A sensitive Salmonella biosensor using platinum nanoparticle loaded manganese dioxide nanoflowers and thin-film pressure detector

https://doi.org/10.1016/j.snb.2020.128616Get rights and content

Highlights

  • Pt@MnO2 nanoflowers were synthesized for dual mimic enzymatic catalysis of H2O2.

  • Piezoresistor was combined with smartphone to real-time monitor pressure change.

  • This pressure biosensor was able to detect Salmonella as low as 13 CFU/mL in 1.5 h.

  • The mean recovery of Salmonella in the spiked chicken samples was 96.8%96.8 %.

Abstract

Salmonella is the leading factor for microbial food poisoning. In this study, a facile pressure biosensor was developed for rapid and sensitive detection of Salmonella using magnetic nanobeads (MNBs) to separate target bacteria, platinum nanoparticle loaded manganese dioxide nanoflowers (Pt@MnO2 NFs) to amplify detection signal, and a thin-film piezoresistor based pressure detector to monitor pressure change. First, the capture antibodies (CAbs) modified MNBs were used to specifically separate Salmonella from sample to form MNB-CAb-Salmonella complexes (magnetic bacteria). Then, the detection antibodies (DAbs) modified Pt@MnO2 NFs were used for labelling magnetic bacteria to form MNB-CAb-Salmonella-DAb-Pt@MnO2 NF complexes (nanoflower bacteria). After nanoflower bacteria were resuspended with H2O2 in a sealed centrifuge tube, H2O2 was catalyzed by Pt@MnO2 NFs to produce O2, resulting in the increase on pressure. Finally, the pressure increase was real-timely monitored by piezoresistor based pressure detector and transferred to smartphone App via Bluetooth for analysis and determination of Salmonella. This biosensor could quantitatively detect Salmonella from 1.5 × 101 to 1.5 × 105 CFU/mL in 1.5 h with low detection limit of 13 CFU/mL. The Pt@MnO2 NFs with high loading capacity of platinum nanoparticles were demonstrated as dual mimic enzymatic catalyst of H2O2 to greatly enhance the sensitivity. More importantly, it is the first time to combine a thin-film piezoresistor with a smartphone App for realtime monitoring of the pressure change resulting from the mimic enzymatic catalysis of H2O2 into O2 by the Pt@MnO2 NFs on the target bacteria to determine pathogenic bacteria in food samples.

Introduction

Salmonella is the primary risk factor for microbial food poisoning, which can be generally transmitted to humans through consuming contaminated animal-origin foods, such as meats, eggs and milk, etc., and has caused millions of people fall into illnesses, sometimes with severe and fatal outcomes [1]. Many countries have zero tolerance for Salmonella in foods, especially ready-to-eat foods, and thus ultrasensitive Salmonella detection methods are needed. However, food samples are always very complex and the concentration of Salmonella in these samples for in-field screening is usually very low. Therefore, it is of great importance to develop rapid, sensitive and cost-effective methods for in-field Salmonella screening.

Biosensors have been widely reported for bacterial detection in recent years [[2], [3], [4], [5], [6]]. Among them, electrochemical biosensors have attracted the most attentions due to their advantages of compact size, rapid response and portable instrumentation. However, they are faced with a big challenge on large difference for inter-batch and intra-batch electrochemical transducers, which greatly limits their practical applications [[7], [8], [9]]. Optical biosensors are also very popular since they have shown the merits of contactless detection, fast analysis and easy integration. However, most of them are still in the stage of laboratory research due to the sophisticated optical instruments and big background interferences [[10], [11], [12]]. In recent years, some new biosensors have been investigated for detection of foodborne bacteria. Among them, pressure biosensors, relying on pressure changes due to gas production in a sealed environment, have received increasing attentions due to low cost, easy implementation and contactless detection [[13], [14], [15], [16]]. A series of interesting studies on the development of various pressure biosensors have been reported by Yang’s group [[17], [18], [19]]. A portable pressure meter was employed to detect the production of oxygen (O2) due to the decomposition of hydrogen peroxide (H2O2) using catalase or its mimic enzymes. Their pressure biosensors could detect miRNA as low as 7.6 fM, which was more sensitive than most reported studies [20]. They also demonstrated a lateral flow assay with a handheld pressure meter for rapid detection of myoglobin, in which the whole detection process could be completed in 20 min with low detection limit of 2.9 ng/mL in diluted serum samples [21]. Besides, some recent novel studies on pressure biosensors were reported by Wan’s group. They innovatively used the disposable syringe and medical infusion extension line for visual readout of pressure change due to different biological reactions, which were related with the concentration of targets [22][23].

