Presentation_1_Phage-Mediated Explosive Cell Lysis Induces the Formation of a Different Type of O-IMV in Shewanella vesiculosa M7T.pdf

• Main Authors: Nicolás Baeza, Lidia Delgado, Jaume Comas, Elena Mercade
• Format: Conference Object
• Language: unknown
• Published: 2021
• Subjects: Microbiology; Microbial Genetics; Microbial Ecology; Mycology; membrane vesicles; gram-negative bacteria; explosive cell lysis; OMV; O-IMV; DNA; flow cytometry; electron microscopy; Antarctic; Antarc*
• Online Access: https://doi.org/10.3389/fmicb.2021.713669.s001; https://figshare.com/articles/presentation/Presentation_1_Phage-Mediated_Explosive_Cell_Lysis_Induces_the_Formation_of_a_Different_Type_of_O-IMV_in_Shewanella_vesiculosa_M7T_pdf/16756846

Shewanella vesiculosa M7 T is a cold-adapted Antarctic bacterium that has a great capacity to secrete membrane vesicles (MVs), making it a potentially excellent model for studying the vesiculation process. S. vesiculosa M7 T undergoes a blebbing mechanism to produce different types of MVs, including outer membrane vesicles and outer-inner membrane vesicles (O-IMVs). More recently, other mechanisms have been considered that could lead to the formation of O-IMVs derived from prophage-mediated explosive cell lysis in other bacteria, but it is not clear if they are of the same type. The bacterial growth phase could also have a great impact on the type of MVs, although there are few studies on the subject. In this study, we used high-resolution flow cytometry, transmission electron microscopy, and cryo-electron microscopy (Cryo-EM) analysis to determine the amount and types of MVs S. vesiculosa M7 T secreted during different growth phases. We show that MV secretion increases during the transition from the late exponential to the stationary phase. Moreover, prophage-mediated explosive cell lysis is activated in S. vesiculosa M7 T , increasing the heterogeneity of both single- and double-layer MVs. The sequenced DNA fragments from the MVs covered the entire genome, confirming this explosive cell lysis mechanism. A different structure and biogenesis mechanisms for the explosive cell lysis-derived double-layered MVs was observed, and we propose to name them explosive O-IMVs, distinguishing them from the blebbing O-IMVs; their separation is a first step to elucidate their different functions. In our study, we used for the first time sorting by flow cytometry and Cryo-EM analyses to isolate bacterial MVs based on their nucleic acid content. Further improvements and implementation of bacterial MV separation techniques is essential to develop more in-depth knowledge of MVs.