Standard methods for calculating microbial growth rates (μ) through the use of proxies, such as
in situ
fluorescence, cell cycle, or cell counts, are critical for determining the magnitude of the role bacteria play in marine carbon (C) and nitrogen (N) cycles. Taxon-specific growth rates in mixed assemblages would be useful for attributing biogeochemical processes to individual species and understanding niche differentiation among related clades, such as found in
Synechococcus
and
Prochlorococcus
. We tested three novel DNA sequencing-based methods (iRep, bPTR, and GRiD) for evaluating growth of light synchronized
Synechococcus
cultures under different light intensities and temperatures. In vivo fluorescence and cell cycle analysis were used to obtain standard estimates of growth rate for comparison with the sequence-based methods (SBM). None of the SBM values were correlated with growth rates calculated by standard techniques despite the fact that all three SBM were correlated with percentage of cells in S phase (DNA replication) over the diel cycle. Inaccuracy in determining the time of maximum DNA replication is unlikely to account entirely for the absence of relationship between SBM and growth rate, but the fact that most microbes in the surface ocean exhibit some degree of diel cyclicity is a caution for application of these methods. SBM correlate with DNA replication but cannot be interpreted quantitatively in terms of growth rate.
Importance
Small but abundant, cyanobacterial strains such as the photosynthetic
Synechococcus
spp. are essential because they contribute significantly to primary productivity in the ocean. These bacteria generate oxygen and provide biologically-available carbon, which is essential for organisms at higher trophic levels. The small size and diversity of natural microbial assemblages means that taxon-specific activities (e.g., growth rate) are difficult to obtain in the field. It has been suggested that sequence-based methods (SBM) may be able to solve this problem. We find, however, that SBM can detect DNA replication and are correlated with phases of the cell cycle but cannot be interpreted in terms of absolute growth rate for
Synechococcus
cultures growing under a day-night cycle, like that experienced in the ocean.
Green fluorescent protein as a marker to investigate targeting of organellar RNA polymerases of higher plants
in vivo