Rising to the surface: capturing and detecting bacteria by rationally-designed surfaces.

Analytical microbiology has made substantial progress since its conception, starting from potato slices, through selective agar media, to engineered surfaces modified with capture probes. While the latter represents the dominant approach in designing sensors for bacteria detection, the importance of sensor surface properties is frequently ignored. Herein, we highlight their significant role in the complex process of bacterial transition from planktonic to sessile, representing the first and critical step in bacteria detection. We present the main surface features and discuss their effect on the bio-solid interface and the resulting sensing capabilities for both flat and particulate systems. The concepts of rationally-designed surfaces for enhanced bacterial detection are presented with recent examples of sensors (capture probe-free) relying solely on surface cues.

Gold Nanorod-Incorporated Halloysite Nanotubes Functionalized with Antibody for Superior Antibacterial Photothermal Treatment.

The global spread of antibiotic-resistant strains, and the need to protect the microflora from non-specific antibiotics require more effective and selective alternatives. In this work, we demonstrate for the first time a superior antibacterial photothermal effect of plasmonic gold nanorods (AuNRs) via their incorporation onto natural clay halloysite nanotubes (HNTs), which were functionalized with anti-E. coli antibodies (Ab-HNTs). AuNRs were incorporated onto the Ab-HNTs through a facile freeze-thaw cycle, and antibody integrity following the incorporation was confirmed via infrared spectroscopy and fluorescence immunolabeling. The incorporation efficiency was studied using UV-Vis absorption and transmission electron microscopy (TEM). Mixtures of E. coli and AuNR-Ab-HNTs hybrids or free AuNRs were irradiated with an 808 nm laser at 3-4 W cm-2, and the resulting photothermal antibacterial activity was measured via plate count. The irradiated AuNR-Ab-HNTs hybrids exerted an 8-fold higher antibacterial effect compared to free AuNR under 3.5 W cm-2; whereas the latter induced a 6 °C-higher temperature elevation. No significant antibacterial activity was observed for the AuNR-Ab-HNTs hybrid against non-target bacteria species (Serratia marcescens and Staphylococcus epidermidis). These findings are ascribed to the localization of the photothermal ablation due to the binding of the antibody-functionalized clay to its target bacteria, as supported through TEM imaging. In the future, the HNTs-based selective carriers presented herein could be tailored with other antibacterial nanoparticles or against another microorganism via the facile adjustment of the immobilized antibody.

Antibody-Functionalized Halloysite Nanotubes for Targeting Bacterial Cells.

Halloysite nanotubes (HNTs) are naturally occurring tubular clay particles which have emerged in recent years as a promising nanomaterial for numerous applications. Specifically, HNTs' large pore volume and high specific surface area in combination with their biocompatibility make them ideal nanocarriers for bioactive compounds. This research aims to design and synthesize functionalized HNTs, which could selectively bind to target bacterial cells in suspension. Such a system can allow us to treat target cells within a challenging heterogeneous population, such as contaminated ecosystems or gut flora. HNTs functionalization is achieved by immobilizing specific antibodies onto the nanotube surface. The synthetic route is realized by the following subsequent steps: acidic etching of the HNTs, silanization of reactive surface hydroxyls, conjugation of protein A, and oriented immobilization of the antibody. HNT functionalization is studied by a set of analytical techniques including attenuated total reflectance Fourier-transform infrared spectroscopy, zeta potential measurements, thermal gravimetric analysis, scanning and transmission electron microscopy, as well as fluorescence microscopy. The selective binding of the functionalized HNTs to their target bacteria is observed upon incubation with live homogenous and heterogeneous cultures using fluorescence microscopy and high-throughput flow cytometry. Plate count and live/dead staining experiments demonstrate the biocompatibility of the antibody-HNT hybrid with its target bacteria. The suggested HNT-based smart carrier constitutes a generic platform for targeted delivery that could be selectively tailored against any microorganism by facile antibody adjustment.

Halloysite nanotubes - the nano-bio interface.

The numerous biological applications of nanoparticles in general and nano-clays in particular are rooted in understanding and harnessing their dynamic nano-bio interface. Among clays, the intrinsically-mesoporous halloysite nanotubes (HNTs) have emerged in recent years as promising nanomaterials. The diverse interactions of these nanotubes with living cells, encompassing electrostatic, van der Waals, and ion exchange, along with cellular response, are crucial in determining the behaviour of HNTs in biological systems. Thus, rational engineering of the nanotube properties allows for vast applications ranging from bacteria encapsulation for bioremediation, through algae flocculation for aquaculture, to intracellular drug delivery. This review summarizes the many aspects of the nano-bio interface of HNTs with different cell types (bacteria, algae and fungi, and mammalian cells), highlighting biocompatibility/bio-adverse properties, interaction mechanisms, and the latest cutting-edge technologies.

