Co-cultivation with Bacillus subtilis promotes biomass production, carotenoid accumulation and settling performance of Dunaliella sp. in a low-cost medium

The high cost of cultivation and biomass recovery continue to limit large-scale production of Dunaliella sp. This study examined whether a fertilizer-based NPK medium at 20 g l‒1 salinity could support Dunaliella sp. cultivation and whether co-cultivation with Bacillus subtilis could improve growth, pigment accumulation and settling. Dunaliella sp. was maintained in the NPK medium for 30 days. OD680 was closely related to dry biomass (R2 = 0.952) and cell density (R2 = 0.959), supporting its use as a rapid growth indicator under the tested conditions. A 7-day co-culture experiment was then performed using Bacillus: Dunaliella ratios of 2:1, 4:1 and 6:1. Inoculated treatments maintained LB-culturable bacterial populations of 9.3–10.3 log10 CFU ml‒1, compared with approximately 7.0–7.7 log10 CFU ml‒1 in the LB control. Relative to the non-inoculated controls, Bacillus addition increased OD680 by 15–23%, with the clearest response in the 4:1 treatment (BS4). Co-cultivation slightly increased total carotenoids and β-carotene. β-Carotene was correlated with OD680 (r = 0.885, p < 0.01), indicating that β-carotene accumulation increased mainly in parallel with biomass development. Bacillus inoculation increased settling efficiency, but biomass recovery remained moderate. Correlation and regression analyses indicated that settling was more closely related to biomass level and cell physiological status than to bacterial abundance alone. Overall, the results suggest that B. subtilis can promote biomass production, carotenoid accumulation and partial biomass recovery of Dunaliella sp. in a low-cost NPK-based culture system.