The abundance and distribution of cyanobacterial diazotrophs were quantified in two regions (Melanesian archipelago, MA; and subtropical gyre, SG) of the western tropical South Pacific using nifH quantitative polymerase chain reaction (qPCR) assays. UCYN-A1 and A2 host populations were quantified using 18S rRNA qPCR assays including one newly developed assay. All phylotypes were detected in the upper photic zone (0-50 m), with higher abundances in the MA region. Trichodesmium and UCYN-B dominated and ranged from 2.18 x 10(2) to 9.41 x 10(6) and 1.10 x 10(2) to 2.78 x 10(6) nifH copies L-1, respectively. Het-1 (symbiont of Rhizosolenia diatoms) was the next most abundant (1.40 x 10(1)-1.74 x 10(5) nifH copies L-1) and co-occurred with het-2 and het-3. UCYN-A1 and A2 were the least abundant diazotrophs and were below detection (bd) in 63 and 79, respectively, of 120 samples. In addition, in up to 39% of samples in which UCYN-A1 and A2 were detected, their respective hosts were bd. Pairwise comparisons of the nifH abundances and various environmental parameters supported two groups: a deep-dwelling group (45 m) comprised of UCYN-A1 and A2 and a surface group (0-15 m) comprised of Trichodesmium, het-1 and het-2. Temperature and photosynthetically active radiation were positively correlated with the surface group, while UCYN-A1 and A2 were positively correlated with depth, salinity, and oxygen. Similarly, in a meta-analysis of 11 external datasets, all diazotrophs, except UCYN-A were correlated with temperature. Combined, our results indicate that conditions favoring the UCYN-A symbiosis differ from those of diatom diazotroph associations and free-living cyanobacterial diazotrophs.

Here, we examine the use of bacterial isolates growing in artificial media or seawater as a means to investigate bacterial activity in the upper ocean. The discovery of a major role of bacteria in the ocean's carbon cycle owes greatly to the development of culture-independent assemblage-level approaches; however, this should not detract from the recognition of model isolates as representing the environmental microbiome. A long-established tool for culturing bacteria, in medicine and general microbiology, has been agar plates. In addition, a great variety of liquid substrates including seawater have been used to successfully identify and cultivate important bacteria such as Pelagibacter ubique. Yet, the discrepancy between microscopic counts and plate counts, the great plate count anomaly, has led to a biased perception of the limited relevance of isolated bacteria. Linking isolates to whole-genome sequencing, phylogenetic analysis and computational modeling will result in culturable model bacteria from different habitats. Our main message is that bacterial ecophysiology, particularly growth rates in seawater, and functionalities inferred through the identity, abundance and expression of specific genes could be mechanistically linked if more work is done to isolate, culture and study bacteria in pure cultures. When we rally behind a strategy aimed at culturing targeted phenotypes, we are not saying that culture independent studies of bacteria in the sea are not informative. We are suggesting that culturebased studies can help integrate the ecological and genomic views.

Diatom diazotroph associations (DDAs) are important components in the world's oceans, especially in the western tropical north Atlantic (WTNA), where blooms have a significant impact on carbon and nitrogen cycling. However, drivers of their abundances and distribution patterns remain unknown. Here, we examined abundance and distribution patterns for two DDA populations in relation to the Amazon River (AR) plume in the WTNA. Quantitative PCR assays, targeting two DDAs (het-1 and het-2) by their symbiont's nifH gene, served as input in a piecewise structural equation model (SEM). Collections were made during high (spring 2010) and low (fall 2011) flow discharges of the AR. The distributions of dissolved nutrients, chlorophyll-a, and DDAs showed coherent patterns indicative of areas influenced by the AR. A symbiotic Hemiaulus hauckii-Richelia (het-2) bloom (> 10(6) cells L-1) occurred during higher discharge of the AR and was coincident with mesohaline to oceanic (30-35) sea surface salinities (SSS), and regions devoid of dissolved inorganic nitrogen (DIN), low concentrations of both DIP (> 0.1 mu mol L-1) and Si (> 1.0 mu mol L-1). The Richelia (het-1) associated with Rhizosolenia was only present in 2010 and at lower densities (10-1.76 x 10(5) nifH copies L-1) than het-2 and limited to regions of oceanic SSS (> 36). The het-2 symbiont detected in 2011 was associated with H. membranaceus (> 10(3) nifH copies L-1) and were restricted to regions with mesohaline SSS (31.8-34.3), immeasurable DIN, moderate DIP (0.1-0.60 mu mol L-1) and higher Si (4.19-22.1 mu mol L-1). The piecewise SEM identified a profound direct negative effect of turbidity on the het-2 abundance in spring 2010, while DIP and water turbidity had a more positive influence in fall 2011, corroborating our observations of DDAs at subsurface maximas. We also found a striking difference in the influence of salinity on DDA symbionts suggesting a niche differentiation and preferences in oceanic and mesohaline salinities by het-1 and het-2, respectively. The use of the piecewise SEM to disentangle the complex and concomitant hydrography of the WTNA acting on two biogeochemically relevant populations was novel and underscores its use to predict conditions favoring abundance and distributions of microbial populations.