Almost 40% of individuals grappling with moderate or severe food insecurity globally experienced severe levels of food insecurity. The incidence of severe food insecurity rose from 9.3% in 2019 to 11.7 percent in 2021, marking an increase equivalent to 207 million additional people over the span of two years.
A crucial action for addressing food security concerns is done by the Q3 molecular workflow developed by STMicroelectronics within the EWA-BELT Consortium, which detects mycotoxins in crops. Through its innovative approach, Q3 aims to reduce costs, enhance sensitivity in mycotoxin detection, and contribute to healthier food production globally.
MARCO CEREDA, STMicroelectronics, Italy: Lab testing of qPCR device at STMicroelectronics, with University of Sassari (QUIRICO MIGHELI, Professor) and at University of Nairobi (Abigael Ouko, Lecturer), Italy & Kenya
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“Falling under task 3.1 on the development and adoption of simple, rapid, and inexpensive ICTs based tools for the detection and quantification of plant pests and pathogens STMicroelectronics has been developing a full molecular workflow on a qPCR portable device called Q3 to test model crops that may be contaminated with mycotoxins produced fungi. Research in the field is crucial in attending to food security because mycotoxins are increasingly acknowledged as significant contributors to foodborne illnesses, especially in Sub-Saharan Africa. But what are they?
Mycotoxins are harmful secondary metabolites produced by filamentous fungi (also known as moulds) that by growing on cereals, fresh and dried fruit, and vegetables may influence grain quality, and crop yield and ultimately cause severe mycotoxicosis in humans and livestock. Among the most harmful mycotoxins, aflatoxins, produced by some Aspergillus species, and fumonisins, produced by some Fusarium species, raise particular concern due to their association with liver and esophageal cancer. But many other mycotoxins exist in nature that have negative impacts on our health status and wellbeing. “It is estimated that about one-third of agricultural commodities produced worldwide are spoiled or wasted during the postharvest stages with increasing numbers admitted for some regions and crops.”
Recognizing the relevance of mycotoxins and the issues these compounds represent for food security in Sub-Saharan Africa, the EWABELT Project is developing innovative approaches, spanning from improved storage technologies and decision support systems, to early detection of mould development in the stored commodities. This is exactly what STMicroelectronics is doing within the Consortium with the development of the portable qPCR tool.
The University of Sassari, along with STMicroelectronics, has selected eight mycotoxin-related genes, considered as specific markers of the ability of fungi to produce major mycotoxins occurring in cereals. Such genes are detected with the portable qPCR tool, which represents an alternative mitigation strategy to cope with the harm posed by these toxic contaminants to food security worldwide. Let’s see how it works.
qPCR is the gold standard molecular technique allowing for specific DNA or RNA sequences in a sample to be analyzed. If such sequences are present, they will be exponentially replicated to billions of copies, so that they can be easily detected, typically through fluorescence measurement. Although qPCR guarantees the best possible sensitivity and specificity, it also has some cons that have limited its diffusion so far. Mainly: - it is quite complex, and - it requires expensive and bulky instrumentation.
Tackling these limitations is the objective of STMicroelectronics within the EWA-BELT project with the development of an innovative molecular analysis workflow, which includes two steps: First, the extraction of DNA from the crop sample. We have set up a simple extraction protocol for maize flour and peanut specimens, which requires very little equipment. This protocol was tested on samples provided by the University of Sassari. In the second step, the extracted DNA is analyzed by qPCR on a system developed by STMicroelectronics which is called Q3. This is composed of three elements:
1. a single-use cartridge,
2. a compact instrument
3. and a software application.
Besides this, specific reagents for each target DNA sequence are needed. The cartridge is the core of the system, where the reactions occur. Since qPCR requires precise heating and cooling cycles, the cartridge is built around a silicon chip that integrates a miniaturized heater and temperature sensor. This chip is produced according to microelectronics standards that guarantee a high-quality level.
The Q3 instrument is far smaller than traditional qPCR equipment; it drives the chip and features a compact fluorescence microscope to measure the optical signals coming from the reactions. Lastly, a dedicated software guides the user with step-by-step instructions and has all the analysis parameters pre-set. At the end of a test, the output files can be uploaded to the PlantHead platform developed by OCCAM, whose demo you saw before. Also, the software displays the results in a very straightforward way, that is, a sample positive or negative for each of the eight mycotoxin-related genes that we analyze.
Thanks to the high sensitivity of qPCR, we can perform early detection of mycotoxin producing fungi, which can promptly trigger containment actions to try and save the crops, or at least limit their waste. We then have a series of advantages compared to traditional qPCR equipment: portability, lower cost, and ease of use, without sacrificing performance. All of this allows taking molecular analyses out of the big, highly equipped laboratories, and making them more accessible. And this is exactly what the University of Nairobi is experimenting with.
Deployment of qPCR tool in the lab at the University of Nairobi: research and testing carried out so far, outcomes, benefits, challenges by Dr. Abigael Ouko. The University of Nairobi was chosen to be the first “end-user” to be trained by ST on all the molecular analysis workflow. The first working session was organized during the WP3 Capacity Building workshop in Kenya in October 2022. A group of researchers and students were trained on the developed DNA extraction and Q3 qPCR procedures. After a few hours of training, they were already able to successfully run the molecular tests. Since then, our laboratory has conducted numerous relevant tests.
Mycotoxin laboratory at the University of Nairobi with bags of peanut samples, peanuts with skin and without skin being pestled for DNA extraction. Peanuts without skin gave the best results. Indeed, when the peanut skin is removed, it is easier to visually distinguish the magnetic beads used for DNA extraction from the peanut debris. Removal of the peanut skin also reduces obstruction of the pipette tips during DNA extraction. We therefore modified the protocol to always use peanut seeds without skin. Then using the Q3 qPCR machine, we were able to detect:
- Peanut control DNA, which indicates that DNA extraction has been successfully performed;
- some mycotoxin-related genes, with clear indication of the positive/negative ‘calls’ for each analyzed mycotoxin, below the qPCR graph.
The plots in our experiments showed clear curves of the peanut DNA controls. In some cases, the curves had an early rise, indicating high DNA concentration, unlike in some experiments where the curves had a slightly later rise, indicating lower DNA concentrations. The report generated after each experiment illustrates detailed but easy-to-understand results from all 4 analysis wells. In doing so, it enables one to tell whether pipetting in each of the wells was precise or not.
Although further testing is required, molecular analyses with the Q3 device can represent a valuable and easy-to-use tool for further accessibility of scientific instruments at all levels, enabling:
- cost reduction of sophisticated molecular tests;
- sensitivity in the detection of mycotoxin-producing fungi
- simultaneous monitoring of many different mycotoxin-producing pathogens
- evaluation of the potential risk for foodstuffs
Combined with traditional and other innovative approaches, like the Decision Support System (DSS) for real-time monitoring in silos and storage facilities under development by CRANFIELD University, these instruments will allow for toxic fungi detection to eventually enrich the existing literature and provide solutions for healthy food for all”
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