Nitrogen cycling in coastal marine ecosystems

Nitrogen cycling in coastal marine ecosystems

R.A. Herbert *Department of Biological Sciences, University of Dundee, Dundee DD1 4HN, Scotland, UK

Received 26 January 1999; revised 15 May 1999; accepted 7 July 1999

Abstract

It is generally considered that nitrogen availability is one of the major factors regulating primary production in temperatecoastal marine environments. Coastal regions often receive large anthropogenic inputs of nitrogen that cause eutrophication.The impact of these nitrogen additions has a profound effect in estuaries and coastal lagoons where water exchange is limited.Such increased nutrient loading promotes the growth of phytoplankton and fast growing pelagic macroalgae while rootedplants (sea-grasses) and benthic are suppressed due to reduced light availability. This shift from benthic to pelagic primaryproduction introduces large diurnal variations in oxygen concentrations in the water column. In addition oxygen consumptionin the surface sediments increases due to the deposition of readily degradable biomass. In this review the physico-chemical andbiological factors regulating nitrogen cycling in coastal marine ecosystems are considered in relation to developing effectivemanagement programmes to rehabilitate seagrass communities in lagoons currently dominated by pelagic macroalgae and/orcyanobacteria. ß 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rightsreserved.

Keywords: N2 ¢xation; Ammoni¢cation; Nitri¢cation; Denitri¢cation; Physico-chemical gradient ; Eutrophication; Anthropogenic input;

Rhizosphere e¡ect; Meiofauna; C/N ratio; Carbon availability

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5642. Nitrogen ¢xation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5653. Ammoni¢cation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5684. Nitri¢cation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5705. Denitri¢cation and nitrate ammoni¢cation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5736. Modelling of nitrogen cycling in coastal marine systems . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5787. E¡ects of anthropogenic inputs on nitrogen cycling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5798. Concluding remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 581Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 582References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 582

0168-6445 / 99 / $20.00 ß 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.PII: S 0 1 6 8 - 6 4 4 5 ( 9 9 ) 0 0 0 2 2 - 4

* Tel. : +44 (1382) 344262; Fax: +44 (1382) 344275.

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1. Introduction

Microbial communities in shallow marine sedi-ments play a key role in the oxidation of complexorganic compounds and regeneration of nutrientsessential for sustaining primary production in theoverlying water column [1^4]. The fundamental sig-ni¢cance and interdependence of these processes iswell exempli¢ed by the biogeochemical cycling ofcarbon and nitrogen. Since these elements are keyconstituents of all living matter it is perhaps notsurprising that the impact of carbon and nitrogenavailability on primary production and mineralisa-tion of organic matter has been the subject of inten-sive study [5^7]. From a biogeochemical viewpointthe two cycles share many common features. Theseinclude the dominance of specialised groups of mi-croorganisms, which carry out speci¢c transforma-tions, and the regulation of these processes by oxy-gen and the prevailing redox regime. Given thatthese cycles are inextricably linked due considerationmust therefore be given to the complex interactionsthat exist between them if any meaningful under-standing of individual transformations is to beachieved. This is well illustrated by nitrogen trans-formations such as heterotrophic nitrogen ¢xationand denitri¢cation that are functionally dependentupon the availability of oxidisable carbon sources[8^10].

It is widely accepted that shallow coastal sedi-ments are important sites for the mineralisation oforganic matter [11,12]. These transformations aremediated principally by bacteria and the resultinggradients of nutrients result in their release to theoverlying water column or adsorption and burial indeeper sediment layers. Several studies of shallowwater coastal ecosystems have indicated that remi-neralisation of nitrogen mediated by the heterotro-phic activity of the microbiota and the larger macro-fauna plays an important role in supporting primaryproduction both in phytoplankton dominated sys-tems and those where macrophytes are the dominantprimary producers [5,13^16]. The driving force forbenthic nitrogen cycling is the degradation of organ-ic matter deposited at the sediment surface or ex-creted by the roots and rhizomes of rooted macro-phytes [17^19].

The major factors controlling the concentrations

of inorganic nitrogen species, principally NO33 and

NH�4 , in the water column of shallow coastal marineecosystems (water depth 0.5^50 m) are inputs arisingfrom £uvial discharges and those resulting from ex-change across the sediment-water interface. Benthicnutrient exchange (benthic £ux) is largely determinedby the rate of detritus sedimentation and decompo-sition and the rate at which nutrients are transportedto or from the overlying water by di¡usion and in-fauna bioturbation [20]. In addition to playing a keyrole in nitrogen recycling, the benthic £ux can beused as a net measure of the individual processesinvolved in sediment nitrogen turnover [21]. Thus,the rates of net ammoni¢cation (NH�4 release fromorganic matter), nitri¢cation (oxidation of NH�4 toNO3

3 ) and denitri¢cation (reduction of NO33 to N2

and N2O) can all be estimated from net benthic£uxes of NO3

3 and NH�4 [21,22].A further factor regulating benthic nutrient regen-

eration is the quantity, quality and spatial distribu-tion of the deposited organic matter in the sediment.When deposition rates are high, heterotrophic micro-organisms are unable to completely degrade the la-bile components before burial or reworking to depthby the benthic infauna. Aerobic respiration whichtakes place in the surface sediment layers (typically0^5 mm depth), results in a rapid depletion of oxy-gen and alternative e3 acceptors if present, such asnitrate, manganese and ferric oxides, sulfate and car-bon dioxide are then sequentially used as oxidants[23,24]. Under these conditions mineralisation pro-ceeds via a sequence of metabolic steps involvingcoupled fermentation and anaerobic respirationprocesses, each of which completes a partial oxida-tion of the organic matter. The result is a spatialand/or temporal succession as successive thermody-namically favourable e3 acceptors are sequentiallydepleted by the indigenous micro£ora. The net e¡ectis that a well-de¢ned vertical biogeochemical zona-tion develops within the sediment except where mac-rofauna burrows allow lateral di¡usion of e3 ac-ceptors from well-irrigated burrow water [25].Concurrent with the oxidation of organic carbon atthese di¡erent depth horizons, organic nitrogen ismineralised principally by deaminitive fermentation.

