|dc.description.abstract||The early Earth oceans of the Archaean Eon (approx. 3.8-2.5 billion years ago) are thought to have been predominantly anoxic with ferruginous waters containing high levels of dissolved ferrous iron and low levels of sulfate. How primary production operated in these ferruginous waters, where early life would have evolved, remains poorly understood. Based on the global occurrence of massive banded iron formations that can be dated to as early as the Mesoarchaean Era, many speculate that biological processes helped to drive the oxidation of iron in the Earth’s oceans, which today are sulfate-rich and iron-poor. Along with oxygenic photosynthesis, where produced molecular oxygen can oxidize iron abiotically, alternative microbial processes such as photoferrotrophy, where ferrous iron serves as the direct electron donor for phototrophy, have been implicated in the deposition of banded iron formations. Other forms of anoxygenic phototrophy, such as phototrophic sulfide oxidation, could have also served as a means of primary production in early ferruginous waters. However, the diversity, ecology, and relative importance of anoxygenic forms of phototrophy in Archaean oceans remain poorly characterized given the current global prevalence of molecular oxygen.
Boreal Shield lakes number in the millions across northern regions. Although most Boreal Shield lakes mix and re-oxygenate seasonally, an estimated 15% of these lakes develop anoxic bottom waters over the summer or winter months that gain high levels of dissolved iron while having low levels of sulfate. As such, these Boreal Shield lakes can potentially complement permanently anoxic and ferruginous lakes that have been used to gain insights about the properties of ferruginous waters but are rare globally. The research presented in this thesis aimed to characterize the diversity and ecology of phototroph communities in the ferruginous water columns of seasonally anoxic Boreal Shield lakes. By performing a multi-year lake survey at the International Institute for Sustainable Development Experimental Lakes Area (IISD-ELA) combining high-throughput DNA/RNA sequencing, enrichment cultivation, physicochemical characterization, and rate measurements, the goal of this research was to clarify the metabolic diversity of phototrophy, physicochemical controls on phototrophs, and the relative importance of different modes of phototrophy in Boreal Shield lake anoxic zones with implications for modern ecology and for study of early Earth biogeochemistry.
Metagenomes were sequenced from preliminary DNA samples collected from Lakes 227 and 442 at the IISD-ELA to analyze the functional gene content of the lake microbial communities (Chapter 2). Metagenomes were assembled and contigs binned to recover high-completeness and low-contamination draft genome bins. The genetic potential for sulfide oxidation, indicated by the dsrA gene, was found in three of the four recovered and curated genome bins classified to the Chlorobia class, but the fourth genome bin was found to contain the cyc2 candidate marker gene for iron oxidation and was unique to Lake 227. Two new species of Chlorobia were enriched in sulfide-containing medium from Lakes 227 and 304. Recovered draft genome bins of both novel Chlorobia members contained both the dsrA and cyc2 genes, indicative of the potential for both ferrous iron and sulfide oxidation. The Lake 304 strain, provisionally named, “Candidatus Chlorobium canadense”, was subsequently incubated in ferrous iron-containing medium yet did not show signs of photoferrotrophic activity. Nevertheless, robust methods for the detection of Chlorobia-associated cyc2 homologs were developed, and analysis of reference genomes indicated that the presence of the cyc2 gene is associated with photoferrotrophic potential. Genomic potential for iron- and sulfur-cycling processes were also detected among several other recovered genome bins outside the Chlorobia. Overall, evidence for sulfide-oxidizing phototrophy was found in the lakes despite low sulfate levels, along with genomic potential for photoferrotrophy, and “Ca. Chl. canadense” was enriched to allow for future study of the functional role of cyc2 among the Chlorobia.
Expanding on initial results from cultivation and metagenome analyses, nine Boreal Shield lakes at the IISD-ELA with diverse physicochemical characteristics were surveyed in the summer and/or fall from 2016-2018 to understand the broader distribution of phototrophs among ferruginous lake systems (Chapter 3). Lake anoxic zones samples generally grouped into two categories based on microbial community composition as inferred from 16S rRNA gene and metagenome data. One of those categories was associated with high relative abundances and diversities of anoxygenic phototrophs within the Chlorobia and Chloroflexota phyla and included three lakes. Lake categories based on microbial community composition generally matched categories determined via hierarchical clustering based on lake physicochemical parameters. Compared to the other surveyed lakes, the three detected “phototroph-rich” lakes tended to have low surface area:depth ratios with high dissolved organic carbon content, high dissolved iron content, and low levels of sulfate in their anoxic waters. Anoxic water column metagenomes from the three lakes contained high relative abundances of Chlorobia members along with Chlorobia-associated cyc2 and dsrA genes. Metatranscriptome sequencing revealed that Chlorobia members comprised nearly 50% of total gene expression in the upper anoxic zones of these three lakes based on the rpoB single-copy taxonomic marker gene. Among Chlorobia members, the dsrA gene for phototrophic sulfide oxidation was highly expressed, whereas the cyc2 gene was expressed at lower levels for two lakes (Lakes 221 and 304) and minimally expressed for the third (Lake 227). Rate incubation data from Lake 227 suggested that oxygenic phototrophy was the dominant contributor to iron oxidation even 1 m below the measured oxic/anoxic zone boundary. Measured iron oxidation rates exceeded measured rates of iron reduction at all measured regions of the upper anoxic water column. Sulfide-oxidizing phototrophy was also detected and was tightly coupled to sulfate reduction. Photoferrotrophy was not detected but may have occurred at rates less than the observed iron reduction rate. Metatranscriptome data also revealed high expression of the particulate methane monooxygenase (pmoA) gene deep in the anoxic zone of Lake 227, implying that novel forms of methane oxidation occur in this lake. Altogether, the data demonstrate that Boreal Shield lakes commonly contain high relative abundance and diverse phototroph communities in their seasonal anoxic zones. In addition, the lake anoxic zones appear to host active and cryptic sulfur cycles, along with the potential for iron oxidation/reduction and methane cycling.
Enrichment cultivation in a ferrous iron-containing medium from Lake 227 allowed for the recovery and characterization of a novel anoxygenic phototrophic Chloroflexota member, provisionally named “Candidatus Chlorohelix allophototropha” (Chapter 4). This organism performs phototrophy using a distinct fourth clade of Type I photosynthetic reaction center (RCI) protein, despite placing sister taxonomically to Type II reaction center (RCII)-utilizing Chloroflexota members. “Ca. Chx. allophototropha” contains chlorosomes, uses bacteriochlorophyll c, and encodes the FMO protein like other RCI-utilizing phototrophs in the Chlorobiales and Chloracidobacterales orders. “Ca. Chx. allophototropha” also encodes the potential for carbon fixation using the Calvin-Benson-Bassham (CBB) cycle, unlike all known RCI-utilizing phototrophs. The discovery of “Ca. Chx. allophototropha”, as the first representative of a novel Chloroflexota order (i.e., “Ca. Chloroheliales”), sheds light on longstanding questions about the evolution of photosynthesis, including the origin of chlorosomes among RCII-utilizing Chloroflexota members. The Chloroflexota is now the only phylum outside the Cyanobacteria containing genomic potential for both major classes of photosynthetic reaction center and can thus serve as an additional system for exploring fundamental questions about the evolution of photosynthesis.||en