This paper presents a portable magnetoelastic (ME) biosensor system that enables rapid, on-site detection of pathogenic bacteria. The system utilizes a patented portable resonant frequency analyzer and two 1 mm long ME biosensors (biosensors coated with and without phage binding a specific pathogen) enabling real-time measurement of resonant frequency changes. By comparing the response of the biosensors, the presence of the specific pathogen can be detected. In this work, detection of Salmonella Typhimurium cells was demonstrated, and it was found that down to 2,500 cfu can be detected in less than 10 min. The detection limit can be improved by using a smaller sensor (e.g., 500 um long sensors) and an optimal chamber design increasing the probability of bacterial cells striking the biosensor surface.
To perform rapid sensing of pathogens on the surface of food or food preparing plates, ME wireless biosensing system was combined with surface swab sampling techniques in this research. The ME biosensors which consist of ME resonators E2 phage was generally used for Salmonella typhimurium direct detections on the surfaces. E2 phage used in this research was designed for Salmonella typhimurium specific binding. Instead of measuring one spot at a time, the desired area or the whole area of a target surface can be swabbed for the inexpensive, rapid and easy-to-use pathogen collections. In this study, we first investigated the efficiency of capture and release of a model pathogen, Salmonella Typhimurium, by swab sampling on wet or dry surfaces. Plate counting was used to identify the recovery rates. The efficiency of capture and release was calculated and compared between various kinds of swabs which were composed of different tip materials, including cotton, rayon, and nylon-flocked ones.
Listeria monocytogenes is the major etiologic agent for foodborne Listeriosis in humans from consumption of readyto- eat (RTE) food. According to Center for Disease Control and Prevention, an estimated 1,600 people contract Listeriosis each year with approximately 260 deaths. This high rate of mortality has alerted the Food Safety Inspection and Services to release the Notice 23-99, Instructions for Verifying the L. monocytogenes Reassessment, on August 3, 1999 for their inspectors. According to the FDA’s Bacterial Analysis Manual Chapter 10, L. monocytogenes in RTE food samples is detected via microbiological culture-based tests, qPCR, pulsed-field gel electrophoresis, and other alternative methods. Unfortunately, these methods are time consuming (48-72 hours) and require dedicated laboratory facility. Thus, to develop a real-time L. monocytogenes biosensor, we isolated L. monocytogenes specific oligopeptides displayed on bacteriophages using modified biopanning procedures. In order to account for major temperature dependent morphological alterations of L. monocytogenes at 4°C versus 37°C, we used bacterial cells adapted to either temperature as the target in our biopanning. To date, we have isolated several candidate probes that can recognize either cold-adapted, warm-adapted L. monocytogenes cells, or both types of bacterial cells. Our isolated probes will be used on the magnetoelastic biosensor platforms for real-time detection of L. monocytogenes in RTE foods stored at 4°C or in samples/fluids for bacterium adapted to human body temperature.
Foodborne illness is a common public health problem because food can be contaminated with pathogens at any point in the farm-to-table continuum. This paper presents a method of capturing a quantity of a specific bacterial pathogen in a large volume of liquid using a biomolecular recognition filter. The filter consists of support frames made of a soft magnetic material and solenoid coils for magnetization/demagnetization of the frames. This filter is a planar, multi-layered arrangement of strip-shaped, phage-immobilized magnetoelastic (ME) biosensors that are magnetically held and arrayed on the filter frames. As a large volume of liquid passes through the biomolecular filter, the pathogen of interest is captured by the phage immobilized ME biosensors. This biomolecular filter is designed to capture a specific pathogen and allow non-specific debris to pass, thus avoiding a common clogging issue in conventional bead filters. In this work, single layer, double layers and triple layers of filter were test to capture Salmonella Typhimurium in a large volume of water. The effects of multiplication of filter layers on Salmonella capture efficiency will be discussed.
This paper investigates the accuracy of surface-scanning measurement of a wireless magnetoelastic (ME) biosensor for direct pathogen detection on solid surfaces. The model experiments were conducted on the surface of a at polyethylene (PE) plate. An ME biosensor (1 mm x 0.2 mm x 30 µm) was placed on the PE surface, and a surface-scanning detector was aligned to the sensor for wireless resonant frequency measurement. The position of the detector was accurately controlled by using a motorized three-axis translation system (i.e., controlled X, Y, and Z positions). The results showed that the resonant frequency variations of the sensor were -125 to +150 Hz for X and Y detector displacements of ± 600 µm and Z displacements of +100 to +500 µm. These resonant frequency variations were small compared to the sensor's initial resonant frequency (˂ 0.007% of 2.2 MHz initial resonant frequency) measured at the detector home position, indicating high accuracy of the measurement. In addition, the signal amplitude was, as anticipated, found to decrease exponentially with increasing detection distance (i.e., Z distance). Finally, additional experiments were conducted on the surface of cucumbers. Similar results were obtained.
