Silver nanoparticle-coated “cyborg” microorganisms: rapid assembly of polymer-stabilised nanoparticles on microbial cells

Abstract

Fabrication of “cyborg” cells (biological cells with surfaces functionalised using a variety of nanomaterials) has become a fascinating area in cell surface engineering. Here we report a simple procedure for fabrication of polycation-stabilised 50 nm silver nanoparticles and application of these nanoparticles for fabrication of viable “cyborg” microbial cells (yeast and bacteria). Cationic polymer-stabilised nanoparticles electrostatically adhere to microbial cells producing an even monolayer on the cell walls, as demonstrated using enhanced dark-field microscopy, atomic force microscopy and microelectrophoresis. Our procedure is exceptionally fast, being completed within 20 min after introduction of cells into nanoparticle aqueous suspensions. Polymer-stabilised silver nanoparticles are highly biocompatible, with viability rates reaching 97%. We utilised “cyborg” cells built using bacteria and silver nanoparticles to deliver nanoparticles into C. elegans microworms. We believe that the technique described here will find numerous applications in cell surface engineering.

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Introduction

Cell surface engineering is a rapidly-growing multidisciplinary area of research aiming at fabrication of artificial functional shells at the surfaces of biological cells.¹ Interfacing cells with nanoscale coatings, such as polymer multilayers, assembled via electrostatic or hydrogen bonding, or encapsulation into spore-mimicking inorganic shells, is regarded as a potent way to attenuate or enhance the intrinsic properties of cells, to control the division of cells or to assemble cells into artificial multicellular clusters.^2–7 Starting from the simple two-component systems (i.e. viable yeast cells coated with a few polyelectrolyte (PE) layers), layer-by-layer (LbL)-deposited live cell-modifying nanocoatings have evolved into sophisticated multifunctional semi-permeable shells which can be precisely controlled during their fabrication and allow for the effective manipulations with the coated cells.^8–11 Briefly, LbL deposition of polymers onto cells practically follows the same procedure as with planar surfaces and colloid microparticles – cells are sequentially incubated in alternating polycations/polyanions solutions, resulting in formation of nanosized layered films of predetermined composition.^12–15 Recently, a great deal of effort has been focused on fabrication of functional coatings on cell surfaces produced using nanoparticles (NPs) or nanorods. Polymer films, sequentially assembled on living cells, have been successfully utilised to immobilise noble metal NPs, carbon nanotubes and graphene oxide particles, among many other examples of LbL-mediated cell surface engineering.^16–19 Fabrication of pure or nanoparticle-doped multilayer polyelectrolyte coatings deposited on microbial or human cells has become a benchmark technology in fabrication of “cyborg” cells, demonstrating its feasibility in a number of applications, including, but not limited to, biosensors, tissue engineering, microfluidics, fabrication of artificial multicellular assemblies and cells protection.^16,20–23 Gold and silver NPs-doped LbL films were effectively used to perform surface-enhanced Raman scattering (SERS)-based characterisation of bacterial species.^24,25 Recently, LbL-coated “cyborg” cells have found applications in nanotoxicity testing as “smart food” particles to deliver metal or oxide nanoparticles and halloysite nanotubes into Caenorhabditis elegans microworms, allowing the control of uptake rates.^26,27

LbL polymer deposition technique, although effective and robust, suffers from prolonged deposition/washing cycles, which seriously limits its applicability to functionalise cells, especially in cases where the rapid deposition and short exposures times are desired. As a result, direct deposition approaches, utilising the polyelectrolyte-coated nanoparticles, have been suggested.²⁸ Briefly, a cationic polymer is employed to stabilise magnetic nanoparticles as well as to secure their straightforward deposition onto the oppositely-charged surfaces of cell walls or plasma membranes of biological cells.^29,30 This strategy benefits from very short exposure times (typically, a deposition procedure is completed within 20 min), which, in turn, reduces the toxic effects and cells loss caused by washing/introduction cycles. However, so far, the application of polyelectrolyte-coated nanoparticles has been limited only to magnetic iron oxide nanospheres and nanorods.³¹ However, other types of nanomaterials, particularly, silver nanoparticles, which are routinely used in LbL-based cell surface engineering, are promising candidates for the direct deposition onto the cells, taking into account the wide variety of their potential applications.³²

Here we report the fabrication of cationic polyelectrolyte-coated silver NPs and their utilisation for the direct single-step deposition onto microbial cells. We selected the popular polycations (poly(allylamine) hydrochloride (PAH), poly(diallyldimethylammonium) chloride (PDADMAC) and poly(ethyleneimine) (PEI)) as stabilisers for citrate-capped silver NPs. Polymer-coated nanoparticles were deposited onto yeast and bacterial cells, resulting in fabrication of uniform nanoparticle coating, similar to that obtained using the conventional LbL approach. We investigated the toxicity of PE-coated silver NPs and compared it with the toxicity of polymer stabilisers alone. Finally, we employed the NPs-coated cells as delivery vehicles to introduce silver NPs into C. elegans nematodes.

