Detection Of Pathogenic Bacteria Research Articles

Magnetic relaxation switch biosensor for detection of Vibrio parahaemolyticus based on photocleavable hydrogel

BackgroundFoodborne pathogens, particularly Vibrio parahaemolyticus (VP) found in seafood, pose significant health risks, including abdominal pain, nausea, and even death. Rapid, accurate, and sensitive detection of these pathogens is crucial for food safety and public health. However, existing detection methods often require complex sample pretreatment, which limits their practical application. This study aims to overcome these limitations by developing a label-free magnetic relaxation switch (MRS) biosensor for the detection of VP, utilizing a photocleavable sol-gel phase transition system for improved efficiency and accuracy. ResultsIn this work, a tag-free magnetic relaxation switch (MRS) biosensor was designed for the detection of Vibrio parahaemolyticus (VP), based on a photocleavable sol-gel phase transition system. A large amount of lithium acyl hypophosphite (LAP), gold nanoparticles (AuNPs), and single-stranded DNA (ssDNA) loaded on the surface of Ti3C2Tx MXene acted as the signal unit LAP-MXene@AuNPs-ssDNA. The pipette tip served as a reaction vessel, and when VP was present, Apt specifically captured VP and released the signal units. The released signal units were then injected into the low-field nuclear magnetic resonance (LF-NMR) test solution, a gel formed by crosslinking of disulfide bonds. The gel was cleaved by LAPs on the signal units under ultraviolet (UV) irradiation, triggering a gel-sol phase transition, which increased transverse relaxation time (T2), thus enabling the detection of VP. Under the optimal experimental conditions, the linear range and detection limit for VP were 102 ∼ 108 CFU/mL and 10 CFU/mL, respectively. Significance and NoveltyThe simplified biometric identification process in the pipette tip reduces errors from multiple sample transfers, enhancing efficiency. The use of photocleavable hydrogel for signal output eliminates issues associated with magnetic material aggregation, significantly improving detection precision. The assay is of good selectivity, stability reproducibility, and convenience, having a broad application prospect in the rapid detection of pathogenic bacteria in the field.

Application of bronchoalveolar lavage fluid cytomorphology in diagnosing Nocardia otitidiscaviarum: a case report

BackgroundNocardiosis is a rare infectious disease, which is frequently underdiagnosed because of the distinct bacterial shape of the causative agent and its developmental history. The morphological analysis of bronchoalveolar lavage fluid can facilitate the detection of pathogenic bacteria by observing cellular–bacterial interactions.Case presentationAn immunocompetent 67-year-old male patient of Asian ethnicity developed a cough without apparent cause 20 days earlier. We initially discovered mycobacteria that appeared to be Nocardia in the cytomorphological examination of bronchoalveolar lavage fluid, which was later determined to be Nocardia otitidiscaviarum through metagenomic next-generation sequencing and microbiological cultures. The patient was eventually diagnosed with Nocardia pneumonitis after other testing and clinical signs were considered. After anti-infective treatment, the patient improved and was discharged.ConclusionSeveral diagnostic approaches were used in this case, and the importance of cell morphology as an early screening method was emphasized for suspected Nocardia infection.

Biosensor‐Based Microfluidic Platforms for Rapid Clinical Detection of Pathogenic Bacteria

AbstractPathogenic bacteria are commonly found in food, water, and soil, posing significant public health challenges globally. Therefore, early, rapid, and highly sensitive strategies for monitoring the bacterial proliferation are crucial for ensuring public health, medical diagnosis, and food safety. Compared to traditional techniques, microfluidic platforms provide powerful detective tools characterized by high integration, high throughput, ease of operation, low reagent consumption, and high sensitivity. Driven by substantial commercial demand, research and development in microfluidic‐based rapid detection methods and technologies has progressed significantly derived by the interdisciplinary integration of multiple disciplines. In this review, progress in clinical detection of pathogenic bacteria with microfluidic biosensors, including microfluidic devices for point‐of‐care (POC) testing, is summarized. Strategies for pathogenic bacteria detection, containing their advantages and disadvantages are discussed in detail. Advanced platforms for capturing and detecting pathogenic bacteria, such as microchannels, microarrays, digital microfluidics (DMF) and paper‐based platforms, are highlighted. The accomplishments and shortcomings of these microfluidic devices are also summarized. Additionally, case studies of biosensor‑based microfluidic devices used for detecting diseases caused by bacterial imbalances are listed. Finally, possible research perspectives for further development in highly effective biosensor‑based microfluidics for clinical detection of pathogenic bacteria are proposed.

