NANOENGINEERED SURFACES AND CARBON NANOTUBE CONJUGATED MICROWIRE BIOSENSOR FOR MICROBIAL CONTROL AND DETECTION

Abstract

Nanotechnology is applied in various fields including the food industry. Nanotechnology integrates several disciplines and uses nanomaterials with size in the range from 1 to 100 nm. In the food industry, nanotechnology has potential to cover many aspects such as product development, food security, and new functional materials. Particularly, nanotechnology is a promising tool to address public food safety concerns by reducing the consumption of contaminated food products. Over the past years, the demand for real-time and sensitive detection of pathogenic bacteria in food has increased significantly. Current detection methods cannot facilitate the needs of food processors due to limitations such as time, cost, and mandatory laboratory settings. Therefore, a biosensor-based detection technology, which has advantages such as high sensitivity and portability, has emerged as an alternative. With the rapid advancement of nanotechnology, various nanomaterials have been integrated into biosensing platforms to address challenges such as sensitivity and rapid response time. In this study, a single-walled carbon nanotube (SWCNT)-based electrochemical impedance immunosensor for on-site detection of Listeria monocytogenes (L. monocytogenes) was developed. The L. monocytogenes immunosensor was functionalized by coating a gold plated tungsten wire with polyethylenimine, SWCNTs, streptavidin, biotinylated L. monocytogenes antibodies, and bovine serum albumin to induce specificity and selectivity. A linear relationship (R2 = 0.982) was observed between the electron transfer resistance measurements and concentrations of L. monocytogenes in the range of 103 - 108 CFU/mL. In addition, the sensor detected L. monocytogenes without significant interference in the presence of other bacterial cells such as Salmonella Typhimurium and Escherichia coli O157:H7. To address the needs of on-site monitoring, the sensor was integrated into a smartphone-controlled biosensor platform. The performance of the smartphone-controlled platform was evaluated with a conventional laboratory instrument. The sensing signals of the sensors immune-reacted with 103 - 105 CFU/mL of L. monocytogenes measured with both devices were not significantly different. The feasibility of the proposed platform for use in real food samples was examined with a lettuce homogenate. The recovery of the lettuce homogenates spiked with 103 - 105 CFU/mL of L. monocytogenes ranged from 90.21% to 93.69%, which proved to be suitable for food samples. Therefore, the developed on-site applicable SWCNT-based immunosensor platform appeared to be a promising tool to be used in field settings for food and agricultural applications. In order to additionally reduce the risk of microbial food contamination, nanotechnology has been extensively utilized to control biofilm formation. Bacterial adhesion on food-contact surfaces results in biofilm formation and imposes a significant challenge to food safety. Current biofilm control strategy is operating routine cleaning using chemical disinfectants. The main limitation of this method is its efficacy is altered by organic materials, pH, and temperature. It has been recognized that surface engineering could mitigate the level of bio-contamination by controlling the topography and physicochemistry of the substrate. As a result, superhydrophobic (SH) surface, which is known to be self-cleanable, has emerged as an alternative. SH surface has a water contact angle (WCA) greater than 150° and can be produced by introducing low surface energy nanoscale roughness on food-contact surface. Although there are many methods to produce SH surface, a combination of electrochemical etching and polytetrafluoroethylene (PTFE) coating has been suggested as an efficient technique due to the possibility of controlling surface morphologies and ease of operation. In this study, surface alterations on stainless steel were performed with electrochemical etching and PTFE film. The substrate was electrochemically etched at various conditions to induce nanoscale roughness and coated with PTFE to lower the surface energy. The nanostructures produced on the stainless steel substrates were characterized by field emission scanning electron microscopy. The stainless steel substrates etched at 10 V for 5 min and 10 V for 10 min with PTFE deposition resulted in an average WCA of 154° ± 4° with pore diameter of 50 nm. The bacterial resistance of these substrates (154° ± 4°) was evaluated by adhering 60 µL of L. monocytogenes (108 CFU/mL) on the substrates for 24 hours. As compared to the bare substrate, these SH surfaces significantly inhibited the bacterial adhesion up to 99%. The anti-biofilm characteristic of the superhydrophobic substrate (10 V 5 min with PTFE) was further evaluated with a CDC biofilm reactor and the bacteria entrapped in the biofilms were reduced by 98.4%. This nanoscale surface modification technique showed the feasibility for use as anti-microbial and anti-biofilm surfaces in the food industry.