Thermo-responsive 3D nanostructures for enhanced performance in food-poisoning bacterial analysis

Abstract

The growing risk of bacterial food poisoning due to global warming has necessitated the development of methods for accurate detection of food-poisoning bacteria. Despite extensive efforts to develop enhanced bacterial-capture methods, challenges associated with the release of the captured bacteria have limited the sensitivity of bacterial detection. In this study, thermo-responsive intelligent 3D nanostructures to improve food-poisoning bacterial analysis performance were fabricated by introducing a thermo-responsive polymer onto an urchin-like 3D nanopillar substrate (URCHANO). A co-polymer of methacryloyl glycinamide and benzyl acrylate (MNAGA-Bn 5%) was introduced as a thermo-responsive co-polymer onto URCHANO using an electron-transfer atom-transfer radical-polymerization method to fabricate Thermo-URCHANO. A temperature-related analysis of the surface properties of Thermo-URCHANO revealed a hydrophobic-to-hydrophilic transition at 37 °C, which facilitated the release of bacteria captured within the nanostructure. In a one-pot analysis to capture and analyze various food-poisoning bacteria in kitchenware (gloves and aprons) and food items (eggs and sausages), mimicking real-life environments, specimens collected using Thermo-URCHANO showed lower C_t values than those collected with uncoated URCHANO, indicating greater bacterial detection. This method could effectively release captured bacteria through temperature changes, improving extraction efficiency during swab collection. While Thermo-URCHANO needs further optimization, it is expected to enhance bacterial analysis performance and sensitivity.

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New concepts

In this manuscript, the concept of thermo-responsive 3D nanostructures is demonstrated to enhance the performance of food-poisoning bacteria detection. The urchin-like 3D nanostructure (URCHANO) was designed to improve bacterial capture ability, while a thermo-responsive co-polymer was utilized to enhance bacterial release properties by altering surface hydrophilicity. Although various types of stimuli responsive polymers have been studied, the majority of the currently reported applications focus on anti-fouling, drug delivery, and cell harvesting, rather than on enhancing the sensitivity of bacterial detection. In this study, enhanced bacterial detection performance was achieved by introducing a thermal responsive polymer coating onto the URCHANO, resulting in Thermo-URCHANO. Thermo-URCHANO can form a hydration layer at 37 °C due to increased surface hydrophilicity from thermo-responsiveness. The formation of this hydration layer effectively releases up to 95% of the bacteria captured in URCHANO, enabling a single-step process that improves bacterial detection accuracy by over 13% compared to existing methods, without the need for additional reagents. This approach holds significant potential for high-performance and sensitive bacterial analysis.

Introduction

Pathogenic bacterial infections cause millions of deaths annually and threaten human health, life, and society, leading to significant economic losses.¹ Food poisoning bacteria are particularly prevalent in daily life, and reportedly one out of ten people experience illness caused by food contamination each year, of which 40% are children [WHO report 2024].¹ The growing global population and rising food consumption have necessitated the development of large-scale production techniques and extensive supply chains, which are more susceptible to bacterial contamination and food spoilage due to global warming.² Therefore, early detection of pathogenic bacteria has gained substantial significance.

To date, various technologies, such as culturing microorganisms, protein-based detection methods (e.g., enzyme-linked immunosorbent assays, rapid kit assays, and mass spectrometry),^3–7 and nucleic acid-based detection methods (e.g., polymerase chain reaction [PCR], quantitative PCR [qPCR], and DNA microarray analyses),^8–11 have been used to detect food-poisoning bacteria, and each of these methods has been evolving to address its limitations. Before incorporating these detection methods, sample collection is essential to determine the presence of food poisoning bacteria. According to the U.S. Food and Drug Administration (FDA)'s Bacteriological Analytical Manual (BAM), a pretreatment step, which includes sampling, homogenization, dilution, selective enrichment, and isolation, is essential before using the bacterial detection methods. Although devices such as stomachers, pulsifiers, blenders, cotton swabs, sterile gloves, and gauze pads are commonly used during sampling procedures, these devices are inefficient for desorption of the collected samples, which adversely affects the accuracy of the analysis results. The resultant sampling errors may yield false-negative results in samples with trace amounts of harmful bacteria, with the test results incorrectly indicating the absence of bacteria when they are present. Such results can cause significant issues in hygiene and food safety, e.g., by allowing distribution of contaminated food.