It is clearly evident from the foregoing section thatnitrogen cycling in marine sediments is subject to acomplex array of regulatory mechanisms involving

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both physico-chemical and biological factors. Theobjective of this review is to provide a synthesis ofrecent developments in our understanding of theprocesses involved in nitrogen cycling in coastal ma-rine sediments.

In order to gain an understanding of the factorsregulating nitrogen cycling in coastal marine sedi-ments there is a need to understand how the individ-ual, microbially mediated processes that make up thenitrogen cycle are controlled. The complexity of thecycle is demonstrated in Fig. 1 which shows thatnitrogen undergoes a series of oxidation/reductionreactions and change in valence state from 33 to+5. These transformations are mediated by a meta-bolically diverse range of autotrophic and heterotro-phic microorganisms and are strongly in£uenced bythe prevailing physico-chemical conditions.

2. Nitrogen ¢xation

The availability of ¢xed nitrogen is considered bymany authorities to be a major factor regulating pri-mary production in shallow marine environments[26^28]. Thus, inputs of `new' nitrogen resultingfrom biological nitrogen ¢xation may enable the pro-ductivity of such ecosystems to increase. Whilst 79%of the earth's atmosphere is composed of molecular

nitrogen (N2), this major nitrogen reservoir is un-available directly to plants and animals. Biologicalnitrogen ¢xation is con¢ned to specialised groupsof prokaryotes which possess the enzyme nitrogenaseand include both autotrophs and heterotrophs. Allmajor groups of cyanobacteria found in the marineenvironment have nitrogen ¢xing representatives, in-cluding unicellular and non-heterocystous species[29^31]. The ability to ¢x nitrogen is also commonamongst members of the Chromatiaceae, Chlorobia-ceae, Chloro£exaceae and Rhodospirillaceae and avariety of chemoautotrophic bacteria [29,30]. Marineheterotrophic nitrogen ¢xing bacteria comprise ataxonomically diverse group (see Table 1) and in-clude aerobes, microaerophiles, facultative and strictanaerobes [29,32]. Since the N7N bond is extremelystable the biological reduction of dinitrogen to am-monia is an energy demanding process estimated to

Fig. 1. The nitrogen cycle showing the chemical forms and keyprocesses involved in the biogeochemical cycling of nitrogen[235].

Table 1Heterotrophic bacteria reported to ¢x nitrogen in marine envi-ronments

Genus Habitat References

AerobesAzotobacter spp. Seagrass sediment [36]

Intertidal sediment [34]Estuarine sediment [37]Salt marsh sediment [38]

MicroaerophilesAzospirillum spp. Spartina roots [39]

Zostera roots [40]Campylobacter spp. Spartina roots [41]Beggiatoa spp. Sediment [42]Facultative anaerobesEnterobacter spp. Beach sediment [43]Klebsiella spp. Intertidal sediment [34]

Estuarine sediment [37]Vibrio spp. Seawater [44]

Zostera roots [45]AnaerobesDesulfobacter spp. Marine sediments [47]Desulfovibrio spp. Seagrass sediment [36]

Intertidal sediment [34]Salt marsh sediment [46]Estuarine sediment [37]

Clostridium spp. Seagrass sediment [36]Intertidal sediment [34]Salt marsh sediment [46]Estuarine sediment [37]

ArchaeaMethanococcus spp. Seawater [48]Methanosarcina spp. Estuarine sediment [49]

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be equivalent to a minimum of 16 ATP per moleculeof N2 ¢xed [33]. Given the high metabolic cost in-volved it is not surprising that diazotrophic photo-autotrophs such as cyanobacteria have a signi¢cantadvantage in photic habitats over their heterotrophiccounterparts whose nitrogen ¢xing capacity is lim-ited by the availability of suitable organic carbonsources [34,35]. Thus, in unvegetated shallow coastallagoons and intertidal sediments where light is notlimiting, dense populations of benthic nitrogen ¢xingcyanobacteria may develop and contribute ¢xed ni-trogen to the local ecosystem. Data presented in Ta-ble 2 show that these cyanobacterial mats in bothtemperate and tropical environments exhibit high ni-trogen ¢xation rates as measured by the acetylenereduction assay (ARA). Whilst the highest rateshave been recorded in the tropics those measuredin temperate salt marsh sediments in the UK andeastern USA are still signi¢cant and substantiallygreater than those recorded for uncolonised sedi-ments (Table 3). However, whilst nitrogen ¢xed bycyanobacterial mats is locally important to the matcommunities themselves their contribution to the to-tal nitrogen budget in most shallow marine ecosys-tems is minor due to their restricted areal distribu-tion. For example, Hanson and Gunderson [56]recorded very high nitrogen ¢xation rates (Table 2)for microbial mats in Kaneohe Bay, Hawaii, yet theyare estimated to contribute only 0.3% of the annualnitrogen input to the Bay. Similarly in salt marshecosystems N-¢xation by cyanobacterial mats is con-sidered to be of less importance as an overall sourceof ¢xed nitrogen than heterotrophic ¢xation [53,67].

