Detection Of Waterborne Pathogens Research Articles

Label-Free Detection of Waterborne Pathogens Using an All-Solid-State Laser-Induced Graphene Potentiometric Ion Flux Immunosensor.

Waterborne pathogens are harmful microorganisms transmitted through water sources. Early and rapid pathogen detection is important for preventing illnesses and implementing stringent water safety measures to minimize the risk of contamination. This work introduces a miniaturized all-solid-state potentiometric ion flux immunosensor for the rapid and label-free detection of waterborne pathogens. A screen-printed silver/silver chloride electrode coated with a reference electrode membrane and polyurethane as an all-solid-state reference electrode was combined with a solid-state contact ion-selective electrode (ISE). An all-solid-state ISE was constructed on laser-induced graphene by coating it with a cationic marker and a carboxylated poly(vinyl chloride)-based membrane for immobilizing antibodies and controlling ion fluxes through the membrane. Proof-of-concept was achieved by detecting Escherichia coli and Salmonella enterica serovar Typhimurium using the assembled immunosensors within 10 min. The potentiometric response shift attributed to the blocking effect in the ion flux caused by pathogen-antibody interaction corresponded to pathogen concentration, indicating detection limits of 0.1 CFU/mL and working ranges of 0.1-105 CFU/mL. Furthermore, the developed sensors revealed high selectivity and were directly applied in groundwater and tap water without any sample preparation, demonstrating high recovery percentages. The simple operation and elimination of sample preparation are key benefits to further usability of the developed immunosensors for efficient pathogen detection.

Development and evaluation of test methods for the detection and enumeration of opportunistic waterborne pathogens from the hospital environment

Many Gram-negative bacteria other than Pseudomonas aeruginosa have been implicated in waterborne outbreaks, but standardized laboratory detection methods for these organisms have not been established. This study aimed to establish laboratory testing methodologies for six waterborne pathogens: Acinetobacter spp., Burkholderia spp., Cupriavidus spp., Delftia acidovorans, Elizabethkingia spp. and Stenotrophomonas maltophilia. Water samples were spiked by UK Health Security Agency laboratories and sent to the Glasgow Royal Infirmary laboratory for analysis. Water samples were spiked with either a pure culture of target organism or the target organism in water containing normal background flora, to ensure that the methodology could identify organisms from a mixed culture. Volumes of 100mL were filtered under negative pressure on to culture media and incubated at 30°C and 37°C. The incubation time was 7 days, with plates read on days 2, 5 and 7. Further identification of colonies was undertaken using matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Optimal recovery of organisms was obtained by culturing water samples on tryptic soy agar, chocolate bacitracin agar and pseudomonas selective agar. The optimal temperature for isolation was 30°C. The optimal incubation time was 5 days, and MALDI-TOF MS identified all test species reliably. The methodology described was able to detect the six tested waterborne pathogens reliably, and can be utilized by laboratories involved in testing water samples during outbreak investigations.

Development of an Immunocapture-Based Polymeric Optical Fiber Sensor for Bacterial Detection in Water.

Conventional methods for pathogen detection in water rely on time-consuming enrichment steps followed by biochemical identification strategies, which require assay times ranging from 24 hours to a week. However, in recent years, significant efforts have been made to develop biosensing technologies enabling rapid and close-to-real-time detection of waterborne pathogens. In previous studies, we developed a plastic optical fiber (POF) immunosensor using an optoelectronic configuration consisting of a U-Shape probe connected to an LED and a photodetector. Bacterial detection was evaluated with the immunosensor immersed in a bacterial suspension in water with a known concentration. Here, we report on the sensitivity of a new optoelectronic configuration consisting of two POF U-shaped probes, one as the reference and the other as the immunosensor, for the detection of Escherichia coli. In addition, another methos of detection was tested where the sensors were calibrated in the air, before being immersed in a bacterial suspension and then read in the air. This modification improved sensor sensitivity and resulted in a faster detection time. After the immunocapture, the sensors were DAPI-stained and submitted to confocal microscopy. The histograms obtained confirmed that the responses of the immunosensors were due to the bacteria. This new sensor detected the presence of E. coli at 104 CFU/mL in less than 20 min. Currently, sub-20 min is faster than previous studies using fiber-optic based biosensors. We report on an inexpensive and faster detection technology when compared with conventional methods.