Recently, we reported on three-dimensionally (3D) structured polyaniline substrates that exhibit enhanced performance in capturing bacteria.^12–16 Bacterial adhesion can be controlled by surface properties such as charge, hydrophilicity, and surface topology;¹⁷ thus, the surface pattern of the nanopillar array in these substrates can influence bacterial adhesion.^18–20 Several reports have confirmed that these substrates can efficiently capture Escherichia coli O157:H7, Staphylococcus aureus, and Salmonella entritidis from kitchenware and foods;¹³ multidrug-resistant bacteria from infected skin via hand contact;¹⁶ and airborne bacteria from air.²¹ Despite these advantages, the structure-swabbed samples were not fully detached, necessitating the use of an additional lysis buffer to detect bacterial strains.

To overcome such challenges, an alternative can include the use of an intelligent 3D nanostructured substrate that is created by introducing an stimuli-responsive polymer into this substrate for high-performance analysis of food poisoning bacteria. Stimuli-responsive polymers change their physical/chemical properties when exposed to specific environmental conditions such as temperature, pH, light, ion strength, and other stimuli.^22–29 For example, phenyl borate is a pH-responsive material that forms borate ester bonds with vicinal diols at an alkaline pH and degrades these bonds when exposed to an acidic environment.²⁵ Poly(2-dimethylaminoethyl methacrylate) is also a pH-responsive polymer that becomes protonated when exposed below its pK_a value, forming a hydration layer and aggregating in alkaline environments.^27,29

Thermally responsive condition can be more easily controlled externally through temperature regulation than other factors. There are two representative types of thermal-responsive polymers that react to surrounding temperature, namely lower critical solution temperature (LCST)-type and upper critical solution temperature (UCST)-type polymers.³⁰ Polymers of the LCST type are predominantly hydrophilic below the LCST because of hydrogen bonds between the polymer chains and water. However, above the LCST, these hydrogen bonds weaken, and intermolecular hydrophobic interactions become the dominant force, rendering the polymer hydrophobic.³¹ Because of these properties, LCST polymers, such as poly(N-isopropylacrylamide) (pNiPAM), can be utilized in cell culture plates to regulate cell detachment and attachment through temperature regulation.^31,32 The LCST of pNiPAM is 32 °C. Consequently, when the cells are cultured on a pNiPAM-coated plate at 37 °C, they adhere properly to the plate and grow. To harvest the cells, the temperature is lowered to 32 °C, causing the intramolecular hydrophobic interactions of pNiPAM to break, extend, and hydrate, thereby repelling and detaching the adhered cells. This allows the separation of cultured cells from the plate solely through temperature regulation without the use of enzymes. Conversely, UCST polymers form hydrophobic and insoluble aggregates at temperatures below the UCST and become hydrophilic when the temperature rises above the UCST.³⁰ Poly(N-acryloyl glycinamide) (pNAGA) is a representative UCST-type polymer. N-Acryloyl glycinamide (NAGA) contains two amide groups that act as H-bond donors and acceptors. Below the UCST, pNAGA forms stable intermolecular interactions through strong H-bonding, leading to aggregation in pure water. Above the UCST, these intramolecular hydrogen bonds are weakened, resulting in hydration and clear dissolution.³³

These thermo-responsive polymers can also be synthesized and applied in various ways to suit the intended purpose since the critical solution temperature can be adjusted by co-polymerization with specific monomers.^30,34,35 However, the majority of the currently reported applications focus on anti-fouling, drug delivery, thermal therapeutics, and cell harvesting, and not on enhancing the sensitivity of bacterial detection.^{23–25,27,32,34,36–38}