Materials and methods

Chemicals, microbial cells and nematode cultures

Ultrapure water purified using a MilliQ (Millipore) system was used throughout. The following chemicals: AgNO₃, sodium citrate, PAH (70 kDa), PDADMAC (medium weight), PEI (600–1000 kDa) were purchased from Sigma-Aldrich (USA) and used as received. Microbial cultures (Escherichia coli bacteria and Saccharomyces cerevisiae yeast) were obtained from Kazan Federal University (Department of Microbiology) cell culture collection and cultivated on Saburo nutrient medium and nutrient broth, respectively. C. elegans wild type strain (N2 Bristol) was maintained at 20 °C on agar-based nematode growth medium (NGM) (US Biological, USA) 15 cm plates with E. coli OP50 bacteria. Worms were synchronised by washing the gravid adults with M9 buffer (22 mM KH₂PO₄, 42 mM Na₂HPO₄, 85.5 mM NaCl, 1 mM MgSO₄), centrifuged and then the pellet was mixed with aqueous 2% NaOCl and 0.45 M NaOH and incubated for 10 min. After the complete destruction of all adult nematodes the preserved eggs were washed with M9 buffer and then transferred into sterile NGM plates. After 24 hours of incubation at 20 °C, the plates. A Carl Zeiss Stemi Div4 stereomicroscope was used to observe the worms.

Synthesis and stabilisation of silver nanoparticles (AgNPs)

Silver NPs have been synthesized following the previously published procedure.³³ Briefly, 36 mg AgNO₃ was dissolved in 20 mL of water and then boiled on a heated magnetic stirrer. Next, 4 mL of 1% sodium citrate was added, followed by the visible colour change (10 minutes after addition). Next, the citrate-coated NPs were removed and gradually cooled down to room temperature. Then, the citrate-capped AgNPs (2 mL) were added to 10 mL of aqueous polyelectrolyte solution (1% PAH, 5% PDADMAC, 5% PEI) and sonicated for 20 min. After sonication, the suspension was slowly stirred for 24 h, then washed thrice with water to remove the unbound polyelectrolyte molecules, followed by centrifugation. The colloid stability of nanoparticles in aqueous suspensions was monitored for 1 month.

Deposition of PE-stabilised AgNPs onto cells

Suspended cells (yeast or bacteria, at concentrations: 3.0 × 10⁷ and 2.5 × 10⁹ cells mL⁻¹, respectively, total volume: 100 μL) were added to a tube containing 1 mL of PE-stabilised AgNP (0.45 mg mL⁻¹), then the tube has been shaken gently for 15 min, then the loose nanoparticles were separated by centrifugation and the cells were washed with water.

Toxicity evaluation

We investigated the influence of pure PE and PE-stabilised AgNPs on the viability of the cells. Cells growth was monitored after inoculating the microplate wells with 10 μL of suspension of yeast cells or bacteria and 10 μL of nanoparticles or polyelectrolytes (at increasing concentrations), next 200 μL of nutrient broth was added. Next, the microwell plate was sealed with a breathable optically-transparent film and incubated at 37 °C. Absorbance values were recorded at 595 nm using a thermostated microplate reader (Termo Scientific Multiscan FC) for 30 hours. Viability assay was also performed using fluorescein diacetate (FDA) viability dye, which stains only viable cells and indicates the intact membranes and enzyme (esterase) activity and propidium iodide (PI), which stains only dead cells. After staining with FDA/PI, fluorescence microscopy images were obtained and the live/dead cell count was obtained.

Delivery of AgNPs into C. elegans nematodes

Synchronised adult nematodes were starved for 24 hours and then supplemented with 100 μL of food (PAH–AgNPs (1 mg mL⁻¹)-coated bacteria at 10¹⁰ cells mL⁻¹) and were allowed to feed freely for 1 hour. After feeding, the worms were collected from the dishes, immobilised with 40 mM aqueous NaN₃, and then further characterised using enhanced dark-field microscopy and hyperspectral imaging.