Signal amplification is crucial to the sensitivity of biosensors. Enzymes are the most often used for the development of various biosensors to effectively amplify their biological signals [[24], [25], [26], [27], [28], [29], [30], [31], [32]]. Compared to the natural enzymes, nanozymes have the merits of low cost, good stability and excellent catalytic ability. Among them, platinum nanoparticles (Pt NPs) and manganese dioxide nanoflowers (MnO2 NFs) are two catalase-mimic nanozymes and have been applied to catalyze the decomposition of H2O2 to produce O2 [[33], [34], [35], [36]]. Besides, MnO2 NFs were also excellent nanocarriers, which could be efficiently loaded with various nanomaterials due to their high specific surface area [23,37,38]. So far, the load of Pt NPs onto MnO2 NFs has rarely been reported for dual amplification of biological signals in the detection of foodborne pathogens. With fast popularization of smartphone, it has often reported to act as data analyzer, result displayer and information transmitter to replace expensive benchtop instrumentation in the development of various biosensors [[39], [40], [41], [42]]. Therefore, the combination of pressure biosensors, manganese dioxide nanoflowers with platinum nanoparticles, and smartphone might be promising to provide rapid, sensitive and low-cost methods for in-field detection of Salmonella.

In this study, a pressure biosensor was developed based on immune magnetic nanobeads (MNBs) for specific bacterial separation, manganese dioxide nanoflowers with platinum nanoparticles (Pt@MnO2 NFs) for dual signal amplification and piezoresistor for sensitive pressure monitoring, and Salmonella was used as research model. As shown in Scheme 1, Salmonella was first separated from sample background using the anti-Salmonella capture antibody (CAb) modified MNBs to form the MNB-CAb-Salmonella complexes (magnetic bacteria). Then, the magnetic bacteria were labeled with the anti-Salmonella detection antibody (DAb) modified Pt@MnO2 NFs to form the MNB-CAb-Salmonella-DAb-Pt@MnO2 NF complexes (nanoflower bacteria). Finally, the Pt@MnO2 NFs on nanoflower bacteria were used to catalyze H2O2 into O2 in a sealed centrifuge tube, leading to the increase on pressure, followed by using the piezoresistor based pressure detector to real-timely monitor the pressure change and transmit the data to the smartphone App via Bluetooth for quantitative detection of Salmonella.

Section snippets

Chemicals and reagents

Chloroplatinic acid (H2PtCl6·6H2O) from Sigma (St. Louis, MO, USA), sodium borohydride (NaBH4) from Aladdin (Shanghai, China), and sodium citrate (Na3C6H5O7) and citric acid (C6H8O7) from Solarbio (Beijing, China) were used to synthesize the Pt NPs. Permanganate potassium (KMnO4) from Sinopharm chemical (Beijing, China), hydrochloride acid (HCl) and Polyvinylpyrrolidone (PVP) from Sigma were used for synthesis of the MnO2 NFs. (3-aminopropyl) triethoxysilane (APTES) from Aladdin was employed

Mechanism of the pressure biosensor

This pressure biosensor was based on the measurement of pressure change due to the produced oxygen in a sealed centrifuge tube using the thin-film piezoresistor. As shown in Fig. 1(a), when H2O2 was catalyzed by the Pt@MnO2 NPs on the nanoflower bacteria to produce a certain volume of O2, resulting in the increase on the pressure (P, unit: Pa) of the sealed detection chamber including the whole 0.2 mL centrifuge tube and the partial syringe cylinder, the rubber in the syringe’s cylinder was

Conclusions

In this study, we have successfully developed a novel pressure biosensor using the MnO2 NFs and the Pt NPs for dual mimic enzymatic catalysis of H2O2, piezoresistor and smartphone App for realtime monitoring of oxygen production, and the MNBs for specific separation and efficient concentration of Salmonella to achieve sensitive and rapid detection of Salmonella. This pressure biosensor was able to determine Salmonella at the range from 1.5 × 101 to 1.5 × 105 CFU/mL in 1.5 h with a low detection

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

CRediT authorship contribution statement

Lei Wang: Conceptualization, Data curation, Formal analysis, Methodology, Validation, Writing - original draft. Li Hao: Resources, Methodology. Wuzhen Qi: Software, Data curation. Xiaoting Huo: Formal analysis, Resources. Li Xue: Resources, Formal analysis. Yuanjie Liu: Supervision, Data curation. Qiang Zhang: Supervision. Jianhan Lin: Conceptualization, Supervision, Project administration, Writing - review & editing.

Declaration of Competing Interest

The authors report no declarations of interest.

Acknowledgments

This study was supported by National Key Research and Development of China (2016YFD0500706), Walmart Foundation (SA17031161) and Walmart Food Safety Collaboration Center.

