Characterized by some of the highest naturally occurring sea surface temperatures, the Red Sea remains unexplored regarding the dynamics of heterotrophic prokaryotes. Over 16 months, we used flow cytometry to characterize the abundance and growth of four physiological groups of heterotrophic bacteria: membrane-intact (Live), high and low nucleic acid content (HNA and LNA) and actively respiring (CTC+) cells in shallow coastal waters. Chlorophyll a, dissolved organic matter (DOC and DON) concentrations, and their fluorescent properties were also measured as proxies of bottom-up control. We performed short-term incubations (6 days) with the whole microbial community (Community treatment), and with the bacterial community only after removing predators by filtration (Filtered treatment). Initial bacterial abundances ranged from 1.46 to 4.80 × 105 cells mL-1. Total specific growth rates in the Filtered treatment ranged from 0.76 to 2.02 d-1. Live and HNA cells displayed similar seasonal patterns, with higher values during late summer and fall (2.13 and 2.33 d-1, respectively) and lower in late spring (1.02 and 1.01 d-1, respectively). LNA cells were outgrown by the other physiological groups (0.33–1.08 d-1) while CTC+ cells (0.28–1.85 d-1) showed weaker seasonality. The Filtered treatment yielded higher bacterial abundances than the Community treatment in all but 2 of the incubations, and carrying capacities peaked in November 2016 (1.04 × 106 cells mL-1), with minimum values (3.61 × 105 cells mL-1) observed in May 2017. The high temperatures experienced from May through October 2016 (33.4 ± 0.4°C) did not constrain the growth of heterotrophic bacteria. Indeed, bacterial growth efficiencies were positively correlated with environmental temperature, reflecting the presence of more labile compounds (high DON concentrations resulting in lower C:N ratios) in summer. The overall high specific growth rates and the consistently higher carrying capacities in the Filtered treatment suggest that strong top-down control by protistan grazers was the likely cause for the low heterotrophic bacteria abundances.

Determination of the abundances of aquatic microbes (i.e., oxygenic and anoxygenic phototrophic and heterotrophic prokaryotes, small phototrophic and heterotrophic eukaryotes and viruses) is nowadays relatively straightforward with the use of flow cytometry. In addition, the technique can be used to test for relative differences in the activity or physiological state of some of these microbial groups, and several indices of community structure can be derived from community composition and flow cytometric signal variability. The technique is sometimes also useful to determine the presence of nonliving organic and inorganic substances and their interaction with the microbes. Here, we provide comprehensive guidance in the use of flow cytometry for these purposes and finally illustrate the usefulness of some of these approaches with data generated in an experiment in which we added oil from a tanker spill to a coastal bacterioplankton community.

We now have a relatively good idea of how bulk microbial processes shape the cycling of organic matter and nutrients in the sea. The advent of the molecular biology era in microbial ecology has resulted in advanced knowledge about the diversity of marine microorganisms, suggesting that we might have reached a high level of understanding of carbon fluxes in the oceans. However, it is becoming increasingly clear that there are large gaps in the understanding of the role of bacteria in regulating carbon fluxes. These gaps may result from methodological as well as conceptual limitations. For example, should bacterial production be measured in the light? Can bacterial production conversion factors be predicted, and how are they affected by loss of tracers through respiration? Is it true that respiration is relatively constant compared to production? How can accurate measures of bacterial growth efficiency be obtained? In this paper, we discuss whether such questions could (or should) be addressed. Ongoing genome analyses are rapidly widening our understanding of possible metabolic pathways and cellular adaptations used by marine bacteria in their quest for resources and struggle for survival (e.g. utilization of light, acquisition of nutrients, predator avoidance, etc.). Further, analyses of the identity of bacteria using molecular markers (e.g. subgroups of Bacteria and Archaea) combined with activity tracers might bring knowledge to a higher level. Since bacterial growth (and thereby consumption of DOC and inorganic nutrients) is likely regulated differently in different bacteria, it will be critical to learn about the life strategies of the key bacterial species to achieve a comprehensive understanding of bacterial regulation of C fluxes. Finally, some processes known to occur in the microbial food web are hardly ever characterized and are not represented in current food web models. We discuss these issues and offer specific comments and advice for future research agendas.

The vast majority of marine dissolved organic carbon (DOC), the largest reservoir of reduced carbon on Earth, is believed to accumulate in the abyssal layers of the ocean over timescales of decades to millennia. However, evidence is growing that small animals that migrate vertically every day from the surface to mesopelagic layers are significantly contributing to the active vertical flux of organic matter. Whether that represents an important source of carbon available for microbial production and respiration at the mesopelagic realm, and its contribution to oceanic carbon budgets and energy flows, is yet to be explored. Here we present data suggesting that Red Sea migrating animals may produce an overlooked source of labile DOC (used at a mean rate of 2.1 µmol C L −1 d −1 ) that does not accumulate but fuels the metabolism in the twilight zone, generating a disregarded hotspot for heterotrophic prokaryotes.

We examined the potential response of Southern Ocean pelagic ecosystems to warming through changes in total primary production (particulate plus dissolved 5 PPP + DPP) and bacterial production (BP), determined simultaneously at ambient temperature (21.4 to 0.4uC) and at 2uC in eight experiments performed near the Antarctic Peninsula in late spring 2002. Short (,6 h) time course experiments of radiocarbon uptake and photosynthesis-irradiance relationships consistently showed that a significant amount of photosynthate appeared as dissolved substances, with a mean 35% extracellular release (PER). Whereas PPP remained virtually unchanged (0.7 mg C m 23 h 21 ), DPP increased significantly at 2uC from 0.5 to 0.9 mg C m 23 h 21 . The corresponding increase in PER (54% on average) was significantly and positively correlated with the temperature difference among treatments, suggesting that an increase in DPP could be expected with a temperature rise in the Southern Ocean. BP, estimated via [ 3 H]leucine incorporation, tended to increase at 2uC only at low absolute values, and this increment was inversely related to PPP. However, our results show that the estimated bacterial carbon demand (BCD) was generally well below concurrent DPP at both treatments (mean BCD:DPP ratios of 0.60 and 0.27 at ambient temperature and 2uC, respectively), indicating that temperature-related extra inputs of organic substrates were not fully and immediately processed by bacteria. To the extent that these results reflect general ecophysiological trends, warming of Southern Ocean surface waters could produce changes in plankton-mediated biogeochemical processes leading to a greater importance of dissolved organic matter fluxes.The crucial role of the oceans in the Earth's climatic system has motivated efforts aimed at evaluating the effects of global warming on the structure and functioning of pelagic ecosystems and their associated biogeochemical fluxes (Sarmiento et al. 2004). One of the strongest evidences of recent warming has been found in the Southern Ocean (Gille 2002), and according to coupled ocean-atmospheric models, this ocean basin will continue to suffer a major temperature rise in the next decades (IPCC 2001). Because of its geographic location, future changes in the Southern Ocean will affect the other three major basins. The Southern Ocean is also one of the largest sinks of anthropogenic CO 2 on Earth, and large increases in phytoplanktonic biomass and production have been recently predicted as a consequence of temperaturemediated major changes in stratification and growth season length (Sarmiento et al. 2004).Macronutrients are not used fully in many parts of the Southern Ocean (e.g., Tréguer and Jacques 1992), the largest high nutrient-low chlorophyll region of the world. Factors such as ambient suboptimal temperatures for phytoplankton growth, intense vertical mixing, predation pressure, and limitation by silica or iron in certain areas have been suggested as possible explanations (see review by Boyd 2002). In fact, e...