Two bacteria incorporated inorganic carbon in lab tests, complicating isotope tracing

An Arizona State University-led study found that two bacteria normally classed as consumers of organic carbon also incorporated dissolved inorganic carbon into their cells under every laboratory condition tested. Published September 28 in Applied and Environmental Microbiology, the result matters because scientists use labeled bicarbonate to trace carbon fixation. Some of that signal could come from heterotrophic bacteria, although this experiment does not establish how much they contribute in any particular soil or seawater sample.

Two strains, different amounts

The researchers grew Pseudomonas putida, a soil bacterium, and Pseudoalteromonas carrageenovora, a marine bacterium. They compared temperatures, supplies of organic carbon and stages of growth, adding carbon-13-labeled bicarbonate to every treatment. The label let them track carbon from an inorganic source into bacterial biomass. Both strains incorporated it in all the conditions tested; neither ceased to depend on organic carbon for growth.

The reported share of biomass carbon originating from labeled dissolved inorganic carbon was 0.006% to 0.029% for P. putida and 0.18% to 1.1% for P. carrageenovora. Incorporation was generally greatest when cells were growing rapidly, at each strain’s preferred temperature and with less organic carbon available. Those are overall patterns rather than rules for every individual sample. The two strains also grew in different media and had different potential carbon-fixing enzymes, so the numerical gap does not identify a single cause.

The team measured isotope incorporation in bulk samples and used single-cell imaging on selected samples of the marine strain. Those finer measurements showed differences among individual cells that a bulk average could conceal. The study’s authors also say respiration may have diluted the labeled inorganic-carbon pool during the experiments, potentially making the reported incorporation percentages underestimates. That caveat does not turn the measured uptake into an estimate of how much carbon bacteria store in the environment.

Why a tracer signal needs care

There is independent precedent in field samples. In a 2024 study, Meyer, Morono and Dekas analyzed 3,203 individual microbial cells from Guaymas Basin sediments 3 to 75 metres below the seafloor. The active cells they analyzed were heterotrophic and drew most of their carbon from organic sources, yet also incorporated inorganic carbon. The researchers estimated that inorganic carbon supplied at least 5% of biomass carbon in the heterotrophs they studied. Their setting and method differed from the new two-strain laboratory experiment, so the percentages are not directly interchangeable.

The Guaymas Basin researchers cautioned that bicarbonate uptake alone cannot definitively identify a cell as autotrophic. The ASU-led results give that warning a controlled laboratory example: even bacteria selected as heterotrophs produced a labeled-bicarbonate signal. Together, the studies give researchers a reason to consider heterotrophic uptake when interpreting stable-isotope measurements. They do not show what proportion of a measured signal in any particular environmental study belongs to heterotrophs.

What remains to test

The new findings cover two cultured strains, not the range of bacteria in soils, oceans or subsurface habitats. They establish incorporation into cells under the tested conditions, but do not measure durable carbon storage or an ecosystem-scale climate effect. Principal investigator Elizabeth Trembath-Reichert described the result to ASU as evidence of a spectrum of bacterial carbon use, rather than two wholly separate groups of carbon makers and consumers.

The study’s authors call for further work pairing isotope probing with gene-expression assays to identify which carbon-incorporating enzymes are involved across different isolates. That could help explain why uptake changes with growth conditions and whether the marked difference between these two strains reflects biology, experimental conditions or both. For now, the measured ranges describe these organisms in the laboratory; the contribution of heterotrophs to carbon-isotope signals in the wider environment remains unquantified.