The Repetitive Detection of Toluene with Bioluminescence Bioreporter Pseudomonas putida TVA8 Encapsulated in Silica Hydrogel on an Optical Fiber

Abstract

Living cells of the lux-based bioluminescent bioreporter Pseudomonas putida TVA8 were encapsulated in a silica hydrogel attached to the distal wider end of a tapered quartz fiber. Bioluminescence of immobilized cells was induced with toluene at high (26.5 mg/L) and low (5.3 mg/L) concentrations. Initial bioluminescence maxima were achieved after >12 h. One week after immobilization, a biofilm-like layer of cells had formed on the surface of the silica gel. This resulted in shorter response times and more intensive bioluminescence maxima that appeared as rapidly as 2 h after toluene induction. Considerable second bioluminescence maxima were observed after inductions with 26.5 mg toluene/L. The second and third week after immobilization the biosensor repetitively and semiquantitatively detected toluene in buffered medium. Due to silica gel dissolution and biofilm detachment, the bioluminescent signal was decreasing 20–32 days after immobilization and completely extinguished after 32 days. The reproducible formation of a surface cell layer on the wider end of the tapered optical fiber can be translated to various whole cell bioluminescent biosensor devices and may serve as a platform for in-situ sensors.

Analysis of Bioavailable Toluene by Using Recombinant Luminescent Bacterial Biosensors with Different Promoters

“…The results demonstrated that luminescence was stable and the highest at 2-2.5 h after incubation for T3-lux-E. coli and T7-lux-E. coli or 1-1.5 h after incubation for SP6-lux-E. coli. The time was equal to or shorter than that previously reported for the lux-based bioluminescent bioreporter P. putida TVA8 (2 h) and luminescence bacterial biosensors without the T7 promoter (3 h) for toluene measurement [6,21]. Therefore, on average, 20-min consecutive measurements were recorded when T3-lux-E. coli and T7-lux-E. coli were cultured for 2 h and when SP6-lux-E. coli was cultured for 1 h. The maximum average luminescence induced by 200 μM toluene for T3-lux-E. coli, SP6-lux-E. coli, and T7-lux-E. coli was 1020 ± 17.6, 510 ± 14.1, and 2120 ± 63.8 RLU, respectively.…”

Analysis of Bioavailable Toluene by Using Recombinant Luminescent Bacterial Biosensors with Different Promoters

“…The results demonstrated that luminescence was stable and the highest at 2-2.5 h after incubation for T3-lux-E. coli and T7-lux-E. coli or 1-1.5 h after incubation for SP6-lux-E. coli. The time was equal to or shorter than that previously reported for the lux-based bioluminescent bioreporter P. putida TVA8 (2 h) and luminescence bacterial biosensors without the T7 promoter (3 h) for toluene measurement [6,21]. Therefore, on average, 20-min consecutive measurements were recorded when T3-lux-E. coli and T7-lux-E. coli were cultured for 2 h and when SP6-lux-E. coli was cultured for 1 h. The maximum average luminescence induced by 200 μM toluene for T3-lux-E. coli, SP6-lux-E. coli, and T7-lux-E.…”

“…Conventional analytical techniques, such as gas chromatography (GC) and high-performance liquid chromatography, are sensitive and reliable for toluene detection but are time-consuming, expensive, and laboratory-bound, and they require large equipment and specialized training [5,6]. By contrast, biological methods can be useful alternatives for organics detection because they are low cost, easy to use, portable, small, and highly specific and can detect bioavailability [7][8][9].…”

Optical Biosensors for Label-Free Detection of Small Molecules

“…Genetically modified organisms have also been coupled to optical fibers for the development of label-free biosensors for the sensitive and selective detection of a wide range of compounds of interest in different fields such as food analysis, environmental monitoring, and clinical diagnostics, among others [49,50,51,52,53]. Sensor performance depends on the introduction of a reporter gene into the host cell, whose expression is modulated by the interaction of the analyte with the molecular recognition element, and promoter sequences [54].…”

“…A robust biomolecule coating on the sensor surface should be stable and, in many cases, preferably regenerable. It is critical to ensure that the surface coating and the immobilization do not degrade the biological activity of the recognition element or result in steric hindrance, and a sufficient number of molecules must be immobilized to ensure reliable signals simultaneously minimizing the non-specific binding to the surface [50]. The following sections discuss SPR-based methods reported for small molecule detection using different recognition elements, and Table 1 summarizes some of the most interesting approaches from recent years.…”