Anhydrobiotic engineering and encapsulation strategies to increase storage stability of microbial control agents

Abstract

Metarhizium is an important group of entomopathogenic fungi that can grow in liquidmedia as blastospores. Blastospores are infective to several agricultural pests.Pseudomonas protegens, a Gram-negative non-sporulating bacterium, holds significantpotential as a biocontrol agent against agricultural pests and diseases. Limitations relatedto low desiccation tolerance and short-term storage stability have constrained thecommercial utilization of Metarhizium blastospores and P. protegens. This two-stepresearch aims to develop pre-conditioning and encapsulation strategies to enhancedesiccation tolerance and sustain the anhydrobiosis state in M. anisopliae blastospores andP. protegens, ensuring storage stability for at least six months. We will formulate cell preconditioning strategies by modifying the culture medium in the initial phase. This will involvevarying water activities, introducing specific intracellular compatible solutes, and applyingthermal treatment during cultivation. For this study, we will cultivate P. protegens using aninnovative microbioreactor platform designed for automated upstream processing,facilitating up to 48 parallel fermentations. Simultaneously, blastospores will be produced inflasks. Furthermore, we will examine intracellular metabolite levels using gaschromatography-mass spectrometry (GC-MS) to discern the physiological adaptations ofM. anisopliae blastospores and P. protegens in response to pre-conditioning treatments. Inthe subsequent phase, we aim to formulate encapsulation strategies for M. anisopliaeblastospores and P. protegens cells. This will involve a combination of externally addedosmo-protectors and co-formulants to the culture medium, along with components creatinga matrix to provid a barrier against gas and moisture exchange.