Classification and Identification of Foodborne Bacteria in Beef by Utilising Surface-Enhanced Raman Spectroscopy Coupled with Chemometric Methods.

The detection and classification of foodborne pathogenic bacteria is crucial for food safety monitoring, consequently requiring rapid, accurate and sensitive methods. In this study, the surface-enhanced Raman spectroscopy (SERS) technique coupled with chemometrics methods was used to detect and classify six kinds of foodborne pathogenic bacteria, including Salmonella typhimurium (S. typhimurium), Escherichia coli (E. coli) O157:H7, Staphylococcus aureus (S. aureus), Listeria monocytogenes (L. monocytogenes), Listeria innocua (L. innocua), and Listeria welshimeri (L. welshimeri). First, silver nanoparticles (AgNPs) with different particle sizes were prepared as SERS-enhanced substrates by changing the concentration of sodium citrate, and the volume ratio of silver nanosol to bacterial solution was optimised to obtain the optimal SERS signal. Then, principal component analysis (PCA) and hierarchical cluster analysis (HCA) were used to classify the SERS spectra of six bacteria at three classification levels (Gram type level, genus level and species level), and appropriate classification models were established. Finally, these models were validated on 540 spectra using linear discriminant analysis (LDA), achieving an average accuracy of 95.65%. Overall, it was concluded that the SERS technique combined with chemometrics methods could achieve the rapid detection and classification identification of foodborne pathogenic bacteria, providing an effective means for food safety monitoring.

“Dandelion” magnetic bead-driven CDs@CeO2 based-multifunctional nanoplatform for tri-modal ultrasensitive detection and efficient eradication of pathogenic bacteria

To address the challenge of foodborne pathogens, we have developed an ultra-sensitive tri-modal detection and effective pathogen eradication strategy utilizing a “dandelion” magnetic bead-driven cerium dioxide doped with carbon dots (CDs@CeO2) multifunctional nanoplatform. Aptamer (Apt) modification CDs@CeO2 specifically recognizes and binds to the outer membrane proteins of Salmonella typhimurium (S. typhimurium). Concurrently, mannose-functionalized dandelion-like magnetic beads selectively recognize adhesins of S. typhimurium, avoiding competition with Apt for binding sites and leveraging the strong bacterial capture capability for high-sensitivity detection. The prepared CDs@CeO2 has been demonstrated to exhibit favourable fluorescence characteristics. In the presence of H2O2, CDs@CeO2 is capable of oxidising colorless 3,3′,5,5′-tetramethylbenzidine (TMB) into blue oxide, displaying exceptional photothermal properties. CDs@CeO2 have the potential to generate •OH through a Fenton-like reaction, which could directly damage cells. Notably, coupling the photosensitizer Ce6 to CDs@CeO2 and subsequent illumination generates 1O2, which synergistically eradicates bacteria with •OH. Detection limits of the fluorescence, colorimetry, and photothermal methods were 3.85 cfu/mL, 9.44 cfu/mL, and 5.00 cfu/mL, respectively. The platform successfully achieves precise detection of S. typhimurium in actual pre-prepared food samples through fluorescence, colorimetry, and photothermal tri-modal signals, and effectively kills bacteria, achieving trace detection and source control.

An Amplification-Free Digital Assay Based on Primer Exchange Reaction-Mediated Botryoidal-Like Fluorescent Polystyrene Dots to Detect Multiple Pathogenic Bacteria.

