In a recent article published in the journal Scientific Reports, researchers from Egypt have synthesized nanomaterials sensitive to N-hexanoyl homoserine lactone (C6-HSL) molecules and evaluated them for their potential as biosensors in industrial environments.
Colorimetric sensing of C6-HSL of SRB bacteria under ambient conditions. + ve (positive control), − ve (Negative control), + 1 (medium intensity of the C6-HSL producing SRB). Image Credit: https://www.nature.com/articles/s41598-024-60093-8
Background
Bacteria employ various chemical signaling mechanisms, such as quorum sensing (QS) molecules and pheromones, to communicate and coordinate their activities within microbial communities. Understanding the signaling molecules involved in bacterial communication, particularly in biofilm formation, is essential for addressing challenges posed by bacteria, such as those encountered in industrial settings like oil and gas fields.
QS molecules, including N-acylated homoserine lactones (AHLs), serve as key signaling molecules for bacteria, regulating gene expression and behaviors critical for community function. Biofilms, characterized by bacterial aggregation and extracellular matrix production, are influenced by factors like nutrient availability, surface attachment, and bacterial movement. Gram-negative bacteria, such as sulfate-reducing bacteria (SRB), produce AHLs like C6-HSL and N-dodecanoyl homoserine lactone (C12-HSL), contributing to microbial-influenced corrosion (MIC) in industrial settings.
Traditional methods for AHL detection, like chromatographic techniques, are often time-consuming and costly. As a result, alternative approaches, such as electrochemical impedance spectroscopy (EIS), have gained attention for their potential to offer rapid and cost-effective detection methods. Integrating metal oxides with conducting polymers to form hybrid nanocomposites (NCs) has shown promise in biosensing applications. These composites offer enhanced sensitivity and selectivity, making them attractive for detecting AHLs produced by bacteria like SRB.
The Current Study
ZnO and Fe2O3 nanoparticles (NPs) were synthesized through co-precipitation, while Polyaniline (PANI) Dodecylbenzene Sulfonic Acid (DBSA) nanocomposites (NCs) with metal oxides were prepared via emulsion polymerization. Aniline monomers underwent polymerization in the DBSA solution, incorporating ZnO or Fe2O3 NPs. The resultant composites were washed to yield ZnO/PANI-DBSA and Fe2O3/PANI-DBSA NC
Next, SRB media were prepared and enriched, then used as inoculum for cultivated reactors. Mild steel coupons were polished, greased, and used as working electrodes. Corrosive activities were then evaluated using various methods, including dissolved sulfide measurement and metal corrosion rate assessment. Scanning electron microscope (SEM) was used to visualize bacterial biofilms.
To assess the biofilm formation, Open Circuit Potential (OCP) and EIS measurements were conducted using glass three-electrode cells under anaerobic conditions. Measurements were performed using the prepared SRB media, and OCP was measured after 30 minutes of soaking.
For the analysis of C6-HSL, ZnO/PANI-DSBA and Fe2O3/PANI-DSBA composites were mixed with carbon paste and deposited onto Screen-Printed Electrode (SPE) sensors. Sensors were evaluated using EIS for measuring bacterial signals in anaerobic cells inoculated with enriched SRB with different C6-HSL concentrations. Colorimetric detection of C6-HSL was performed using A. tumefaciens KYC55 biosensor strain. Positive C6-HSL production was indicated by blue coloration on agar plates stained with X-Gal.
Results and Discussion
The prepared PANI-DBSA/metal oxide NCs underwent extensive characterization. X-ray diffraction (XRD) analysis revealed the amorphous nature of PANI-DBSA and the crystalline structure of the metal oxide NPs, with ZnO/PANI-DBSA exhibiting a crystallite size of 86 nm and Fe2O3/PANI-DBSA a size of 78 nm. High-resolution transmission electron Microscopy (HR-TEM) images confirmed the morphology and size distribution of the NPs, with ZnO displaying hexagonal and rod-like shapes and Fe2O3 showing spherical morphology.
The average molecular weight was observed as 15,080 g/mol for PANI-DBSA. Electrochemical assessment of enriched-SRB biofilm using EIS revealed changes in charge transfer resistance (Rct) and biofilm resistance (Rbf) over time, indicating biofilm formation and corrosion kinetics.
Electrochemical assessment of C6-HSL using Cyclic Voltammetry (CV) demonstrated the sensitivity of ZnO/PANI-DBSA and Fe2O3/PANI-DBSA sensors towards C6-HSL, with ZnO-based sensors exhibiting higher cathodic peak currents. The sensors showed a quasi-linear correlation between Rct and C6-HSL concentration, with ZnO/PANI-DBSA and Fe2O3/PANI-DBSA sensors having a limit of detection (LOD) of 739 ppm and 484 ppm, respectively.
Conclusion
In summary, this study emphasizes the synthesis and evaluation of two nanocomposites, namely ZnO/PANI-DBSA and Fe2O3/PANI-DBSA, for the detection of C6-HSL, a pivotal molecule in environmental monitoring. The ZnO/PANI-DBSA sensor exhibited heightened sensitivity compared to its Fe2O3 counterpart, showcasing superior detection limits. Moreover, enhanced conductivity, particularly at elevated C6-HSL concentrations, was observed, corroborated by colorimetric assays.
According to the authors, using ZnO/PANI-DBSA and Fe2O3/PANI-DBSA in sensors could offer a simpler and cheaper way to measure C6-HSL levels in oil and gas wells. Both sensors operate through a diffusion-controlled system, with ZnO/PANI-DBSA showing better sensitivity and stability. This research opens up avenues for practical environmental monitoring in the oil and gas industry.
Journal Reference
Gado, W.S., Al-Gamal, A.G., Badawy, M.S.E.M. et al. (2024). Detectable quorum signaling molecule via PANI-metal oxides nanocomposites sensors. Scientific Reports 14, 10041. https://doi.org/10.1038/s41598-024-60093-8, https://www.nature.com/articles/s41598-024-60093-8