Visualization of Extracellular Vesicles of Prokaryotes and Eukaryotic Microbes

Ki Woo Kim

doi:10.9729/AM.2018.48.4.96

https://doi.org/10.9729/AM.2018.48.4.96

Visualization of Extracellular Vesicles of Prokaryotes and Eukaryotic Microbes

Applied Microscopy 2018;48:96-101

Published online December 28, 2018

Ki Woo Kim

¹School of Ecology and Environmental System, Kyungpook National University, Sangju 37224, Korea, ²Tree Diagnostic Center, Kyungpook National University, Sangju 37224, Korea

Correspondence to: Kim KW, http://orcid.org/0000-0002-7010-0336, Tel: +82-54-530-1246, Fax: +82-54-530-1248, E-mail: kiwoo@knu.ac.kr

Received December 19, 2018; Revised December 27, 2018; Accepted December 27, 2018.

his is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted noncommercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract: The release of nanoscale membrane-bound vesicles is common in all three domains of life. These vesicles are involved in a variety of biological processes such as cell-to-cell communication, horizontal gene transfer, and substrate transport. Prokaryotes including bacteria and archaea release membrane vesicles (MVs) (20 to 400 nm in diameter) into their extracellular milieu. In spite of structural differences in cell envelope, both Gram-positive and negative bacteria produce MVs that contain the cell membrane of each bacterial species. Archaeal MVs characteristically show surface-layer encircling the vesicles. Filamentous fungi and yeasts as eukaryotic microbes produce bilayered exosomes that have varying electron density. Microbes also form intracellular vesicles and minicells that are similar to MVs and exosomes in shape. Electron and fluorescence microscopy could reveal the presence of DNA in MVs and exosomes. Given the biogenesis of extracellular vesicles from the donor cell, in situ high-resolution microscopy can provide insights on the structural mechanisms underlying the formation and release of microbial extracellular vesicles.; Keywords : Archaea, Bacteria, Fungi, Membrane vesicle

Figures

Fig. 1. Schematics of extracellular vesicles of microbial cells. ERG, ergosterol; GlcConj, glycoconjugates; GSL, glycosphingolipids; LPS, lipopolysaccharide; OMP, outer membrane protein. From with permission from the publisher.

Fig. 2. Membrane vesicles of Gram-negative bacteria. (A) Transmission electron micrograph of Escherichia coli (b) labeled with anti-lipopolysaccharide antibody and Protein A Gold. Subsets depict membrane vesicles. Bar=150 nm. (B) Scanning electron micrograph of Xylella fastidiosa having membrane vesicles (arrows). Bar=200 nm. A: from ; B: from with permission from the publisher.

Fig. 3. Transmission electron micrographs of membrane vesicles of Staphylococcus aureus. (A) Membrane vesicles (arrow) released from the cell. (B) Membrane vesicles pelleted by ultracentrifugation from the bacterial culture. Bars=100 nm. A and B: from with permission from the publisher.

Fig. 4. Transmission electron micrograph of membrane vesicles of Sulfolobus solfataricus. These vesicles are surrounded with surface-layer (arrow). From with permission from the publisher.

Fig. 5. Transmission electron micrographs of membrane vesicles of Saccharomyces cerevisiae. (A–D) Exosomes. They are bilayered structures with varying electron density. Bar=100 nm. A–D: from with permission from the publisher.

Fig. 6. Transmission electron micrographs of Agrobacterium tumefaciens. (A) Cell having the partially filled acidocalcisome (arrow). (B) Cell having the empty acidocalcisome (arrow). A and B: from with permission from the publisher.

Fig. 7. Localization of DNA in membrane vesicles. (A–C) Super resolution micrographs of membrane vesicles of Pseudomonas aeruginosa. (A) Membrane vesicles stained with the lipophilic fluorescent dye, DiO (green). Bar=1 μm. (B) Membrane vesicles stained with the DNA-specific dye, 5- ethynyl-2-deoxuridine (EdU). (C) Merged image of (A) and (B). (D) Transmission electron micrograph of membrane vesicles of Shewanella vesiculosa. Electron-dense particles indicate gold particles with an antibody specific for DNA. OM, outer membrane; PM, plasma membrane. A–C: from ; D: from with permission from the publisher.

