Mycotoxin contamination in rice is less reported, compared to that in wheat or maize, however, some Fusarium fungi occasionally infect rice in the paddy field. Fumonisins are mycotoxins mainly produced by Fusarium verticillioides, which often ruins maize. Rice adherent fungus Gibberella fujikuroi is taxonomically near to F. verticillioides, and there are sporadic reports of fumonisin contamination in rice from Asia, Europe and the United States. Therefore, there exists the potential risk of fumonisin contamination in rice as well as the need for the validated analytical method for fumonisins in rice. Although both natural and spiked reference materials are available for some Fusarium mycotoxins in matrices of wheat and maize, there are no reference materials for Fusarium mycotoxins in rice. In this study, we have developed a method for the preparation of a reference material containing fumonisins in Thai rice. A ShakeMaster grinding machine was used for the preparation of a mixed material of blank Thai rice and F. verticillioides-infected Thai rice. The homogeneity of the mixed material was confirmed by one-way analysis of variance, which led this material to serve as an in-house reference material. Using this reference material, several procedures to extract fumonisins from Thai rice were compared. Accordingly, we proved the applicability of an effective extraction procedure for the determination of fumonisins in Japanese rice.
The identification of rice bacterial leaf blight disease requires a simple, rapid, highly sensitive, and quantitative approach that can be applied as an early detection monitoring tool in rice health. This paper highlights the development of a turn-off fluorescence-based immunoassay for the early detection of Xanthomonas oryzae pv. oryzae (Xoo), a gram-negative bacterium that causes rice bacterial leaf blight disease. Antibodies against Xoo bacterial cells were produced as specific biorecognition molecules and the conjugation of these antibodies with graphene quantum dots and gold nanoparticles was performed and characterized, respectively. The combination of both these bio-probes as a fluorescent donor and metal quencher led to changes in the fluorescence signal. The immunoreaction between AntiXoo-GQDs, Xoo cells, and AntiXoo-AuNPs in the immunoaggregation complex led to the energy transfer in the turn-off fluorescence-based quenching system. The change in fluorescence intensity was proportional to the logarithm of Xoo cells in the range of 100-105 CFU mL−1. The limit of detection was achieved at 22 CFU mL−1 and the specificity test against other plant disease pathogens showed high specificity towards Xoo. The detection of Xoo in real plant samples was also performed in this study and demonstrated satisfactory results.
Blooms of toxic and harmful microalgae, known as "red tides", represent a significant and tremendous threat to human health and fisheries resources throughout the Southeast Asia region and the world. A harmful microalga which produces secondary metabolite known as saxitoxin (STX) is a neurotoxin produced by a variety of genera of dinoflagellates. The ingestion of this toxin through contaminated shellfish will inhibit the neuron depolarizations and action potentials which result in respiratory failure in human and known as PSP (paralytic shellfish poisoning). PSP takes effect as soon as five minutes depending on the species and the concentration consumed. Currently, the identification of dinoflagellates cells is done by taxonomy, which is based on a broad expertise of specially trained staff, expensive equipment like electron microscopes and is very time consuming. In many countries, for the toxin monitoring, they use mouse bio-assay which is bulky, expensive, loss of animal life and high variability and higher limit of detection (40 µg per 100 g meat) and HPLC methods required many pre-treatment steps for the analysis. Therefore, in order to overcome this problem, development of early detection of harmful algae bloom and bio-toxin in aquatic ecosystem is needed. The detection is based on screen-printed carbon working electrode with onboard carbon counter and silver-silver chloride pseudo-reference electrode. A direct ELISA (enzyme-linked immunosorbent assay) format was developed and optimized on the surface of a carbon screen-printed working electrode by immobilizing the capture antibody using electro-deposition of gold nanoparticles conjugated with polyclonal anti-toxin antibodies. The detection reagent, toxin-HRP (horseradish peroxidase) conjugate was used as an enzyme label. Electrochemical detection was then carried out using 3,3',5,5'-tetramethylbenzidine dihydrochloride (TMB)/H 2 O 2 as the enzyme mediator/substrate system and conducted using chronoamperometry at 100 mV vs. onboard screen-printed Ag-AgCl pseudo-reference electrode. The minimum detection limits for harmful micro algae about 1 cell per mL. The immunosensor is very selective which is able to detect Alexandrium minitum with 100% selectivity and minimal cross reaction with others nontoxic algae (< 2%).
Aflatoxin B1 (AFB1) and ochratoxin A (OTA) are potent mycotoxins produced by the fungal genus Aspergillus. Their occurrence in grain corn is alarming hence the need for rapid on-site detection. An immuno-based biosensor technique for detection of the aforementioned toxins is described here. Highly specific in-house polyclonal antibodies against AFB1 and OTA were employed as bioreceptors in a label-free electrochemical biosensor; immobilized on modified screen-printed carbon electrodes (SPCEs). The immuno-functionalized SPCEs were first characterized on a laboratory electrochemical workstation for proof-on-concept study using differential pulse voltammetry (DPV) electrochemical technique. An Android-based device is improvised as a portable electrochemical reader integrated with internet of thing (IoT) features which include cloud server and a dedicated website. Sensitivity achieved by the modified SPCEs on the portable device is superior compared to enzyme-linked immunosorbent assay (ELISA) method and lab-based electrochemical workstation. The miniaturized biosensor system has been successfully tested on cornfield for in-situ mycotoxins detection with simple sample extraction. Analysis performed on twenty samples were validated using chromatographic analysis. This biosensor-IoT system offers a potential application for real-time detection and the portable reader serves as an excellent tool for point-of-care in routine monitoring of harmful mycotoxins.
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