Molecular detection of waterborne pathogens in infants' drinking water and their relationship with water quality determinants in eastern Ethiopia: loop-mediated isothermal amplification (LAMP)-based study.

Cryptosporidium, Shigella, toxin-producing Escherichia coli, and rotavirus were reported to be the most responsible for severe and fatal diarrhea among infants. This study aimed to investigate the presence of these pathogens in infants' drinking water samples and analyzing using water quality determinants in eastern Ethiopia. A molecular (LAMP)-based cross-sectional study design was employed. A total of 410 and 37 water samples were tested from infant point-of-use at household and corresponding water source, respectively, from June 2020 to May, 2021. Cryptosporidium, Shigella, toxin-producing E. coli, and rotavirus were detected in 28.5, 30.0, 26.3, and 32.2%, of water samples tested from infant point-of-use, respectively. About 13.2% of the water samples were positive for all (four) pathogens together. Cryptosporidium, Shigella, toxin-producing E. coli, and rotavirus were detected in 27.0, 32.4, 29.7, and 37.8%, of water samples tested from water sources, respectively. Positive significant correlation was observed between infant point-of-consumption and water sources from which it is drawn toward the presence of each targeted pathogen. Unimproved water source showed a strong significant association with the presence of Cryptosporidium, Shigella and toxin-producing E. coli. Therefore, efforts should be made in development of improved water sources, source protection safety and health education to caretakers of infants.

On-site monitoring of Escherichia coli O157:H7 in drinking water based on rapid detection of the rfbE gene at the single copy level

The sensitive and on-site detection of highly pathogenic Escherichia coli O157:H7 (E. coli O157:H7) is of great importance to ensure public health. However, conventional methods for E. coli O157:H7 detection may suffer disadvantages such as high expense and time consumption. Herein, we reported a cost-effective colorimetric platform based on loop-mediated isothermal amplification (LAMP) for ultrasensitive and rapid detection of E. coli O157:H7. In this study, a novel quantitative colorimetric LAMP assay was established by in-situ measurement of hydroxy naphthol blue (HNB) color change with a designed portable device, which was applied to the amplification of the specific rfbE gene of E. coli O157:H7. As expected, this method featured a wide linear range (100-104 copies µL−1) with excellent linear regression analysis (R2 > 0.99) and possessed a record-low detection limit of 1 copy µL−1 to lower the risk of missed detection. The proposed strategy could be employed on drinking water samples with favorable recoveries and also demonstrated potential in the analysis of complex samples. This work exhibits the promising application prospect in on-site detection without the need for expensive instrumentation and seeks to provide an original paradigm for the rapid and sensitive detection of waterborne pathogens.

A novel culture-enriched metagenomic sequencing strategy effectively guarantee the microbial safety of drinking water by uncovering the low abundance pathogens

Assessing the presence of waterborne pathogens and antibiotic resistance genes (ARGs) is crucial for managing the environmental quality of drinking water sources. However, detecting low abundance pathogens in such settings is challenging. In this study, a workflow was developed to enrich for broad spectrum pathogens from drinking water samples. A mock community was used to evaluate the effectiveness of various enrichment broths in detecting low-abundance pathogens. Monthly metagenomic surveillance was conducted in a drinking water source from May to September 2021, and water samples were subjected to five enrichment procedures for 6 h to recover the majority of waterborne bacterial pathogens. Oxford Nanopore Technology (ONT) was used for metagenomic sequencing of enriched samples to obtain high-quality pathogen genomes. The results showed that selective enrichment significantly increased the proportions of targeted bacterial pathogens. Compared to direct metagenomic sequencing of untreated water samples, targeted enrichment followed by ONT sequencing significantly improved the detection of waterborne pathogens and the quality of metagenome-assembled genomes (MAGs). Eighty-six high-quality MAGs, including 70 pathogen MAGs, were obtained from ONT sequencing, while only 12 MAGs representing 10 species were obtained from direct metagenomic sequencing of untreated water samples. In addition, ONT sequencing improved the recovery of mobile genetic elements and the accuracy of phylogenetic analysis. This study highlights the urgent need for efficient methodologies to detect and manage microbial risks in drinking water sources. The developed workflow provides a cost-effective approach for environmental management of drinking water sources with microbial risks. The study also uncovered pathogens that were not detected by traditional methods, thereby advancing microbial risk management of drinking water sources.

