Rapid Detection Of Foodborne Pathogens Research Articles

Research progress on the detection of foodborne pathogens based on aptamer recognition.

Foodborne diseases caused by bacterial contamination are a serious threat to food safety and human health. The classical plate culture method has the problems of long detection cycle, low sensitivity and specificity, and complicated operation, which cannot meet the growing demand for rapid quantitative detection of pathogenic bacteria. The frequent outbreak of foodborne diseases has put forward higher requirements for rapid and simple detection technology of foodborne pathogens. Aptamer is a kind of oligonucleotide fragment that can recognize targets with the advantages of high affinity and good specificity. The target can be range from proteins, small molecules, cells bacteria, and even viruses. Herein, the latest advances in sensitive and rapid detection of foodborne pathogens based on aptamer recognition was reviewed. Special attention has been paid to the obtained sequences of aptamers to various foodborne pathogens, the optimization of sequences, and the mechanism of aptamer recognition. Then, the research progress of biosensors for the detection of pathogenic bacteria based on aptamer recognition were summarized. Some challenges and prospects for the detection of foodborne pathogens based on aptamer recognition were prospected. In summary, with the further deepening of aptamer research and improvement of detection technology, aptamer-based recognition can meet the needs of rapid, sensitive, and accurate detection in practical applications.

Boron nitride nanoplate-based strand exchange amplification with enhanced sensitivity and rapidity for quantitative detection of Staphylococcus aureus in food samples.

Staphylococcus aureus is one of the most common foodborne pathogens that can cause serious food poisoning and infectious diseases in humans. Standard identification approaches include nucleic acid amplification, but current amplification tools suffer from low amplification efficiency, resulting in the risk of low sensitivity and long detection time. Herein, boron nitride nanoplates (BNNPs) were chosen as an additive for enhancing the sensitivity and rapidity of strand exchange amplification (SEA), thereby successfully expanding the application of nucleic acid detection for detecting Staphylococcus aureus in food samples. As a result, SEA based on boron nitride nanoplates (BNNP-SEA) was employed for sensitive and rapid detection of foodborne pathogen Staphylococcus aureus. Compared with classical SEA, the BNNP-based SEA assay was more than 10-fold sensitive, and the detection time was reduced by 15 minutes. The optimized BNNP-based SEA shows a wide linear range from 40 pg to 50 ng in a diluted solution of the target DNA with a low detection limit of 40 pg. Moreover, the BNNP-based SEA achieves the quantitative detection of Staphylococcus aureus in different food samples (pork, beef, mutton, duck, milk and shrimp). In contrast to the classical SEA, the BNNP-based SEA method enabled sensitive and rapid detection of Staphylococcus aureus in the above food samples at concentrations as low as 5 × 103 CFU mL-1. The BNNP-based SEA assay is specific, sensitive and reliable, offering a valuable diagnostic technology for routine analysis in food safety research.

A convenient electrochemical biosensor mediated by filtration-assisted pretreatment and multi-TEs amplification system for Staphylococcus aureus detection in complex matrices

Rapid detection of foodborne pathogens plays a more and more significant role in the field of public health. In this research, we developed a convenient electrochemical biosensor mediated by filtration-assisted pretreatment and multiple thermostatic enzymes (Multi-TEs) amplification system for the detection of Staphylococcus aureus (S. aureus) in complex matrices. Firstly, the designed filtration-assisted pretreatment could simultaneously collect and enrich target bacterium from various actual samples within 5 min. Then, the target bacterial genome template was reacted at 37 °C in Multi-TEs amplification system to obtain specific double stranded products. Finally, the specific double stranded products were fixed on the surface of the electrode and the electrochemical indicators (Methylene blue, MB) was combined for the quantitative one-step electrochemical analysis of S. aureus. This assay has great linear calibration curves with the concentration ranging from 101 CFU/mL to 106 CFU/mL. The limit of detections (LOD) was defined as 1.28 CFU/mL, which performance is superior to some commercial ELISA kits. Meanwhile, amazingly, it has excellent specificity for S. aureus against other foodborne pathogens and could be applied for S. aureus analysis in human serum samples with the great recoveries and associated relative standard deviations (RSDs). These features do make the ideal facile electrochemical analytical platform for on-site detection of S. aureus.

