An Agar-Water-Assisted OD₆₅₀ Calibration Model for Standardized Quantification of Beauveria bassiana Conidia in...

Beauveria bassiana is one of the most widely used entomopathogenic fungi in insect pest management, and the need for rapid and reproducible quantification of fungal conidia to monitor process performance and to quality control products during biopesticide production is imperative. Conventional methodologies, such as hemocytometer counting and plate dilution assays, are time consuming, laborious and subject to significant operator-to-operator variability. Although optical methods have been increasingly explored for estimating fungal propagule concentrations, species-specific calibration, suspension stability, wavelength selection, and independent validation remain important for routine applications. In this study, we developed an agar-water-assisted UV-visible spectrophotometric calibration protocol for estimating conidial concentration using B. bassiana as a model entomopathogenic fungus. A 0.1% (w/v) agar-water suspension was used in order to get homogeneous, stable dispersions of conidia for optical measurements. Calibration of conidia concentration was accomplished through reliable optical density (OD) values measured at wavelengths 500 nm, 530 nm, 560 nm, 600 nm, and 650 nm. Linear correlations were observed across the tested wavelengths, with the highest goodness of fit for the model at 650 nm (R² = 0.9907). The resulting regression equation, conidia concentration (×10⁷ mL^-1) = 4.184 × OD₆₅₀-0.12450, has been independently verified with separate conidia batches, resulting in acceptable relative errors ranging from 13.78% and 18.98%. This agar-water-assisted OD₆₅₀ calibration model provides a practical and species-specific tool for the standardization of conidial dosages in biopesticide research, facilitating the reliable evaluation and application of entomopathogenic fungi within integrated pest management systems.