EFSA approves Rhizomucor pusillus mycoprotein as novel food

Production and characterisation

The novel food is produced through controlled fermentative cultivation of the thermophilic filamentous fungus R. pusillusstrain CBS 143028, followed by heat inactivation, washing, and drying to yield an off-white powder. The applicant, The Protein Brewery B.V., demonstrated reproducible manufacturing through analysis of five independent industrial-scale batches, revealing a composition comprising approximately 51% crude protein, 31% dietary fibre, and 5% total fat (EFSA NDA Panel, 2025). Whole genome sequencing confirmed taxonomic identity with 99.1% similarity to reference strains, while comprehensive in silico analyses detected no genes associated with mycotoxin biosynthesis, antimicrobial resistance, or known virulence factors (Scaife et al., 2023).

Methodological approach

EFSA’s evaluation followed established guidance for novel food applications, encompassing identity verification, production process assessment, compositional characterisation, toxicological testing, and dietary exposure modelling (EFSA NDA Panel, 2021). Notably, the Panel requested additional genotoxicity testing on culture supernatants and a 90-day subchronic toxicity study in rats, reflecting the absence of R. pusillus from the Qualified Presumption of Safety (QPS) list for filamentous fungi (EFSA BIOHAZ Panel, 2021). The intake assessment utilised individual consumption data from the EFSA Comprehensive European Food Consumption Database, applying maximum proposed use levels across 11 food categories, including cereal-based products, dairy alternatives, chocolate confectionery, and functional beverages.

Toxicological and nutritional findings

The genotoxicity test battery, comprising bacterial reverse mutation assays and in vitro mammalian cell micronucleus tests, yielded uniformly negative results for both the biomass powder and culture supernatant (EFSA NDA Panel, 2025). The 90-day dietary toxicity study in Wistar rats, conducted according to OECD Test Guideline 408, established a no-observed-adverse-effect level (NOAEL) of 300,000 ppm – the highest concentration tested – corresponding to mean intakes of 19,668 mg/kg body weight per day in males and 22,461 mg/kg bw per day in females (Scaife et al., 2025). Hepatocellular vacuolisation observed in treated animals was minimal in severity, non-progressive, and attributed to dietary overfeeding rather than substance-specific toxicity, as evidenced by absence of hepatocellular necrosis, inflammation, or enzyme elevations.

Nutritional evaluation revealed that while the protein quality, assessed through Digestible Indispensable Amino Acid Score (DIAAS), fell below the FAO threshold of 75% (achieving 61% with leucine as the limiting amino acid), this was deemed acceptable given that the novel food would not constitute the sole protein source in varied European diets (FAO, 2013). The presence of chitin (approximately 9.1% w/w) and chitosan (9.2% w/w) as inherent cell wall components raised theoretical concerns regarding mineral bioavailability; however, the Panel concluded that consumption under proposed conditions would not substantially increase exposure to these antinutrients beyond background dietary levels (EFSA NDA Panel, 2025).

Exposure assessment and risk characterisation

Dietary exposure modelling yielded highest mean and 95th percentile intake estimates ranging from 539 to 1,013 mg/kg bw per day across population groups, with young children (1–3 years) representing the highest consumers when whey protein powder was included as a surrogate category (EFSA NDA Panel, 2025). The resulting margins of exposure – calculated by dividing the NOAEL by estimated intakes – ranged from 19 for young children to 37 for infants, values the Panel deemed sufficient considering the production process, compositional data, and absence of toxicity in provided studies. For adults consuming the novel food as a food supplement (maximum 6 g/day) or in meal replacement products (up to 40 g per meal), the combined intake estimate reached 626.6 mg/kg bw per day, maintaining an adequate margin of exposure of 31.

Allergenicity considerations

The allergenicity assessment integrated systematic literature review, in silico homology searches against the AllergenOnline database, proteomic analysis, and in vitro IgE binding assays using sera from 106 food-allergic individuals (EFSA NDA Panel, 2025). While bioinformatic analysis identified 23 proteins sharing >50% sequence identity with known allergens from crustaceans, fish, and plant sources – including enolases, triosephosphate isomerases, and cyclophilins – five of six homologous proteins were confirmed present in the final product through LC–MS/MS. IgE binding studies revealed predominantly carbohydrate-mediated responses attributable to cross-reactive carbohydrate determinants (CCDs) rather than protein-specific epitopes, leading the Panel to conclude that the likelihood of clinically relevant allergic reactions remains low, though potential cross-reactivity in individuals with existing shellfish or fish allergies cannot be entirely excluded.

Regulatory implications and conclusions

The EFSA Panel concluded that R. pusillus biomass powder is safe under the proposed conditions of use, namely as a food ingredient or whole food for the general population, in food supplements for adults over 18 years (excluding pregnant and lactating women), and in meal replacement products for weight control in adults (EFSA NDA Panel, 2025). This positive opinion facilitates regulatory authorisation under Regulation (EU) 2015/2283, potentially expanding the portfolio of sustainable protein sources available to European consumers and food manufacturers.