E. coli O157:H7 Detection Using Surface Plasmon Resonance Based Biosensor

The detection of foodborne pathogenic bacteria remains a significant challenge, and the need for fast and sensitive detection methods is becoming increasingly important. Escherichia coli is a prevalent bacteria associated with foodborne illness, and this study aimed to evaluate the ability of a surface plasmon resonance (SPR) based biosensor to detect E. coli O157:H7 at low levels in pure culture and artificially contaminated bay leaves (Laurus nobilis) using different injection methods. To develop a biological sensing surface, the sensor surface was functionalized with 3-aminopropyltriethoxysilane (APTES), and polyclonal antibodies were immobilized on the surface for bacteria detection. Bacterial attachment to the antibodies resulted in a change in resonance angle. The biosensor was able to discriminate between cellular concentrations of 103 to 107 CFU/mL and showed potential in detecting different pathogens in various food samples. Before the SPR detection, the sample preparation step was optimized to ensure complex food matrices were suitable for SPR analysis. The results suggest that the SPR based biosensor is a promising tool for the rapid detection of foodborne pathogens in complex food matrices.

Capture antibody imitator MnO2 nanozyme-based dual-signal immunochromatographic assay for rapid detection of Salmonella enteritidis

The sensitive and rapid detection of foodborne pathogens is of great significance to global public health security. Here, with two-dimensional nanomaterial MnO2 NSs as the capture antibody imitator and signal reporter molecules, based on its enzyme-catalyzed signal amplification capability, a label-free, dual-signal output immunochromatographic biosensor was constructed to achieve the sensitive detection of S. enteritidis. The capture antibody imitator of MnO2 capture probe shows an outstanding capture ability, forming a stable immune complex with S. enteritidis. As an in-situ catalytic amplification colorimetric signal trigger, MnO2 NSs exert oxidase-like enzyme activity and obtain satisfactory detection sensitivity. It is worth noting that the proposed dual-signal ICA has high analytical performances for S. enteritidis during the concentration range of 103-107 cfu mL−1 both in standard bacterial solutions and food matrixes.

Rapid detection of foodborne pathogens in diverse foodstuffs by universal electrochemical aptasensor based on UiO-66 and methylene blue composites.

Rapid and sensitive detection of foodborne pathogens in complex environments is essential for food protection. A universal electrochemical aptasensor was fabricated for the detection of three common foodborne pathogens, including Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), and Salmonella typhimurium (S. typhimurium). The aptasensor was developed based on the homogeneous and membrane filtration strategy. Zirconium-based metal-organic framework (UiO-66)/methylene blue (MB)/aptamer composite was designed as a signal amplification and recognition probe. Bacteria were quantitatively detected by the current changes of MB. By simply changing the aptamer, different bacteria could be detected. The detection limits of E. coli, S. aureus and S. typhimurium were 5, 4 and 3CFU·mL-1, respectively. In humidity and salt environments, the stability of the aptasensor was satisfactory. The aptasensor exhibited satisfactory detection performance in different real samples. This aptasensor has excellent potential for rapid detection of foodborne pathogens in complex environments.

Preparation of a multifunctional and ultrasensitive Au@AgNPs-Van/PDMS film SERS substrate for rapid detection of foodborne pathogens in beef

We present a novel and super-sensitive vancomycin (Van) modified Au@AgNP self-assembly with polydimethylsiloxane (PDMS) (Au@AgNPs/Van-PDMS) film surface-enhanced Raman scattering (SERS) substrate for identification of foodborne pathogens in beef, without the need for in vitro bacterial culture. The results illustrated that the Au@AgNPs/Van-PDMS film exhibited high reproducibility and Raman enhancement effect (1.09 × 105). Clostridium perfringens, Bacillus subtilis and Staphylococcus aureus isolated from beef captured by Au@AgNPs/Van-PDMS film exhibited high reproducibility and significant differences. Principal component analysis score plots distinguished classes of foodborne pathogens, and the qualitative identification of linear discriminant analysis was correct at 100%. The detection limit of S. aureus in beef was as low as 3 CFU/mL. This method could provide an effective means and technical support for realizing the application of SERS technology for quick and highly sensitive detection of foodborne pathogens in complex environments.