Spectroscopic Methods for the Detection of Microbial Pathogens and Diagnostics of Infectious Diseases—An Updated Overview

Microbial pathogens cause a quarter of all deaths worldwide annually due to deadly infectious diseases. Nevertheless, the fast and precise identification of pathogens remains one of the most challenging tasks in the medical sector. Early identification and characterization of microbes through medical diagnosis could pave the way for specific treatment strategies that could dramatically improve infection management, reduce healthcare costs, mitigate increasing antimicrobial resistance, and save numerous lives. To date, numerous traditional and molecular methods have been employed to diagnose illnesses with proven accuracy, reliability, and efficiency. Here, we have reviewed the most reliable tools that are prerequisites for the rapid detection of microbes. In particular, the remarkable roles of surface-enhanced Raman scattering, Fourier-transform infrared, electrochemical impedance, near-infrared, and MALDI-TOF/TOF in the identification and characterization of pathogenic microbes are discussed in detail. The approaches described herein cover broad ranges of biomedical applications, including the diagnosis of clinical infectious diseases, epidemiology, detection of vector-borne diseases, food security, phytosanitary monitoring, biosensing, and food- and waterborne pathogen detection. Considering the current pandemic outbreak, this review briefly emphasizes the importance of rapid detection and upgraded tools for early diagnosis to prevent the loss of lives.

Low complexity and accurate Machine learning model for waterborne pathogen classification using only three handcrafted features from optofluidic images

Pathogenic microorganism can cause human diseases and even death, so waterborne pathogen detection is essential for water quality monitoring. Conventional microorganism detection/classification through biochemical approaches is expensive, time-consuming, labour-intensive and leaves chemical pollutions. Recently, many image processing algorithms are developed for cheaper, faster, more automatic, and more environmentally friendly microorganism classification. However, most existing classification algorithms are designed for microscopic images and focus only on the classification performance, not the computational cost. Alternatively, the optofluidic systems provide a smaller and cheaper way to capture microorganism images than conventional microscopes. Thus, in this paper, an image classification algorithm based on optofluidic images is proposed for portable, real-time water quality monitoring applications. With a comprehensive investigation on different categories of features and careful analysis on our optofluidic images, 2 morphological features were selected to classify the waterborne pathogens with a proposed feature, EHHOG (entropy of histogram of HOG) (HOG: histogram of oriented gradients). The proposed algorithm can achieve higher or similar classification accuracy as existing methods, while the computational cost is much lower with only 3 features.

A microfluidic biosensor for rapid simultaneous detection of waterborne pathogens

A microfluidic based biosensor was investigated for rapid and simultaneous detection of Salmonella, Legionella, and Escherichia coli O157:H7 in tap water and wastewater. The biosensor consisted of two sets of focusing electrodes connected in parallel and three sets of interdigitated electrodes (IDE) arrays. The electrodes enabled the biosensor to concentrate and detect bacteria at both low and high concentrations. The focusing region was designed with vertical metal sidewall pairs and multiple tilted thin-film finger pairs to generate positive dielectrophoresis (p-DEP) to force the bacteria moving toward the microchannel centerline. As a result, the bacterial pathogens were highly concentrated when they reached the detection electrode arrays. The detection IDE arrays were coated with three different antibodies against the target bacterial pathogens and a cross-linker to enhance the binding of antibodies to the detection electrode. As the binding of bacterial pathogen to its specific antibodies took place, the impedance value changed. The results demonstrated that the biosensors were capable of detecting Salmonella, Legionella, and E. coli 0157:H7 simultaneously with a detection limit of 3 bacterial cells/ml in 30 − 40 min.