The assessment underscores the importance of comprehensive molecular characterisation, including whole genome sequencing, for novel foods derived from microorganisms not holding QPS status, while demonstrating that appropriate margins of exposure can be established even for products with incomplete toxicological datasets when supported by mechanistic evidence and robust intake modelling.

Regulatory timeline and comparison with GRAS-based approaches

The protracted approval timeline for this novel food application – spanning from initial submission in May 2020 to scientific opinion adoption in September 2025, and still pending formal authorisation – underscores fundamental structural differences between the European Union’s mandatory pre-market authorisation system and alternative regulatory frameworks such as the United States’ Generally Recognized As Safe (GRAS) mechanism. While the EU’s precautionary approach ensures comprehensive safety evaluation through centralised scientific assessment, the five-year duration from application to scientific approval significantly disadvantages European food innovation, particularly in the rapidly evolving alternative protein sector where accelerated market entry can determine commercial viability (EFSA NDA Panel, 2025).

The US GRAS paradigm historically permitted manufacturers to self-determine safety status based on scientific consensus without FDA notification; however, growing concerns regarding transparency and potential conflicts of interest prompted significant regulatory reform culminating in the FDA’s 2016 final rule establishing a voluntary GRAS notification programme with enhanced documentation requirements, independent expert panel evaluation, and public dossier accessibility (FDA, 2016). This evolved GRAS framework now requires comprehensive safety data submission, methodological transparency, and FDA acknowledgement letters, effectively creating a middle ground between unfettered self-certification and mandatory pre-market approval.

A comparable tiered authorisation system for the EU could incorporate lessons from both the original GRAS weaknesses and subsequent US reforms, establishing a ‘fast-track procedure’ for qualifying novel foods that mandates:

• independent expert panel assessment by food safety professionals without financial conflicts of interest;
• public registration and dossier publication ensuring stakeholder scrutiny;
• EFSA verification authority with power to elevate applications to full assessment when safety questions arise; and
• robust post-market surveillance including adverse event reporting and periodic safety reviews.

Products eligible for this streamlined track would include those derived from well-characterised organisms with QPS status or equivalent safety recognition, substances substantially equivalent to authorised foods with only minor modifications, and ingredients supported by comprehensive toxicological datasets meeting defined adequacy criteria.

Traditional novel foods presenting genuine scientific uncertainty – such as entirely new protein sources from non-QPS organisms, foods involving novel processing technologies, or products with limited consumption history – would continue through existing full EFSA evaluation procedures to maintain consumer protection standards (EFSA NDA Panel, 2021).

Such differentiated regulation would enhance proportionality by allocating intensive regulatory resources to products presenting greater uncertainty while facilitating innovation for lower-risk applications, potentially reducing timelines to 12–18 months for expedited track foods compared to current 4–5 year cycles for standard applications.

Dario Dongo

References

• EFSA BIOHAZ Panel (EFSA Panel on Biological Hazards). (2021). Statement on the update of the list of QPS-recommended biological agents intentionally added to food or feed as notified to EFSA 14: Suitability of taxonomic units notified to EFSA until March 2021. EFSA Journal, 19(7), e06689. https://doi.org/10.2903/j.efsa.2021.6689

• EFSA NDA Panel (EFSA Panel on Dietetic Products, Nutrition and Allergies). (2021). Guidance on the preparation and submission of an application for authorisation of a novel food in the context of Regulation (EU) 2015/2283 (Revision 1). EFSA Journal, 19(3), e06555. https://doi.org/10.2903/j.efsa.2021.6555

• EFSA NDA Panel (EFSA Panel on Nutrition, Novel Foods and Food Allergens). (2025). Safety of Rhizomucor pusillusbiomass powder as a novel food pursuant to Regulation (EU) 2015/2283. EFSA Journal, 23(12), e9707. https://doi.org/10.2903/j.efsa.2025.9707

• FAO (Food and Agriculture Organization). (2013). Dietary protein quality evaluation in human nutrition: Report of an FAO expert consultation (FAO Food and Nutrition Paper No. 92). FAO. https://www.fao.org/4/i3124e/i3124e.pdf

• FDA (U.S. Food and Drug Administration). (2016). Substances generally recognized as safe: Final rule. Federal Register, 81(159), 54960–55055. https://www.federalregister.gov/documents/2016/08/17/2016-19164/substances-generally-recognized-as-safe

• Scaife, K., Phipps, K. R., Scalise, D., & Polgár, O. (2025). Subchronic toxicity evaluation of dietary administration of a fungal biomass from Rhizomucor pusillus. Journal of Applied Toxicology, 45(3), 400–417. https://doi.org/10.1002/jat.4684

• Scaife, K., Vo, T. D., Dommels, Y., Leune, E., Albermann, K., & Pařenicová, L. (2023). In silico and in vitro safety assessment of a fungal biomass from Rhizomucor pusillus for use as a novel food ingredient. Food and Chemical Toxicology, 179, Article 113972. https://doi.org/10.1016/j.fct.2023.113972