Induction Techniques | FusaPrime™ Inoculant | FusaPrime™ Formulation | FusaPrime™ Development | FusaPrime™ Profile
Standard Operating Procedure (SOP) for the Preparation and Application of Fusarium oxysporum Inoculant for agarwood resin induction, formatted for use by plantations or research teams:
Trade Name: BarIno™ FusaPrime™
Target Species: Aquilaria malaccensis
Purpose: Biological resin induction using a non-lethal strain of Fusarium oxysporum
I. Preparation of Inoculant Culture
A. Materials Needed
- Pure culture of Fusarium oxysporum (resin-inducing strain)
- Potato Dextrose Agar (PDA) or cooked rice substrate (for mass production)
- Sterile distilled water
- Blender or shaker
- Measuring cylinder
- Sterile containers
- Laminar flow hood or sterile workspace
- PPE: gloves, lab coat, face mask
B. Liquid Suspension Preparation (Small Scale)
- Subculture the Fungus:
- Grow Fusarium oxysporum on PDA plates for 5–7 days at room temperature (25–28°C).
- Spore Harvesting:
- Add 10–20 mL sterile water to PDA plates.
- Scrape gently to collect conidia/spores.
- Suspension:
- Filter and transfer the suspension to sterile container.
- Adjust spore concentration to ~10⁶ CFU/mL.
- Optional: Add 1% molasses or brown sugar as carbon source to stimulate colonization.
- Storage:
- Use immediately or store refrigerated (4°C) for up to 2 weeks.
C. Stick Inoculant Preparation (Solid Carrier)
- Prepare Substrate:
- Autoclave rice or sawdust-filled bamboo sticks.
- Inoculate:
- Inject with F. oxysporum spore slurry and incubate in dark, humid area for 7–10 days.
- Storage:
- Air-dry for long-term storage or use fresh for higher activity.
II. Tree Selection and Site Preparation
- Select Aquilaria trees aged 4+ years with healthy bark and vascular activity.
- Choose the dry season or after rainfall breaks for optimal stress response.
III. Application Procedure
A. Drilling Protocol
- Drill Hole:
- Diameter: 8–10 mm
- Depth: 3–5 cm
- Angle: Downward (45–60° angle)
- Spacing: 15–30 cm apart in spiral or radial pattern
- Sanitize:
- Wipe drill bit and hole area with 70% ethanol to minimize contamination.
B. Inoculation
| Inoculant Type | Dose/Injection |
|---|---|
| Liquid Suspension | 5–10 mL per hole using sterile syringe |
| Solid Stick | 1 colonized stick per hole, tightly inserted |
C. Sealing the Hole
- Plug holes with cotton and seal with wax, mud, or sterile clay to retain moisture and prevent contamination.
IV. Post-Inoculation Care
- Avoid pruning or harvesting leaves within 1 month.
- Monitor tree health: Watch for over-pathogenicity signs (e.g., yellowing, leaf drop).
- Inspect holes monthly for fungal colonization and resin response.
V. Resin Development Timeline
- Initial signs: 1–2 months
- Visible resin formation: 3–6 months
- Peak yield window: 6–18 months
VI. Safety and Handling
- Handle live cultures in sterile conditions.
- Use PPE at all times.
- Dispose of contaminated media or tools through sterilization (autoclaving or burning).
- Prevent fungal escape into non-target environments.
VII. Storage and Shelf Life
| Formulation | Storage Conditions | Shelf Life |
|---|---|---|
| Liquid Suspension | 4°C, sealed sterile bottle | 2–4 weeks |
| Solid Stick (dry) | Cool, dry, shaded area | 2–3 months |