Intellectual merit: The reactive oxygen species (ROS), superoxide, hydrogen peroxide, and hydroxyl, can have significant effects on the cycling of metals and organic compounds in aquatic systems. Although it is known that fungi, algae and bacteria are all capable of extracellular ROS production, it has generally been assumed that biological ROS production is negligible in freshwater systems, compared to photo-oxidation of natural organic matter. However, recent results from the Voelker lab indicate that dark biological production of H2O2 is more important than photochemical production. The overall goal of this research is to elucidate the biological role in ROS production in freshwater systems. The specific objectives are to determine which types of organisms are significant sources of reactive oxygen species to freshwater systems, which environmental variables affect ROS production rates, and which biogeochemical processes are likely to be affected by biological ROS production. The PIs will focus on three hypotheses: (i) Biological production of ROS is widespread, and specific microbial groups (which may be bacterial, algal or fungal) are responsible. (ii) Biological production rates by a specific organism can change as a result of changes in the environmental conditions such as available nutrients and light intensity. (iii) Superoxide and Fenton products are generated as a result of biological ROS production, at rates sufficient to affect the cycling of organic compounds and metals. The PIs will cultivate and isolate organisms responsible for ROS production at selected field sites, targeting algal, bacterial, and/or fungal groups. The PIs will then use their work on the cultivated organisms to design molecular techniques to examine the relationship between ROS production rates by natural assemblages of organisms.
Broader Impacts. The proposed work will provide an educational opportunity for several graduate students and undergraduates. The PIs have a strong track record in advising minorities and women, and student recruitment efforts for this proposal will continue to be focused on underrepresented groups in science and engineering. In addition, the PIs will draw on the results of this research to develop a series of lectures on the role of microorganisms in the production of reactive oxygen species and their impact on the health of microbial ecosystems and aquatic biogeochemistry. The PIs will present the lectures at the annual Professional Development Workshop for Science Educators entitled ?Windows on the Invisible World? coordinated by the Microbial Sciences Initiative at Harvard University.
Normal 0 false false false EN-US X-NONE X-NONE Reactive oxygen species (ROS) are key components controlling the chemistry and biology of the natural systems. They can also be toxic. ROS degrade DNA, proteins, and even cell membranes. Formation of ROS is an unfortunate consequence of breathing oxygen, so all oxygen-breathing organisms must strictly control the concentration of ROS inside our cells in order to survive. This is accomplished by producing enzymes (proteins) that degrade ROS to benign products so they can be excreted. If cells don’t properly remove these ROS, there are consequences – cancer and premature aging in humans, cell lysis and death in microorganisms. Yet, high concentrations of the ROS superoxide and hydrogen peroxide are also ubiquitous in the environment, in both fresh and marine waters for instance. (A)biotic reactions that are directly or indirectly coupled to sunlight have been the only recognized sources of environmentally relevant ROS. Photo-excitation of dissolved organic matter has been presumed the primary source of ROS in surface environments. More recently, phytoplankton (algae, diatoms) have also been shown to produce ROS – yet, as with abiotic ROS sources, their abundance and distribution in nature is fundamentally constrained by sunlight. More recent discoveries have indicated that the majority of ROS production in the ocean is produced biologically within the dark – yet the sources and reasons for this production were unknown. The goals of this research were to determine if bacteria are sources of ROS to natural systems, identify the means (e.g., proteins) by which they are making this ROS, and identify the relative contribution of abiotic and biotic source of the ROS hydrogen peroxide within natural freshwater systems. To meet these goals, we conducted both field and laboratory studies. In brief, we have discovered that extracellular production of the ROS superoxide is widespread among a large diversity of marine and freshwater bacteria obtained from a variety of natural habitats. These organisms, that were not regarded as ROS sources previously, produce ROS at rates comparable with phytoplankton, yet are not constrained by light and thus now provide a source of ROS in the dark. In fact, we have measured high concentrations of biologically produced hydrogen peroxide (a derivative of superoxide) in the dark (photic zone) in freshwater ponds on Cape Cod, MA pointing to the importance of bacteria in these fluxes. We also found the production of hydrogen peroxide varies seasonally, shifting from primarily abiotic sources to solely biological depending on the time of year. The biological source also shifts from phytoplankton to bacterial driven seasonally illustrating a complex dynamic in ROS production within freshwater systems. Lastly, as ROS are key components driving the cycling of metals, we also found that these ROS fluxes impact the cycling of the micronutrient manganese. By cycling between two oxidation states, manganese appears to be an important control on the degradation of these ROS in natural systems. In summary, the ROS hydrogen peroxide is ubiquitous in freshwater systems, involving a number of sources that fluctuate seasonally. Identifying bacteria as sources of ROS, including superoxide, has immense implications for the cycling of metals, degradation of carbon, and health of organisms in "dark" environments – a region that comprises ~95% of our global habitat. This research takes us closer to understanding the dynamics of these critical chemicals that will aid in our understanding of the chemistry of natural systems. As ROS are also toxic as observed in massive fish kills during red tides, this research also has implications in defining the controls on environmental health and by extension human health. This research supported the educational and scientific development of two postdoctoral scientists and one undergraduate student at Harvard University and the Woods Hole Oceanographic Institution. This research was also part of a collaborative effort with Dr. Tina Voelker and her students at the Colorado School of Mines. Outreach associated with this research included presentations to elementary school children and K-12 teachers in Colorado and undergraduate students from underrepresented groups in STEM in Massachusetts.
