Evologic’s Formulation Technology Is Unlocking New Microbial Products for the Ag Market

Date:06-05-2024

Alejandro del Barrio 

Lead Scientist at Evologic Technologies GmbH

One of the main advantages of microbial products in agriculture is their low persistence in the environment, which minimizes residues on crops and negative effects on non-target organisms. However, this advantage is also their greatest weakness, creating significant efficacy and stability challenges in product development ^[1]. The decline of viable cells in a product based on living microbes results in a loss in product efficacy, causing supply chain & commercial risks, as well as lost opportunities in resource efficiency and food safety.

Supposedly, performance of microbial products in the field is rather inconsistent across different soils, crops and climates but how many studies control for stability over time, and compatibility with other ag input materials？^[2]. All these factors are related to limitations in formulation development and challenges that come with conservation of stability, efficacy and quality of live microbes from manufacturing to application.

Formulation development as a solution 

The most successful microbial products in the agricultural input market are based on a single Gram-positive species of bacteria – Bacillus thuringiensis (Bt). Bt-based products occupy about 95% percent of the biopesticide market ^[3], with CAGR of 6.4% during the forecast period 2024-2032 ^[4]. This might not come as a surprise looking from the formulation perspective, as Bt is a highly stable organism, with considerable tolerance to various stressors. Formulation of Gram-negative bacteria and fungi poses a more demanding engineering challenge due to the sensitive nature of their cell structures. The lack of formulation expertise and capacity for these organisms, as well as high production costs, greatly limit the introduction of new microbial products to the market.

Evologic has identified this gap and created BioShield^® – a platform technology to economically stabilize microbes in capsules that meet the specifications required by the industry and farmers.

BioShield^® – novel approach to microbial formulation

Conventional microbial formulations in agriculture – solid and liquid formulations – face significant challenges in maintaining microbial cell stability and viability ^[2,5]. Encapsulation of microbes in natural biopolymers creates a protective microenvironment within the capsule that shields microbial cells from stressful conditions encountered at each step – from manufacturing to the field. 

Mostly encapsulation processes start with fermentation of microbes in bioreactors – a pivotal step that yields high-cell densities with customized metabolic states required for subsequent processing. Fermented microbes are then mixed with a safe, fully degradable biopolymer, yielding a fluid ready to be encapsulated. Droplets are formed and polymerized, forming hydrogel capsules containing microbes ^[6] (Figure 1). The capsules are finally dried and employed in subsequent applications. The protective microenvironment inside the capsule leads to an extended shelf life, gradual release of microbes, and extended persistence after application.

Figure 1. Encapsulation based on ionic gelation.  Adapted from Vemmer et al. (2013) ^[6]

High cost of goods (COGs) has been hindering adoption of this promising technology at commercially-relevant production scales – a challenge solved with BioShield^®. With an integrated approach to upstream processing, tailored growing media, and optimized drying procedures, BioShield^® achieves high throughput and yields microbes of desired metabolic state and cell density at competitive costs.

The optimized upstream fermentation and customized drying procedures prime the microbes for stress adaptation, triggering the modulation of cell physiology to adapt to the perceived environmental stress. As a result, the microbial cells are more tolerant to desiccation, stable, have a prolonged shelf-life, and provide more consistent effects upon application.

The key effect of increased stability and shelf life is the reduction of application dose while having greater density of viable microbial cells – lowering costs without compromising quality.

Compatibility with application – custom capsule size

The ultimate challenge of encapsulation is producing capsules compatible with agricultural application processes, which most commonly include employment of various spraying technologies. Capsule size must be small to easily flow though narrow nozzles – larger particles clog them, resulting in subpar delivery (Figure 2). 

Figure 2. Capsules generated with standard encapsulation technologies (a) and tiny nanocapsules generated with the BioShield® encapsulation technology (b)

To meet this technical challenge, Evologic’s BioShield^® is developed to achieve manufacturing of nanocapsules as tiny as 20 μm. These nanocapsules can deliver a highly concentrated product to a wide range of environments, even small-size surfaces, like plant seeds. Aside from the custom capsule size, the technology also enables a wide range of different physical properties to tailor the compatibility of industrial application processes. 

BioShield^® for Gram-negative bacteria

Gram-negative bacteria encompass promising solutions for agricultural applications, both as biostimulants (e.g. Rhizobia, Azotobacter - N-fixing endosymbiotic bacteria) and biocontrol agents (e.g. Pseudomonas – root pathogen control). However, their specific cell wall structures and sensitivity to environmental stressors pose significant formulation challenges. Through BioShield^®, Evologic has developed a superior drying process to the current state-of-the-art (freeze-drying), enhancing the viability and stability of Gram-negative bacteria and prolonging their shelf-life up to 12 months, meeting the demands of agricultural applications (Figure 3&4).

Figure 3. Comparison of viability loss of a Gram-negative bacterium. BioShield® vs freeze-drying	Figure 4. Storage stability of Gram-negative bacteria represented as LOG10 of the CFU/g over a period of 180 days stored at 4°C. The total viability loss is 0.65 LOG10

BioShield^® for fungi

Fungal cells have more complex structures and larger cell size compared to bacteria, making their formulation more technically demanding. BioShield^® enables adjustment of capsule size to encapsulate the large fungal cells, while keeping the capsules small enough to be compatible with agricultural application technologies. As proof of concept, Evologic has achieved encapsulation of AMF spores with prolonged viability (Figure 5). 

Figure 5. AMF spores encapsulated with BioShield^®

Conclusion

Evologic’s Bioshield technology serves to solve the critical challenges in stabilization and application of difficult-to-produce microbes like Gram-negative bacteria and fungi. Delivering a highly concentrated, stable product with customizable particle size, Bioshield enables seamless incorporation of microbial products to a wide range of agricultural applications. Bioshield effectively stabilizes even the most difficult-to-produce microbes and achieves up to 12 months of shelf life.

References

1. The development, regulation and use of biopesticides for integrated pest management. (2011). David Chandler. doi: 10.1098/rstb.2010.0390.

2. Realities and hopes in the application of microbial tools in agriculture. (2021). Bruna D. Batista. doi: 10.1111/1751-7915.13866.

3. Bacillus thuringiensis-based nanopesticides for crop protection. (2019). P.S. Vimala Devi. doi:10.1016/B978-0-12-815829-6.00010-3.

4. Global Bacillus thuringiensis Market Research Report 2024. Market Growth Reports.

5. Maintenance and assessment of cell viability in formulation of non‐sporulating bacterial inoculants. (2018). Berninger, Teresa, et al. doi: 10.1111/1751-7915.12880

6. Review of encapsulation methods suitable for microbial biological control agents. (2013). Marina Vemmer, and Anant V. Patel. Biological Control 67.3 (2013): 380-389. doi:10.1016/j.biocontrol.2013.09.003.

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