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Interaction of Bacterial Membrane Vesicles with Specific Species and Their Potential for Delivery to Target Cells

View Article: PubMed Central - PubMed

ABSTRACT

Membrane vesicles (MVs) are secreted from a wide range of microbial species and transfer their content to other cells. Although MVs play critical roles in bacterial communication, whether MVs selectively interact with bacterial cells in microbial communities is unclear. In this study, we investigated the specificity of the MV-cell interactions and evaluated the potential of MVs to target bacterial cells for delivery. MV association with bacterial cells was examined using a fluorescent membrane dye to label MVs. MVs derived from the enterobacterium Buttiauxella agrestis specifically interacted with cells of the parent strain but interacted less specifically with those of other genera tested in this study. Electron microscopic analyses showed that MVs were not only attached on B. agrestis cells but also fused to them. The interaction energy, which was characterized by hydrodynamic diameter and zeta potential based on the Derjaguin–Landau–Verwey–Overbeek (DLVO) theory, was significant low between MVs and cells in B. agrestis, compared to those between B. agrestis MVs and cells of other genera. Similar specific interaction was also occurred between B. agrestis MVs and cells of six other species belonging to Buttiauxella spp. B. agrestis harboring plasmid pBBR1MCS-1 secreted plasmid-containing MVs (p-MVs), and plasmid DNA in p-MVs was transferred to the same species. Moreover, antibiotic-associated MVs enabled effective killing of target species; the survival rate of B. agrestis was lower than those of Escherichia coli and Pseudomonas aeruginosa in the presence of gentamicin-associated MVs derived from B. agrestis. Altogether, we provide the evidence that MVs selectively interact with target bacterial cells and offer a new avenue for controlling specific bacterial species using bacterial MVs in microbial communities.

No MeSH data available.


Detection and observation of the association of MVs derived from B. agrestis CUETM77-167 with CUETM77-167 and E. coli MG1655 cells. FM4-64-labeled MVs (20 μg/mL of phospholipids) were incubated with FITC-labeled bacterial cells for 30 min at 30°C. (A,B) Flow cytometry analysis of CUETM77-167 (A) and MG1655 cells (B) associated with MVs. FITC-labeled cells (green plots) and FITC-labeled cells incubated with FM4-64-labeled MVs (red plots) underwent flow cytometry independently, and the plots are shown in the same chart. (C,D) Fluorescence microscopy observation of FITC-labeled CUETM77-167 (C) and MG1655 (D) cells incubated with FM4-64-labeled MVs derived from CUETM77-167. The bars indicate 5 μm.
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Figure 2: Detection and observation of the association of MVs derived from B. agrestis CUETM77-167 with CUETM77-167 and E. coli MG1655 cells. FM4-64-labeled MVs (20 μg/mL of phospholipids) were incubated with FITC-labeled bacterial cells for 30 min at 30°C. (A,B) Flow cytometry analysis of CUETM77-167 (A) and MG1655 cells (B) associated with MVs. FITC-labeled cells (green plots) and FITC-labeled cells incubated with FM4-64-labeled MVs (red plots) underwent flow cytometry independently, and the plots are shown in the same chart. (C,D) Fluorescence microscopy observation of FITC-labeled CUETM77-167 (C) and MG1655 (D) cells incubated with FM4-64-labeled MVs derived from CUETM77-167. The bars indicate 5 μm.

Mentions: To further corroborate the specific interaction of MVs derived from B. agrestis with the same species, the association between MVs and bacterial cells was analyzed at the single-cell level. B. agrestis cells labeled with FITC were incubated with FM4-64-labeled MVs for 30 min, and washed cells were analyzed by flow cytometry. As a control, FITC-labeled cells, which were not treated with MVs, were also assessed by flow cytometry. The fluorescence intensities of both FM4-64 and FITC were detected in MV-reacted cells (approximately 1 × 102 RFU), while only the intensity of FITC was detected in the control sample (Figure 2A). Then, the same experiment was conducted using E. coli and MVs derived from B. agrestis. The fluorescence of FM4-64 was not detected in E. coli either with or without B. agrestis MVs (Figure 2B). Similar results were also obtained by fluorescence microscopy analysis; the fluorescence of both FM4-64 and FITC was observed in MV-treated B. agrestis (Figure 2C), but the fluorescence of FM4-64 was not observed in E. coli, which was treated identically (Figure 2D). These data confirm the hypothesis that MVs derived from B. agrestis specifically interact with bacterial cells of the same species.