In this study, enhanced bacterial detection performance was achieved by introducing a poly(methacryloyl glycinamide) (pMNAGA)-based thermo-responsive co-polymer with UCST behavior onto an urchin-like 3D nanopillar substrate (URCHANO) to capture bacteria. Methacryloyl glycinamide (MNAGA), which consists of an additional methyl group on the backbone of NAGA, disrupts strong intermolecular hydrogen bonding, thereby enhancing its flexibility and fast-recovery properties.³⁹ These flexible and hydrophilic polymer chains exhibit anti-fouling properties owing to the formation of a hydration layer and their repelling mobilities. First, the MNAGA monomer was synthesized and coated onto URCHANO via a surface-initiated activator generated by electron-transfer atom-transfer radical-polymerization (SI-ARGET-ATRP) method to fabricate thermo-responsive URCHANO (Thermo-URCHANO). The ARGET-ATRP method used in this study reduces the amount of catalyst and has high tolerance to oxygen, allowing polymerization to be conducted under open-air conditions in aqueous solutions, making it an environmentally friendly method with high accessibility.^40–42 Using Thermo-URCHANO, food-poisoning bacteria were captured from food or kitchenware, and the performance of Thermo-URCHANO in relation to the thermal response was analysed (Fig. 1).

Fig. 1 Schematic illustration of the overall process of analysing food-poisoning bacteria using a thermo-responsive copolymer coating on an urchin-like 3D nanostructure substrate (Thermo-URCHANO).

Results and discussion

To fabricate Thermo-URCHANO for enhanced performance in food-poisoning bacterial analysis, MNGA monomers as thermo-responsive copolymer precursors were first synthesized using a one-pot amidation reaction. Glycinamide was deprotonated using K₂CO₃, and methacryloyl chloride was gradually added dropwise to form an amide bond. The synthesized MNAGA was purified by recrystallization and confirmed by evaluating the ¹H-nuclear magnetic resonance (NMR) spectra (Fig. S3, ESI†). Subsequently, the MNAGA monomer and different hydrophobic acrylate monomers were co-polymerized onto URCHANO using the SI-ARGET-ATRP method to generate Thermo-URCHANO.

In previous research, bacterial fibrils such as pili, curli, and fimbriae have been shown attach to the nanopillar structure of this URCHNANO or become trapped between spacing areas, enabling the capture of various types of bacteria.^12,13 In this study, by introducing thermo-responsiveness to this substrate, we developed a structure that facilitated bacterial capture and promoted bacterial release in response to temperature changes, thereby simplifying the analytical process. Thermo-URCHANO was generated by three-step procedure. Briefly, URCHANO was functionalized with an amine, and the polymerization initiator 2-bromoisobutryl bromide (BiBB) was immobilized onto the amine-functionalized URCHANO using vacuum-phase deposition. Subsequently, the MNAGA and acrylate monomers were co-polymerized onto URCHANO, yielding a thermo-responsive co-polymer coating on URCHANO (Thermo-URCHANO) (Fig. 2).

Fig. 2 The synthesis of the thermo-responsive pMNAGA-Bn co-polymer 5% and its coating on URCHANO for fabrication of Thermo-URCHANO.

In a previous study, Xue et al.³⁴ reported that the UCST of pNAGA increased from 24 °C to 28 °C and 34 °C as the percentage of the hydrophobic monomer phenylacrylamide increased. Thus, the addition of a hydrophobic moiety can increase the UCST of thermo-responsive co-polymers. Therefore, three types of hydrophobic acrylate monomers were used in this study: ethyl (Et), butyl (Bu), and benzyl (Bn) acrylates. The different MNAGA co-polymer-coated substrates were designated as MNAGA-XX y%, where XX and y represented the acrylated monomer and feed molar ratio, respectively. The fabricated Thermo-URCHANO was characterized by measurement of the static water contact angle and X-ray photoelectron spectroscopy (XPS) spectra. Although polyaniline (PANI) exhibits a hydrophobic nature, its nanostructured topology can make it appear hydrophilic, as indicated by the water contact angle results.^13,43–45 Therefore, to clearly confirm the presence of the thermo-responsive co-polymer coating and the thermo-responsiveness of MNAGA-XX y%, this polymer was first introduced onto the Au substrate for analysis before introduction of PANI.

