Nitrogen (N2) fixation by heterotrophic non-cyanobacterial diazotrophs is common in marine deep-sea sediments. However, in shallow coastal areas, where resuspension of sediments is extensive, the ...magnitude of sediment-associated N2 fixation during resuspension is unknown. We examined nitrogen fixation in dark slurry incubations with sediments (0–5 cm and 0–10 cm depths) under simulated resuspension from five shallow stations (water depth <1 m) and an anoxic site in the Bay of Gdansk (109 m) in the Baltic Sea. Abiotic variables and the composition of nitrogen fixing organisms (diazotrophs) were measured at the study sites. To estimate the contribution of nitrogen fixing sulfate reducing bacteria to total nitrogen fixation, parallel incubations with sodium molybdate as inhibitor were performed. Our data show low but variable nitrogen fixation rates (n.d. - 23.7 nmol N g−1 d−1), promoted in small-grained sediments associated with increased organic carbon content and high nutrient concentrations in pore waters. Highest nitrogen fixation at the shallow sites was encountered in the upper 0–5 cm of the sediments while rates were negligible below. Sulfate reducing bacteria (e.g. Desulfobacterales and Desulfovibrionales) were responsible for most of the heterotrophic nitrogen fixation and appear as key players for pelagic N2 fixation during resuspension. Our study reveals an important sediment – water coupling, which may be accentuated by the increased storms and resuspension events predicted for the Baltic Sea region.
•Heterotrophic N2 fixation occurs in shallow coastal sediments of the Baltic Sea.•N2 fixation appears to be associated with small-grained sediments, possibly due to elevated levels of organic carbon and nutrients in the pore waters.•Diazotrophic SRB (Desulfobacterales and Desulfovibrio) perform most of dark N2 fixation under simulated resuspension.
The contribution of non-cyanobacterial diazotrophs (NCDs) to total N
fixation in the marine water column is unknown, but their importance is likely constrained by the limited availability of ...dissolved organic matter and low O
conditions. Light could support N
fixation and growth by NCDs, yet no examples from bacterioplankton exist. In this study, we show that the phototrophic NCD,
sp. BAL398, which is a member of the diazotrophic community in the surface waters of the Baltic Sea, can utilize light. Our study highlights the significance of biofilm formation for utilizing light and fixing N
under oxic conditions and the role of cell plasticity in regulating these processes. Our findings have implications for the general understanding of the ecology and importance of NCDs in marine waters.
We investigated dinitrogen (N
2
) fixation activity and diazotroph community composition across the Cape Verde Frontal Zone (CVFZ), from photic epipelagic waters (0-200 m) to aphotic meso- ...(200–1000 m) and bathypelagic (> 1000 m) waters. The highest N
2
fixation rates of 4.1 ± 2.2 nmol N l
− 1
day
− 1
and 7.8 ± 2.3 nmol N l
− 1
day
− 1
were in epipelagic waters south of the front. We detected aphotic N
2
fixation in 5 out of 32 samples, primarily south of the front, and sporadically down to 3,000 m, with rates ranging from 0.03 ± 0.01 nmol N l
− 1
day
− 1
to 0.07 ± 0.01 nmol N l
− 1
day
− 1
. Cyanobacteria dominated the diazotroph community and nitrogenase gene (
nifH
) expression profiles in surface waters and, surprisingly, in aphotic waters. The detection of cyanobacterial
nifH
genes by DNA sequencing and quantitative PCR in the aphotic zone, together with
nifH
expression in meso- and bathypelagic waters, indicates a downward flux of metabolically active cyanobacteria, and points to a contribution to the observed aphotic N
2
fixation rates. In the photic zone, UCYN-A dominated north of the front, whereas
Trichodesmium
was mainly found in the southern region. However, our results also show that cross-frontal advection of cyanobacterial diazotrophs can occur via intrusions of surface water. Salinity, temperature, and mixed layer depth were the main determinants of the diazotroph composition and distribution of the key cyanobacteria. Thus, the front appeared to act as a dynamic barrier controlling the distribution of cyanobacterial diazotrophs.
