Iron (Fe)-oxidizing bacteria have the potential to produce morphologically unique structures that may be used as biosignatures in geological deposits. One particular example is Mariprofundus ferrooxydans, which produces extracellular twisted ribbon-like stalks consisting of ferrihydrite, co-located with organic and inorganic elements. It is currently thought that M. ferrooxydans excrete and co-precipitate polysaccharides and Fe simultaneously; however, the cellular production of these polysaccharides has yet to be confirmed. Here, we report on a time-series study that used scanning transmission X-ray microscopy and C 1s and Ca 2p near-edge X-ray adsorption fine structure spectroscopy to investigate production of polysaccharides over the growth cycle of M. ferrooxydans. The production and morphology of twisted iron stalks were consistent with previous observations, but unexpectedly, in the log phase, the carbon content of the stalks was extremely low. It was not until stationary growth phase that a significant component of carbon was detected on the stalks. During the log phase, low levels of carbon, only detectable when the stalks were thin, suggested that M. ferrooxydans produce an extracellular polysaccharide template onto which the Fe precipitates. By stationary phase, the increased carbon association with the stalks was a result of adsorption of organic compounds that were released during osmotic shock post-stalk production. In the environment, elevated concentrations of DOC could adsorb onto the Fe stalks as well as a number of other elements, for example, Si, P, Ca, which, by preventing chemical interactions between the Fe nanoparticles, will prevent structural deformation during recrystallization and preserve the structure of these filaments in the rock record.