The water contact angle of the Au substrate was 110.9° ± 5.3°, and it decreased to 16.9° ± 2.4° after amine functionalization. Subsequently, it increased to 127.0° ± 1.8° after BiBB initiator immobilization. After coated solely with MNAGA, the water contact angle at 25 °C was 12.0° ± 0.2° (Fig. S4 and S5 and Table S3, ESI†). Upon co-polymerization with hydrophobic Et or Bu acrylate monomers, the water contact angle increased with increasing feed molar ratios (Fig. S4 and S5 and Table S3, ESI†). For MNAGA-Bn, the water contact angle decreased at a feed ratio of 20% at 25 °C. However, the water contact angle of MNAGA-Bu 20% was measured asymmetrically. These results could be attributed to the presence of a bulky hydrophobic portion, which led to an uneven coating on the surfaces (Fig. S6, ESI†). Next, the thermo-responsiveness of MNAGA-XX y% coatings on Au substrates under various temperature conditions (25 °C, 37 °C, and 45 °C) were investigated. A heating block was placed on the sample stage of the static water contact angle goniometer, and MNAGA-XX y%-coated Au substrates were placed on it. The water contact angles of the MNAGA-Bn 5%- and MNAGA-Bu 5%-coated substrates decreased significantly at 37 °C, while those of the MNAGA-Et 5%-, MNAGA-Et 10%-, and MNAGA-Bu 10%-coated substrates decreased at 45 °C, with no significant changes observed with the other coatings. These phenomena were attributed to an increase in hydrophobic components, which affected the UCST of the thermo-responsive polymers, consistent with previous reports.³⁴ Among these substrates, the MNAGA-Bn 5%-coated Au substrate exhibited the most distinctive change at 37 °C. Therefore, with the aim of facilitating the capture and release of bacteria for effective analysis of food-poisoning bacteria, subsequent experiments were conducted using MNAGA-Bn 5% by coating it onto URCHANO using the same method used for the Au substrate. The water contact angle of uncoated and amine-functionalized URCHANO was 31.2° ± 2.2° and 28.8° ± 2.7° respectively, and it increased to 115.1° ± 1.2° after initiator immobilization. The water contact angle of the MNAGA-Bn 5%-coated URCHANO at 25 °C was 19.2° ± 0.1° (Fig. 3).

Fig. 3 Static water contact angles of uncoated URCHANO, amine-functionalized URCHANO, BiBB initiator-immobilized URCHANO, and Thermo-URCHANO at 25 °C and 37 °C. Each value represents the mean of nine measurements from three replicates, and the standard deviation (±) values represent the 95% confidence limit.

XPS spectra were also analyzed at every surface-modification step (Fig. S7 and Table S4, ESI†). The Si 2p peak was observed after amine functionalization using 3-aminopropyltriethoxysilane (APTES), and the Br 3d peak was observed after BiBB initiator immobilization, which originated from the respective representative atoms. After MNAGA-Bn 5% coating, both Br 3d and Si 2p peaks decreased, whereas the N 1s peak increased owing to the amine-rich MNAGA components. In addition, the high-resolution XPS spectra of the C 1s peak showed 287.8-eV and 291-eV peaks, which corresponded to N–C=O and pi–pi stacking of the benzene ring, respectively, that originated from the MNAGA-Bn 5% co-polymer. The Fourier-transform infrared (FT-IR) spectra were also analyzed to confirm the MNAGA-Bn 5% coating on URCHANO (Fig. 4). Peaks at 1146 cm⁻¹ (C–H aromatic stretching and –NH⁺= stretching of aniline), 1240 cm⁻¹ (C–O stretching of benzenoid), 1297 cm⁻¹ (C–N stretching of secondary aromatic amine), and 1487 cm⁻¹ (C=C stretching of benzenoid) were observed in all substrates, which were attributed to the PANI backbone.^12,13 However, the C=C stretching peak of quinoid shifted slightly from 1560 cm⁻¹ to 1587 cm⁻¹ as the surface-modification step proceeded, indicating that the surface chemical structure was different. After initiator immobilization, a new peak at 1718 cm⁻¹ was observed, which originated from the carbonyl group of BiBB, and an amide peak at 1681 cm⁻¹ appeared after amine-rich MNAGA-Bn 5% coating.