The CRISPR (clustered regularly interspaced short palindromic repeats) system protects archaea and bacteria by eliminating nucleic acid invaders in a crRNA-guided manner. The Sulfolobus islandicus ...type III-B Cmr-α system targets invading nucleic acid at both RNA and DNA levels and DNA targeting relies on the directional transcription of the protospacer in vivo. To gain further insight into the involved mechanism, we purified a native effector complex of III-B Cmr-α from S. islandicus and characterized it in vitro. Cmr-α cleaved RNAs complementary to crRNA present in the complex and its ssDNA destruction activity was activated by target RNA. The ssDNA cleavage required mismatches between the 5΄-tag of crRNA and the 3΄-flanking region of target RNA. An invader plasmid assay showed that mutation either in the histidine-aspartate acid (HD) domain (a quadruple mutation) or in the GGDD motif of the Cmr-2α protein resulted in attenuation of the DNA interference in vivo. However, double mutation of the HD motif only abolished the DNase activity in vitro. Furthermore, the activated Cmr-α binary complex functioned as a highly active DNase to destroy a large excess DNA substrate, which could provide a powerful means to rapidly degrade replicating viral DNA.
Estuaries receive substantial anthropogenic nitrogen loading and are mainly considered net nitrogen sinks. While several studies have identified diverse diazotrophic communities in estuarine ...sediments, the role of pelagic diazotrophs in these systems is not well understood. We investigated the links between diazotrophic community composition, nitrogenase (nifH) gene expression, N2 fixation, and environmental conditions in Narragansett Bay (USA). Pelagic N2 fixation rates ranged between 0.02 and 9.41 nmol N L−1 d−1 and correlated significantly with fluctuations in diazotroph community composition. These fluctuations were also correlated with temperature, salinity, and mean sea level. The dominant sequences in our pelagic samples were related to sequences previously detected in the bay's sediments and were dominated by nifH gene Clusters I and III. We interpret this as a coupling between sediment and pelagic diazotroph communities and speculate that resuspension plays an important role for pelagic N2 fixation in shallow estuarine environments such as Narragansett Bay. For instance, the finding of active sulfate reducers in the oxygenated water illustrates that the sediment‐pelagic coupling can impact nutrient cycling in shallow environments. The pelagic N2 fixation measured during our study period showed only a minor contribution (< 1%) to the total estimated nitrogen load to Narragansett Bay. However, with intensifying nitrogen management in estuaries, the need to constrain the rates of pelagic N2 fixation in these systems will be essential for estimating nitrogen fluxes within the bay and to the adjacent coastal ecosystems.
Seagrasses can enhance nutrient mobilization in their rhizosphere via complex interactions with sediment redox conditions and microbial populations. Yet, limited knowledge exists on how ...seagrass-derived rhizosphere dynamics affect nitrogen cycling. Using optode and gel-sampler-based chemical imaging, we show that radial O
loss (ROL) from rhizomes and roots leads to the formation of redox gradients around below-ground tissues of seagrass (
), which are co-localized with regions of high ammonium concentrations in the rhizosphere. Combining such chemical imaging with fine-scale sampling for microbial community and gene expression analyses indicated that multiple biogeochemical pathways and microbial players can lead to high ammonium concentration within the oxidized regions of the seagrass rhizosphere. Symbiotic N
-fixing bacteria (
) were particularly abundant and expressed the diazotroph functional marker gene
in
rhizosphere areas with high ammonium concentrations. Such an association between
and
can facilitate ammonium mobilization, the preferred nitrogen source for seagrasses, enhancing seagrass productivity within nitrogen-limited environments. ROL also caused strong gradients of sulfide at anoxic/oxic interfaces in rhizosphere areas, where we found enhanced
transcription by sulfate-reducing bacteria. Furthermore, we found a high abundance of methylotrophic and sulfide-oxidizing bacteria in rhizosphere areas, where O
was released from seagrass rhizomes and roots. These bacteria could play a beneficial role for the plants in terms of their methane and sulfide oxidation, as well as their formation of growth factors and phytohormones. ROL from below-ground tissues of seagrass, thus, seems crucial for ammonium production in the rhizosphere via stimulation of multiple diazotrophic associations.
