Ishimota_2026_Sci.Total.Environ_1014_181349

Ongoing climate change and chemical pollution are major stressors that threaten the stability and biodiversity of ecosystems and long-term environmental health worldwide. In this study, we evaluated the acquisition of chemical tolerance by Daphnia magna under thermal stress and used RNA sequencing to determine the genes associated with chemically tolerant clones. D. magna was continuously exposed to sublethal concentrations of the carbamate pirimicarb at two temperatures (20 degreesC and 25 degreesC) over 15 generations. Consequently, the 48 h median effective concentration in the pirimicarb-exposed group was twofold higher than that in the control group at 20 degreesC, whereas the values were not different between the groups at 25 degreesC. Therefore, pirimicarb tolerance acquisition under continuous exposure to the carbamate was inhibited by the higher temperature. Additionally, the intrinsic growth rate of the continuous exposure group was significantly lower when exposed to pirimicarb than that of the control group at 25 degreesC, whereas no growth difference was observed between the groups at 20 degreesC. In line with this, only in the pirimicarb-tolerant Daphnia at 20 degreesC the mRNA levels of oxidative stress-related genes (including cytochrome P450 and peroxidase) and acetylcholinesterase were significantly upregulated. Real-time quantitative PCR analysis validated the results, revealing that peroxidase may be an essential factor for pirimicarb tolerance. Furthermore, the carapace of D. magna was thicker in the pirimicarb-exposed groups than in the controls at both temperatures. These findings underscore the importance of temperature for the buildup of tolerance to pesticides under multigenerational exposure of Cladocerans.