Imaging of nanoparticles and cells

Optical microscopy images were obtained using a Carl Zeiss Imager Z2 microscope (Germany) equipped with an AxioCam HRC CCD camera operated using ZEN software. Enhanced dark field (EDF) microscopy images, spectral libraries and hyperspectral images were obtained using a CytoViva® enhanced dark-field condenser attached to an Olympus BX51 upright microscope equipped with fluorite 100× objective and DAGE CCD camera. Extra clean dust-free Nexterion® glass slides and coverslips (Schott, Germany) were used for EDF microscopy and hyperspectral imaging to minimise dust interference. A Carl Zeiss Libra instrument was used to obtain TEM images of Ag nanoparticles. A drop of NPs was deposited onto formvar-coated copper TEM grids and left to evaporate. Atomic force microscopy (AFM) images were collected using a Dimension Icon microscope (Bruker, USA) using ScanAsyst PeakForce Tapping (in air) mode. Cells were fixated, washed with water to remove salt and debris, dehydrated and imaged in air with ScanAsyst-Air cantilevers (tip radius – 2 nm, spring constant – 0.4 N m⁻¹) cantilevers (Bruker).

Results and discussion

Stabilisation of Ag NPs with PE

Silver NPs have been synthesized using the citrate reduction method and then stabilized using PAH, PDADMAC and PEI – positively-charged synthetic polymers frequently used in fabrication of LbL films, including cell surface modifying films.¹⁹ We followed the previously reported protocol, where we coated iron oxide NPs with PAH and then successfully applied the cationic magnetic NPs for the direct functionalisation of microbial and human cells.^28–30 Here the citrate-stabilised silver NPs (AgNPs) were introduced and dispersed into the concentrated polyelectrolyte aqueous solution, incubated for 24 hours, resulting in the replacement of citrate molecules with polymers at the surfaces of NPs. After removing the unbound polymer, we investigated the changes in zeta-potential and hydrodynamic sizes of the PE-coated AgNPs. As expected (Table 1), zeta-potential has been reversed after deposition of polycations if compared with originally negative zeta-potential of citrate-coated AgNPs. Hydrodynamic diameters of PE-coated nanoparticles measured using dynamic light scattering were considerably larger than the core sizes evaluated using TEM images (Fig. 1). We attribute this effect to both aggregation of AgNPs and the hydrated polymer coating encapsulating the nanoparticles, which is consistent with the previous results obtained using magnetic iron oxide NPs.³⁰ The nanoparticles formed stable colloid suspensions in water, no aggregation was observed for at least 1 month. In addition, hydrodynamic diameters measurements performed 2 and 4 weeks after synthesis also confirm the stability of nanoparticles (data not shown).

Table 1 Size and zeta-potential distribution of PE-coated AgNPs

Nanoparticles	Zeta-potential, mV	Hydrodynamic diameter, nm	Metal core size (TEM), nm
AgNPs	−21.6 ± 0.2	71.05 ± 0.5	43.6 ± 12.7
PAH–AgNPs	79.0 ± 1.1	176.2 ± 6.9	46.5 ± 14.1
PDADMAC–AgNPs	67.6 ± 3.1	183.7 ± 0.5	66.3 ± 12.5
PEI–AgNPs	41.9 ± 1.6	127.8 ± 1.1	47.8 ± 11.1

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Fig. 1 TEM images of PE-coated AgNPs: PAH–AgNPs (a), PDADMAC–AgNPs (b) and PEI–AgNPs (c); hyperspectral images of PAH–AgNPs (d), PDADMAC–AgNPs (e) and PEI–AgNPs (f), the corresponding spectral libraries are given below each image.

TEM images (Fig. 1) reveal the overall near-spherical geometry of silver NPs characteristic to citrate-mediated synthesis, suggesting that PE-coating does not affect the metal core morphology. We further employed hyperspectral microscopy to collect hyperspectral images and corresponding spectral libraries of PE-coated AgNPs. As shown in Fig. 1, nanoparticles are clearly seen on hyperspectral images, whereas their reflected light spectral profiles differ significantly, allowing to distinguish the different types of coatings.

Spectral libraries taken from PE-coated nanoparticles will further allow to detect the NPs in complex environments (i.e. inside C. elegans nematodes).

