# Effects of Culture Aeration and the C:N Ratio on Propagule Production by Submerged Cultivation of Clonostachys rosea and Its Antifungal Metabolite Profiling

## TL;DR

This study shows that increasing aeration during submerged cultivation of Clonostachys rosea boosts production of disease-fighting propagules and antifungal compounds, which can help control tomato gray mold.

## Contribution

The study identifies optimal aeration and C:N ratio conditions for maximizing antifungal propagules and metabolites in Clonostachys rosea.

## Key findings

- High aeration and a 50:1 C:N ratio significantly increased submerged conidia production.
- Microsclerotia formation was favored under low C:N (10:1) with high aeration.
- Propagules and sorbicillinoids from C. rosea effectively reduced gray mold on cherry tomatoes.

## Abstract

Clonostachys rosea is a necrotrophic mycoparasite studied for biocontrol of plant pathogenic fungi, including Botrytis cinerea, the causal agent of gray mold that causes economic losses in several common fruits and vegetables. This study evaluated how the culture aeration, manipulated through the medium‐to‐flask volume ratio, affects the submerged production of conidia and microsclerotia, key propagules for disease control. A low medium‐to‐flask ratio (1:5), which enhances aeration, significantly increased propagule yields. A high C:N ratio (50:1) favored submerged conidia production under elevated aeration, while microsclerotia formed only with low C:N (10:1) and boosted under high aeration. These propagules, along with cell‐free culture filtrates, were formulated into water‐dispersible microgranules and tested for efficacy against gray mold on cherry tomatoes. All formulations reduced disease incidence. UPLC ESI–QTOF–MS analysis of the organic extract from the culture filtrate revealed sorbicillinoids as the major antifungal metabolites. Overall, this study highlights the role of aeration in optimizing C. rosea submerged cultivation and supports the potential of its propagules and metabolites for use in biocontrol strategies against postharvest disease induced by B. cinerea.

Optimized submerged fermentation of Clonostachys rosea under high aeration enhances production of submerged conidia and microsclerotia. Formulated propagules and antifungal metabolites, including sorbicillinoids, effectively control tomato gray mold, offering a promising strategy for biocontrol product development.