In contrast to cyanobacterial mats, N-¢xationrates in unvegetated marine sediments are low rang-ing from 0.002^0.65 g N m32 year31 (Table 3). TheN2 ¢xing community responsible comprises a diversearray of heterotrophic and chemolithoautotrophicbacterial genera with physiologies ranging from strictanaerobes to obligate aerobes (Table 1). The highestrates have been reported in organically rich sedi-ments such as those found in the Waccasassa estu-ary, Bank End salt marsh and Flax Pond mud £ats.This ¢nding is unsurprising given that metabolisablecarbon is required to support heterotrophic N-¢xa-tion and in oligotrophic marine environments theavailability of organic carbon is probably the factorlimiting the nitrogen ¢xing potential of unvegetatedsediments. Herbert [34] reported that amending car-bon depleted sediments from the Tay estuary, Scot-land, with 5 mM glucose stimulated heterotrophic N-¢xation rates under both aerobic (U3) and anaerobic(U2.5) conditions. Similarly, Tibbles et al. [68] dem-onstrated that the plant structural polysaccharidesxylan and alginate stimulated nitrogenase activity5^18-fold when added to salt marsh sedimentsfrom the Langebaan Lagoon, South Africa. Evenhigher levels of stimulation were recorded, 19- to92-fold respectively, when plant storage polysacchar-ides, laminarin and glycogen were added to sedimentsamples compared with untreated controls. Thesedata indicate whilst the potential for high nitrogen¢xing activity in unvegetated coastal marine ecosys-tems exists it is rarely realised due to the lack of

Table 2Nitrogen ¢xation rates reported for cyanobacterial mats

System N-¢xation rate(g N m32 year31)

Reference

Island of Mellum, Germany 0.8^1.5 [50]Sippewisset salt marsh,Massachusetts

1.42 [51]

Colne Point marsh, UK 5.99 [35]Hiddensee Island, Baltic 7.60 [52]Flax Pond salt marsh,New York

13.44 [53]

Bank End, UK 10.06 [54]Enewatak Atoll, MarshallIslands

65.70 [55]

Kaneohe Bay, Hawaii 76.00 [56]

Table 3Nitrogen ¢xation rates reported for uncolonised marine sedi-ments

System Nitrogen ¢xation rate(g N m32 year31)

Reference

Vostok Bay, Japan 0.002 [57]Narragansett Bay,Rhode Island

0.03 [58]

Rhode River Estuary,Maryland

0.13 [59]

Lune Estuary, England 0.14 [37]Waccasassa Estuary,Florida

0.37 [60]

Bank End, England 0.43 [54]Kaneohe Bay, Hawaii 0.60 [56]Flax Pond mud £ats,New York

0.65 [53]

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suitable oxidisable carbon substrates. As a conse-quence, whilst unvegetated sediments are extensivewhen measured on an areal basis their contributionto the overall nitrogen budgets of temperate estua-ries, salt marshes and coastal lagoons is relativelysmall. Nixon [69] estimated that in NarragansettBay, sediment nitrogen ¢xation accounted forV4% of the total annual nitrogen input. This is asimilar value to that reported by Marsho et al. [59]for the Rhode River estuary in Chesapeake Bay. Intropical coastal marine lagoons however, nitrogen¢xation in unvegetated sediments may account forsigni¢cant inputs of ¢xed nitrogen. In KaneoheBay, Hawaii sediment nitrogen ¢xation has been cal-culated to contribute 11% of the annual nitrogeninput [56]. Nitrogen budgets calculated by Smith[70] for Shark Bay, Australia and two Paci¢c islandatolls suggest that nitrogen ¢xation is the principalnitrogen input into these oligotrophic ecosystems.However, no direct measurements of N-¢xationhave yet been made to con¢rm the validity of theestimated rates used to calculate the nitrogen budg-ets for these lagoons. Hence the values of 56^97% ofthe net nitrogen input should be treated with cau-tion. From the foregoing section it is apparent that,with the exception of oligotrophic tropical lagoons,nitrogen ¢xation contributes a relatively small per-centage of the annual nitrogen input to non-vege-tated shallow coastal marine environments.

Many shallow coastal marine environments arecharacterised by the presence of extensive meadowsof rooted macrophytes. These include Spartina alter-ni£ora, Zostera marina, Zostera noltii, Zostera capri-corni and Thalassia testudinium [61,63,65]. In orderto achieve and sustain such high levels of primaryproduction substantial inputs of ¢xed nitrogen arerequired [71]. Whilst e¤cient recycling of organicnitrogen in the sediment can supply a large propor-tion of this ¢xed nitrogen [72^74], it is insu¤cient tomeet the growth requirements of these plant com-munities [71,75]. Data presented in Table 4 showthat high rates of nitrogen ¢xation have been re-corded in seagrass colonised sediments. It has beenestimated that in tropical seagrass meadows nitrogen¢xation can supply up to 50% of the nitrogen re-quirements of the plant communities [36,63,65,76].In temperate seagrass meadows such as those inthe Bassin d'Arcachon, southwest France the contri-

bution from nitrogen ¢xation to the ¢xed nitrogenrequirements of the plants is markedly lower. Welshet al. [61] have estimated that in this shallow lagoonsystem nitrogen ¢xation provides between 6 and 12%of the annual nitrogen requirement of the Z. noltiimeadow. These values are similar to those recordedfor a Z. marina bed in the Limfjorden, Denmark byRisgaard-Petersen and co-workers [77]. These inves-tigators concluded that in this system nitrogen ¢xa-tion was unimportant due to the high nitrogen avail-ability in the water column and sediment. The highrates of nitrogen ¢xation associated with salt marshgrass and seagrass colonised sediments have beendemonstrated to be intimately associated with theexcretion of organic compounds from the plant rootsand closely coupled to the photosynthetic activity ofthe plants [61,64,76]. Several authors have speculatedthat speci¢c symbiotic associations between the het-erotrophic diazotrophs and the macrophyte may ex-ist but to date none have been demonstrated [30,78].A number of studies have demonstrated that diazo-trophic sulfate reducing bacteria are responsible forthe bulk of nitrogen ¢xing activity associated withthe roots/rhizomes of S. alterni£ora, Z. noltii andZ. marina [30,61,79^81]. When 20 mM sodium mo-lybdate, a speci¢c inhibitor of sulfate reducing bac-teria was added to sediment cores nitrogen ¢xation,measured as acetylene reduction rates, was inhibitedby 70 to 90%. Acetylene reduction rates were alwaysgreater in the light than in the dark, indicating a

Table 4Nitrogen ¢xation rates recorded for temperate and tropical sea-grass meadows

Seagrass species Nitrogen ¢xation rate(mg N m32 day31)