Fabrication of AgNPs-coated “cyborg” cells

We employed the PE-coated AgNPs to fabricate “cyborg” cells – hybrid colloid microparticles consisting of microbial cell cores and nanoparticles coating attached to the cell walls.¹⁵ Typically, a range of nanoparticles is used to produce the nanoparticulate layer at the cellular surfaces. Silver nanoparticles are particularly interesting in view of their plasmonic properties, extensively utilised in surface-enhanced Raman scattering.³⁴ Previously, AgNPs have been deposited via a time-consuming LbL-based procedure.²⁴ Here we followed a direct route schematically shown in Scheme 1. This approach is based on the direct introduction of microbial cells into the excessive amount of PE-coated cationic AgNPs (at 0.45 mg mL in water), followed by the brief (15 min) incubation, after which the unattached AgNPs are removed by centrifugation/washing (5 min).

Scheme 1 The one-step direct functionalisation of microbial cells using polyelectrolyte-coated AgNPs.

We have selected yeast and E. coli bacteria as the important models for the deposition of Ag-NPs. Native (uncoated) cells exhibit negative zeta-potential in water, therefore positively-charged PE-coated AgNPs readily adhere to the cell walls of yeast and bacteria, effectively reversing the zeta-potential (Table 2). Previously, we have demonstrated that cationic PAH-coated magnetic NPs, although producing a fairly uniform monolayer on cell membranes of human cells, did not reverse the native zeta-potential.³⁰ This was attributed to the intercalation of the nanoparticles into the microvilli network on human cells. However, in case if relatively smooth surface topography of carbohydrate-coated microbial cells cationic nanoparticles deposition facilitates the change of the original negative surface charge, which is especially prominent in case of PEI-stabilised nanoparticles (41 mV and 45 mV for S. cerevisiae and E. coli, respectively).

Table 2 Zeta-potential of native and PE-stabilised AgNPs-coated cells

Cells (addlayer)	Zeta-potential, mV
Native S. cerevisiae	−21.2 ± 1.2
PAH–AgNPs@S. cerevisiae	8.4 ± 0.7
PDADMAC–AgNPs@S. cerevisiae	10.1 ± 1.1
PEI–AgNPs@S. cerevisiae	41.1 ± 2.5
Native E. coli	−25.4 ± 3.3
PAH–AgNPs@E. coli	13.8 ± 0.4
PDADMAC–AgNPs@E. coli	32.4 ± 1.2
PEI–AgNPs@E. coli	45.6 ± 0.9

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In addition, we investigated the concentration-dependent electrostatic adsorbtion of PE-stabilised AgNPs onto yeast cells by monitoring the zeta-potential changes during the automated titration with aqueous dispersions of AgNPs. We found (Fig. 2) that relatively low concentrations of PE-stabilised AgNPs are required to reverse zeta-potential of the yeast cell wall, we attribute this effect to the high affinity of the polycation-coated nanoparticles. This suggests that cations used in this study strongly adhere to the cell walls of microbial cells, which can be further employed in functionalisation of other nanomaterials (i.e. mesoporous nanoparticles, nanotubes, etc.) for the subsequent application in fabrication of “cyborg” cells. In our experiments we used higher concentrations of PE-stabilised AgNPs (0.45 mg mL⁻¹), however we suppose that for certain applications (such as directed controllable delivery of nanoparticles into target multicellular organisms) lower concentrations can be applied as well.

Fig. 2 Zeta-potential changes during the titration of S. cerevisiae yeast cells with polymer-stabilised AgNPs.

Next, we employed enhanced dark-field microscopy to visualise the polymer-stabilised nanoparticles deposited on the cell walls of yeast and bacteria (Fig. 3) in aqueous media. EDF microscopy allows imaging nanosized particles without any special sample preparation, therefore here we took advantage from its capability to demonstrate the effective deposition of AgNPs on live microbial cells in their native environments. As shown in Fig. 3, PAH–AgNPs, PDADMAC–AgNPs and PEI–AgNPs were effectively immobilised on the cell walls, as has been previously shown with LbL deposition and direct deposition of PAH-coated magnetic NPs.¹⁶ EDF microscopy images clearly demonstrate that the PAH-coated AgNPs exhibit a higher affinity towards microbial cells, whereas PEI and PDADMAC-stabilised silver nanoparticles are less adhesive, which is supported by presence of free AgNPs seen in EDF images (Fig. 3). This can be explained by electrostatic repulsion of AgNPs at higher concentrations (0.45 mg mL⁻¹), which was avoided by using lower concentrations (0.1 mg mL⁻¹) (data not shown).

