Profiling Gene Expression to Distinguish the Likely Active Diazotrophs from a Sea of Genetic Potential in Marine Sediments

Nitrogen (N) cycling microbial communities in marine sediments are extremely diverse, and it is unknown whether this diversity reflects extensive functional redundancy. Sedimentary denitrifiers remove significant amounts of N from the coastal ocean and diazotrophs are typically regarded as inconsequential. Recently, N fixation has been shown to be a potentially important source of N in estuarine and continental shelf sediments. Analysis of expressed genes for nitrite reductase (nirS) and a nitrogenase subunit (nifH) was used to identify the likely active denitrifiers and nitrogen fixers in surface sediments from different seasons in Narragansett Bay (Rhode Island, USA). The overall diversity of diazotrophs expressing nifH decreased along the estuarine gradient from the estuarine head to an offshore continental shelf site. Two groups of sequences related to anaerobic sulphur/iron reducers and sulphate reducers dominated libraries of expressed nifH genes. Quantitative polymerase chain reaction (qPCR) and quantitative reverse transcription polymerase chain reaction (qRT-PCR) data shows the highest abundance of both groups at a mid bay site, and the highest nifH expression at the head of the estuary, regardless of season. Several potential environmental factors, including water temperature, oxygen concentration and metal contamination, may influence the abundance and nifH expression of these two bacterial groups

Database of diazotrophs in global ocean: abundance, biomass and nitrogen fixation rates

Marine N2 fixing microorganisms, termed diazotrophs, are a key functional group in marine pelagic ecosystems. The biological fixation of dinitrogen (N2) to bioavailable nitrogen provides an important new source of nitrogen for pelagic marine ecosystems and influences primary productivity and organic matter export to the deep ocean. As one of a series of efforts to collect biomass and rates specific to different phytoplankton functional groups, we have constructed a database on diazotrophic organisms in the global pelagic upper ocean by compiling about 12 000 direct field measurements of cyanobacterial diazotroph abundances (based on microscopic cell counts or qPCR assays targeting the nifH genes) and N2 fixation rates. Biomass conversion factors are estimated based on cell sizes to convert abundance data to diazotrophic biomass. The database is limited spatially, lacking large regions of the ocean especially in the Indian Ocean. The data are approximately log-normal distributed, and large variances exist in most sub-databases with non-zero values differing 5 to 8 orders of magnitude. Reporting the geometric mean and the range of one geometric standard error below and above the geometric mean, the pelagic N2 fixation rate in the global ocean is estimated to be 62 (52–73) Tg N yr?1 and the pelagic diazotrophic biomass in the global ocean is estimated to be 2.1 (1.4–3.1) Tg C from cell counts and to 89 (43–150) Tg C from nifH-based abundances. Reporting the arithmetic mean and one standard error instead, these three global estimates are 140 ± 9.2 Tg N yr?1, 18 ± 1.8 Tg C and 590 ± 70 Tg C, respectively. Uncertainties related to biomass conversion factors can change the estimate of geometric mean pelagic diazotrophic biomass in the global ocean by about ±70%. It was recently established that the most commonly applied method used to measure N2 fixation has underestimated the true rates. As a result, one can expect that future rate measurements will shift the mean N2 fixation rate upward and may result in significantly higher estimates for the global N2 fixation. The evolving database can nevertheless be used to study spatial and temporal distributions and variations of marine N2 fixation, to validate geochemical estimates and to parameterize and validate biogeochemical models, keeping in mind that future rate measurements may rise in the future. The database is stored in PANGAEA (doi:10.1594/PANGAEA.774851)

Doing synthetic biology with photosynthetic microorganisms

The use of photosynthetic microbes as synthetic biology hosts for the sustainable production of commodity chemicals and even fuels has received increasing attention over the last decade. The number of studies published, tools implemented, and resources made available for microalgae have increased beyond expectations during the last few years. However, the tools available for genetic engineering in these organisms still lag those available for the more commonly used heterotrophic host organisms. In this mini-review, we provide an overview of the photosynthetic microbes most commonly used in synthetic biology studies, namely cyanobacteria, chlorophytes, eustigmatophytes and diatoms. We provide basic information on the techniques and tools available for each model group of organisms, we outline the state-of-the-art, and we list the synthetic biology tools that have been successfully used. We specifically focus on the latest CRISPR developments, as we believe that precision editing and advanced genetic engineering tools will be pivotal to the advancement of the field. Finally, we discuss the relative strengths and weaknesses of each group of organisms and examine the challenges that need to be overcome to achieve their synthetic biology potential.Peer reviewe

Cyanobacterial nitrogen fixation in the ocean: Diversity, regulation and ecology

Nitrogen is an essential and major component of biomass. While virtually all life depends on combined forms of nitrogen that are usually limited in availability, some prokaryotes, including many groups of cyanobacteria, can use the ubiquitous atmospheric dinitrogen (N2). As photoautotrophic bacteria they can easily meet the energy demand that is required by nitrogenase, the enzyme that reduces N2to NH3. However, nitrogenase is very sensitive to oxygen and the oxygenic cyanobacteria have evolved various strategies to cope with this paradox. Primary production in the ocean is generally considered to be limited by nitrogen. In recent years it has become clear that N2-fixing cyanobacteria are important in the nitrogen budget of the surface oceans. Estimates of N2 fixation indicate that approximately half of global N2 fixation occurs in the sea. N2 fixation is not distributed homogenously throughout the oceans. Pelagic diazotrophic cyanobacteria are only found in (sub)tropical oceans and are notably absent in temperate and colder seas. However, at lower salinities in estuaries and other brackish environments, N2-fixing cyanobacteria can be abundant. N2-fixing cyanobacteria are also abundant in benthic mats in coastal and aquatic environments all over the globe, including polar regions. This demonstrates that N2-fixing cyanobacteria are not excluded from temperate and cold marine environments, even though they are only found in the water column of warm oceans. In this chapter we will discuss these aspects and review the existing knowledge of the diversity of N2-fixing cyanobacteria and the factors that determine their global distribution.

Cyanobacterial nitrogen fixation in the ocean : diversity, regulation and ecology

Nitrogen is an essential and major component of biomass. While virtually all life depends on combined forms of nitrogen that are usually limited in availability, some prokaryotes, including many groups of cyanobacteria, can use the ubiquitous atmospheric dinitrogen (N2). As photoautotrophic bacteria they can easily meet the energy demand that is required by nitrogenase, the enzyme that reduces N2 to NH3. However, nitrogenase is very sensitive to oxygen and the oxygenic cyanobacteria have evolved various strategies to cope with this paradox. Primary production in the ocean is generally considered to be limited by nitrogen. In recent years it has become clear that N2-fixing cyanobacteria are important in the nitrogen budget of the surface oceans. Estimates of N2 fixation indicate that approximately half of global N2 fixation occurs in the sea. N2 fixation is not distributed homogenously throughout the oceans. Pelagic diazotrophic cyanobacteria are only found in (sub)tropical oceans and are notably absent in temperate and colder seas. However, at lower salinities in estuaries and other brackish environments, N2-fixing cyanobacteria can be abundant. N2-fixing cyanobacteria are also abundant in benthic mats in coastal and aquatic environments all over the globe, including polar regions. This demonstrates that N2-fixing cyanobacteria are not excluded from temperate and cold marine environments, even though they are only found in the water column of warm oceans. In this chapter we will discuss these aspects and review the existing knowledge of the diversity of N2-fixing cyanobacteria and the factors that determine their global distribution