Development of a Multiplex Lateral Flow Assay for Rapid Detection of Waterborne Pathogens

Development of a Multiplex Lateral Flow Assay for Rapid Detection of Waterborne Pathogens

Microorganism image classification with circle-based Multi-Region Binarization and mutual-information-based feature selection

BackgroundAn important part of water quality control is waterborne pathogen detection. Conventional microorganism detections are done by costly and complicated biochemical approaches, which leave chemical pollutions and require professional laboratories, technicians, and much time. Recently, machine-learning-based image classification algorithms are developed for faster, cheaper, simpler, and more environmentally friendly microorganism detection. Most of them are designed for conventional microscopic images. On the other hand, the optofluidic system is a smaller and cheaper alternative to conventional microscopes, so we aim to propose a microorganism classification algorithm for optofluidic images. MethodIn our previous study, the Binarized-Greyscale-Hybrid Algorithm with Multi-Region Binarization (BiGHAM) algorithm was developed for our optofluidic image dataset. In this paper, a new circle-based region partition method for the Multi-Region Binarization and a new mutual-information-based feature selection method were proposed to improve the BiGHAM method. Additionally, an implementation example of how to adapt and apply our method on an open dataset was presented. Results and conclusionWith the circle-based region partition method and the new feature combination selected based on mutual information, the improved-BiGHAM algorithm achieved higher classification accuracies, and the algorithm is simpler than the previous BiGHAM method during training. For our optofluidic image dataset, the accuracy improvement for the 1000-image-per-class case was from 94.8 to 96.3%, and for the 30-image-per-class case was from 77.8 to 85.6% with the support vector machine as the classifier. The adapted algorithm also works well on the open dataset, showing the potential to be extended to applications on different microorganism images.

Principles and Recent Advances in Biosensors for Pathogens Detection

AbstractFoodborne and waterborne diseases caused by pathogens pose a serious threat to human life, and the detection of such pathogens in food, water, and beverages has gained paramount importance in view of the increasing number of pathogenic diseases in recent times. Standard and traditional analytical techniques employed for the recognition of deadly pathogens, including polymerase chain reaction‐based techniques, immunology‐based assays, culture and colony counting techniques, require several hours or perhaps even a few days for the results. All these techniques, despite being quite sensitive, are time‐intensive. Therefore, several researchers are focusing on the development of rapid pathogen detection techniques. Although the most advanced biosensing technologies exhibit potential approaches, significant research and testing is an urgent need to design innovative biosensors. Novel bioanalytical approaches for the recognition and quantification of target pathogens are being designed and developed to enhance the characteristics of biosensors, including rapid response, selectivity, and sensitivity. In this context, we not only provide an overview of the types of biorecognition elements employed for the detection of pathogens, but also discuss in detail the traditional approaches, biomolecular techniques, the latest advances in the detection and quantification of foodborne and waterborne pathogens, with particular emphasis on biosensors.

A Whole-Cell Biosensor for Point-of-Care Detection of Waterborne Bacterial Pathogens.

Water contamination by pathogenic bacteria is a major public health concern globally. Monitoring bacterial contamination in water is critically important to protect human health, but this remains a critical challenge. Engineered whole-cell biosensors created through synthetic biology hold great promise for rapid and cost-effective detection of waterborne pathogens. In this study, we created a novel whole-cell biosensor to detect water contamination by Pseudomonas aeruginosa and Burkholderia pseudomallei, which are two critical bacterial pathogens and are recognized as common causative agents for waterborne diseases. The biosensor detects the target bacterial pathogens by responding to the relevant quorum sensing signal molecules. Particularly, this study constructed and characterized the biosensor on the basis of the QscR quorum sensing signal system for the first time. We first designed and constructed a QscR on the basis of the sensing module in the E.coli host cell and integrated the QscR sensing module with a reporting module that expressed an enhanced green fluorescent protein (EGFP). The results demonstrated that the biosensor had high sensitivity in response to the quorum sensing signals of the target bacterial pathogens. We further engineered a biosensor that expressed a red pigment lycopene in the reporting module to produce a visible signal readout for the pathogen detection. Additionally, we investigated the feasibility of a paper-based assay by immobilizing the lycopene-based whole-cell biosensor on paper with the aim to build a prototype for developing portable detection devices. The biosensor would provide a simple and inexpensive alternative for timely and point-of-care detection of water contamination and protect human health.