Fig. 3 Representative EDF microscopy images of S. cerevisiae yeast (upper row) and E. coli bacteria coated with polymer-stabilised AgNPs.

The coating density was visibly higher in case of bacteria, which we attribute to the higher overall negative charge of bacteria (Table 2) and the spatial matching of NPs aggregated with the cells. AFM images (Fig. 4) of AgNPs-coated bacteria taken in air confirm the deposition of nanoparticles, although the apparent density of is lower, which is likely to be caused by partial removal of nanoparticles during sample preparation (dehydration and drying).

Fig. 4 AFM images of E. coli cells coated with PAH–AgNPs (a); PDADMAC–AgNPs (b) and PEI–AgNPs (c).

AFM images also demonstrate that the morphology of the AgNPs-coated cells is preserved, the nanoparticles are localised on the cell walls of the cells. The similar results were obtained for yeast cells (data not shown).

Toxicity of PE-stabilised AgNPs and PEs

Next, to determine the biocompatibility of polymer-stabilised NPs, we employed several representative viability tests to evaluate the toxicity of PE-coated AgNPs. Previously, serious concerns on the potential toxicity of the commercially available polyelectrolytes, including PAH, PDADMAC and PEI used in this study, have been raised.³ The effects of PE–AgNPs (at 0.45 mg mL⁻¹) on yeast cells were tested using FDA/PI viability dyes (Fig. 5).

Fig. 5 Viability of AgNPs-coated cyborg yeast cells (FDA/PI viability stain): (A) native cells (98% viable); (B) – PAH–AgNPs@yeast (97% viable); (C) – PDADMAC–AgNPs@yeast (94% viable); (D) – PEI–AgNPs@yeast (63% viable). Insets demonstrate budding cells.

We found that PAH and PDADMAC-coated AgNPs did not induce any significant toxic effect on yeast cells if compared with control samples. We detected budding cells, which indicate that the PE-coated AgNPs do not form solid shells preventing the cells from budding. PEI-coated AgNPs were more toxic, although still ∼63% of coated yeast was viable. Next, we tested the long-term viability of bacterial cells coated with the increasing concentrations of PE–AgNPs. Cell were grown in media supplemented with nanoparticles at normal conditions, and the cell growth curves were plotted using real-time absorbency measurements during 31 hours. The results (Fig. 6) demonstrate that the toxicity of PE-coated AgNPs is negligible, nanoparticles-treated cells grow normally if compared with the control samples.

Fig. 6 E. coli growth curves demonstrating the effects of PAH (A); PDADMAC (B) and PEI (C) on bacterial proliferation.

Further, we investigated the toxicity of pure polyelectrolytes (Fig. 7), finding out that PAH was not toxic within 0.5–2% (which is typical for cells surface treatment techniques) and even stimulated the growth rate of E. coli cells, whereas pure PDADMAC and PEI severely suppressed cells division.

Fig. 7 E. coli growth curves demonstrating the effects of PE-stabilised AgNPs on bacterial proliferation: (A) – PAH–AgNPs@E. coli; (B) – PDADMAC–AgNPs@E. coli; (C) – PEI–AgNPs@E. coli.

Similar tests were performed with AgNPs and pure PE-treated yeast with similar results (data not shown). The results obtained suggest that PE at lower concentrations (needed to effectively coat AgNPs) are not toxic towards bacteria and yeast, and can be applied for cell surface engineering of viable proliferating cells. Importantly, we did not observe any retardation of growth presumably induced by the full-scale LbL multi-layered coatings, when the grow of microbial cells is delayed for 2–3 hours required for cells to pierce the polymer layers.³⁵ Among the polymers tested here, PAH appears to be the most biocompatible one, whereas PDADMAC and PEI are more toxic, which should be taken into account in cell surface engineering applications. The overall evaluation of toxicity induced by PE-stabilised silver nanoparticles employing an enzymatic assay/membrane integrity test and the more complex growth analysis suggests that the biocompatibility is very high, even if compared with principally less toxic coatings (such as silk fibroin shells).²³