Multiple and ultrasensitive detection of pathogenic bacteria is critical but remains a challenge. Here, we introduce a digital assay for multiplexed and target DNA amplification-free detection of pathogenic bacteria using botryoidal-like fluorescent polystyrene dots (PS-dots), which were first prepared through the hybridization reaction between primer exchange reaction chains and polystyrene nanospheres that encapsulated polymer dots for signal preamplification. The pathogenic bacteria's DNA was cleavaged by the argonaute (Ago) protein-mediated multiple and precise cleavage reactions, where the obtained target sequences bridged the magnetic beads (MBs) and botryoidal-like PS-dots via a hybridization reaction, and the fluorescent MB-botryoidal PS-dot complexes were utilized as digital probes based on colors and sizes for digital encoding. An artificial-intelligence-fluorescent microsphere counting algorithm was applied to identify and count the fluorescent MBs for digital readout. This digital assay combined the ultrabright botryoidal-like PS-dots with Clostridium butyricum Ago's precise enzyme cleavage properties, achieving simultaneous detection of three pathogenic bacteria with a linearity range from 102 to 106 CFU/mL without target DNA amplification within 1.5 h. This digital assay has also been applied to detect aquatic and clinical samples with accepted accuracy (98%), which offers an avenue for a next-generation multiplexed digital platform for pathogenic bacteria analysis.

Enzyme-Responsive Chitosan-Based Electrospun Nanofibers for Enhanced Detection of β-Glucuronidase from Pathogenic E. coli

Antimicrobial resistance (AMR) poses a significant global health challenge, leading to the ineffectiveness of numerous conventional antibiotics against various bacterial infections. Hence the rapid detection and identification of pathogenic bacteria are imperative for managing AMR and implementing suitable treatment approaches. To improve rapid detection, a biopolymer-based autonomously reporting enzyme-sensitive biopolymer material has been developed for detecting β-glucuronidase (β-Gus) from pathogenic E. coli. In the presence of enzyme β-Gus a blue-colored fluorophore is released from functionalized electrospun chitosan-polyethylene oxide nanofibers, which is monitored via fluorescence spectroscopy. The nanofibers exhibited a 3.4 times enhanced sensitivity compared to neat hydrogels and also to related chromogenic sensing materials. For an observation time of 60 minutes, a limit of detection (LOD) for β-Gus was determined to be 4.7 nM. This nanofiber sensing substrate was then studied with the pathogenic E. coli strain NCTC 10418, showing a three times greater sensitivity compared to the hydrogel substrate. These results are attributed to a larger surface to volume ratio of the electrospun chitosan nanofibers compared to the neat swollen hydrogel.

Functional Nucleic Acids as Bacterial Biosensors: A Focus on Clostridioides difficile Infection

Clostridioides difficile (CD) is a Gram‐positive, anaerobic, and spore‐forming bacillus that colonizes the human gut and causes a range of diseases, such as pseudomembranous colitis and antibiotic‐associated diarrhea, that are generally known as CD infection (CDI). Rapid and accurate detection of CDI with high sensitivity and specificity is crucial for patient treatment, infection control, and epidemiological monitoring. Current diagnostic methods for CDI have several limitations, such as high cost, long turnaround time, suboptimal sensitivity, and the need for specialized equipment. Hence, novel detection methods that can overcome these limitations are needed. Functional nucleic acids (FNAs) are a promising class of molecular recognition element (MRE) that can be incorporated into biosensors for detecting infectious pathogens. Several FNAs have been developed for detecting CD. In this review, an overview of CD, CDI, and current diagnostic methods for CDI and their drawbacks are provided. Furthermore, the design principles and working mechanisms of FNAs as well as their applications for the detection of pathogenic bacteria, including CD, are discussed. The potential for developing point‐of‐care paper sensors using currently available CD‐selective FNAs is also highlighted.