Reference

TemperateZostera noltii (winter) 0.1^0.2 [61]Zostera noltii (summer) 2.0^7.3 [61]Zostera marina 5 [30]Zostera marina 1^6 [62]TropicalThalassia hemprichii (summer) 16 [63]Enhalus acoroides (summer) 25 [63]Zostera capricorni (summer) 25^40 [64]Zostera capricorni (winter) 10 [64]Thalassia testudinium 27^140 [36]Thalassia testidunium 5^24 [65]Hadodule beaudetti 28 [66]

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photosynthetically driven input of organic carbon tothe sediment as the driving force for heterotrophicnitrogen ¢xation in the rhizosphere. Under oxygenlimiting conditions Z. marina roots switch to fermen-tative metabolism producing ethanol, CO2 and lac-tate with the latter accumulating in the root tissue[82]. Lactate accumulated in the plant roots repre-sents a signi¢cant pool of organic carbon. Capone[30] elegantly demonstrated that following the expo-sure of Z. marina roots to 15N2 there was a rapidtranslocation of the ¢xed nitrogen to the distal leaftissues. These data suggest a mutualistic relationshipbetween Zostera and sulfate reducing bacteria in therhizosphere: the bacteria bene¢tting from the supplyof organic carbon by the plant roots and in turn theplant is provided with a supply of ¢xed nitrogen. Byadopting such a strategy the diazotrophs are able toovercome the constraint of carbon availability andthus contribute to the overall productivity of theseseagrass and salt marsh grass communities.

In addition to carbon availability a broad range ofphysico-chemical parameters can also in£uence ni-trogen ¢xation activity in benthic sediments. Theseinclude temperature, light, pH, O2, inorganic nitro-gen, salinity and trace metal availability [30,83,84].None of these factors have been systematically inves-tigated in situ to determine how they regulate/in£u-ence nitrogen ¢xation in coastal marine systems.Whilst molecular oxygen is inhibitory to nitrogenaseit is unlikely to be a major factor modulating nitro-gen ¢xing activity in marine sediments, since oxygenconcentrations decrease rapidly with depth and evenwithin a few millimetres of the sediment surface, oxy-gen is undetectable [85]. Porewater ammonium con-centrations of 50^100 WM have been shown toseverely inhibit nitrogen ¢xation in salt marshsediments [30,51,86] but it is still not clear whetherammonium is a major factor modulating nitrogen¢xation in these systems. Capone and Carpenter[87] showed that the removal of interstitial ammo-nium by perfusion stimulated nitrogen ¢xation 7^8-fold. However, in a recent study Welsh et al. [88]showed that the addition of 1 mM ammonium chlor-ide to Z. noltii colonised sediments only reduced ni-trogen ¢xation to 30% of the rate in the unamendedcontrol sediment [88]. The authors concluded thatphotosynthetically driven release of carbon fromthe plant roots was the dominant factor regulating

nitrogen ¢xation in this system. Similar results havebeen reported by McGlathery et al. [62] for Z. mari-na. These results highlight the di¤culty in unequiv-ocally establishing the role of porewater ammoniumin regulating nitrogen ¢xation and stem in part fromthe di¤culty in accurately measuring in situ concen-trations of this nitrogen species. In shallow marinesediments ammonium released from sedimented or-ganic matter by ammoni¢cation can be assimilatedby the seagrass/salt marsh grass roots and the indig-enous micro£ora. Alternatively ammonium may beadsorbed onto sediment particles or di¡use upwardinto the surface oxic zone where it is oxidised byautotrophic nitrifying bacteria to nitrate. Given thecomplexity of these interactions it is perhaps notsurprising that the role of ammonium in regulatingheterotrophic nitrogen ¢xation in these systems stillremains equivocal.

3. Ammoni¢cation

In shallow coastal marine environments benthicnutrient regeneration and metabolism are regulatedby the quantity and quality of the organic mattersupplied to the sediment. As a result of the closeproximity of the sediment to the productive photiczone the time-scale of benthic-pelagic coupling isshort compared to oceanic systems except where or-ganic matter is advected away from the area of pro-duction. Deposition of organic matter in these eco-systems can result from episodic events such asthe rapid sedimentation of annual phytoplanktonblooms or in systems where rooted macrophytesare dominant deposition of moribund plant materialoccurs throughout the year [22,95]. The quality ofthe deposited organic matter i.e. whether it is labileor highly refractory determines how rapidly it is min-eralised and this in turn is dependent upon its origin[96,97]. Seagrass detritus consists of 25^30% ¢brewith a lignin content of V8% and its mineralisationrate is low compared to phytoplankton cells whichcontain more labile nitrogenous material. Irrespec-tive of its origin, all living matter contains nitroge-nous macromolecules, such as nucleic acids, proteinsand polyamino-sugars as well as low molecular masscompounds and these become available upon deathof the cells to decomposer organisms. The release of

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ammonium from this nitrogenous matter is termedammoni¢cation. Depending upon the structuralcomplexity of the organic matter ammoni¢cationcan be either a simple deamination reaction or acomplex series of metabolic steps involving a numberof hydrolytic enzymes during which N-containingpolymers are broken down to their soluble mono-meric sub-units. The low molecular mass forms oforganic N forming the dissolved organic nitrogenpool (DON) are still poorly characterised. The iden-ti¢ed components comprise amino acids, short poly-peptides, amines, nucleic acids and urea [98^102].Components such as amino acids, purines, pyrimi-dines and urea are rapidly degraded by the indige-nous bacterial £ora [94,100,102^104]. Boon et al. [94]demonstrated that between 35 and 65% of 15N-gly-cine added to sediment cores containing intact Z.capricorni was deaminated within 12 h. These obser-vations are consistent with those of Jorgensen et al.[104] who reported amino acid deamination ac-counted for up to 25% of the ammonium regeneratedin sediments colonised by Posidonia oceanica andCymodocea nodosa. Urea is another important or-ganic nitrogen compound present in coastal marinesediments and is produced as a degradation productof nucleic acids [103,105,106]. Therkildsen et al. [103]showed that urea production was stimulated whenAMP, CMP and 16S ribosomal RNA were addedto an anoxic, defaunated marine sediment. Subse-quent laboratory studies using enriched cultures ofaerobic, fermentative and sulfate reducing bacteriashowed that urea could be produced from RNAunder both oxic [106] and anoxic conditions (Lyng-aard, unpublished results). Whilst few studies havebeen undertaken it is now recognised that urea ef-£uxing from the sediment may be an importantsource of regenerated nitrogen available to primaryproducers in the overlying water column [106^109].Urea, since it can be rapidly hydrolysed by bacterialureases is also an important source of ammonium insediments particularly those with a high macrofaunalbiomass. Lomstein et al. [109] showed that in sedi-ments of the productive Bering Sea Shelf urea hy-drolysis could be responsible for up to 80% of thegross production of ammonium. Urease activity is awidely distributed property of many Gram-negativeand Gram-positive bacteria, including aerobes, fac-ultative anaerobes and obligate anaerobes [110,111]