Polydopamine-coated two-dimensional nanomaterials as high-affinity photothermal signal tag towards dual-signal detection of Salmonella typhimurium by lateral flow immunoassay

Lateral flow immunoassay (LFIA) with gold nanoparticles (AuNPs) as signal reporters has been extensively utilized for the rapid detection of foodborne pathogens. Nevertheless, it’s challenging for LFIA to obtain sensitive detection, because of the inefficient coupling of AuNPs and antibodies and the poor colorimetric signals. To improve on this, we used a two-dimensional nanomaterial (Cu2MoS4) modified with PDA (CMS@PDA) that provides superior bio-affinity and excellent photothermal signal qualities. With the CMS@PDA material, we successfully constructed a portable LFIA device for specific detection of Salmonella typhimurium (S. typhimurium) using a dual-signal probe that combines colorimetric and photothermal signals (CM/PT). The dynamic detection range of the CMS@PDA-LFIA is 103 to 107 cfu mL−1. Notably, the detection limit of 103 cfu mL−1 is 100 times more sensitive compared to AuNPs-based LFIA. Moreover, the assay exhibits satisfactory recovery rates for S. typhimurium detection in food samples. In this research, the coating of PDA solves the problem of low water-solubility of CMS, making the proposed CMS@PDA nanocomposite showcase better performance in the CM/PT mode detection by LFIA. Beyond all doubt, this work offers a novel supplement for applying multifunctional two-dimensional nanomaterials in LFIAs.

A New Duplex Recombinase Polymerase Amplification (D-RPA) Method for the Simultaneous and Rapid Detection of Shigella and Bacillus cereus in Food.

Shigella and Bacillus cereus are two common foodborne pathogens that cause intestinal diseases and seriously affect human life and health. Traditional microbiological culture methods are time-consuming and laborious, and polymerase chain reaction (PCR)-based methods rely on expensive thermal cyclers and lengthy reaction times. In this study, on the basis of the specific gene ipaH7 of Shigella and the virulence gene nheABC of B. cereus, a duplex detection system was established for the first time by using the recombinase polymerase amplification technique (D-RPA). After optimization, D-RPA could be effectively amplified at 42 °C for 25 min with excellent specificity, and the detection limits of D-RPA for Shigella and B. cereus in artificially contaminated samples were 2.7 × 101 and 5.2 × 102 CFU/mL, respectively. This study provides a certain research basis for multiple detection with RPA, an isothermal amplification technology. Furthermore, it lays a good foundation for high-throughput rapid detection of foodborne pathogens.

Influence of Biological and Environmental Factors in the Extraction and Concentration of Foodborne Pathogens using Glycan-Coated Magnetic Nanoparticles

Rapid detection of foodborne pathogens is essential to preventing foodborne illness outbreaks. Before detection can occur, however, it is often necessary to extract and concentrate bacteria. Conventional methods such as centrifugation, filtration, and immunomagnetic separation can often be time-consuming, ineffective, or costly when working with complex food matrices. This work used cost-effective glycan-coated magnetic nanoparticles (MNPs) for rapid concentration of Escherichia coli O157, Listeria monocytogenes, and Staphylococcus aureus. Glycan-coated MNPs were used to concentrate bacteria from both buffer solution and food matrices while examining the effect of factors including solution pH, bacterial concentration, and target bacterial species. In both pH 7 and reduced pH experiments, successful extraction of bacterial cells occurred in all food matrices and bacteria tested. In neutral pH buffer solution, bacteria were concentrated to 4.55 ± 1.17, 31.68 ± 6.10 and 64.27 ± 16.78 times their initial concentration (mean ± standard deviation) for E. coli, L. monocytogenes and S. aureus, respectively. Successful bacterial concentration occurred in several food matrices, including S. aureus in milk (pH 6), L. monocytogenes in sausage (pH 7), and E. coli O157 in flour (pH 7). The insights gained may facilitate future applications of glycan-coated MNPs to extract foodborne pathogens.