Dual-electrode signal amplification self-powered biosensing platform based on nanozyme boosting target-induced DNA nanospace array for ultrasensitive detection of sugarcane Pokkah Boeng disease pathogenic bacteria

Sugarcane is a crop with significant economic importance worldwide. However, pokkah boeng disease poses a serious threat to its production and the sustainable development. There is a pressing necessity for precise and portable detection methods. We develop a dual-electrode signal amplification biosensing platform, for highly sensitive detection of sugarcane pokkah boeng disease pathogenic bacteria. This innovative platform integrates highly catalytic AuNPs/Mn3O4 nanozymes with N-GDY, along with a target-induced development of DNA nanostructure arrays. AuNPs/N-GDY serves as dual electrode substrates, and AuNPs/Mn3O4 nanozymes are surface-loaded as the bioanode. The biocathode is constructed by introducing DNA nanospace arrays onto the electrode through target-induced methods. [Ru(NH3)6]3+ is embedded into the nucleic acid double-helix scaffold via electrostatic adsorption, generating an EOCV signal that is strongly correlated with the target concentration. To further enhance sensitivity, the detection platform is combined with a capacitor to amplify the detection signal, utilizing its high power density, which results in a 22.5-fold increase in sensitivity. The method offers a linear detection range of 0.0001 to 10,000 pM and an detection limit of 32.5 aM (S/N = 3). This method supplies a novel approach for real-time monitoring and competent oversight of pokkah boeng disease pathogenic bacteria.

CRISPR-Cas12a-based nanoparticle biosensor for detection of pathogenic bacteria in food

Pathogenic bacterial contamination of food is a major challenge in food safety and public health management. To monitor pathogenic bacteria in food, a rapid, sensitive, and specific pathogen detection method is needed. CRISPR-Cas12a has been developed as a sensitive method for detecting pathogenic bacteria, but its output signal is fluorescent, which need equipment to detect. Here, the nanoparticle probe was designed to detect the activity of Cas12a and were able to visualize the assay results by observing the color change by naked eyes. The target sequences of pathogenic bacteria were amplified by recombinase polymerase, then recognized and bound by the crRNA and Cas12a complex. The activated Cas12a cleaved the linker-ssDNA connecting the nanoparticles, causing the nanoparticles to transition from aggregation to dispersion, and the color showed as purple. The sensitivity analysis showed that the limits of detection (LOD) reached 1 × 10°CFU/mL, 1 × 104 CFU/mL, 1 × 102 CFU/mL, 1 × 103 CFU/mL, 1 × 101 CFU/mL for Escherichia coli, Salmonella typhimurium, Vibrio parahaemolyticus, Staphylococcus aureus, and Listeria monocytogenes, which were all specific for five bacteria in the milk samples. Therefore, this method is a dependable, fast, sensitive and specific method, which make the CRIPSR-Cas12a detection method more useful in bacterial pathogenic detection.

Enzyme-accelerated catalytic DNA circuits enable rapid and one-pot detection of bacterial pathogens

Catalytic DNA circuits, serving as signal amplification strategies, can enable simple and accurate detection of pathogenic bacteria in complex matrices but suffer from low reaction rates and depths. Herein, we design an enzyme-accelerated catalytic hairpin assembly (EACHA) in which duplex DNA products are converted into hairpin reactants to continue participating in the next circuit reaction with the assistance of RNase H. Profiting from the high recyclability of the reactants, EACHA exhibits an approximately 37.6-fold enhancement in the rate constant and a two-order-of-magnitude improvement in sensitivity compared to conventional catalytic hairpin assembly (CHA). By integrating an allosteric probe with EACHA, a one-pot method is developed for rapid and direct detection of S. enterica Enteritidis (S. Enteritidis). This method is capable of detecting 15 CFU mL−1 of S. Enteritidis within 20 min, which is superior to that of real-time PCR. By testing 60 milk samples, we demonstrate this method's high accuracy in discriminating contaminated samples, with an area under the curve (AUC) of 0.997. Moreover, this method can be employed to accurately diagnose early-stage infected mice, with an AUC of 1.00 for feces samples and 0.986 for serum samples. Therefore, this study offers a simple and feasible method for identifying pathogens in complex matrices.

