Does metabolic water control the isotopic composition of water in microbial cells?

Metabolic water, the water that is produced from O₂ during respiration, carries an isotopic signature that can be different from that of the water the cell is growing in. It has been well known that for large land organisms, like birds and mammals, metabolic water contributes significantly to the water balance and has an important control on the signature of the oxygen-stable-isotopes of the water inside the organism. This isotopic signature is then carried over through isotopic equilibrium to other oxygen-bearing species like phosphate. However, for small organisms like bacteria, it has been widely assumed for decades, that the large surface area to volume ratio enables a fast exchange of the cell water with the ambient water. As a result, the isotopic signature of the metabolic water will be heavily diluted and erased. In contrast, a recent work reported indirect evidence of significant control of metabolic water on the oxygen isotopes inside microbial cells. This indirect evidence is based on deviations of oxygen isotopes in phosphate from the expected equilibrium with the ambient water. Here we report the results of experiments that directly tested the possible contribution of metabolic water to phosphate oxygen isotopes in bacteria. We found that ambient water did control the oxygen isotopes in the phosphate. However, there were large deviations from the expected equilibrium. Nevertheless, we found that these deviations were not correlated with the isotopic composition of metabolic water. Hence, other mechanisms, which will be discussed, are responsible for these deviations.