An integrated colorimetric and photothermal lateral flow immunoassay based on bimetallic Ag–Au urchin-like hollow structures for the sensitive detection of E. coli O157:H7

Lateral flow immunoassay (LFIA) with gold nanoparticles (AuNPs) as the signal reporter is widely used for the rapid detection of food-borne pathogens. However, it is difficult for LFIA to achieve sensitive detection due to the insufficient colorimetric signal brightness of AuNPs. Herein, we developed a bimetallic Ag–Au urchin-like hollow nanospheres (BUHNPs) based on the simple and rapid synthesis through co-reduction and galvanic replacement reactions. The BUHNPs exhibit superb colorimetric signal brightness, strong photothermal signals and high antibody coupling efficiency. The obtained colorimetric and photothermal LFIA (BUHNPs-CM-LFIA and BUHNPs-PT-LFIA) based on BUHNPs were used for the sensitive detection of a pathogenic bacterium (Escherichia coli O157:H7). The limit of detection values of the colorimetric and photothermal quantitative analysis were 2.48 × 103 and 5.50 × 102 CFU mL−1, which were approximately 4-fold and 18-fold lower than that of AuNPs-LFIA (9.92 × 103 CFU mL−1), respectively. This work suggests that a dual-readout LFIA with BUHNPs as a promising signal reporter can be used to improve the detection performance of LFIA and construct a more accurate and sensitive detection platform.

Magnetorheological elastomer and smartphone enable microfluidic biosensing of foodborne pathogen

Rapid detection of foodborne pathogens is crucial to prevent the outbreaks of foodborne diseases. In this work, we proposed a novel microfluidic biosensor based on magnetorheological elastomer (MRE) and smartphone. First, micropump and microvalves were constructed by deforming the MRE under magnetic actuation and integrated into the microfluidic biosensor for fluidic control. Then, the micropump was used to deliver immune porous gold@platinum nanocatalysts (Au@PtNCs), bacterial sample, and immunomagnetic nanoparticles (MNPs) into a micromixer, where they were mixed, incubated and magnetically separated to obtain the Au@PtNC-bacteria-MNP complexes. After 3,3′,5,5′-tetramethylbenzidine and hydrogen peroxide were injected and catalyzed by the Au@PtNCs, smartphone was used to measure the color of the catalysate for quantitative analysis of target bacteria. Under optimal conditions, this biosensor could detect Salmonella typhimurium quantitatively and automatically in 1 h with a linear detection range of 8.0 × 101 CFU/mL to 8.0 × 104 CFU/mL and a detection limit of 62 CFU/mL. The microfluidic biosensor was compact in size, simple to use, and efficient for detection, and might be used for in-field screening of foodborne pathogens to prevent food poisoning.

Recent Progress in Spectroscopic Methods for the Detection of Foodborne Pathogenic Bacteria.

Detection of foodborne pathogens at an early stage is very important to control food quality and improve medical response. Rapid detection of foodborne pathogens with high sensitivity and specificity is becoming an urgent requirement in health safety, medical diagnostics, environmental safety, and controlling food quality. Despite the existing bacterial detection methods being reliable and widely used, these methods are time-consuming, expensive, and cumbersome. Therefore, researchers are trying to find new methods by integrating spectroscopy techniques with artificial intelligence and advanced materials. Within this progress report, advances in the detection of foodborne pathogens using spectroscopy techniques are discussed. This paper presents an overview of the progress and application of spectroscopy techniques for the detection of foodborne pathogens, particularly new trends in the past few years, including surface-enhanced Raman spectroscopy, surface plasmon resonance, fluorescence spectroscopy, multiangle laser light scattering, and imaging analysis. In addition, the applications of artificial intelligence, microfluidics, smartphone-based techniques, and advanced materials related to spectroscopy for the detection of bacterial pathogens are discussed. Finally, we conclude and discuss possible research prospects in aspects of spectroscopy techniques for the identification and classification of pathogens.

Biofunctionalized Magnetic Nanoparticles with Multiplex Touchdown PCR for Simultaneous and Rapid Detection/Identification of Campylobacter jejuni and Campylobacter coli

