UC Santa Cruz

Photoarsenotrophy is an anoxygenic photosynthesis-dependent arsenite oxidation pathway encoded by the Arx gene cluster and is linked to light-dark cycling of arsenic in freshwater environments.

This dissertation uses techniques from microbiology, molecular genetics, DNA sequencing, and analytical chemistry to characterize photoarsenotrophy in freshwater environments. The hypothesis is that photoarsenotrophy occurs in freshwater environments and is associated with light-dark cycling of arsenic, which may have a different effect on arsenic biogeochemical cycling when compared to Arx-type arsenotrophy. Light-dark arsenic redox cycling is defined herein as variations in arsenic species, arsenite and arsenate, that correlate to light or dark phases.

Since the discovery of photoarsenotrophy in 2008, less than a dozen Arx-dependent arsenotrophs have been isolated, and light-dependent arsenite oxidation has only been detected in three genera (Ectothiorhodospira sp. strains MLW-1, PHS-1, BSL-9, Ect. shaposhnikovii strains DSM 243 and DSM 2001, Halorhodospira halophila SL-1, and now Cereibacter azotoformans str. ORIO). The other studied Arx-dependent arsenotrophs couple arsenite oxidation to anaerobic respiration (i.e., nitrate reduction) instead of anoxygenic photosynthesis (Alkalilimnicola ehrlichii str. MLHE-1, Azoarcus sp. CIB, Sterolibacteraceae strain M52, Desulfotomaculum strain TC-1, and Halomonas sp. ANAO440). In this thesis, I define the light requirement by referring to photosynthesis-dependent arsenite-oxidation as photoarsenotrophy, and photosynthesis-independent Arx-type arsenite-oxidation as Arx-type arsenotrophy. This difference in light requirement is important as it introduces the possibility that light-dark cycling of arsenic could occur in environments containing a photoarsenotroph and arsenate reducer, since Arx-type arsenotrophs are able to oxidize arsenite in the dark so long there is availability of a terminal electron acceptor.

The first question investigated was: Does photoarsenotrophy occur in freshwater environments? This question was studied through the isolation and genetic characterization of a novel photoarsenotroph from freshwater sediments in Owens River, CA, USA. Our results show the photoarsenotrophs are present in freshwater environments and harbor the Arx genes required for photoarsenotrophy.

The second question investigated was: Can arsenic be light-dark cycled in the environment? In other words, do concentrations of different arsenic forms correlate to the light phase or dark phase of a day? To answer this question, we performed anaerobic microcosm studies with sediment collected from Owens River. Arsenic speciation was measured over light-dark cycles and was followed by metagenome sequencing and analysis. The results provide evidence of light-dark cycling in freshwater sediments and the potential genes involved in the cycle.

The third question investigated was: Can we develop a new model organism for studying photoarsenotrophy? We successfully determined that Cereibacter azotoformans str. ORIO is genetically malleable using traditional cloning techniques and can serve as a model for studying the biological mechanism underlying photoarsenotrophy. This was achieved by validating the role of the arxA gene in ORIO and characterizing the physiology surrounding arsenite oxidation.Taken together, these studies show photoarsenotrophy occurs in freshwater environments, ORIO can serve as a model organism for studying photoarsenotrophy, and evidence of light-dark arsenic redox cycling in freshwater sediments.