Evaluation of aflatoxin contamination in protein-rich pulses using a GFP-expressing Aspergillus flavus strain

Authors: BRANSTAD-SPATES, EMILY - Oak Ridge National Laboratory; Sickler, Christine; Lebar, Matthew; Carter Wientjes, Carol; Rajasekaran, Kanniah

Submitted to: Frontiers in Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/13/2025
Publication Date: 5/29/2025
Citation: Branstad-Spates, E.H., Sickler, C.M., Lebar, M.D., Carter Wientjes, C.H., Rajasekaran, K. 2025. Evaluation of aflatoxin contamination in protein-rich pulses using a GFP-expressing Aspergillus flavus strain. Frontiers in Microbiology. 16. Article 1587035. https://doi.org/10.3389/fmicb.2025.1587035.
DOI: https://doi.org/10.3389/fmicb.2025.1587035

Interpretive Summary: The plant-based protein industry is rapidly expanding and is projected to become a $27 billion industry by 2030. Including pulses or legumes would aid in diversifying cropping systems, providing a sustainable nutrition source, and developing a resilient food and feed supply chain for the growing population. Despite being labeled as “underutilized” legume crops, pulses are widely used in many countries including USA. These plant-rich protein sources need to be evaluated for food and feed safety and quality due to their inclusion in human and livestock diets. This research explores the safety of protein-rich pulses such as chickpeas, lentils, and peas in comparison to corn kernels. We conclude that pulses do not produce higher amounts of aflatoxins compared to corn and other grains. Importance of screening pulse crops for potential contamination with aflatoxins is emphasized. This study is useful to food industry, consumers, and import/export industry.

Technical Abstract: Pulse crops, part of the Fabaceae family, are important agricultural products that are of high nutritional value, sustainable, and increasing in relevance as a food and feed product globally. Like other plant commodities such as corn or cottonseed, pulses can be affected by mycotoxigenic fungi throughout the supply chain (from field to postharvest). These fungi, specifically Aspergillus spp., can cause serious consequences for human and animal health, and lead to economic losses due to reduced yield and marketability from contamination by aflatoxin, a carcinogenic and mutagenic mycotoxin. The resistance or susceptibility of protein-rich pulse crops to infection by A. flavus, namely chickpeas [Cicer arietinum L.], lentils [Lens culinaris Medik], peas [Pisum sativum L.], and corn [Zea mays L.], as a positive control, were evaluated by a kernel screening assay (KSA). A. flavus strain 70 (AF-70) expressing a green-fluorescent protein (GFP) was used to quantify fungal spread, aflatoxin levels, cyclopiazonic acid (CPA), and a-aflatrem in real-time following fungal infection of seeds on a 2-day interval basis up to 10 days post inoculation. Although all seeds exhibited A. flavus infection, corn produced significantly higher levels of AFB1 and AFB2 compared to pulses, which produced higher levels of CPA and a-aflatrem. Given the agricultural and nutritional importance of pulses, further research is needed to investigate mechanisms of resistance to A. flavus from field to storage.