and would account for the rapid hydrolysis of ureaand ammonium production observed in these sedi-ments.

The mineralisation of complex nitrogenous macro-molecules in sediments is still poorly understood.The initial step in the degradation of these complexpolymers is hydrolysis to their monomeric compo-nents. Proteins are hydrolysed by proteinases andpeptidases to their constituent amino acids whichin turn are deaminated to release ammonium. Theoverall reaction can be summarised as:

Proteinproteinases

Peptidespeptidases

Amino acids

Amino acidsdeamination

Organic acid�NH�4

A diverse range of microorganisms which produceactive proteinases are present in marine sediments.These include representatives of the genera Pseudo-monas, Vibrio, Proteus, Serratia, Bacillus and Clos-tridium as well as many actinomycetes and fungi[110,112,113]. Donnelly and Herbert [114] showedthat in the shallow northern Adriatic populationsof ammonifying bacteria increased rapidly to4.7U109 ml sediment31 in the sur¢cial sedimentlayer following the collapse of the spring phyto-plankton bloom and was correlated with a markedincrease in proteolytic activity as measured usingazocasein as a model substrate. These data showthat in this dynamic shallow marine system strongbenthic-pelagic coupling was occurring with a rapidrelease of ammonium to the water column. Whilstthere have been a limited number of studies inves-tigating the mineralisation of proteins in marine sedi-ments our knowledge of the degradation of othernitrogen containing macromolecules such as DNA,RNA and chitin is almost unknown. There is anurgent need for a systematic investigation of theprocesses involved.

In the absence of speci¢c methods to determinethe mineralisation of nitrogenous organic mattermost investigators have used ammonium productionas a measure of organic N mineralisation rates.These methods range from simple sediment incuba-tion experiments in which ammonium accumulationis measured over time [115,116] to more sophisti-cated methods such as the 15N-NH�4 isotope dilution

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technique in which intact sediment cores or mixedanoxic slurries are injected with 15N-NH�4 [117].The advantage of this method is that it enablesboth gross and net rates of mineralisation to be cal-culated. Other methods that have been extensivelyused include measuring the exchange of ammonium,nitrate and nitrite across the sediment/water inter-face. In shallow lagoons and intertidal sedimentsthese measurements can be performed in situ usingbenthic chambers and provide a more realistic esti-mate of whole community activity than laboratoryincubated sediment cores [118]. This method can alsobe used to determine the e¡ects of seasonal inputs oforganic matter on the magnitude of N-£uxes to theoverlying water column. Data presented in Table 5show reported rates of ammoni¢cation in a range ofunvegetated and vegetated coastal marine sediments.As might be anticipated ammoni¢cation rates in sea-grass colonised sediments are substantially higherthan for bare sediment due to continual depositionof plant detritius to the sediment surface. Dennisonet al. [74] showed that the rapid regeneration of am-monium in Great Harbour, Massachusetts, ensuredthat nitrogen was always available in excess of theeelgrass requirements. Similarly, Iizumi et al. [72]demonstrated that ammonium regeneration in thesediments of the Izembek Lagoon, Alaska, balanced

the nitrogen requirements of the eelgrass communitypresent in this shallow marine environment.

Whilst ammoni¢cation rates are substantially low-er in uncolonised sediments this is probably attrib-utable to the lower organic nitrogen input these en-vironments receive. Laboratory studies by Hansenand Blackburn [119] show that the addition of themarine diatom Dithylum brightwellii to sedimentcores from Aarhus Bay stimulated ammoni¢cationwith 62 to 68% of the ammonium £uxes occurringin the ¢rst ¢ve days. These workers estimated thatthe `half-life' of the added algal material was 2 to 3weeks and are similar to these reported by Graf et al.[120] and Gaber [121].

Although the processes whereby ammonium is re-leased from organic N are still poorly de¢ned it isclearly evident that ammoni¢cation plays a centralrole in nitrogen recycling in coastal marine environ-ments. In these shallow water ecosystems (6 50 mdepth) benthic recycling may account for between20^80% of the nitrogen requirements of the phyto-plankton [22,69,119]. However, not all the ammo-nium produced during the deamination of organicN in sediments is available to the primary producers.A proportion, which will vary depending upon thephysico-chemical characteristics of the sediment,may be oxidised to nitrate in the sur¢cial oxiczone. This process of ammonia oxidation, termednitri¢cation, is mediated by specialist chemolitho-trophs. Nitri¢cation provides a link between the re-duced and oxidised sides of the nitrogen cycle and istherefore of fundamental importance in all ecosys-tems.

4. Nitri¢cation

Our understanding of the signi¢cance of nitri¢ca-tion in shallow coastal sediments has advanced con-siderably over the past decade. It is now recognisedthat oxidation of ammonium plays a pivotal role ingenerating a source of nitrate for denitrifying bacte-ria. The coupling of this obligately aerobic process(nitri¢cation) with an anaerobic process (denitri¢ca-tion) leads to the loss of nitrogen to the atmosphereas nitrous oxide and/or dinitrogen.

