In the context of global warming, the frequency of severe weather occurrences, such as unexpected cold spells and heat waves, will grow, as well as the intensity of these natural disasters. Lizards, as a large group of reptiles, are ectothermic. Their body temperatures are predominantly regulated by their environment and temperature variations directly impact their behavior and physiological activities. Frequent cold periods and heat waves can affect their biochemistry and physiology, and often their ability to maintain their body temperature. Mitochondria, as the center of energy metabolism, are crucial for maintaining body temperature, regulating metabolic rate, and preventing cellular oxidative damage. Here, we used RT-qPCR technology to investigate the expression patterns and their differences for the 13 mitochondrial PCGs in Sphenomorphus incognitus (Squamata:Scincidae), also known as the brown forest skink, under extreme temperature stress at 4 °C, 8 °C, 34 °C, and 38 °C for 24 h, compared to the control group at 25 °C. In southern China, for lizards, 4 °C is close to lethal, and 8 °C induces hibernation, while 34/38 °C is considered hot and environmentally realistic. Results showed that at a low temperature of 4 °C for 24 h, transcript levels of ATP8, ND1, ND4, COI, and ND4L significantly decreased, to values of 0.52 ± 0.08, 0.65 ± 0.04, 0.68 ± 0.10, 0.28 ± 0.02, and 0.35 ± 0.02, respectively, compared with controls. By contrast, transcript levels of COIII exhibited a significant increase, with a mean value of 1.86 ± 0.21. However, exposure to 8 °C for 24 h did not lead to an increase in transcript levels. Indeed, transcript levels of ATP6, ATP8, ND1, ND3, and ND4 were significantly downregulated, to 0.48 ± 0.11, 0.68 ± 0.07, 0.41 ± 0.08, 0.54 ± 0.10, and 0.52 ± 0.07, respectively, as compared with controls. Exposure to a hot environment of 34 °C for 24 h led to an increase in transcript levels of COI, COII, COIII, ND3, ND5, CYTB, and ATP6, with values that were 3.3 ± 0.24, 2.0 ± 0.2, 2.70 ± 1.06, 1.57 ± 0,08, 1.47 ± 0.13, 1.39 ± 0.56, and 1.86 ± 0.12, respectively, over controls. By contrast, ND4L exhibited a significant decrease (to 0.31 ± 0.01) compared with controls. When exposed to 38 °C, the transcript levels of the 13 PCGs significantly increased, ranging from a 2.04 ± 0.23 increase in ND1 to a 6.30 ± 0.96 rise in ND6. Under two different levels of cold and heat stress, the expression patterns of mitochondrial genes in S. incognitus vary, possibly associated with different strategies employed by this species in response to low and high temperatures, allowing for rapid compensatory adjustments in mitochondrial electron transport chain proteins in response to temperature changes. Furthermore, this underscores once again the significant role of mitochondrial function in determining thermal plasticity in reptiles.

Heptageniidae are known for their flat heads and bodies and are divided into three subfamilies. Despite the extensive diversity within this group and considerable efforts made to understand their evolutionary history, the internal classifications and origin time of Heptageniidae remains controversial. In this study, we newly sequenced 17 complete mitogenomes of Heptageniidae to reconstruct their phylogenetic positions within this family. Because of the ambiguous time of origin, our study also estimated the divergence time within Heptageniidae based on five fossil calibration points. The results of BI and ML trees all highly supported the monophyly of Heptageniidae and three subfamilies. The phylogenetic relationship of Rhithrogeninae + (Ecdyonurinae + Heptageniinae) was also recovered. The divergence time showed that Heptageniidae originated from 164.38 Mya (95% HPD, 150.23–181.53 Mya) in the mid-Jurassic, and Rhithrogeninae originated from 95.54 Mya (95% HPD, 73.86–120.19 Mya) in the mid-Cretaceous. Ecdyonurinae and Heptageniinae began to diverge at 90.08 Mya (95% HPD, 68.81–113.16 Mya) in the middle Cretaceous. After morphological identification, analysis of the mitogenome’s composition, genetic distance calculation, phylogenetic analysis, and divergence time calculation, we suggest that two different populations of Epeorus montanus collected from Aksu, Xinjiang Uygur Autonomous Region (40°16′ N, 80°26′ E) and Xinyuan, Xinjiang Uygur Autonomous Region (43°20′ N, 83°43′ E) in China are cryptic species of E. montanus, but further detailed information on their morphological characteristics is needed to fully identify them.

Potamanthidae belongs to the superfamily Ephemeroidea but has no complete mt genome released in the NCBI (except for two unchecked and one partial mt genome). Since the sister clade to Potamanthidae has always been controversial, we sequenced seven mt genomes of Potamanthidae (two species from Rhoenanthus and five species from Potamanthus) in order to rebuild the phylogenetic relationships of Potamanthidae in this study. The divergence time of Potamanthidae was also investigated by utilizing five fossil calibration points because of the indeterminate origin time. In addition, because Rhoenanthus coreanus and Potamanthus luteus are always in low-temperature environments, we aimed to explore whether these two species were under positive selection at the mt genome level. Amongst the 13 PCGs, CGA was used as the start codon in COX1, whereas other genes conformed to initiating with an ATN start codon. From this analysis, UUA (L), AUU (I), and UUU (F) had the highest usage. Furthermore, the DHU arm was absent in the secondary structure of S1 in all species. By combining the 13 PCGs and 2 rRNAs, we reconstructed the phylogenetic relationship of Potamanthidae within Ephemeroptera. The monophyly of Potamanthidae and the monophyly of Rhoenanthus and Potamanthus were supported in the results. The phylogenetic relationship of Potamanthidae + (Ephemeridae + Polymitarcyidae) was also recovered with a high prior probability. The divergence times of Potamanthidae were traced to be 90.44 Mya (95% HPD, 62.80–121.74 Mya), and the divergence times of Rhoenanthus and Potamanthus originated at approximately 64.77 Mya (95% HPD, 43.82–88.68 Mya), thus belonging to the late Pliocene Epoch or early Miocene Epoch. In addition, the data indicated that R. coreanus was under negative selection and that ATP8 and ND2 in Potamanthidae had a high evolutionary rate.