Department of Microbiology, National Center of Infectious and Parasitic Diseases, Sofia, Bulgaria.
Faculty of Chemistry and Pharmacy, Sofia University, Sofia, Bulgaria.
Front Cell Infect Microbiol. 2023 Jan 18;12:1091341. doi: 10.3389/fcimb.2022.1091341. eCollection 2022.
The blood microbiome is still an enigma. The existence of blood microbiota in clinically healthy individuals was proven during the last 50 years. Indirect evidence from radiometric analysis suggested the existence of living microbial forms in erythrocytes. Recently targeted nucleic acid sequencing demonstrated rich microbial biodiversity in the blood of clinically healthy individuals. The morphology and proliferation cycle of blood microbiota in peripheral blood mononuclear cells (PBMC) isolated from freshly drawn and cultured whole blood are obscure.
To study the life cycle of blood microbiota we focused on light, and electron microscopy analysis. Peripheral blood mononuclear cells isolated from freshly drawn blood and stress-cultured lysed whole blood at 43°C in presence of vitamin K from healthy individuals were studied.
Here, we demonstrated that free circulating microbiota in the PMBC fraction possess a well-defined cell wall and proliferate by budding or through a mechanism similar to the extrusion of progeny bodies. By contrast, stress-cultured lysed whole blood microbiota proliferated as cell-wall deficient microbiota by forming electron-dense or electron-transparent bodies. The electron-dense bodies proliferated by fission or produce in chains Gram-negatively stained progeny cells or enlarged and burst to release progeny cells of 180 - 200 nm size. On the other hand, electron-transparent bodies enlarged and emitted progeny cells through the membrane. A novel proliferation mechanism of blood microbiota called by us "a cell within a cell" was observed. It combines proliferation of progeny cells within a progeny cell which is growing within the "mother" cell.
The rich biodiversity of eukaryotic and prokaryotic microbiota identified in blood by next-generation sequencing technologies and our microscopy results suggest different proliferation mechanisms in whole and cultured blood. Our documented evidence and conclusions provide a more comprehensive view of the existence of normal blood microbiota in healthy individuals.
血液微生物组仍然是一个谜。在过去的 50 年里，已经证明在临床健康个体中存在血液微生物群。放射性分析的间接证据表明，在红细胞中存在有生命的微生物形式。最近的靶向核酸测序技术证明，在临床健康个体的血液中存在丰富的微生物多样性。从刚抽出并培养的全血中分离出的外周血单核细胞（PBMC）中，血液微生物群的形态和增殖周期尚不清楚。
为了研究血液微生物群的生命周期，我们专注于光镜和电子显微镜分析。从新鲜抽取的血液和在 43°C 下于含维生素 K 的裂解全血中培养的压力培养的外周血单核细胞中分离出 PBMC，并对其进行研究。
在这里，我们证明了在 PMBC 部分中自由循环的微生物群具有明确的细胞壁，并通过出芽或通过类似于挤出后代体的机制进行增殖。相比之下，压力培养的裂解全血微生物群通过形成无细胞壁的微生物群进行增殖，形成电子致密或电子透明的体。电子致密体通过分裂或形成革兰氏阴性染色的后代细胞链进行增殖，或者增大并爆裂以释放 180-200nm 大小的后代细胞。另一方面，电子透明体通过膜增大并释放后代细胞。我们观察到一种称为“细胞内的细胞”的血液微生物群新的增殖机制。它结合了在“母”细胞内生长的后代细胞内的后代细胞的增殖。
下一代测序技术和我们的显微镜结果在血液中鉴定出的真核生物和原核生物微生物群的丰富多样性表明，全血和培养血中存在不同的增殖机制。我们的有文件证明和结论提供了对健康个体中正常血液微生物群存在的更全面的认识。