Antimicrobial Carvacrol-Containing Polypropylene Films: Composition, Structure and Function.

Significant research has been directed toward the incorporation of bioactive plant extracts or essential oils (EOs) into polymers to endow the latter with antimicrobial functionality. EOs offer a unique combination of having broad antimicrobial activity from a natural source, generally recognized as safe (GRAS) recognition in the US, and a volatile nature. However, their volatility also presents a major challenge in their incorporation into polymers by conventional high-temperature-processing techniques. Herein, antimicrobial polypropylene (PP) cast films were produced by incorporating carvacrol (a model EO) or carvacrol, loaded into halloysite nanotubes (HNTs), via melt compounding. We studied the composition-structure-property relationships in these systems, focusing on the effect of carvacrol on the composition of the films, the PP crystalline phase and its morphology and the films' mechanical and antimicrobial properties. For the first time, molecular dynamics simulations were applied to reveal the complex interactions between the components of these carvacrol-containing systems. We show that strong molecular interactions between PP and carvacrol minimize the loss of this highly-volatile EO during high-temperature polymer processing, enabling semi-industrial scale production. The resulting films exhibit outstanding antimicrobial properties against model microorganisms (Escherichia coli and Alternaria alternata). The PP/(HNTs-carvacrol) nanocomposite films, containing the carvacrol-loaded HNTs, display a higher level of crystalline order, superior mechanical properties and prolonged release of carvacrol, in comparison to PP/carvacrol blends. These properties are ascribed to the role of HNTs in these nanocomposites and their effect on the PP matrix and retained carvacrol content.

Antimicrobial LDPE/EVOH Layered Films Containing Carvacrol Fabricated by Multiplication Extrusion.

This work describes the fabrication of antimicrobial multilayered polymeric films containing carvacrol (used as a model essential oil) by co-extrusion and multiplication technique. The microlayering process was utilized to produce films, with up to 65 alternating layers, of carvacrol-containing low-density polyethylene (LDPE) and ethylene vinyl alcohol copolymer (EVOH). Carvacrol was melt compounded with LDPE or loaded into halloysite nanotubes (HNTs) in a pre-compounding step prior film production. The detailed nanostructure and composition (in terms of carvacrol content) of the films were characterized and correlated to their barrier properties, carvacrol release rate, and antibacterial and antifungal activity. The resulting films exhibit high carvacrol content despite the harsh processing conditions (temperature of 200 °C and long processing time), regardless of the number of layers or the presence of HNTs. The multilayered films exhibit superior oxygen transmission rates and carvacrol diffusivity values that are more than two orders of magnitude lower in comparison to single-layered carvacrol-containing films (i.e., LDPE/carvacrol and LDPE/(HNTs/carvacrol)) produced by conventional cast extrusion. The (LDPE/carvacrol)/EVOH and (LDPE/[HNTs/carvacrol])/EVOH films demonstrated excellent antimicrobial efficacy against E. coli and Alternaria alternata in in vitro micro-atmosphere assays and against A. alternata and Rhizopus in cherry tomatoes, used as the food model. The results presented here suggest that sensitive essential oils, such as carvacrol, can be incorporated into plastic polymers constructed of tailored multiple layers, without losing their antimicrobial efficacy.

The effect of sugar stereochemistry on protein self-assembly: the case of ß-casein micellization in different aldohexose solutions.

Protein self-assembly applications, such as nanoencapsulation of drugs and nutraceuticals, require deep understanding of the parameters governing the micellization process, including the effects of ionic and non-ionic co-solutes, like salts and sugars respectively, which is often overlooked. Herein, with the aim of shedding light on the effect of nonionic cosolute stereochemistry on protein self-assembly, we studied the ternary system of water-protein-sugar by examining the concentration-dependent effects of three aldohexoses, d-glucose (Glu), d-galactose (Gal) and d-mannose (Man) and that of urea, on the micellization of beta casein (ß-Cas), using pyrene as a fluorescent probe for the formation of hydrophobic domains. Pyrene's excitation spectra were recorded for several sets of samples with rising protein concentration (0-5 mg ml(-1)), each set with a different co-solute type and concentration. Critical micellization concentration (CMC) and cooperativity of micellization were evaluated according to changes in pyrene spectra as it partitioned from the aqueous environment to the hydrophobic cores of ß-Cas micelles. All sugars examined lowered the CMC of ß-Cas with increasing sugar concentration and with a diminishing degree of effectiveness (Glu > Gal > Man) which correlated well with the sugars' dynamic hydration number, defined by Uedaira, and correlated negatively with their hydrophobic to hydrophilic molecular surface ratio. These results support the hypothesis that sugars affect protein self-assembly through both changes in water structure and by hydrophobic interactions, both of which are evidenced to be highly sensitive to sugar stereochemistry.