The simple, accurate, and rapid detection of foodborne pathogens is essential for public health. Development of an immunomagnetic separation (IMS) multiplex touchdown PCR (IMS–multiplex TD–PCR) assay for simultaneous detection and distinguishing of C. jejuni and C. coli is reported herein. Polyclonal antibody (pAb) against multiepitope antigen (MEA) was conjugated to ferromagnetic nanoparticles (FMNs) to produce anti-MEA FMNs. Optimal anti-MEA FMNs loading yielded 26.7 μg of immunoglobulin G (IgG) molecules per mg of FMNs with an average size of 72 ± 9 nm, corresponding to an 83% rate of pAb conjugation. Anti-MEA FMNs (20 μg) for IMS captured culturable C. jejuni cells at 3.54 × 10 2 colony-forming unit (CFU)/mL in pure culture, while higher amounts (40 and 60 μg) reduced the recovery. The scanning electron microscope (SEM) analysis revealed the attachment of anti-MEA FMNs to target bacteria, forming aggregated cells and magnetic nanoparticles in ellipse-like shapes. The subsequent multiplex TD–PCR assay simultaneously detected and distinguished C. jejuni and C. coli at 104 CFU/mL in mixed culture and at 103 CFU/mL for each individual species. Furthermore, the limit of detection (LOD) of the IMS–multiplex TD–PCR assay was 104 CFU/g in spiked chicken breast samples. Specificity was 100% for both C. jejuni and C. coli as none of the amplicons were detected in control samples where Campylobacter was absent. This assay is able to detect and distinguish C. jejuni and C. coli simultaneously and is simple, accurate, and rapid with a time to result of 4 h without an enrichment step, making it a promising approach for rapid and culture-free detection of Campylobacter in chicken products.

Multiplex detection of foodborne pathogens by real-time loop-mediated isothermal amplification on a digital microfluidic chip

Rapid detection of foodborne pathogens is critical to assure food quality. In this study, a digital microfluidic (DMF) chip in combination with loop-mediated isothermal amplification (LAMP) was developed for multiplex detection of four common foodborne pathogens. The DMF chip contains ten reaction chambers with dehydrated primers targeting different genes for simultaneous amplification by LAMP in the volume of 2 μl/reaction with a detection limit of 102 CFU/ml for all target bacteria according to the 3σ criteria. Using spiked milk for validation, as low as 103 CFU/ml of all four pathogenic bacteria were detected simultaneously in a single test within 50 min. The proposed method exhibited higher sensitivity to the commercial benchtop LAMP method with the advantages of simpler operation, high efficiency, and miniaturization, showing great potential for point-of-care (POC) testing of various foodborne pathogens.

Listeria monocytogenes의 검출을 위한 loop-mediated isothermal amplification, real-time PCR, 배지 도말법의 검출 성능 및 특성 비교

Listeria monocytogenes is a foodborne pathogen causing listeriosis, which can be fatal in specific high-risk groups. The aim of this study was to compare the performance (accuracy, sensitivity, and specificity) of 3M™ Molecular Detection System (3M™ MDS) and Korean Food Codex [real-time PCR (RT-PCR) and selective agar] for the detection of L. monocytogenes in various food matrices. The detection performance of the three methods was determined against 100-103 CFU/mL of L. monocytogenes in vitro and showed high accuracy in the order of RT-PCR, 3M™ MDS, and selective agar. There was no difference in sensitivity and specificity of the three methods. Eleven food matrices, selected from agricultural, livestock, and seafood products, were artificially inoculated with 100-103 CFU/25 g of L. monocytogenes and enriched in 3M™ Demi-Fraser Broth. None of the three methods could completely detect low concentrations of L. monocytogenes in a food matrix. However, 3M™ MDS, which is a loop-mediated isothermal amplification (LAMP)-based technology for rapid detection, showed a higher positive detection rate than RT-PCR did, but lower than that of selective agar. These data indicated that 3M™ MDS was superior for the rapid detection of L. monocytogenes, compared to RT-PCR in food matrices containing various inhibitors. Consequentially, the study findings suggest that the LAMP method is a promising alternative to RT-PCR for the rapid detection of foodborne pathogens.

A microfluidic chemiluminescence biosensor based on multiple signal amplification for rapid and sensitive detection of E.coli O157:H7

E. coli O157:H7 has become a crucial foodborne pathogen with certain morbidity and mortality. However, the available methods still face the significant challenges including the improvement of detection specificity and sensitivity for real sample analysis. Herein, a microfluidic chemiluminescence biosensor based on a multiple signal amplification strategy was developed for rapid and ultrasensitive detection of E.coli O157:H7. The microfluidic device was consisted of an upstream snake-shape mixing channel and a downstream ship-shape microcavity with micropillar array for high efficient mixing of CL reagents, sufficient CHA amplification reaction and CL reaction. The multiple signal amplification was achieved by bacteria competitively bind to trigger catalytic hairpin assembly (CHA) and HRP-Au NPs catalyzed CL reaction. Based on the platform, the detection of target bacteria is converted to the detection of nucleic acid with a limit detection down to 130 CFU/mL, which is better or comparable to previously reported biosensors. Moreover, the whole analysis time was about 1.5 h with only 10 μL of sample. The biosensor exhibited an excellent recovery rate ranged from 90.0% to 110.0% and good selectivity to the target bacteria. Overall, this developed biosensor is a promising tool for ultrahigh sensitive and rapid detection of foodborne pathogens.

