaalia odontolytica). This protocol was monitored by TaqMan real-time quantitative PCR (qPCR) using a system specific for designed in this study, as well as by electron microscopy and sequencing. We succeeded in coculturing and sequencing the complete genomes of two new species, " Minimicrobia naudis" and " Minimicrobia vallesae." In addition, we noticed a decrease in the values of and a significant multiplication through their physical association with Schaalia odontolytica strains in the enriched medium that we developed. This work may help bridge gaps in the genomic database by providing new CPR members, and in the future, their currently unknown characteristics may be revealed. In this study, the first TaqMan real-time quantitative PCR (qPCR) system, targeting phylum, has been developed. This technique can specifically quantify members in any sample of interest in order to investigate their prevalence. In addition, another easy, specific, and sensitive protocol has been developed to maintain the viability of cells in an enriched medium with their bacterial host. The use of this protocol facilitates subsequent studies of the phenotypic characteristics of CPR and their physical interactions with bacterial species, as well as the sequencing of new genomes to improve the current database.
近年来，宏基因组学在不同的生态和微生物生境中的应用不断增加，提高了我们对全球微生物生物多样性的认识。在这些丰富而广泛存在的微生物中，候选门辐射（CPR）群被认为代表了微生物王国的很大一部分（>26%）。CPR 的特点是与其他微生物宿主（主要是细菌）有共生或外寄生的活动。目前，微生物学家分离 CPR 仍然具有挑战性。本研究的目的是开发一种与合适的细菌宿主共培养 CPR 的适应方案。基于各种痰液样本，我们试图富集 CPR（成员）并与纯宿主（Schaalia odontolytica）共培养。该方案通过使用我们在这项研究中设计的特定于系统的 TaqMan 实时定量 PCR（qPCR）进行监测，以及通过电子显微镜和测序进行监测。我们成功地共培养和测序了两个新的 物种，“Minimicrobia naudis”和“Minimicrobia vallesae”的完整基因组。此外，我们注意到与 Schaalia odontolytica 菌株在我们开发的富集培养基中的物理关联，导致 值下降，并且通过它们的物理关联显著增殖。这项工作可能有助于通过提供新的 CPR 成员来弥合基因组数据库中的空白，并且在未来，它们目前未知的特征可能会被揭示。 在这项研究中，开发了第一个针对 门的 TaqMan 实时定量 PCR（qPCR）系统。该技术可以特异性地定量任何感兴趣的样本中的 成员，以调查它们的流行程度。此外，还开发了另一种简单、特异、灵敏的方案，以维持富含其细菌宿主的培养基中 CPR 细胞的活力。该方案的使用便于随后研究 CPR 的表型特征及其与细菌物种的物理相互作用，以及测序新的基因组以改进当前数据库。
