Isotopic Signatures of Carbon Transfer in a Proterozoic Analogue Microbial Mat.

Modern microbial mats are potential analogues for Proterozoic ecosystems, yet only a few studies have characterized mats under low-oxygen conditions that are relevant to Proterozoic environments. Here, we use protein-stable isotope fingerprinting (P-SIF) to determine the protein carbon isotope (δ13C) values of autotrophic, heterotrophic, and mixotrophic organisms in a benthic microbial mat from the low-oxygen Middle Island Sinkhole, Lake Huron, USA (MIS). We also measure the δ13C values of the sugar moieties of exopolysaccharides (EPS) within the mat to explore the relationships between cyanobacterial exudates and heterotrophic anabolic carbon uptake. Our results show that Cyanobacteria (autotrophs) are 13C-depleted, relative to sulfate-reducing bacteria (heterotrophs), and 13C-enriched, relative to sulfur oxidizing bacteria (autotrophs or mixotrophs). We also find that the pentose moieties of EPS are systematically enriched in 13C, relative to the hexose moieties of EPS. We hypothesize that these isotopic patterns reflect cyanobacterial metabolic pathways, particularly phosphoketolase, that are relatively more active in low-oxygen mat environments, rather than oxygenated mat environments. This results in isotopically more heterogeneous C sources in low-oxygen mats. While this might partially explain the isotopic variability observed in Proterozoic mat facies, further work is necessary to systematically characterize the isotopic fractionations that are associated with the synthesis of cyanobacterial exudates. IMPORTANCE The δ13C compositions of heterotrophic microorganisms are dictated by the δ13C compositions of their organic carbon sources. In both modern and ancient photosynthetic microbial mats, photosynthetic exudates are the most likely source of organic carbon for heterotrophs. We measured the δ13C values of autotrophic, heterotrophic, and mixotrophic bacteria as well as the δ13C value of the most abundant photosynthetic exudate (exopolysaccharide) in a modern analogue for a Proterozoic environment. Given these data, future studies will be better equipped to estimate the most likely carbon source for heterotrophs in both modern environments as well as in Proterozoic environments preserved in the rock record.

Absence of canonical trophic levels in a microbial mat.

In modern ecosystems, the carbon stable isotope (δ13 C) ratios of consumers generally conform to the principle "you are what you eat, +1‰." However, this metric may not apply to microbial mat systems where diverse communities, using a variety of carbon substrates via multiple assimilation pathways, live in close physical association and phagocytosis is minimal or absent. To interpret the δ13 C record of the Proterozoic and early Paleozoic, when mat-based productivity likely was widespread, it is necessary to understand how a microbially driven producer-consumer structure affects the δ13 C compositions of biomass and preservable lipids. Protein Stable Isotope Fingerprinting (P-SIF) is a recently developed method that allows measurement of the δ13 C values of whole proteins, separated from environmental samples and identified taxonomically via proteomics. Here, we use P-SIF to determine the trophic relationships in a microbial mat sample from Chocolate Pots Hot Springs, Yellowstone National Park (YNP), USA. In this mat, proteins from heterotrophic bacteria are indistinguishable from cyanobacterial proteins, indicating that "you are what you eat, +1‰" is not applicable. To explain this finding, we hypothesize that sugar production and consumption dominate the net ecosystem metabolism, yielding a community in which producers and consumers share primary photosynthate as a common resource. This idea was validated by confirming that glucose moieties in exopolysaccharide were equal in δ13 C composition to both cyanobacterial and heterotrophic proteins, and by confirming that highly 13 C-depleted fatty acids (FAs) of Cyanobacteria dominate the lipid pool, consistent with flux-balance expectations for systems that overproduce primary photosynthate. Overall, the results confirm that the δ13 C composition of microbial biomass and lipids is tied to specific metabolites, rather than to autotrophy versus heterotrophy or to individual trophic levels. Therefore, we suggest that aerobic microbial heterotrophy is simply a case of "you are what you eat."