Seagrasses are important marine habitats providing several ecosystem services in coastal waters worldwide, such as enhancing marine biodiversity and mitigating climate change through efficient carbon sequestration. Notably, the fitness of seagrasses is affected by plant-microbe interactions. However, these microscale interactions are challenging to study and large knowledge gaps prevail. Our study shows that redox microgradients in the rhizosphere of seagrass select for a unique microbial community that can enhance the ammonium availability for seagrass. We provide first experimental evidence that
, including the symbiotic N
-fixing bacteria
, can contribute to the bacterial ammonium production in the seagrass rhizosphere. The release of O
from rhizomes and roots also caused gradients of sulfide in rhizosphere areas with enhanced nifH transcription by sulfate-reducing bacteria. O
release from seagrass root systems thus seems crucial for ammonium production in the rhizosphere via stimulation of multiple diazotrophic associations.
Linezolid resistance in
Enterococcus
spp. is increasingly considered critically important and a public health threat which mandates the need to understand their genomic contents and dissemination ...patterns. Here, we used whole-genome sequencing to characterize the resistome, virulome and mobile genetic elements of nine linezolid-resistant (LZD
R
) enterococci (seven
optrA
-
E. faecalis,
one
poxtA
-
E. faecium
and one
optrA
-
E. casseliflavus
) previously obtained from the nares of healthy dogs, pigs, pig farmers and tracheal samples of nestling storks in Spain. Also, the relatedness of the isolates with publicly available genomes was accessed by core-genome single nucleotide polymorphism (SNP) analysis. The
optrA
gene of the
E. faecalis
and
E. casseliflavus
isolates was located downstream of the
fexA
gene. The
optrA
gene in the
E. casseliflavus
isolate was carried in a plasmid (pURX4962), while those in the seven
E. faecalis
isolates were chromosomally located. The OptrA proteins were mostly variants of wild type (DP-2: Y176
D
/T481
P
; RDK: I104
R
/Y176
D
/E256K; DD-3: Y176
D
/G393
D;
and EDD: K3
E
/Y176
D
/G393
D
), except two that were wild type (one
E. faecalis
and one
E. casseliflavus
). The
poxtA
gene in the
E. faecium
isolate was found alone within its contig. The
cfrD
was upstream of
ermB
gene in the
E. casseliflavus
isolate and flanked by
IS
NCY and
IS
1216
.
All the LZD
R
enterococci carried plasmid
rep
genes (2–3) containing tetracycline, chloramphenicol and aminoglycoside resistance genes. All isolates except
E. casseliflavus
carried at least one intact prophage, of which
E. faecalis
-ST330 (X4957) from a pig carried the highest (
n
= 5). Tn
6260
was associated with
lnuG
in
E. faecalis
-ST330 while Tn
554
was with
fexA
in
E. feaecalis-
ST59 isolates. All except
E. casseliflavus
(
n
= 0) carried at least two metal resistance genes (MRGs), of which
poxtA
-carrying
E. faecium
-ST1739 isolate contained the most (
arsA, copA, fief, ziaA, znuA, zosA, zupT,
and
zur
). SNP-based analyses identified closely related
optrA
-
E. faecalis
isolates from a pig and a pig farmer on the same farm (SNP = 4). Moreover,
optrA
- carrying
E. faecalis
-ST32, -ST59, and -ST474 isolates from pigs were related to those previously described from humans (sick and healthy) and cattle in Spain, Belgium, and Switzerland (SNP range 43–86). These findings strongly suggest the transmission of LZD
R
-
E. faecalis
between a pig and a pig farmer and potential inter-country dissemination. These highlight the need to strengthen molecular surveillance of LZD
R
enterococci in all ecological niches and body parts to direct appropriate control strategies.
Abstract The photosynthetic cyanobacterium Trichodesmium is widely distributed in the surface low latitude ocean where it contributes significantly to N2 fixation and primary productivity. Previous ...studies found nifH genes and intact Trichodesmium colonies in the sunlight-deprived meso- and bathypelagic layers of the ocean (200–4000 m depth). Yet, the ability of Trichodesmium to fix N2 in the dark ocean has not been explored. We performed 15N2 incubations in sediment traps at 170, 270 and 1000 m at two locations in the South Pacific. Sinking Trichodesmium colonies fixed N2 at similar rates than previously observed in the surface ocean (36–214 fmol N cell−1 d−1). This activity accounted for 40 ± 28% of the bulk N2 fixation rates measured in the traps, indicating that other diazotrophs were also active in the mesopelagic zone. Accordingly, cDNA nifH amplicon sequencing revealed that while Trichodesmium accounted for most of the expressed nifH genes in the traps, other diazotrophs such as Chlorobium and Deltaproteobacteria were also active. Laboratory experiments simulating mesopelagic conditions confirmed that increasing hydrostatic pressure and decreasing temperature reduced but did not completely inhibit N2 fixation in Trichodesmium. Finally, using a cell metabolism model we predict that Trichodesmium uses photosynthesis-derived stored carbon to sustain N2 fixation while sinking into the mesopelagic. We conclude that sinking Trichodesmium provides ammonium, dissolved organic matter and biomass to mesopelagic prokaryotes.