AgNPs-coated “cyborg” cells as “smart food” particles for microworms

Recently, we demonstrated that LbL-coated nanoparticle-carrying microbial cells, i.e. bacteria and microscopic algae, can be applied as “nanobaits” to deliver magnetic and silver nanoparticles²⁶ or halloysite nanotubes²⁷ into microscopic C. elegans worms. The use of nanocoated cells allows for controllable delivery of nanoparticles, when one can precisely tune the concentrations of nanoparticles taken up by bacteria-feeding worms. Potentially, this method may find applications in surface-enhanced Raman scattering characterisation techniques, etc. However, LbL deposition of nanoparticles suffers from time-consuming surface functionalisation step and lengthy washing/centrifugation cycles. Typically, the simplest cell surface architecture consisting of PAH/AgNPs/PAH/PSS requires at least 2 hours for the deposition. Obviously, the direct deposition of nanoparticles would be beneficial to make the preparation of nanobaits more rapid, in addition, in this case PE layers would not interfere and affect the interaction of nanoparticles with cells. Here we tested if PE-stabilised silver nanoparticles can be applied to prepare “nanobaits” for the delivery of AgNPs into C. elegans microworms. We used PAH-AgNPs to coat E. coli cells and then fed adult synchronised nematodes with the resulting “nanobaits” as described previously.^26,27 Next, the worms were immobilised and visualised using EDF microscopy and hyperspectral imaging (Fig. 8).

Fig. 8 Delivery of nanoparticles into C. elegans nematodes using bacteria coated with PAH-stabilised AgNPs: (a) – optical microscopy and (b) – corresponding EDF microscopy image of AgNPs distribution in the nematode; (c) – hyperspectral image of AgNPs-fed nematode (grinder area) and (d) – corresponding spectral library (indicates the boxed areas in (c)).

We found that the nanoparticles were effectively delivered into the nematodes. As shown in Fig. 8a and b, PAH–AgNPs are evenly distributed inside the intestines of the microworms, indicating that the NPs were delivered during the ingestion of nanoparticle-coated bacteria. Even on bright-field optical microscopy images dense brown spots representing the sites there AgNPs are concentrated can be clearly seen. Moreover, EDF microscopy images (Fig. 8b) show the aggregated AgNPs inside the nematodes along the whole intestine, starting from buccal cavity to the anus, with prominent aggregations in interior bulb and terminal bulb. More clearly the distribution of PAH-stabilised AgNPs in a whole fixated animal is shown in Video 1 (ESI†), where the focal plane is being changed at several regions to demonstrate the spatial distribution of the nanoparticles. We were curious to see if the nanoparticles are firmly attached to the nematode's guts or persist as free-standing nanoparticles. To check this we imaged the living anesthetized animals using EDF microscopy and collected real time footages (Video 2, ESI†), which unquestionably show that most of the nanoparticles have been adsorbed by intestinal cells, whereas several nanoparticles are actively moving inside the lumen. This suggests that the PE-mediated attachment of nanoparticles onto microbial cells is reversible and once the carrier cells are grinded and digested the released NPs can be distributed freely in the target organism. We also applied hyperspectral imaging combined with spectral mapping (Fig. 8c and d), to demonstrate that PAH–AgNPs can be visualized inside the bodies of nematodes and detected using hyperspectral mapping. Potentially, biocompatible AgNPs having sizes around 50 nm delivered inside nematodes might be a powerful tool in studying biochemical reactions in worms using SERS.³⁴

Conclusions

Here we demonstrated for the first time a straightforward technique to fabricate cationic polyelectrolyte-stabilised ∼50 nm silver nanoparticles. These nanoparticles rapidly adhere to yeast and bacteria cell walls allowing reducing the deposition time six fold. Atomic force microscopy and enhanced dark-field microscopy confirm the formation of an even nanoparticulate layer, clearly detectable and stable. The viability of the microorganisms was preserved at very high level (93–97%), suggesting that the PE-stabilised nanoparticles are non-toxic within a range of concentrations. We employed PAH-stabilised AgNPs to fabricate “nanobait” bacterial cells for the directed delivery of nanoparticles into C. elegans nematodes. The technique described here can also be extended using other metal nanoparticles (i.e. gold NPs). We believe that the direct approach reported here will be appreciated in a number of practical applications, including such as fabrication of multicellular clusters,³⁶ and could be further extended to produce polyelectrolyte-coated nanotubes,³⁷ thus helping to extend the rapidly growing field of cell surface engineering.³⁸ Future work will be focused on application of PE-stabilised AgNPs for fabrication of human “cyborg” cells.

Acknowledgements

We thank Dr A. Noskov and Dr E. Nuzhdin for help with TEM imaging of AgNPs. This study was supported by Russian Scientific Fund grant no. 14-14-00924.

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Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c4ra15857a

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