Sulfur Isotope Fractionation during the Evolutionary Adaptation of a Sulfate-Reducing Bacterium

Abstract

c Dissimilatory sulfate reduction is a microbial catabolic pathway that preferentially processes less massive sulfur isotopes relative to their heavier counterparts. This sulfur isotope fractionation is recorded in ancient sedimentary rocks and generally is considered to reflect a phenotypic response to environmental variations rather than to evolutionary adaptation. Modern sulfate-reducing microorganisms isolated from similar environments can exhibit a wide range of sulfur isotope fractionations, suggesting that adaptive processes influence the sulfur isotope phenotype. To date, the relationship between evolutionary adaptation and isotopic phenotypes has not been explored. We addressed this by studying the covariation of fitness, sulfur isotope fractionation, and growth characteristics in Desulfovibrio vulgaris Hildenborough in a microbial evolution experiment. After 560 generations, the mean fitness of the evolved lineages relative to the starting isogenic population had increased by ϳ17%. After 927 generations, the mean fitness relative to the initial ancestral population had increased by ϳ20%. Growth rate in exponential phase increased during the course of the experiment, suggesting that this was a primary influence behind the fitness increases. Consistent changes were observed within different selection intervals between fractionation and fitness. Fitness changes were associated with changes in exponential growth rate but changes in fractionation were not. Instead, they appeared to be a response to changes in the parameters that govern growth rate: yield and cell-specific sulfate respiration rate. We hypothesize that cell-specific sulfate respiration rate, in particular, provides a bridge that allows physiological controls on fractionation to cross over to the adaptive realm. Dissimilatory sulfate reduction (DSR) is a microbial metabolism that consumes sulfate and utilizes this sulfur as a terminal electron acceptor, excreting sulfide. This process creates characteristic enrichments and depletions in the stable isotopes of sulfur that are preserved in sediments and sedimentary rocks as a legacy of the metabolic processing (1). In this way, sulfur isotope fractionation can be thought of as a phenotypic trait of the specific microbes that perform DSR. When the rock record is examined like this, the S isotope phenotype has been interpreted to be continually present in ancient sediments back to at least Ϸ3.5 billion years ago (2). However, the interpretation of S isotope fractionation as a phenotypic trait that can be preserved in ancient rocks opens up a basic question: does evolutionary adaptation influence the S isotope phenotype?Evolution-driven modifications to lineages of sulfate reducers (3, 4) may be capable of influencing the isotope phenotype by modifying the relative processing rates within the DSR pathway. If growth and, in turn, the energy supplied by sulfate respiration influence survival, then the controls on sulfate uptake, the internal regulation of concentrations of metabolites, and the struct...

Experimental Design, Population Dynamics, and Diversity in Microbial Experimental Evolution

“…While founded by a common ancestor, separate ancestral clones are preferentially used when starting these parallel lines to avoid a potential skew toward mutations that might already be present initially by chance in the founding clone. These ancestral clones are often also genetically labeled, expressing either a fluorescent protein (42), antibiotic resistance (43,44), or a specific pattern of resistance to phages or displaying a specific colony color (45). The label allows the detection of external contamination or cross-contamination between lines in…”

Quantification and isotopic analysis of intracellular sulfur metabolites in the dissimilatory sulfate reduction pathway

“…Given the large variations in natural abundance of sulfur isotopes up to several percent, numerous attempts have been made to relate the magnitude of isotopic fractionation to their genetic (Detmers et al, 2001), evolutionary (Pellerin et al, 2015), and environmental controls, including levels of electron acceptor or donor (Harrison and Thode, 1958;Chambers et al, 1975;Habicht et al, 2005;Hoek et al, 2006;Sim et al, 2011a;Leavitt et al, 2013), limitation of other nutrients (Sim et al, 2012), and temperature (Canfield et al, 2006;Mitchell et al, 2009). These studies provide a general consensus that limiting the supply of electron donor to the sulfate reduction pathway leads to larger sulfur isotope effects, while depletion of terminal electron acceptor, sulfate, decreases the magnitude of fractionation (see Figure 6 of Bradley et al, 2016).…”

Diversity decoupled from sulfur isotope fractionation in a sulfate‐reducing microbial community

“…x is elemental sulfur (S 0 ; blue data points), chromium‐reducible sulfur (CRS; green data points), or hydrogen sulfide (H 2 S; red data points). Small black data points are assembled from literature of cultured sulfate reducers, and all values are 34 ε sulfide‐sulfate (Sim, Ono, Donovan, Templer, & Bosak,; Leavitt, Halevy, Bradley, & Johnston, ; Pellerin, Anderson‐Trocmé, et al, ). Isotopic notation ( 33 λ, 34 ε) is described in Methods.…”

“…Tracking biological fractionation of sulfur isotopes through interpretation of mineralized sulfur compounds provides a metric of biogeochemical activity and the role of microbial metabolisms in the cycling of sulfur, both in modern and ancient environments (Canfield, 2004). Strikingly, measured differences in environmental sulfur isotope fractionation are only ever interpreted in terms of environmental controls, despite strong evidence for species-specific isotope enrichment effects (Figure 1b; Detmers, Bruchert, Habicht, & Kuever, 2001;Pellerin, Anderson-Trocmé, et al, 2015;Zaarur, Wang, Ono, & Bosak, 2017). There is no known relationship between microbial diversity and isotopic fractionation.…”