Dinitrogen fixation in the unicellular diazotroph Crocosphaera watsonii

Crocosphaera watsonii is an abundant organism in the tropical and subtropical ocean and is considered to be an important contributor to the marine nitrogen cycle due to its ability to fix dinitrogen (N2). Light-dark cycles were used to determine the diel variation in N2 fixation and photosynthesis in the unicellular diazotrophic cyanobacterium C. watsonii WH8501. A first set of experiments showed that N2 fixation and photosynthesis were separated temporally with N2 fixation during the dark and photosynthesis during the light periods. Due to the storage of carbon reserves during the day and subsequent respiration at night, C. watsonii links its cellular nitrogen and carbon metabolism. N2 fixation and photosynthesis appear to be regulated and optimized by a circadian rhythm. Gene expression analysis demonstrated cyclic patterns of the major genes involved in nitrogen and carbon metabolism. However, the patterns of expression did not coincide with patterns of activity with the gene expression peaking several hours prior to the activity. In a second set of experiments, N2 fixation and photosynthesis were simultaneously measurable at population level in the respective dark period, i.e. an artificial light period instead of a dark period. Single-cell analysis using nanoSIMS revealed that the co-occurrence of N2 fixation and photosynthesis at population level was probably due to each individual cell’s capability of coping with the O2-sensitivity of the nitrogenase enzyme. In addition, single-cell rates showed large variability with the highest rate about six times higher than the mean rate during the normal dark N2 fixation. The experimental work with a pure culture of a diazotroph provided the opportunity for a re-assessment of the two commonly used methods for the measurement of N2 fixation in both field and laboratory studies, i.e. the 15N2-tracer addition method and the acetylene reduction assay.