Bromodeoxyuridine labelling and fluorescence‐activated cell sorting of polyamine‐transforming bacterioplankton in coastal seawater

Abstract

Polyamines (PAs) are a group of nitrogen-rich dissolved organic nitrogen (DON) compounds that are ubiquitously distributed in marine environments. To identify bacteria that are involved in PA transformations, coastal bacterioplankton microcosms were amended with a single PA model compound, i.e. putrescine (PUT) or spermidine (SPD), or with no addition as controls (CTRs). Bromodeoxyuridine (BrdU) was added to all the microcosms to label newly synthesized DNAs. Fluorescence-activated cell sorting (FACS) analysis indicated significant increases in numbers of total cells and cells with both high and low levels of BrdU incorporation in the PUT and SPD microcosms, but not in the CTRs. 16S rDNA pyrotag sequencing of FACS-sorted cells indicated that PUT- and SPD-transforming bacteria were composed similarly of a diverse group of taxa affiliated with Actinobacteria, Bacteroidetes, Firmicutes and Proteobacteria (especially Roseobacter of its alpha lineage). Broad taxonomic distribution of PA-transforming bacteria was also indicated by the abundance and distribution of PA transporter gene homologues in a survey of sequenced marine bacterial genomes. Our results suggest that PAs may be common DON substrates for marine bacterioplankton, in line with the hypothesis that bacterially mediated PA transformation accounts for an important proportion of marine DON flux.

Identification of polyamine‐responsive bacterioplankton taxa in South Atlantic Bight

“…D). The importance of Vibrionaceae , and of Gammaproteobacteria in general, in PA utilization is in accordance with the common occurrence of PA metabolizing genes among genomes of marine Gammaproteobacteria (Mou et al ., ).…”

“…However, the relative abundance of SAR11 did not increase significantly relative to CTRs in any of the PUT or SPD treatments. Similarly, SAR11 did not respond strongly in a recent study of PA‐responsive bacterioplankton at an inshore site (Mou et al ., ). These observations do not necessarily exclude SAR11 from PA‐using assemblages because the growth rate of SAR11 is between 0.13 and 0.72 per day (Eilers et al ., ; Yokokawa et al ., ; Malmstrom et al ., ) and the incubation time (2 days) used in this and the previous study may be too short to allow the relative abundance of SAR11 and other slow‐growing bacterioplankton taxa to increase significantly in PUT‐ and SPD‐amended treatments.…”

“…A). The dominance of Rhodobacteraceae is in agreement with prior studies (Mou et al ., , ) and an in silico study of the distribution of PA‐metabolizing genes among marine bacterial genomes (Mou et al ., ). Rhodobacteraceae , especially the roseobacter clade, represents a numerically (up to 25% of bacterioplankton) and ecologically important bacterial lineage with broad capacity for processing plankton‐derived DOC in coastal marine environments (Buchan et al ., ; Brinkhoff et al ., ).…”

“…However, the two studies yielded contrasting results concerning the importance of SAR11 in PA transformation, probably due to the method difference. The metatranscriptomic study suggested that SAR11 were important PA transformers (Mou et al ., ), but the 16S rRNA gene pyrotag sequencing‐based study suggested they were not (Mou et al ., ).…”

“…Direct investigations of PA metabolizing bacterioplankton communities are just emerging. So far, only two such studies have been reported and they both were conducted at the same inshore site on Sapelo Island, Georgia (Mou et al ., , ). Both studies found that PUT and SPD are used by similar bacterial taxa, with roseobacters as an important functional lineage (Mou et al ., ).…”