Ammonia oxidation to nitrate is a two stage proc-ess. The ¢rst step mediated by ammonia oxidising

Table 5Ammoni¢cation rates reported for unvegetated and vegetatedcoastal sediments

System Ammoni¢cation rate(mg N m32 day31)

Reference

Unvegetated sedimentsAarhus Bay, Denmark 7a [22]Kaltegat, Denmark (summer) 34 [22]Limfjorden, Denmark 30^95 [89]Southern North Sea 11^74 [90, 91]Narragansett Bay, USA 106 [92]Patuxent Estuary, USA 157 [93]Chesapeake Bay, USA 120^430 [73]Vegetated sedimentsOyster Bay, Jamaica 229 [66]Izembek Lagoon, Alaska 396 [72]Crane Cove, Alaska 508 [72]Mangoku-Ura Bay, Japan 644 [72]Moreton Bay, Australia 50^490 [94]Great Harbour, Woods Hole 225^1125 [74]

aAverage daily rate.

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bacteria produces nitrite which in turn is oxidised tonitrate catalysed by nitrite oxidisers. Until recentlythe classi¢cation of nitrifying bacteria was based pri-marily on morphological characteristics. Nitrosomo-nas, Nitrosococcus, Nitrosospira, Nitrosolobus andNitrosovibrio are generally accepted as ammonia ox-idisers and Nitrobacter, Nitrosococcus, Nitrospinaand Nitrospira as nitrite oxidising genera [122,123].Recent phylogenetic analysis of pure cultures, basedon 16S and RNA sequence data has demonstratedthat ammonia oxidising bacteria can be sub-dividedinto two distinct groups. The ¢rst contains Nitroso-coccus oceanus and forms a deep branch within the Q-proteobacteria [124]. The second group forms a tightcluster within the L-proteobacteria and can be sub-divided into two clades corresponding to Nitrosomo-nas spp. and Nitrosospira spp. [125,126]. Since am-monia oxidisers previously classi¢ed as Nitrosovibriosp., Nitrosolobus sp. and Nitrosospira sp. exhibit veryhigh levels of 16S and RNA gene sequence homol-ogy they are now accommodated within a single ge-nus for which the name Nitrosospira has priority.Phylogenetic analysis of nitrite oxidising species ofthe genus shows that they form a group within theO sub-division. In shallow coastal systems Nitroso-monas spp. and Nitrobacter spp. are the principalorganisms responsible for the two steps, respectively,of nitri¢cation although recently Stephen et al. [127]have provided the ¢rst evidence for the existence ofmarine Nitrosospira spp.

Nitrifying bacteria are notoriously di¤cult to iso-late and grow and this has prevented, until recently,meaningful study of their community structure anddiversity. Most probable number methods have beenwidely used to enumerate nitrifying bacteria[128,129]. The e¤ciency of this technique is lowand it has an inherently low statistical precision.Typically population densities range from 102^104

per ml31 sediment and exceptionally may be ashigh as 107 per ml31 sediment using this method[129^131]. The introduction of improved enumera-tion techniques based on immuno£uorescence hasyielded population densities substantially higherthan MPN counts [132]. Whilst immuno£uorescenceis a considerable improvement on conventionalMPN methods it nonetheless underestimates thetrue abundance of nitri¢ers because of the speci¢cityof £uorescent antibodies [133]. As yet it is not

proved possible to obtain a true estimate of theabundance or diversity of nitri¢ers in marine envi-ronments. The recent development of speci¢c geneprobes for di¡erent ammonia oxidisers will howeverprovide a powerful tool to elucidate the communitystructure of nitrifying bacteria in marine sediments[127,134].

A number of di¡erent methods have been devel-oped to estimate nitri¢cation rates in marine sedi-ments. One of the most widely used is the 15N-NO3 isotope dilution technique which can be em-ployed with either sediment slurries or intact cores[135]. Nitri¢cation rates have also been determinedby measuring 15N-N2 production from sedimentcores amended with 15N-NH�4 [130]. This techniquehas the advantage that it enables both nitri¢cationand denitri¢cation rates to be measured and the de-gree of coupling between the two processes deter-mined.

Alternatively, non-isotopic methods using speci¢cnitri¢cation inhibitors can be employed. Three inhib-itors (nitrapyrin, allylthiourea and chlorate) havebeen widely used to measure nitri¢cation rates incoastal marine sediments [20,129,136,137]. Inhibitorssuch as nitrapyrin (also called N-serve) and allyl-thiourea prevent the oxidation of ammonia to hy-droxylamine by inhibiting ammonia monoxygenase,the enzyme catalysing this reaction [138]. The nitri¢-cation rate is determined by measuring the di¡erencein ammonium accumulation in the presence and ab-sence of the inhibitor. A variation on this techniqueis to measure the dark incorporation of 14C-bicar-bonate into the cells of nitrifying bacteria in thepresence and absence of N-serve [139]. The nitri¢ca-tion rate is calculated by relating CO2 incorporationto ammonium oxidation using an N:C ratio of 8.3.A particular problem of this method is that N:Cratio can vary from 4 to 40 depending upon thegrowth conditions and oxygen tension thus makinginterpretation of the results di¤cult [141].

All the above methods for determining nitri¢ca-tion rates have inherent limitations and assumptionsbuilt into them and these have been excellently re-viewed by Henriksen and Kemp [140] and Ward[141]. Data presented in Table 6 show that irrespec-tive of the geographical location, di¡erent sedimenttypes and methods used to measure nitri¢cation, therates are remarkably similar. However, when ana-

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lysed on a seasonal basis a di¡erent pattern emerges.In Danish coastal waters nitri¢cation rates show aminimum during the summer months and similarobservations have also been reported for the Provi-dence river station in Narragansett Bay [136,142] andthe upper reaches of Chesapeake Bay. These minimahave been attributed to a combination of reduced O2

penetration into the sediment, greater competitionfor ammonium and elevated sul¢de levels which areinhibitory to nitrifying bacteria [136]. In other ma-rine systems such as the Tay Estuary, Scotland andthe middle reaches of Narragansett Bay, USA theconverse has been observed and summer maximafor nitri¢cation have been recorded [92,136]. Inboth these cases seasonal patterns of nitri¢cationfollow the annual temperature cycle with maximumrates in June^July. In these systems temperaturerather than O2 di¡usion into the sediment appearsto be a more important factor regulating nitrifyingactivity.

