Improved High-Throughput Raman-Activated Cell Sorting Yields a High-DHA <i>Aurantiochytrium</i> Mutant with Enhanced Lipid Recycling

Industrial microbial strain breeding remains hindered by the inherent bottleneck of low-throughput phenotypic screening. Here, a high-throughput Raman-activated cell sorting strategy was employed to directly quantify the intracellular docosahexaenoic acid (DHA) content in single cells of the thraustochytrid Aurantiochytrium sp. SD116, eliminating the need for prolonged cultivation and lipid extraction. From a mutant library of 50,000 cells, a strain designated ABBS26, exhibiting a DHA content exceeding 65%, was isolated. Omics analysis revealed that the high DHA content of ABBS26 stems from robust lipid recycling, particularly targeting saturated fatty acids in triacylglycerol. After process optimization during fed-batch fermentation, ABBS26 achieved a final biomass of 147.3 g/L, a DHA production of 47.4 g/L, and a DHA content of 67.6% of total lipids. This work provides a promising approach for the improvement of PUFA-producing thraustochytrid strains and yields strain ABBS26, which exhibits great potential for applications in the food and pharmaceutical fields.