Long-Term Lithium Exposure Induces Persistent Physiological and Methylation-Associated Alterations in Chromochloris...

Lithium (Li), a critical component of Li-ion batteries, persistently accumulates in aquatic environment, posing growing ecotoxicological risks. This study investigated the toxicological effects of long-term Li⁺ exposure and the persistent post-stress responses after Li⁺ removal in the microalga Chromochloris zofingiensis. The results showed that growth and chlorophyll content were markedly inhibited by long-term exposure to 50 mg/L Li⁺, whereas the contents of storage components-including starch, total fatty acid, and carotenoids-were significantly increased. Besides, the antioxidant system was suppressed, leading to enhanced levels of reactive oxygen species and malondialdehyde. After a 28-day Li⁺-free recovery period, the dry cell weight and cell number were reduced by 26.4% and 18.0%, respectively, relative to the control, indicating persistent post-stress growth inhibition rather than complete physiological recovery. Integrated omics analysis showed that persistent transcriptional changes in pathways associated with fatty acid elongation, starch and sucrose metabolism, and photosynthesis were accompanied by methylation alterations in promoter and gene body regions, suggesting a potential association between DNA methylation changes and post-stress physiological persistence. These findings provide insights into the molecular responses associated with long-term Li⁺ exposure and highlight the potential involvement of DNA methylation changes in persistent post-stress responses.