A number of physico-chemical and biological fac-tors are important in regulating nitrifying activity incoastal marine sediments. These include temperature,NH�4 concentration, O2 tension, pH, dissolved CO2

concentration, salinity, presence of inhibitory com-pounds, light, macrofaunal activity and presence ofmacrophyte roots. Most of the data reported in theliterature are derived from pure culture studies andtherefore extrapolation of how these parameters af-fect in situ nitrifying activity must be interpretedwith care.

The optimum temperature for the growth of purecultures of nitrifying bacteria isolated from temper-ate environments is in the range 25^35³C [143] andbelow 15³C growth rates decline sharply. Hansen[131] showed that nitri¢cation rates in Danish coast-al sediments increased 5-fold when the sediment tem-perature increased from 2³C in spring to 22³C inautumn. However, shallow coastal sediments, partic-ularly intertidal sediments are subject to both sea-sonal and diurnal changes in temperatures and henceit may be expected that nitrifying bacteria wouldexhibit optimal growth and/or activity during thesummer months when temperatures are maximal.The little quantitative evidence that is available iscontradictory. Macfarlane and Herbert [129] re-ported that in the Tay estuary, Scotland maximalnitri¢cation rates were recorded in summer whensediment temperatures reached 19^21³C. Similar sea-sonal patterns have been reported for the middleregion of Narragansett Bay [92]. However, temper-ature also a¡ects other parameters, most notably O2

solubility and this coupled with increased benthicrespiration during the summer months, as a resultof higher ambient temperatures, means that thedownward di¡usion of oxygen is limited to the top1^2 mm of the sediment. Thus, in organically richsediments oxygen availability rather than tempera-ture per se is the factor most probably limiting ni-trifying activity, and would explain the recordedsummer nitri¢cation minima recorded in Danishcoastal sediments and Chesapeake Bay [130,136].

Nitrifying bacteria are obligate aerobes and thusthe depth distribution of nitrifying bacteria is ulti-mately constrained by the limits of downward O2

di¡usion which is typically 1^6.5 mm dependingupon sediment type, organic matter content, temper-ature and degree of mixing and bioturbation [85].Nitrifying bacteria therefore have to compete withother heterotrophs for the limited supplies of dis-solved oxygen. Laboratory studies show that hetero-trophic bacteria have a much higher a¤nity for oxy-gen (Km 6 1 WM O2) and are likely to outcompetenitrifying bacteria at low oxygen concentrations. Thereported dissolved oxygen concentrations at whichammonium oxidation is inhibited range from 1.1 to6.2 WM O2 [146^148]. At low oxygen concentrations(6 10 WM O2) ammonia oxidisers produce nitrousoxide in addition to nitrite [147,148]. Laboratory

Table 6Estimated nitri¢cation rates reported for di¡erent coastal marineand estuarine sediments

System Nitri¢cation rate(mg N m32 day31)

Reference

North Sea sediments 2^9 [20]Kingoodie Bay, UK 20 [129]Limjforden sediments,Denmark

38 [11]

Normsinde Fjord, Denmark 112 [144]Ochlockonee Bay, Florida 84 [145]Southern North Sea sediments 26 [90]Narragansett Bay, USA 23 [92]Chesapeake Bay, USA 14^23 [140]Patuxent River Estuary, USA 26 [130]Kysing Fjord, Denmark 23^27 [136]Odawa Bay, Japan 38^42 [135]

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studies have shown that under conditions of oxygenlimitation Nitrosomonas europaea functions as a de-nitri¢er, using nitrite generated as the end-productof ammonium oxidation as terminal e3 acceptor[148,149]. In coastal marine sediments dissolved O2

concentrations undergo very sharp diel cycles: fromsuper-saturating conditions resulting from benthicmicroalgal photosynthesis during daytime to com-plete anoxia at night as a consequence of high res-piratory demand [150]. The ability of ammonia oxi-disers to grow at low dissolved O2 tensions usingnitrite as terminal e3 acceptor may be a mechanismwhereby these bacteria survive such rapidly changingconditions. At the other end of the spectrum almostnothing is known about the e¡ect of elevated oxygenconcentrations on nitrifying activity. Yet in shallowwater sediments oxygen levels can reach 2^3 timesair saturation values during daytime as a result ofbenthic primary production [85]. Henriksen andKemp [140] reported that increasing O2 concentra-tions to 2 and 2.6 times air saturation values resultedin a 15% and 25% inhibition of nitrifying activity inestuarine sediment slurries. However, no systematicinvestigations have been undertaken to evaluate thee¡ects of elevated oxygen concentrations on nitrify-ing activity.

In sediment systems colonised by emergent andsubmersed vascular plants the release of oxygeninto the rhizosphere zone may stimulate nitri¢cationactivity [151,152]. However, few direct measurementsof nitri¢cation rates have been made in macrophytecolonised sediments. Kemp et al. [153] reported thatnitri¢cation rates in sediments colonised by Potamo-geton perfoliatus were 20-fold higher than in un-colonised sediments. Equally, discontinuities arisingfrom the e¡ects of macrofaunal activity signi¢cantlyalter the spatial gradients of O2 and NH�4 in sedi-ments. Several studies have demonstrated higher ni-tri¢cation activity in the lining of infauna burrowsthan in defaunated sediments [154^156]. This may beexplained by the higher ammonium concentrationsfound in the deeper parts of burrows which togetherwith ammonium excreted by their macrofauna in-habitants provides a higher substrate availability.The downward transport of oxygen into the burrowsis dependent on the ventilation activity of the burrowinhabitants. Whilst burrows are di¡erent from sur-face sediment they can in some respects be consid-