Abstract Heterotrophic bacterial diazotrophs (HBDs) are ubiquitous in the pelagic ocean, where they have been predicted to carry out the anaerobic process of nitrogen fixation within low-oxygen ...microenvironments associated with marine pelagic particles. However, the mechanisms enabling particle colonization by HBDs are unknown. We hypothesized that HBDs use chemotaxis to locate and colonize suitable microenvironments, and showed that a cultivated marine HBD is chemotactic toward amino acids and phytoplankton-derived DOM. Using an in situ chemotaxis assay, we also discovered that diverse HBDs at a coastal site are motile and chemotactic toward DOM from various phytoplankton taxa and, indeed, that the proportion of diazotrophs was up to seven times higher among the motile fraction of the bacterial community compared to the bulk seawater community. Finally, three of four HBD isolates and 16 of 17 HBD metagenome assembled genomes, recovered from major ocean basins and locations along the Australian coast, each encoded >85% of proteins affiliated with the bacterial chemotaxis pathway. These results document the widespread capacity for chemotaxis in diverse and globally relevant marine HBDs. We suggest that HBDs could use chemotaxis to seek out and colonize low-oxygen microenvironments suitable for nitrogen fixation, such as those formed on marine particles. Chemotaxis in HBDs could therefore affect marine nitrogen and carbon biogeochemistry by facilitating nitrogen fixation within otherwise oxic waters, while also altering particle degradation and the efficiency of the biological pump.
Nitrogen (N2) fixation rates in the brackish Baltic Sea are among the highest per unit of area in the world. However, beyond the filamentous heterocyst-forming cyanobacteria, knowledge about the ...composition and distribution of N2-fixing microbes (diazotrophs) is limited. To address this, we investigated nitrogenase gene (nifH) composition and expression at coastal (<10 km offshore) and offshore (>10 km offshore) stations, at surface (avg. 1.8 m) and at depth (avg. 24 m) and in free-living (0.2–3.0 μm) and particle-associated size fractions (>3 μm). Surprisingly, nifH genes affiliated with Pseudanabaena and non-cyanobacterial diazotrophs (NCDs) dominated the composition whereas filamentous heterocyst-forming cyanobacteria accounted for almost 80% of the nifH transcripts. Salinity had a minor influence on the composition, but Aphanizomenon and Nodularia showed increased relative nifH gene expression at low and higher salinity, respectively. Pseudanabaena only accounted for up to 5% of the nifH transcripts and nifH gene expression by Candidatus Atelocyanobacterium thalassa (sublineage UCYN-A2) was mainly observed in the most saline western part of the Baltic. The only notable expression by NCDs (up to 15% of nifH transcripts at a given station) coincided with an upwelling event at the southern coast and was largely accounted for by a Pseudomonas-like nifH phylotype, recurrently found in the Baltic Sea. NCD relative abundances were dominant in coastal stations, presumably driven by sediment resuspension as evidenced by higher turbidity and DOC levels and the recovery of sediment diazotrophs in the pelagic zone. This study reveals the heterogeneity of the composition and activity of diazotrophs in the Baltic Sea, and underscores the need for future N2 fixation studies that include coastal and offshore Baltic waters.
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•Pseudanabaena and non-cyanobacteria dominate the nitrogenase (nifH) gene pool.•Heterocyst-forming cyanobacteria accounted for 80% of the nifH gene transcripts.•Coastal upwelling affected composition and activity of diazotrophs.•The composition and activity of diazotrophs in the Baltic Sea is heterogeneous.•Future N2 fixation studies should include coastal and offshore Baltic waters.
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