Magnetic Fe3O4@MOF@aptamer-mediated entropy-driven fluorescence biosensor for multiplexed and DNA extraction- and amplification-free detection of pathogenic bacteria

Rapid and early identification of pathogenic bacteria in food and clinical samples is critical for infectious disease control and precision medicine. In this study, we present a novel Magnetic Fe3O4@metal–organic framework (MOF)@Aptamer-based Entropy-driven fluorescence biosensor combined with mesophilic Clostridium butyricumArgonaute (CbAgo) for multiplexed and DNA extraction- and amplification-free detection of pathogenic bacteria, termed the MAEA biosensor. In this study, magnetic zirconium-based MOF nanoparticles were modified with a phosphorylated aptamer as an efficient signal converter, which recognized the pathogens and released the complementary DNA to activate the entropy-driven circuit reaction. The highly efficient catalytic DNA circuit and isothermal enzyme-free format produce large amounts of guide DNA. CbAgo exerts precise and highly efficient endonuclease activity under the guidance of guide DNA, which makes the MAEA biosensor useful for multiplexed and DNA extraction- and amplification-free detection of pathogenic bacteria with high sensitivity (<102 CFU/mL). This study demonstrates a highly selective and reliable magnetic Fe3O4@MOF@aptamer-based biosensor using CbAgo for simultaneously detecting multiple pathogenic bacteria, providing a potential tool for other infectious disease pathogens with convenient operation and high sensitivity.

Bioreceptor modified electrochemical biosensors for the detection of life threating pathogenic bacteria: a review.

The lack of reliable and efficient techniques for early monitoring to stop long-term effects on human health is an increasing problem as the pathogenesis effect of infectious bacteria is growing continuously. Therefore, developing an effective early detection technique coupled with efficient and continuous monitoring of pathogenic bacteria is increasingly becoming a global public health prime target. Electrochemical biosensors are among the strategies that can be utilized for accomplishing that goal with promising potential. In recent years, identifying target biological analytes by interacting with bioreceptors modified electrodes is among the most commonly used detection techniques in electrochemical biosensing strategies. The commonly employed bioreceptors are nucleic acid molecules (DNA or RNA), proteins, antibodies, enzymes, organisms, tissues, and biomimetic components such as molecularly imprinted polymers. Despite the advancement in electrochemical biosensing, developing a reliable and effective biosensor for detecting pathogenic bacteria is still in the infancy stage with so much room for growth. A major milestone in addressing some of the issues and improving the detection pathway is the investigation of specific bacterial detection techniques. The present study covers the fundamental concepts of electrochemical biosensors, human PB illnesses, and the latest electrochemical biosensors based on bioreceptor elements that are designed to detect specific pathogenic bacteria. This study aims to assist researchers with the most up-to-date research work in the field of bio-electrochemical pathogenic bacteria detection and monitoring.

Rapid detection and occurrence of foodborne pathogens in minimally processed vegetables: a review

SummaryThe increased consumption of minimally processed vegetables (MPV) in various countries is related to the continued interest of consumers in seeking practical and healthy food items. Due to multiple processing steps, MPV can be contaminated by several foodborne pathogens that pose significant health risks to consumers. The use of rapid techniques to detect pathogens in ready‐to‐eat (RTE) foods such as MPV is therefore essential to provide high quality and safe products. This review aims to provide a comprehensive description of molecular‐based techniques for rapid detection of pathogenic bacteria in MPV, and their occurrence reported in studies published in the last 10 years. The main pathogens detected using rapid methods were Salmonella spp., Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, Shigella spp., and Campylobacter jejuni. Molecular‐based techniques included real‐time polymerase chain reaction (PCR), multiplex PCR, matrix‐assisted laser desorption ionisation time of flight mass spectrometry (MALDI‐TOF MS), and denaturing gradient gel electrophoresis (DGGE). The data indicate high incidences of pathogenic bacteria in MPV, stressing the need for their rapid detection in these products to prevent associated health risks. Further studies should be carried out to increase the sensitivity of molecular‐based techniques and prevent false positives due to undesirable non‐specific PCR amplifications.