Interaction of Bacterial Membrane Vesicles with Specific Species and Their Potential for Delivery to Target Cells
Detection and observation of the association of MVs derived from B. agrestis CUETM77-167 with CUETM77-167 and E. coli MG1655 cells. FM4-64-labeled MVs (20 μg/mL of phospholipids) were incubated with FITC-labeled bacterial cells for 30 min at 30°C. (A,B) Flow cytometry analysis of CUETM77-167 (A) and MG1655 cells (B) associated with MVs. FITC-labeled cells (green plots) and FITC-labeled cells incubated with FM4-64-labeled MVs (red plots) underwent flow cytometry independently, and the plots are shown in the same chart. (C,D) Fluorescence microscopy observation of FITC-labeled CUETM77-167 (C) and MG1655 (D) cells incubated with FM4-64-labeled MVs derived from CUETM77-167. The bars indicate 5 μm.
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Figure 2: Detection and observation of the association of MVs derived from B. agrestis CUETM77-167 with CUETM77-167 and E. coli MG1655 cells. FM4-64-labeled MVs (20 μg/mL of phospholipids) were incubated with FITC-labeled bacterial cells for 30 min at 30°C. (A,B) Flow cytometry analysis of CUETM77-167 (A) and MG1655 cells (B) associated with MVs. FITC-labeled cells (green plots) and FITC-labeled cells incubated with FM4-64-labeled MVs (red plots) underwent flow cytometry independently, and the plots are shown in the same chart. (C,D) Fluorescence microscopy observation of FITC-labeled CUETM77-167 (C) and MG1655 (D) cells incubated with FM4-64-labeled MVs derived from CUETM77-167. The bars indicate 5 μm.
Mentions: To further corroborate the specific interaction of MVs derived from B. agrestis with the same species, the association between MVs and bacterial cells was analyzed at the single-cell level. B. agrestis cells labeled with FITC were incubated with FM4-64-labeled MVs for 30 min, and washed cells were analyzed by flow cytometry. As a control, FITC-labeled cells, which were not treated with MVs, were also assessed by flow cytometry. The fluorescence intensities of both FM4-64 and FITC were detected in MV-reacted cells (approximately 1 × 102 RFU), while only the intensity of FITC was detected in the control sample (Figure 2A). Then, the same experiment was conducted using E. coli and MVs derived from B. agrestis. The fluorescence of FM4-64 was not detected in E. coli either with or without B. agrestis MVs (Figure 2B). Similar results were also obtained by fluorescence microscopy analysis; the fluorescence of both FM4-64 and FITC was observed in MV-treated B. agrestis (Figure 2C), but the fluorescence of FM4-64 was not observed in E. coli, which was treated identically (Figure 2D). These data confirm the hypothesis that MVs derived from B. agrestis specifically interact with bacterial cells of the same species.

View Article: PubMed Central - PubMed

ABSTRACT

Membrane vesicles (MVs) are secreted from a wide range of microbial species and transfer their content to other cells. Although MVs play critical roles in bacterial communication, whether MVs selectively interact with bacterial cells in microbial communities is unclear. In this study, we investigated the specificity of the MV-cell interactions and evaluated the potential of MVs to target bacterial cells for delivery. MV association with bacterial cells was examined using a fluorescent membrane dye to label MVs. MVs derived from the enterobacterium Buttiauxella agrestis specifically interacted with cells of the parent strain but interacted less specifically with those of other genera tested in this study. Electron microscopic analyses showed that MVs were not only attached on B. agrestis cells but also fused to them. The interaction energy, which was characterized by hydrodynamic diameter and zeta potential based on the Derjaguin–Landau–Verwey–Overbeek (DLVO) theory, was significant low between MVs and cells in B. agrestis, compared to those between B. agrestis MVs and cells of other genera. Similar specific interaction was also occurred between B. agrestis MVs and cells of six other species belonging to Buttiauxella spp. B. agrestis harboring plasmid pBBR1MCS-1 secreted plasmid-containing MVs (p-MVs), and plasmid DNA in p-MVs was transferred to the same species. Moreover, antibiotic-associated MVs enabled effective killing of target species; the survival rate of B. agrestis was lower than those of Escherichia coli and Pseudomonas aeruginosa in the presence of gentamicin-associated MVs derived from B. agrestis. Altogether, we provide the evidence that MVs selectively interact with target bacterial cells and offer a new avenue for controlling specific bacterial species using bacterial MVs in microbial communities.

No MeSH data available.