Fig. 4 Fourier transform-infrared spectra of uncoated URCHANO (black), amine-functionalized URCHANO (red), BiBB initiator immobilized-URCHANO (blue), and MNAGA-Bn 5%-coated URCHANO (Thermo-URCHANO).

When the morphologies of URCHANO before and after coating with the thermo-responsive co-polymer MNAGA-Bn 5% were analyzed (Fig. 5), the urchin shape of the coated URCHANO appeared relatively smoother than that of uncoated URCHANO (Fig. 5a and d). This was because MNAGA-Bn 5% was coated onto URCHANO. Specifically, the cross-sectional images confirmed that MNAGA-Bn 5% formed a layer approximately 50 nm thick on URCHANO (Fig. 5e, f and Fig. S8, ESI†). In previous study, a polymer coating layer exceeding 70 nm was shown to alter the morphology of URCHANO,¹⁵ potentially compromising its bacterial capture ability. Therefore, a 50 nm thickness of MNAGA-Bn5% coating is considered appreciate for enhancing both bacterial capture and release properties. These results indicate that the MNAGA-Bn 5% coating was successfully applied to URCHANO, yielding Thermo-URCHANO.

Fig. 5 Scanning electron microscopy (SEM) images of uncoated URCHANO (above) and Thermo-URCHANO (below). (A) and (D) Top-view SEM images and cross-sectional images of (B) and (C) uncoated URCHANO and (E) and (F) MNAGA-Bn 5% coated-URCHANO (Thermo-URCHANO) (scale bar: 1 μm). The yellow arrows indicate the MNAGA-Bn 5% coating layer.

The thermo-responsiveness of Thermo-URCHANO (MNAGA-Bn 5%-coated URCHANO) at 25 °C and 37 °C was investigated using water contact angle measurements and the captive bubble method (Fig. 3 and 6). The water contact angle of Thermo-URCHANO decreased from 19.2° ± 0.1° at 25 °C to 14.0° ± 1.8° at 37 °C (Fig. 3). Although the water contact angle changed, the degree of change was too small to confirm thermo-responsiveness; therefore, additional measurements were conducted using the captive bubble method. Thermo-URCHANO was fixed upside down on the sample stage, immersed in a chamber containing deionized water, and the air bubbles on it were removed. An empty syringe with a curved needle was used to generate air bubbles under the water. The air bubble was then raised and captured on Thermo-URCHANO, and the contact angles were measured immediately. In contrast to the water contact angle measurements, a high contact angle of a captive bubble indicates a hydrophilic state, whereas a low contact angle indicates a hydrophobic state. The captive bubble contact angle of Thermo-URCHANO increased from 129.9° ± 0.8° to 139.0° ± 3.1° after heating from 25 °C to 37 °C, indicating that its surface properties had changed to become more hydrophilic due to its thermo-responsiveness (Fig. 6).

Fig. 6 Captive bubble contact angle captured by Thermo-URCHANO at (A) 25 °C and (B) 37 °C. Each value is the mean of nine measurements from three replicates, and the standard deviation (±) value represents the 95% confidence limit.