Lei Wang is now a Ph.D. student at the Department of Electronic Engineering, China Agricultural University in Beijing, China. She got her bachelor degree of Electrical Engineering in June, 2016 in Hebei Agricultural University. She started her doctoral study on biosensors with Dr. Jianhan Lin at China Agricultural University since September 2016. Her current research interests include various biosensors for rapid and sensitive detection of pathogenic microorganisms in food or animal samples,

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    Lei Wang is now a Ph.D. student at the Department of Electronic Engineering, China Agricultural University in Beijing, China. She got her bachelor degree of Electrical Engineering in June, 2016 in Hebei Agricultural University. She started her doctoral study on biosensors with Dr. Jianhan Lin at China Agricultural University since September 2016. Her current research interests include various biosensors for rapid and sensitive detection of pathogenic microorganisms in food or animal samples, etc.

    Li Hao is now a master student at the Department of Electronic Engineering, China Agricultural University in Beijing, China. She got her bachelor degree in Electronic Engineering at Sichuan Agricultural University in June 2018. She started her research on biosensor with Dr. Jianhan Lin at China Agricultural University since July 2018. Her current research interests include optical biosensors for rapid detection of pathogenic bacteria in foods.

    Wuzhen Qi is now a Ph.D. student at the Department of Electronic Engineering, China Agricultural University in Beijing, China. He got his bachelor degree of Agricultural Mechanical Engineering in June 2015 and master degree of Mechanical Engineering in June 2018 at Shandong Agricultural University. He started his doctoral study on biosensors with Dr. Jianhan Lin at China Agricultural University since September 2018. His current research interests include various biosensors for rapid and sensitive detection of pathogenic microorganisms in food or animal samples, etc.

    Xiaoting Huo is now a master student at the Department of Electronic Engineering, China Agricultural University in Beijing, China. She got her bachelor degree in Communication Engineering at Hunan University of Technology in June 2018. She started her research on microfluidic biosensors with Dr. Jianhan Lin at China Agricultural University since July 2018. Her current research interests include microfluidic biosensors for rapid and automatic detection of pathogenic bacteria in foods.

    Li Xue is now a Ph.D. student at the Department of Electronic Engineering, China Agricultural University in Beijing, China. She got her bachelor degree of Electrical Engineering in 2015 at Shandong University of Science and Technology and master degree of Agricultural Engineering in 2018 at China Agricultural University. She started her doctoral study on optical biosensors and biosensing methods for bacteria detection with Dr. Jianhan Lin since July 2019. Her current research interests include optical biosensors, nanomaterials and magnetic separation for detection of pathogenic microorganisms in foods, as well as portable bio-instruments for in-field detection of foodborne pathogens, etc.

    Dr. Yuanjie Liu is now an Associate Professor at the Department of Computer Science, China Agricultural University in Beijing, China. He got his bachelor degree of Mathematics and Applied Mathematics at Nanjing University in 2007, and Ph. D. degree of Applied Mathematics at University of Chinese Academy of Sciences in 2012. He started to work as Assistant Professor in the Institute of Semiconductors, Chinese Academy of Sciences from August 2012 to August 2018, and joined China Agricultural University since September 2018. His current research interests include big data and cloud computing for food safety monitoring and animal disease prevention and data mining and processing for biosensing, etc.

    Dr. Qiang Zhang is now a full Professor at the Department of Biosystems Engineering, University of Manitoba, Canada. He got his bachelor degree in Mechanical Engineering at Hefei Polytechnic University in 1982, his Master and Ph. D. degrees in Agricultural Engineering at Pennsylvania State University in 1985 and 1987, respectively. His current research interests include animal production environment sensing and control, airborne transmission of disease pathogens in animal facilities, and bulk solids handling and storage, etc.

    Dr. Jianhan Lin is now a Professor at the Department of Electronic Engineering, China Agricultural University in Beijing, China. He got his bachelor degree of Electronic Engineering in 2001, master degree and Ph. D. degree of Agricultural Engineering in 2004 and 2007 at China Agricultural University. He started his Postdoctoral Research in Biological Engineering at the University of Arkansas at Fayetteville, Arkansas, US from October 2007 to May 2009, and worked as Research Associate at the University of Arkansas from June 2009 to June 2011. Then, he was appointed as Chief Technology Officer by Aibit Biotech LLC at Wuxi, China in July 2011 and joined China Agricultural University since September 2012. His current research interests include biosensors for rapid detection of foodborne pathogens and animal diseases, microfluidic chip for separation and detection of pathogens, food supply chain management and traceability, as well as infectious animal disease prevention and control, etc.

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