References

Bielaszewska, M, Ruter, C, Bauwens, A, Greune, L, Jarosch, KA, Steil, D, Zhang, W, He, X, Lloubes, R, Fruth, A, Kim, KS, Schmidt, MA, Dobrindt, U, Mellmann, A, and Karch, H (2017). Host cell interactions of outer membrane vesicle-associated virulence factors of enterohemorrhagic Escherichia coli O157: intercellular delivery, trafficking and mechanisms of cell injury. PLoS Pathog. 13, e1006159.
Bitto, NJ, Chapman, R, Pidot, S, Costin, A, Lo, C, Choi, J, D’Cruze, T, Reynolds, EC, Dashper, SG, Turnbull, L, Whitchurch, CB, Stinear, TP, Stacey, KJ, and Ferrero, RL (2017). Bacterial membrane vesicles transport their DNA cargo into host cells. Sci Rep. 7, 7072.
Bohuszewicz, O, Liu, J, and Low, HH (2016). Membrane remodeling in bacteria. J Struct Biol. 196, 3-14.
Chernov, VM, Mouzykantov, AA, Baranova, NB, Medvedeva, ES, Grygorieva, TY, Trushin, MV, Vishnyakov, IE, Sabantsev, AV, Borchsenius, SN, and Chernova, OA (2014). Extracellular membrane vesicles secreted by mycoplasma Acholeplasma laidlawii PG8 are enriched in virulence proteins. J Proteom. 110, 117-128.
De Silva, RP, Puccia, R, Rodrigues, ML, Oliveira, DL, Joffe, LS, Cesar, GV, Nimrichter, L, Goldenberg, S, and Alves, LR (2015). Extracellular vesicle-mediated export of fungal RNA. Sci Rep. 5, 7763.
Deatherage, BL, and Cookson, BT (2012). Membrane vesicle release in bacteria. eukaryotes, and archaea: a conserved yet underappreciated aspect of microbial life. Inf Immun. 80, 1948-1957.
Ellen, AF, Albers, S-V, Huibers, W, Pitcher, A, Hobel, CFV, Schwarz, H, Folea, M, Schouten, S, Boekema, EJ, Poolman, B, and Driessen, AJM (2009). Proteomic analysis of secreted membrane vesicles of archaeal Sulfolobus species reveals the presence of endosome sorting complex components. Extremophiles. 13, 67-79.
Giessen, TW, and Silver, PA (2016). Encapsulation as a strategy for the design of biological compartmentalization. J Mol Biol. 428, 916-927.
Ionescu, M, Zaini, PA, Baccari, C, Tran, S, Da Silva, AM, and Lindow, SE (2014). Xylella fastidiosa outer membrane vesicles modulate plant colonization by blocking attachment to surfaces. PNAS. 111, E3910-3918.
Joffe, LS, Nimrichter, L, Rodrigues, M, and Del Poeta, M (2016). Potential roles of fungal extracellular vesicles during infection. mSphere. 1, e00099-16.
Knox, KW, Vesk, M, and Work, E (1966). Relation between excreted lipopolysaccharide complexes and surface structures of a lysine-limited culture of Escherichia coli. J Bacteriol. 92, 1206-1217.
Liu, Y, Defourny, KAY, Smid, EJ, and Abee, T (2018). Gram-positive bacterial extracelluar vesicles and their impact on health and disease. Front Microbiol. 9, 1502.
Marguet, E, Gaudin, M, Gauliard, E, Fourquaux, I, Le, Blond, du Plouy, S, Matsui, I, and Forterre, P (2013). Membrane vesicles, nanopods and/or nanotubes produced by hyperthermophilic archaea of the genus Thermococcus. Biochem Soc Trans. 41, 436-442.
Martínez-Penafiel, E, Fernandez-Ramirez, F, Ishida, C, Reyes-Cortes, R, Sepulveda-Robles, O, Guarneros-Pena, G, Bermudez-Cruz, RM, and Kameyama, L (2012). Overexpression of Ipe protein from the coli-phage mEp021 induces pleiotropic effects involving haemolysis by HlyE-containing vesicles and cell death. Biochimie. 94, 1262-1273.
Oliveira, DL, Nakayasu, ES, Joffe, LS, Guimaraes, AJ, Sobreira, TJP, Nosanchuk, JD, Cordero, RJB, Frases, S, Casadevall, A, Almeida, IC, Nimrichter, L, and Rodrigues, ML (2010). Characterization of yeast extracellular vesicles: evidence for the participation of different pathways of cellular traffic in vesicle biogenesis. PLoS ONE. 5, e11113.
Pérez-Cruz, C, Carrión, O, Delgado, L, Martinez, G, López-Iglesias, C, and Mercade, E (2013). New type of outer membrane vesicle produced by the Gram-negaive bacterium Shewanella vesiculosa M7^T: implications for DNA content. Appl Environ Microbiol. 79, 1874-1881.
Pérez-Cruz, C, Delgado, L, López-Iglesias, C, and Mercade, E (2015). Outer-inner membrane vesicles naturally secreted by Gram-negative pathogenic bacteria. PLoS ONE. 10, e0116896.
Raposo, G, and Stoorvogel, W (2013). Extracellular vesicles: exosomes, microvesicles, and friends. J Cell Biol. 200, 373-383.
Schwechheimer, C, and Kuehn, MJ (2015). Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions. Nat Rev Microbiol. 13, 605-619.
Seufferheld, MJ, Kim, KM, Whitfield, J, Valerio, A, and Caetano-Anolles, G (2011). Evolution of vacuolar proton pyrophosphatase domains and volutin granules: clues into the early evolutionalry origin of the acidocalcisome. Biol Direct. 6, 50.
Silva, BMA, Prados-Rosales, R, Espadas-Moreno, J, Wolf, JM, Luque-Garcia, JL, Goncalves, T, and Casadevall, A (2014). Characterization of Alternaria infectoria extracelluar vesicles. Med Mycol. 52, 202-210.
Sjöström, AE, Sandblad, L, Uhlin, BE, and Wai, SN (2015). Membrane vesicle-mediated release of bacterial RNA. Sci Rep. 5, 15329.
Soler, N, Marguet, E, Verbavatz, J-M, and Forterre, P (2008). Virus-like vesicles and extracellular DNA produced by hyperthermophilic archaea of the order Thermococcales. Res Microbiol. 159, 390-399.
Thery, C, Ostrowski, M, and Segura, E (2009). Membrane vesicles as conveyors of immune responses. Nat Rev Immunol. 9, 581-593.
Toyofuku, M, Tashiro, Y, Hasegawa, Y, Kurosawa, M, and Nomura, N (2015). Bacterial membrane vesicles, an overlooked environmental colloid: biology, environmental perspectives and applications. Adv Colloid Interface Sci. 226, 65-77.
Wang, X, Thompson, CD, Weidenmaier, C, and Lee, JC (2018). Release of Staphyloccous aureus extracellular vesicles and their application as a vaccine platform. Nat Commun. 9, 1379.