The conformational changes due to the thermo-responsiveness were analyzed using AFM measurements with tapping mode. The uncoated URCHANO and Thermo-URCHANO were submerged in deionized water with varied temperature (Fig. S9, ESI†). As shown in Fig. S9 (ESI†), topological changes were observed in Thermo-URCHANO as the temperature increased from R.T to 37 °C, while no significant difference was observed in uncoated-URCAHNO. In detail, the height of uncoated URCHANO at both R.T and 37 °C was measured as 450 nm, while Thermo-URHCANO at R.T was 600 nm and increased to 750 nm at 37 °C (Fig. S9b, ESI†). In addition, the RMS roughness of uncoated URCHANO was slightly changed from 79.7 nm to 94 nm as the temperature increased, while Thermo-URCAHNO was increased from 123 nm to 165 nm (Fig. S9a, ESI†). This observation indicates that the increase in temperature disturbed intermolecular hydrogen bonding, hydrated and extended that the Thermo-responsive coating has brush type conformation. On the basis of these unique properties, Thermo-URCHANO was used in a one-pot analysis to capture and detect food-poisoning bacteria. The common food poisoning bacterium S. aureus was used for testing (Fig. 7 and Table S5, ESI†). S. aureus suspensions were prepared at concentrations ranging from 10² to 10⁵ colony-forming units CFU mL⁻¹. After dropping this suspension onto Thermo-URCHANO, the bacteria released with and without thermal treatment were analyzed by real-time PCR (RT-PCR), and uncoated URCHANO was used as the control. In the RT-PCR analysis, an increase in the C_t value indicated a decrease in the concentration of the target bacteria. As shown in Fig. 7, over the entire range of bacterial concentrations, the C_t value after thermal treatment of Thermo-URCHANO at 37 °C was similar to the C_t value of the bacteria initially dropped onto the Thermo-URCHANO. However, without thermal treatment, the C_t value was at a barely identifiable level at 25 °C. Thus, while bacteria were adequately captured in the urchin-shaped structure of Thermo-URCHANO at 25 °C, as the temperature increased, the release of bacteria was accelerated owing to changes in the properties of the thermo-responsive co-polymer on the surface of the substrate. Compared to Thermo-URCHANO treated at 25 °C, the number of remaining bacteria were considerably reduced after thermal-treatment (Fig. S11, ESI†). This may explain why the C_t value was confirmed to similarly regardless of thermal treatment of uncoated URCHANO, which was not coated with MNAGA-Bn 5% (Fig. 7 and Fig. S4, ESI†). This suggests that when the temperature increased above the transition temperature of the MNAGA-Bn 5% coating, the intermolecular hydrogen bonding of the amide moieties broke and became hydrated, similar to the UCST type polymers. Consequently, the immobilized MNAGA-Bn 5% co-polymer exhibited brush-like mobility, and the hydrated layer repelled the attached bacteria. Thus, the release of the captured bacteria was accelerated, and the released bacteria were detected at levels similar to those of the originally treated bacterial concentration, confirming the recovery of most of the captured bacteria. In contrast, the uncoated URCHANO did not exhibit any release effects with temperature changes. The reversibility of the thermo-responsiveness of Thermo-URCHANO was also confirmed. Static water contact angles were measured over five repetitive cooling and heating cycles at 25 °C and 37 °C, respectively (Fig. S12a, ESI†). The thermo-responsiveness of Thermo-URCHANO was maintained over at least five cycles of repetitive thermal treatment. In addition, the major peaks remained in the FT-IR spectra, and no significant changes were observed after five thermal cycles (Fig. S12b, ESI†).

Fig. 7 RT-PCR analysis of S. aureus captured by Thermo-URCHANO and uncoated URCHANO and released with and without thermal treatment at different concentrations of S. aureus cells (10²–10⁵ CFU mL⁻¹). Each value is the mean of 18 measurements from six replicates. The error bars indicate the standard deviations (n.s.: not significant; **P < 0.01, ***P < 0.001, gray: origin, red: Thermo-URCHANO at 37 °C, pink: Thermo-URCHANO at 25 °C, blue: uncoated URCHANO at 37 °C, and sky blue: uncoated URCHANO at 25 °C.).

