How do you culture Sanmite for tick control? - briefly
To produce Sanmite for tick management, inoculate a sterile grain or liquid substrate (e.g., rice, corn steep liquor) with a pure culture of Metarhizium anisopliae and incubate at 25‑28 °C for 10‑14 days until conidia mature. Harvest the spores, dry them to 5‑10 % moisture, and store in airtight containers at 4 °C until use.
How do you culture Sanmite for tick control? - in detail
Culturing Sanmite for tick management requires a sterile environment, defined growth media, and precise environmental parameters. Begin by selecting a reputable Sanmite strain known for acaricidal activity. Obtain a lyophilized or frozen stock from a certified supplier and store it at –80 °C until use.
Prepare a nutrient broth containing 2 % glucose, 0.5 % yeast extract, 0.2 % peptone, and 0.05 % magnesium sulfate. Adjust pH to 6.8 ± 0.2 with sterile 1 M NaOH or HCl. Autoclave the medium at 121 °C for 15 min; cool to 30 °C before inoculation.
Inoculate the cooled medium with 1 mL of thawed Sanmite stock under a laminar flow hood. Mix gently to distribute cells evenly. Transfer the culture to a shaking incubator set at 28 °C and 150 rpm. Monitor optical density (OD₆₀₀) every 12 h; target OD₆₀₀ ≈ 0.8–1.0, which corresponds to the exponential growth phase and maximum toxin production.
For scale‑up, transfer 10 % of the seed culture into larger fermenters (e.g., 5 L). Maintain dissolved oxygen above 30 % saturation and temperature at 28 °C ± 0.5 °C. Add antifoam (0.02 % v/v) as needed. Harvest cells after 48 h by centrifugation at 4,000 g for 10 min, then resuspend the pellet in sterile water to a concentration of 10⁸ CFU/mL.
Quality control steps include:
- Plate counts on selective agar to verify viable cell density.
- PCR confirmation of Sanmite-specific genes.
- Bioassay against laboratory‑reared tick larvae to assess mortality rates (target > 80 % after 24 h exposure).
Store the final formulation at 4 °C for up to 30 days. Apply the product to vegetation or directly onto host animals using a calibrated sprayer, delivering 1 mL m⁻² or 0.5 mL kg⁻¹, respectively. Reapply at 2‑week intervals during peak tick activity periods.