Endoprotein-activating DNAzyme assay for nucleic acid extraction- and amplification-free detection of viable pathogenic bacteria

Pathogenic bacteria in food or environment, can pose threats to public health, highlighting the requirement of tools for rapid and accurate detection of viable pathogenic bacteria. Herein, we report a sequential endoprotein RNase H2-activating DNAzyme assay (termed epDNAzyme) that enables nucleic acid extraction- and amplification-free detection of viable Salmonella enterica (S. enterica). The direct detection allows for a rapid detection of viable S. enterica within 25 min. Besides, the assay, based on sequential reporting strategy, circumvents internal modifications in the DNAzyme's active domain and improve its catalytic activity. The multiple-turnover DNAzyme cutting and the enhanced catalytic activity of DNAzyme render the epDNAzyme assay to be highly sensitive, and enables the detection of 190 CFU/mL and 0.1% viable S. enterica. The assay has been utilized to detect S. enterica contamination in food and clinical samples, indicating its potential as a promising tool for monitoring pathogen-associated biosafety.

Precise, Sensitive Detection of Viable Foodborne Pathogenic Bacteria with a 6-Order Dynamic Range via Digital Rolling Circle Amplification.

The presence of viable pathogenic bacteria in food can lead to serious foodborne diseases, thus posing a risk to human health. Here, we develop a digital rolling circle amplification (dRCA) assay that enables the precise and sensitive quantification of viable foodborne pathogenic bacteria. Directly targeting pathogenic RNAs via a ligation-based padlock probe allows for precisely discriminating viable bacteria from dead one. The one-target-one-amplicon characteristic of dRCA enables high sensitivity and a broad quantitative detection range, conferring a detection limit of 10 CFU/mL and a dynamic range of 6 orders. dRCA can detect rare viable bacteria, even at a proportion as low as 0.1%, which is 50 times more sensitive than the live/dead staining method. The high sensitivity for detecting viable bacteria accommodates dRCA for assessing sterilization efficiency. Based on the assay, we found that, for pasteurization, slightly elevating the temperature to 68 °C can reduce the heating time to 10 min, which may minimize nutrient degradation caused by high-temperature exposure. The assay can serve as a precise tool for estimating the contamination by viable pathogenic bacteria and assessing sterilization, which facilitates food safety control.

An NMR immunosensor based on streptavidin-biotin signal amplification for rapid detection of Salmonella in milk

Sensitive and accurate detection of harmful pathogenic bacteria in food is of great significance to public health. In this paper, a nuclear magnetic resonance (NMR) sensor based on biological signal amplification was constructed. The carboxymethyl dextran (CMD) was used as both the probe skeleton and the chelating agent to realize the fast chelating adsorption of the signal source gadolinium ion. Finally, the magnetic complex and biotinylated antibody formed an immune probe through the biological bridge to achieve rapid capture of Salmonella. In addition, non-target bacteria such as Proteus vulgaris and Listeria monocytogenes were used to test the specificity of the constructed sensor. The sensor can detect Salmonella within 1.5 h under optimal conditions, the detection limit in milk is 7.4 × 103 CFU mL−1, and the RSD (Relative standard deviation) of reproducibility experiment is 2.35%, which has good reproducibility.

UV-ozone treated glass fiber based lateral flow DNA extraction platform integrated with LAMP for rapid detection of pathogen bacteria in whole blood

The need for a fast and reliable platform for the detection of pathogens in point-of-care (POC) applications is increasing day by day. Molecular detection of pathogens is made possible by a good DNA isolation method, which allows for efficient and pure extraction from the cell. To address this need, our work presents a new lateral flow platform based on UV ozone-treated glass fiber pads for high-throughput DNA isolation from very low volumes of bacterial samples. The platform we developed mainly consists of UV ozone-treated glass fiber, which serves as the essential component binding DNA, and a cellulose-based absorbent pad acting as a reservoir for washing buffers and undesired substances from bacterial cells. UV ozone treatment increased the DNA recovery efficiency of glass fiber by 2.5 times compared to untreated glass fiber. qPCR results confirmed the isolation of pure DNA from both gram-negative and gram-positive bacteria samples performed on the developed platform. For five different bacteria concentrations (10–109 cfu/mL), a good linearity between Cq values and bacteria concentrations was obtained. A minimum concentration of 10 cfu/mL could be detected for both bacterial samples of 10 µL. The developed DNA extraction platform was also integrated with a portable heater for the LAMP-based detection application. We successfully demonstrated that the proposed extraction platform, combined with the colorimetric LAMP-based system, can detect Staphylococcus aureus bacteria spiked in whole blood samples in under one hour, including both extraction and detection processes. Our simplified paper-based platform will be a good alternative for the DNA extraction and molecular detection of pathogens without the need for complex laboratory equipment in POC applications.

