The downstream processing of efficient biomass-based microbial biopesticides is heavily reliant on obtaining the largest concentration of viable cells in the most cost-effective manner. The goal of this research was to assess the ability of chitosan flocculation to recover bacterial Bacillus sp. BioSol021 biomass from the broth after biological treatment of wastewaters from the dairy and wine industries. Second-order factorial design models were used to estimate the effect of chitosan concentration and mixing speed on flocculation efficiency, settling velocity, and antimicrobial activity against Aspergillus flavus, i.e., inhibition zone diameter. Response surface methodology was followed by multi-objective optimization by applying the desirability function (DF) and genetic algorithm (GA). The optimum values for flocculation efficiency, settling velocity, and inhibition zone diameter for cheese whey effluent were 88%, 0.10 mm/s, and 51.00 mm, respectively. In the case of winery flotation effluent, the optimum values were flocculation efficiency 95% and settling velocity 0.05 mm/s, while the inhibition zone diameter was 48.00 mm. These results indicate that utilizing chitosan as a flocculation agent not only fits the criteria for effective downstream processing, but also has a synergistic effect on Bacillus sp. antibacterial activity.