In this paper, a novel device named as phage filter is designed and presented to capture and identify a small number of Salmonella Typhimurium cells from large volumes of water. This phage filter is constructed from a filter chamber, filter frames on a spindle, strip-shape magnetoelastic filter elements, and a spinning speed control unit. The filter elements are made from Metglas 2826MB and coated with a specifically designed phage that only binds with Salmonella Typhimurium. These phage-coated filter elements can be held and arranged on the filter frames by magnetic force produced from couples of permanent magnets in the frame. Layers of filter frames are fixed on the spindle. The spindle with filter frames and filter elements can spin in the filter chamber and the spinning speed can be continuously adjusted. When the filter works, the tested water passes through the filter frame, and Salmonella Typhimurium cells striking on the filter elements can be bound by the phage on the element surfaces and removed from the tested water.
This paper demonstrates a highly sensitive surface-scanning detector used for magnetoelastic (ME) biosensors for the detection of Salmonella on the surface of a polyethylene (PE) food preparation surface. The design and fabrication methods of the new planar spiral coil are introduced. Different concentrations of Salmonella were measured on the surface of a PE board. The efficacy of Salmonella capture and detection is discussed.
This paper investigates the effects of surface-scanning detector position on the resonant frequency and signal amplitude of a wireless magnetoelastic (ME) biosensor for direct pathogen detection on solid surfaces. The experiments were conducted on the surface of a flat polyethylene (PE) plate as a model study. An ME biosensor (1 mm × 0.2 mm × 30 μm) was placed on the PE surface, and a surface-scanning detector was brought close and aligned to the sensor for wireless resonant frequency measurement. The position of the detector was accurately controlled by using a motorized three-axis translation system (i.e., controlled X, Y, and Z positions). The results showed that the resonant frequency variations of the sensor were -125 to +150 Hz for X and Y detector displacements of ±600 μm and Z displacements of +100 to +500 μm. These resonant frequency variations were small compared to the sensor's initial resonant frequency (< 0.007% of 2.2 MHz initial resonant frequency) measured at the detector home position, indicating high accuracy of the measurement. In addition, the signal amplitude was, as anticipated, found to decrease exponentially with increasing detection distance (i.e., Z distance). Finally, additional experiments were conducted on the surface of cucumbers. Similar results were obtained.
Phage based magneto-elastic (ME) biosensors have been shown to be able to rapidly detect Salmonella in various food systems to serve food pathogen monitoring purposes. In this ME biosensor platform, the free-standing strip-shaped magneto-elastic sensor is the transducer and the phage probe that recognizes Salmonella in food serves as the bio-recognition element. According to Sorokulova et al. at 2005, a developed oligonucleotide probe E2 was reported to have high specificity to Salmonella enterica Typhimurium. In the report, the specificity tests were focused in most of Enterobacterace groups outside of Salmonella family. Here, to understand the specificity of phage E2 to different Salmonella enterica serotypes within Salmonella Family, we further tested the specificity of the phage probe to thirty-two Salmonella serotypes that were present in the major foodborne outbreaks during the past ten years (according to Centers for Disease Control and Prevention). The tests were conducted through an Enzyme linked Immunosorbent Assay (ELISA) format. This assay can mimic probe immobilized conditions on the magnetoelastic biosensor platform and also enable to study the binding specificity of oligonucleotide probes toward different Salmonella while avoiding phage/ sensor lot variations. Test results confirmed that this oligonucleotide probe E2 was high specific to Salmonella Typhimurium cells but showed cross reactivity to Salmonella Tennessee and four other serotypes among the thirty-two tested Salmonella serotypes.
This paper investigates a phage-based biomolecular filter that enables the evaluation of large volumes of liquids for the presence of small quantities of bacterial pathogens. The filter is a planar arrangement of phage-coated, strip-shaped magnetoelastic (ME) biosensors (4 mm × 0.8 mm × 0.03 mm), magnetically coupled to a filter frame structure, through which a liquid of interest flows. This "phage filter" is designed to capture specific bacterial pathogens and allow non-specific debris to pass, eliminating the common clogging issue in conventional bead filters. ANSYS Maxwell was used to simulate the magnetic field pattern required to hold ME biosensors densely and to optimize the frame design. Based on the simulation results, a phage filter structure was constructed, and a proof-in-concept experiment was conducted where a Salmonella solution of known concentration were passed through the filter, and the number of captured Salmonella was quantified by plate counting.
This paper presents a method for detection of a few pathogenic bacteria and determination of live versus dead cells. The method combines wireless phage-coated magnetoelastic (ME) biosensors and a surface-scanning dectector, enabling real-time monitoring of the growth of specific bacteria in a nutrient broth. The ME biosensor used in this investigation is composed of a strip-shaped ME resonator upon which an engineered bacteriophage is coated to capture a pathogen of interest. E2 phage with high binding affinity for Salmonella Typhimurium was used as a model study. The specificity of E2 phage has been reported to be 1 in 105 background bacteria. The phage-coated ME biosensors were first exposed to a low-concentration Salmonella suspension to capture roughly 300 cells on the sensor surface. When the growth of Salmonella in the broth occurs, the mass of the biosensor increases, which results in a decrease in the biosensor's resonant frequency. Monitoring of this mass- induced resonant frequency change allows for real-time detection of the presence of Salmonella. Detection of a few bacteria is also possible by growing them to a sufficient number. The surface-scanning detector was used to measure resonant frequency changes of 25 biosensors sequentially in an automated manner as a function of time. This methodology offers direct, real-time detection, quantification, and viability determination of specific bacteria. The rate of the sensor's resonant frequency change was found to be largely dependent on the number of initially bound cells and the efficiency of cell growth.