Aptamer-based approaches for the detection of waterborne pathogens.

Waterborne ailments pose a serious threat to public health and are a huge economic burden. Lack of hygiene in drinking and recreational water is the chief source of microbial pathogens in developing countries. Poor water quality and sanitation account for more than 3.4 million deaths a year worldwide. This has urged authorities and researchers to explore different avenues of pathogen detection. There is a growing demand for rapid and reliable sensor technologies, in particular those that can detect in situ and perform in harsh conditions. Some of the major waterborne pathogens include Vibrio cholerae, Leptospira interrogans, Campylobacter jejuni, Shigella spp., enterotoxigenic Escherichia coli, Clostridium difficile, Cryptosporidium parvum, Entamoeba histolytica, and Hepatitis A virus. While conventional methods of pathogen detection like serodiagnosis and microbiological methods have been superseded by nucleic acid amplification methods, there is still potential for improvement. This review provides an insight into aptamers and their utility in the form of aptasensors. It discusses how aptamer-based approaches have emerged as a novel strategy and its advantages over more resource-intensive and complex biochemical approaches.

Wastewater-based epidemiology-surveillance and early detection of waterborne pathogens with a focus on SARS-CoV-2, Cryptosporidium and Giardia.

Waterborne diseases are a major global problem, resulting in high morbidity and mortality, and massive economic costs. The ability to rapidly and reliably detect and monitor the spread of waterborne diseases is vital for early intervention and preventing more widespread disease outbreaks. Pathogens are, however, difficult to detect in water and are not practicably detectable at acceptable concentrations that need to be achieved in treated drinking water (which are of the order one per million litre). Furthermore, current clinical-based surveillance methods have many limitations such as the invasive nature of the testing and the challenges in testing large numbers of people. Wastewater-based epidemiology (WBE), which is based on the analysis of wastewater to monitor the emergence and spread of infectious disease at a population level, has received renewed attention in light of the current coronavirus disease 2019 (COVID-19) pandemic. The present review will focus on the application of WBE for the detection and surveillance of pathogens with a focus on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the waterborne protozoan parasites Cryptosporidium and Giardia. The review highlights the benefits and challenges of WBE and the future of this tool for community-wide infectious disease surveillance.

Αίτια και Επιδημιολογικά Δεδομένα Υδατογενών Λοιμώξεων

Waterborne infections are caused due to pathogenic microorganisms, mainly by bacteria, parasites and viruses. As an epidemiological criterion, we evaluate the presence in two or more individuals with similar symptoms after con- sumption of potable water or exposure to water that is used for relaxation purposes. Most waterborne epidemies (outbreaks) caused by microorganisms show a seasonal distribution, with a higher incidence in summer. Worldwide, the observed increase in waterborne infections has been attributed to factors such as overcrowding and urbanization, mass displacement of societies, aging of the population, increase of immunosuppressed individuals, changes in life style, as well as the climate changes. More-over, reasons are the aging water-distribution, wastewater, and sanitation systems, and evidently the improvement of methods in recording and laboratory detection of water-borne pathogens. The effort that has taken place over the past years for more thorough laboratory testing of clinical and environmental samples taken in the framework of epidemies should be go on with undeniable interest.

Simultaneous detection of Salmonella spp., Pseudomonas aeruginosa, Bacillus cereus, and Escherichia coli O157:H7 in environmental water using PMA combined with mPCR.