Thus, the properties of bacterial release were not due to the polishing or degradation of the polymer layers, but were rather caused by the thermo-responsiveness of the MNAGA-Bn 5% coating. Using this confirmed Thermo-URCHANO, we performed a one-pot analysis of the enhanced performance for capturing and analyzing various species of food-poisoning bacteria found in various environments such as kitchenware and foods (Fig. 8a). Gloves and aprons were chosen as representative kitchenware items, and eggs and sausages were chosen as representative food items. Common bacteria that cause food poisoning in daily life were used in the study, which included Staphylococcus aureus, Salmonella entritidis, Listeria monocytogenes, and Bacillus cereus. The bacteria mixed with different compositions were applied to kitchenware and food surfaces to mimic real-life environments (Fig. 8b). Specimens were collected from each item using Thermo-URCHANO swabs and then analyzed after thermal treatment. As a result, in all samples, when Thermo-URCHANO was used, a lower C_t value, i.e., a larger number of bacteria, was detected in comparison with that detected with uncoated URCHANO (Fig. 8c). Although slight differences in efficiency were observed depending on the bacterial strain, when Thermo-URCHANO was applied, the captured bacteria were effectively released owing to the temperature changes and efficiently analyzed. Thus, the use of this material is expected to solve the problems caused by low extraction efficiency when collecting swabs, even in the presence of food-poisoning bacteria.

Fig. 8 Bacterial capture and release from actual samples. (A) Bacterial capture from (i) gloves, (ii) aprons, (iii) eggs, and (iv) sausages. (B) Composition of the bacterial mixtures used in this study (+: positive and −: negative). (C) RT-PCR analysis results of bacteria captured by Thermo-URCHANO (red) in (i) glove, (ii) apron, (iii) egg, and (iv) sausage. Uncoated URCHANO (blue) was used as the control. Each value is the mean of nine measurements from three replicates. The error bars show the standard deviation values (green area: detected and yellow area: not detected).

Conclusions

Thermo-URCHANO, which contained thermo-responsive intelligent 3D nanostructures, was fabricated to enhance the performance of food-poisoning bacterial analysis, enabling one-pot analysis for efficient capture and detection. To achieve these characteristics and performance, a thermo-responsive co-polymer based on MNAGA was introduced onto URCHANO, and its properties were analyzed. The findings confirmed the optimal thermo-responsiveness of URCHANO coated with MNAGA-Bn 5%. MNAGA-Bn 5%-coated URCHANO, which we named Thermo-URCHANO, could form a hydration layer by increasing the hydrophilicity of the surface at 37 °C in comparison with that at 25 °C due to changes in thermo-responsiveness. The formation of the hydration layer effectively released up to 95% of the bacteria captured in the URCHANO, enhancing bacterial detection accuracy by over 13% in comparison with existing methods in a one-pot process without usage of additional reagents. This thermo-responsive intelligent 3D nanostructures created using this method could be broadly applicable for high-performance and sensitive bacterial analysis.

Author contributions

The manuscript was written through contributions of all authors. All authors have given approval for the final version of the manuscript.

Data availability

The data supporting this article have been included as part of the ESI.†

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgements

This research was supported by NRF and NST grants funded by Korea government (MSIT) (2021M3H4A1A02051048, 2023R1A2C2005185, 2021M3E5E3080844, 2022R1C1C1008815, RS-2024-00348576, RS-2024-00438316, RS-2024-00439931 and RS-2024-00459749), Ministry of Education (RS-2023-00275869), KEIT grants funded by Korea government (MOTIE) (RS-2022-00154853, RS-2024-00403563, and RS-2024-00432382), KEITI grant funded by Korea government (ME) (2021003370003), IPET grant funded by Korea government (MAFRA) (RS-2024-00401639), Nanomedical Devices Development Program of National Nano Fab Center, Technology Innovation Program (2410004004) funded By the Ministry of Trade, Industry & Energy (MOTIE, Korea), Research Supporting Program of OTTOGI Ham Taiho Foundation, and KRIBB Research Initiative Program (KGM5472413).

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Footnote

† Electronic supplementary information (ESI) available. See DOI: https://doi.org/10.1039/d4mh01062k

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