Antimicrobial Peptide-Based Sensor for the Electrochemical Detection of Methicillin-Resistant Staphylococcus Aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is a gram-positive bacterium that colonizes on the skin and nasal track of humans, which can lead to serious illnesses such as skin and soft tissue infections, pneumonia, and septicemia.MRSA is resistant to an entire class of antibiotics called beta-lactams, including penicillin, amoxicillin, and oxacillin, making it more difficult to treat. Culture based methods and molecular based methods like PCR are currently being used for the detection of MRSA, but these methods possess the drawbacks of being slow and expensive, respectively. Thus, we focused on developing a reagentless, cost-effective, near real-time detection method for intact MRSA cells.Antimicrobial peptides (AMPs) possess the ability to interact with bacterial cells by inhibiting their growth or by lysing them.However, this initial interaction between the cationic AMPs and the negatively charged bacterial cells can be used for the detection of pathogens like MRSA. Here, we used a MRSA-specific antimicrobial peptide, DP7, to fabricate an electrochemical peptide-based (E-PB) sensor. The DP7 probe was further modified with a methylene blue redox label and an alkanethiol chain which allows for the formation of a thiol-gold self-assembled monolayer. The sensor surface was passivated with both 6-mercapto-1-hexanol and a thiolated tetra-thymine sequence to prevent surface fouling. Electrochemical detection of MRSA was accomplished using alternating current voltammetry. Control experiments were performed using two gram-positive bacteria, Bacillus subtilis and Staphylococcus epidermidis, and two gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa. The DP7 sensor responded to MRSA in a concentration-dependent manner and was proven to be selective for MRSA over the other four microbes. Specifically, the % signal suppression obtained for MRSA was much larger when compared to those obtained for the other microbes. The same behavior was observed in pure buffer and 5% synthetic wound fluid. In addition, the detection time was relatively short, with signal saturation in less than 60 min. Overall, the DP7 sensor has demonstrated to possess attributes similar to other E-PB sensors. It has the potential to be developed as a rapid, reliable, and cost-effective diagnostic tool for MRSA, which will be useful in preventing the spread of the pathogen in the community. Furthermore, this sensing platform is versatile and can be redesigned for detection of other pathogenic bacteria.

Swift and precise detection of unlabeled pathogens using a nanogap electrode impedimetric sensor facilitated by electrokinetics

For the protection of human health and environment, there is a growing demand for high-performance, user-friendly biosensors for the prompt detection of pathogenic bacteria in samples containing various substances. We present a nanogap electrode-based purely electrical impedimetric sensor that utilizes the dielectrophoresis (DEP) mechanism. Our nanogap sensor can directly and sensitively detect pathogens present at concentrations as low as 1–10 cells/assay in buffers and drinking milk without the need for separation, purification, or specific ligand binding. This is achieved by minimizing the electrical double-layer effect and electrode polarization in nanogap impedance sensors, reducing signal loss. In addition, even at low DEP voltages, nanogap sensors can quickly establish strong DEP forces between the nanogap electrodes to control the spatial concentration of pathogens around the electrodes. This activates and stabilizes inter-electrode signal transmission along the nanogap-aligned pathogens, increasing sensitivity and reducing errors during repeated measurements. The DEP-enabled nanogap impedance sensor developed in this study is valuable for a variety of pathogen detection and monitoring systems including point-of-care testing (POCT) as it can detect pathogens in diverse samples containing multiple substances quickly and with high sensitivity, is compatible with complex solutions such as food and beverages, and provides highly reproducible results without the need for separate binding and separation processes.