A multiplex polymerase chain reaction (mPCR) with propidium monoazide (PMA) and internal amplification control (IAC) for the simultaneous detection of waterborne pathogens Salmonella spp., Pseudomonas aeruginosa, Bacillus cereus, and Escherichia coli O157:H7, was developed. This PMA-IAC-mPCR assay used four new specific primers based on the genes for invA, ecfX, cesB, and fliC, respectively. A 16S rRNA primer was chosen for IAC to eliminate false negative results. The photosensitive dye, propidium monoazide (PMA) was used to exclude signals from dead bacteria that could lead to false positive results. In pure culture, the limits of detection (LOD) were 101 CFU/ml for P. aeruginosa, 102 CFU/ml for both Salmonella spp. and E. coli O157:H7, and 103 CFU/ml for B. cereus, respectively. In addition, with a 6-8 h enrichment of all four bacteria that were combined in a mixture that was spiked in water sample matrix, the LOD was 3 CFU/ml for Salmonella spp., 7 CFU/ml for E. coli O157:H7, 10 CFU/ml for B. cereus and 2 CFU/ml for P. aeruginosa. This PMA-IAC-mPCR assay holds potential for application in the multiplex assay of waterborne pathogens.

Recent Progress on the Electrochemical Biosensing of Escherichia coli O157:H7: Material and Methods Overview.

Escherichia coli O157:H7 (E. coli O157:H7) is a pathogenic strain of Escherichia coli which has issued as a public health threat because of fatal contamination of food and water. Therefore, accurate detection of pathogenic E. coli is important in environmental and food quality monitoring. In spite of their advantages and high acceptance, culture-based methods, enzyme-linked immunosorbent assays (ELISAs), polymerase chain reaction (PCR), flow cytometry, ATP bioluminescence, and solid-phase cytometry have various drawbacks, including being time-consuming, requiring trained technicians and/or specific equipment, and producing biological waste. Therefore, there is necessity for affordable, rapid, and simple approaches. Electrochemical biosensors have shown great promise for rapid food- and water-borne pathogen detection. Over the last decade, various attempts have been made to develop techniques for the rapid quantification of E. coli O157:H7. This review covers the importance of E. coli O157:H7 and recent progress (from 2015 to 2020) in the development of the sensitivity and selectivity of electrochemical sensors developed for E. coli O157:H7 using different nanomaterials, labels, and electrochemical transducers.

Ultrafast detection of infectious bacteria using optoelectronic nose based on metallic nanoparticles

Developing an accurate and sensitive method for detection of waterborne pathogens and urinary tract infection are important environmental and clinical challenges. We report a colorimetric assay based on arrays of nanoparticles for identification of 10 infectious bacteria strains belonging to both gram positive and gram-negative species. The detection system works based on the interaction of the volatile metabolites emitted from bacteria with the arrays of metallic nanoparticles deposited on paper substrate. The color changes, monitored by a flatbed scanner, are analyzed by statistical pattern recognition methods. A unique colorimetric pattern is achieved for each bacteria strain. We could detect bacteria at very low concentrations (around 100 bacteria/mL) in less than 10 min, which is the fastest method reported ever. The sensor can be used successfully for detection of bacteria in drinking water as well as human urine (containing 177 healthy and 123 patient samples) in less than 50 min, which is dramatically faster than urine culture. The provided detection system opens a new way to develop simple, small, sensitive and rapid devices for detecting pathogens in various clinical samples.

Integrated Electrochemical Biosensors for Detection of Waterborne Pathogens in Low-Resource Settings.

More than 783 million people worldwide are currently without access to clean and safe water. Approximately 1 in 5 cases of mortality due to waterborne diseases involve children, and over 1.5 million cases of waterborne disease occur every year. In the developing world, this makes waterborne diseases the second highest cause of mortality. Such cases of waterborne disease are thought to be caused by poor sanitation, water infrastructure, public knowledge, and lack of suitable water monitoring systems. Conventional laboratory-based techniques are inadequate for effective on-site water quality monitoring purposes. This is due to their need for excessive equipment, operational complexity, lack of affordability, and long sample collection to data analysis times. In this review, we discuss the conventional techniques used in modern-day water quality testing. We discuss the future challenges of water quality testing in the developing world and how conventional techniques fall short of these challenges. Finally, we discuss the development of electrochemical biosensors and current research on the integration of these devices with microfluidic components to develop truly integrated, portable, simple to use and cost-effective devices for use by local environmental agencies, NGOs, and local communities in low-resource settings.