Efforts were made to incorporate the phage Magnetoelastic (ME) biosensor in FDA's Spinach Soaking procedures according to FDA 2015 BAM method. Three soaking materials (Lactose broth, LB broth, and Peptone water) and various soaking times were investigated. Using merely 100 Salmonella cells spiked on the produce surfaces, the phage biosensors detected Salmonella within 5 hours when soaking tomatoes in LB broth as opposed to taking up to 24 hours. Salmonella was detected on spinach leaves within 7 hours. These phage ME biosensors provide a promising rapid detection platform using LB broth in FDA's soaking procedures while shortening the incubation period.
Real-time in-situ detection of pathogenic bacteria on fresh food surfaces was accomplished with phage-based magnetoelastic (ME) biosensors. The ME biosensor is constructed of a small rectangular strip of ME material that is coated with a biomolecular recognition element (phage, antibodies or proteins, etc.) that is specific to the target pathogen. This mass-sensitive ME biosensor is wirelessly actuated into mechanical resonance by an externally applied time-varying magnetic field. When the biosensor binds with target bacteria, the mass of the sensor increases, resulting in a decrease in the sensor's resonant frequency. In order to compensate for nonspecific binding, control biosensors without phage were used in this experiment. In previous research, the biosensors were measured one by one. However, the simultaneous measurement of multiple sensors was accomplished in this research, and promises to greatly shorten the analysis time for bacterial detection. Additionally, the use of multiple biosensors enables the possibility of simultaneous detection of different pathogenic bacteria. This paper presents results of experiments in which multiple phage-based ME biosensors were simultaneously monitored. The E2 phage and JRB7 phage from a landscape phage library served as the bio-recognition element that have the capability of binding specifically with Salmonella typhimurium and B. anthracis spores, respectively. Real-time in-situ detection of Salmonella typhimurium and B. anthracis spores on food surfaces are presented.
KEYWORDS: Sensors, Biosensors, Magnetic sensors, Scanning electron microscopy, Liquids, Particles, Microfabrication, Chemical elements, Magnetism, Gold
Food borne illnesses from the ingestion of S. typhimurium have been of primary concern due to their common
occurrence in food products of daily consumption. In this paper, micron size, magnetoelastic (ME) biosensors for the
detection of S. typhimurium were fabricated and tested in liquid solutions containing known concentrations of S.
typhimurium cells. The biosensors are comprised of a ME sensor platform and immobilized bio-molecular recognition
element (E2 phage) that has been engineered to bind the S. typhimurium multi-valently. The micron size ME sensor
platforms were manufactured using microelectronics fabrication techniques. Phage was engineered at Auburn
University and immobilized onto all surfaces of the sensor. The ME biosensor oscillates with a characteristic resonance
frequency when subjected to a time varying magnetic field. Binding between the phage and bacteria, adds mass to the
sensor that causes a shift in the sensor's resonance frequency. Sensors with the dimension of 500x100x4 μm were
exposed to S. typhimurium with increasing known concentrations ranging from 5 x101 to 5 x 107 cfu/ml. The ME
biosensor exhibited high sensitivity and a detection limit better than 50 cfu/ml.
In this paper the performance of a magnetoelastic biosensor detection system for the simultaneous identification of B.
anthracis spores and S. typhimurium was investigated. This system was also designed for selective in-situ detection of
B. anthracis spores in the presence a mixed microbial population. The system was composed of a reference sensor
(devoid of phage), an E2 phage sensor (coated with phage specific to S. typhimurium) and a JRB7 phage sensor (coated
with phage specific to B. anthracis spores). When cells/spores are bound to the specific phage-based ME biosensor
surface, only the resonance frequency of the specific sensor changed. The instantaneous response of the multiple
sensor system was studied by exposing the system to B. anthracis spores and S. typhimurium suspensions sequentially.
A detection limit of 1.6×103 cfu/mL and 1.1×103 cfu/m was observed for JRB7 phage sensor and E2 phage sensor,
respectively. Additionally, the performance of the system was also evaluated by exposure to a flowing mixture of B.
anthracis spores (5×101-5×108 cfu/ml) in the presence of B. cereus spores (5×107 cfu/ml). Only the JRB7 phage
biosensor responded to the B. anthracis spores. Moreover, there was no appreciable frequency change due to
non-specific binding when other microorganisms (spores) were in the mixture. A detection limit of 3×102 cfu/mL was
observed for JRB7 phage sensor. The results show that the multi-sensor detection system offers good performance,
including good sensitivity, selectivity and rapid detection.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.