Detection of Unamplified E. coli O157 DNA Extracted from Large Food Samples Using a Gold Nanoparticle Colorimetric Biosensor.

Rapid detection of foodborne pathogens such as E. coli O157 is essential in reducing the prevalence of foodborne illness and subsequent complications. Due to their unique colorimetric properties, gold nanoparticles (GNPs) can be applied in biosensor development for affordability and accessibility. In this work, a GNP biosensor was designed for visual differentiation between target (E. coli O157:H7) and non-target DNA samples. Results of DNA extracted from pure cultures indicate high specificity and sensitivity to as little as 2.5 ng/µL E. coli O157 DNA. Further, the biosensor successfully identified DNA extracted from flour contaminated with E. coli O157, with no false positives for flour contaminated with non-target bacteria. After genomic extraction, this assay can be performed in as little as 30 min. In addition, food sample testing was successful at detecting approximately 103 CFU/mL of E. coli O157 magnetically extracted from flour after only a 4 h incubation step. As a proof of concept, these results demonstrate the capabilities of this GNP biosensor for low-cost and rapid foodborne pathogen detection.

Recent advancements in microfluidic chip biosensor detection of foodborne pathogenic bacteria: a review.

Foodborne diseases caused by pathogenic bacteria pose a serious threat to human health. Early and rapid detection of foodborne pathogens is an urgent task for preventing disease outbreaks. Microfluidic devices are simple, automatic, and portable miniaturized systems. Compared with traditional techniques, microfluidic devices have attracted much attention because of their high efficiency and convenience in the concentration and detection of foodborne pathogens. This article firstly reviews the bio-recognition elements integrated on microfluidic chips in recent years and the progress of microfluidic chip development for pathogen pretreatment. Furthermore, the research progress of microfluidic technology based on optical and electrochemical sensors for the detection of foodborne pathogenic bacteria is summarized and discussed. Finally, the future prospects for the application and challenges of microfluidic chips based on biosensors are presented.Graphical abstract

Development of modified enrichment broth for short enrichment and recovery of filter-injured Salmonella Typhimurium

The filter concentration method facilitates the rapid detection of foodborne pathogens. The filter concentration method lowered the limit of detection (LOD) of artificially inoculated cabbage with Salmonella Typhimurium; however, the procedure injured foodborne pathogens during filtering procedure. Thus, to detect injured pathogens under the detection limit, an enrichment broth promoting pathogen resuscitation and growth is required. To rapidly recover, cultivate and lower the time to result (TTR) of S. Typhimurium detection after filter concentration method, a brain heart infusion (BHI) broth-based modified enrichment broth (MEB) was developed. The MEB was developed by fitting growth curves to a modified Gompertz model; 1.00 g/L of sodium pyruvate, 0.20 g/L proline and 2.0 g/L magnesium sulphate additives were optimized as additional components to rapidly grow filter-injured S. Typhimurium. As a result, the rate of filter-injured S. Typhimurium went from 100% to 0.0% using MEB within 3.5 h. In contrast, BHI required 4 h and buffered peptone water (BPW) required more than 4 h to decrease the injury rate to 0.0%. Using MEB, BHI and BPW, filter-injured S. Typhimurium in cabbages were enriched to 4.056 ± 0.026 Log CFU/25 g, 3.571 ± 0.187 Log CFU/25 g and 3.708 ± 0.156 Log CFU/25 g, respectively. Additionally, 1–9 CFU/mL S. Typhimurium in cabbage was detected within 3.0 h, including 1 h enrichment with MEB, whereas 5.0 h was required for BHI and BPW. Thus, the MEB developed in this study showed great potential as a short enrichment broth for the rapid detection of filter-injured S. Typhimurium.