The Effects of Hydrogen Peroxide on the Circadian Rhythms of Microcystis aeruginosa Haifeng Qian1., Baolan Hu2., Shuqiong Yu 1 , Xiangjie Pan1, Tao Wu 1 , Zhengwei Fu1* 1 College of Biological and Environmental Engineering, Zhejiang University of Technology, Hangzhou, People’s Republic of China, 2 College of Environmental and Resource Sciences, Zhejiang University, Hangzhou, People’s Republic of China Abstract Background: The cyanobacterium Microcystis aeruginosa is one of the principal bloom-forming cyanobacteria present in a wide range of freshwater ecosystems. M. aeruginosa produces cyanotoxins, which can harm human and animal health. Many metabolic pathways in M. aeruginosa, including photosynthesis and microcystin synthesis, are controlled by its circadian rhythms. However, whether xenobiotics affect the cyanobacterial circadian system and change its growth, physiology and biochemistry is unknown. We used real-time PCR to study the effect of hydrogen peroxide (H2O2) on the expression of clock genes and some circadian genes in M. aeruginosa during the light/dark (LD) cycle. Results: The results revealed that H2O2 changes the expression patterns of clock genes (kaiA, kaiB, kaiC and sasA) and significantly decreases the transcript levels of kaiB, kaiC and sasA. H2O2 treatment also decreased the transcription of circadian genes, such as photosynthesis-related genes (psaB, psbD1 and rbcL) and microcystin-related genes (mcyA, mcyD and mcyH), and changed their circadian expression patterns. Moreover, the physiological functions of M. aeruginosa, including its growth and microcystin synthesis, were greatly influenced by H2O2 treatment during LD. These results indicate that changes in the cyanobacterial circadian system can affect its physiological and metabolic pathways. Conclusion: Our findings show that a xenobiotic can change the circadian expression patterns of its clock genes to influence clock-controlled gene regulation, and these influences are evident at the level of cellular physiology. Citation: Qian H, Hu B, Yu S, Pan X, Wu T, et al. (2012) The Effects of Hydrogen Peroxide on the Circadian Rhythms of Microcystis aeruginosa. PLoS ONE 7(3): e33347. doi:10.1371/journal.pone.0033347 Editor: Brett Neilan, University of New South Wales, Australia Received October 26, 2011; Accepted February 14, 2012; Published March 7, 2012 Copyright: 2012 Qian et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was financially supported by the National Basic Research Program of China (No. 2010CB126100), the Natural Science Foundation of China (21077093), Qianjiang talents project of the technology office in Zhejiang province (2010R10033), and the Science Foundation of Chinese University. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: azwfu2003@yahoo.com.cn . These authors contributed equally to this work. Introduction With the exacerbation of eutrophication and chemical pollu- tion, water blooms are increasingly frequent in lakes around the world, which has resulted in severe environmental problems. These include the release of repugnant odors and bottom-layer anoxia, which have significant adverse impacts on aquatic environments. Cyanobacterial blooms also pose a serious health hazard to humans (via water consumption) because they release cyanotoxins, which are toxic or carcinogenic [1]. Cyanotoxins have appeared in many freshwater lakes, such as Lake Erie (North America), Lake Winnipeg (Canada), Lake Victoria (the largest of the African rift lakes), Lakes Biwa and Kasimagaura (one of Japan’s largest lakes) and Lake Taihu (the third-largest freshwater lake in China) [2]. More than 60,000 incidents of intoxication per year have been reported worldwide [3], and millions of dollars have been lost [4]. Furthermore, the development of urban and industrial agriculture is rapidly accelerating the input of nutrient sources into water systems. Thus, it is difficult to control the formation of water blooms only via the input of nutrients, such as nitrogen (N) and phosphorus (P) [5]. Some studies have researched chemical algaecides that inhibit the growth of cyanobacteria. Copper Sulfate (CuSO4) is an effective and cheap algaecide, which was used in reservoirs, lakes and ponds. However, copper compound is difficult to biodegrade, and it is easy to accumulate in organisms’ bodies or sediments. The worry is that the broad application of CuSO4 may cause metal compound secondary pollution. Hydrogen peroxide (H2O2) is a simple oxidizing agent by quickly releasing a single oxygen atom. H2O2 exists in natural freshwater and is photochemically generated from organic constituents present in the water under the sunlight [6]. It is reported that the natural levels of H2O2 can reach up to1025 M (0.34 mg L21) [6,7]. Recently, H2O2 was demonstrated as an effective chemical algaecide to limit cyanobacterial and green algal growth [8,9]. Our previous report also demonstrated that H2O2 inhibits Microcystis aeruginosa growth, and changes physiological and biochemistry process [10]. Given that H2O2 is considered to be benign to the environment, only forming water upon degradation, we selected H2O2 as a potential algaecide to research its toxic mechanism. M. aeruginosa is a unicellular species of cyanobacteria that is commonly involved in the formation of blue-green algae blooms and is widely distributed in eutrophic lakes, ponds and reservoirs PLoS ONE | www.plosone.org 1 March 2012 | Volume 7 | Issue 3 | e33347