ered as an extension of the sediment-water interface.Kristensen [156] estimated that an estuarine popula-tion of the polychaete Nereis virens (700 individualsper m32) increased the contact zone between sedi-ment and water by V150% and oxic sediment vol-ume by 30^50%. Kristensen et al. [157] estimatedthat in sediments colonised by N. virens nitri¢cationin the burrows accounted for between 10 and 70%(mean value 40%) of the bulk sediment nitri¢cation.These data are similar to these reported by Henrik-sen et al. [158] and Blackburn and Henriksen [21].Thus, oxygenated burrows increase the potential fornitri¢cation in a sediment by providing additionalsites for ammonium oxidation and stimulating bac-terial activity. Nitrate produced by the oxic sedimentlayers can either di¡use into the overlying water col-umn as a source of `regenerated' nitrogen or enterthe anoxic sediment where it can be reduced to ni-trous oxide and/or dinitrogen by denitri¢cation orNH�4 by nitrate ammonifying bacteria.

5. Denitri¢cation and nitrate ammoni¢cation

Whereas nitri¢cation involves the oxidation of re-duced nitrogen mediated by obligate aerobes, deni-tri¢cation is a reductive process, whereby heterotro-phic bacteria utilise nitrate as a terminal e3 acceptorin respiration and reduce it to either gaseous prod-ucts (denitri¢cation) or ammonium (nitrate ammoni-¢cation). Denitri¢cation is a key process in the sedi-ment nitrogen cycle since it decreases the amount ofnitrogen available to the primary producers as thegaseous end-products (N2O and N2) di¡use intothe atmosphere. It also provides a mechanism, incoastal marine systems that receive large quantitiesof nitrogen from anthropogenic sources, to removeexcess nitrogen and therefore help control the rate ofeutrophication of these environments [172,173,180,181]. The ability to denitrify is widely distributedamongst di¡erent taxonomic groups of heterotrophicbacteria [182^184]. In the presence of molecular oxy-gen these bacteria grow aerobically but under oxygendepleted conditions they are able to maintain respi-ratory activity using nitrate as the terminal e3 ac-ceptor. The most frequently isolated denitrifyingbacteria belong to the genus Pseudomonas whichproduce dinitrogen as the end-product of nitrate res-

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piration [185,186]. In addition to denitri¢cation anumber of studies have shown that nitrate can bereduced to ammonium by a number of fermentativeand strictly anaerobic bacteria [185^189]. In contrastto denitri¢cation where nitrogen is lost from the eco-system nitrate ammoni¢cation results in the conser-vation of nitrogen in an available form. A furtheraspect of denitri¢cation that has been intensivelystudied is the coupling of nitri¢cation to denitri¢ca-tion since this provides a mechanism whereby sub-stantial quantities of nitrogen can be removed frommarine ecosystems [130,140,144].

A wide range of experimental methodologies havebeen developed to estimate denitri¢cation rates inshallow marine environments. These include massbalance methods which estimate denitri¢cation rateson the di¡erence between N-inputs and outputs[102,145,181,190], the acetylene inhibition technique[191,192], N2 production measurements [92,130,171],diagenetic models of pore water pro¢les [92], nitrateconsumption measurements [193], microelectrodemethods using either acetylene inhibition in conjunc-tion with O2/N2O microelectrodes or by calculatingdenitri¢cation rates from nitrate pro¢les measuredusing a nitrate microsensor [144,194,195] and 15N-tracer techniques [4,130,174,176,196,197]. The acety-lene inhibition technique (AIT) is based on the in-hibition of nitrous oxide reductase by acetylene andis a simple, sensitive and inexpensive method[180,192]. However, it su¡ers a number of disadvan-tages, most notably that at the low nitrate concen-trations often found in marine sediments (6 10 WM)inhibition is incomplete [180,192]. As a consequenceof the incomplete inhibition of nitrous oxide reduc-tase by acetylene, denitri¢cation rates may be under-estimated by as much as 30^50% [19,198]. The limi-tations of the AIT method have been clearlydemonstrated by Lohse et al. [91]. These investiga-tors simultaneously measured denitri¢cation rates inthe southern North Sea by the AIT technique andisotope pairing method (IPM) and showed that ratesrecorded using acetylene inhibition were 45% lowerthan those obtained by the IPM method (see Table 7for details). Furthermore acetylene severely inhibitsnitrifying activity [199] and in marine sediments sul-¢de reverses the acetylene block of nitrous oxide re-ductase activity [180,192]. As a consequence of theseexperimental limitations direct measurements using

15N-tracer techniques are now the preferred methodfor measuring denitri¢cation rates. The recently de-veloped 15N isotope pairing method is a powerfulanalytical technique which not only enables therate of denitri¢cation to be accurately determinedbut also the proportion arising from nitrate in thewater column and that produced by nitri¢cation inthe sediment [176,177,196]. This experimental ap-proach has the major advantage that no inhibitoris required and that nitri¢cation, denitri¢cation, ni-trate ammoni¢cation and N-mineralisation rates canall be determined in a single experiment if the nitrateand ammonium £uxes have been measured [91,177].

Data presented in Table 7 show reported denitri-¢cation rates for a range of coastal and estuarinesediments using 15N isotope and acetylene inhibitiontechniques. The sediments show a wide range of ac-tivities with a trend to higher rates in shallow near-shore waters e.g. Lendrup Vig, Kysing Fjord, Nors-minde Fjord, Chesapeake Bay and Narragansett Baywhere the supplies of organic carbon and nitrate run-o¡ are higher. High denitri¢cation rates have alsobeen recorded in Z. marina colonised sediments inChesapeake Bay (Table 7). These probably arise asa result of the macrophytes increasing the organiccontent of the sediment by trapping detritus and/orexcreting organic carbon from the roots [200,201].

A number of studies have shown in temperate ma-rine environments that denitri¢cation rates show dis-tinct seasonal patterns governed principally by tem-perature, supply of nitrate and availability of organiccarbon [172,179,202]. In systems where there is asubstantial input of nitrate throughout the year thereis a good correlation between denitri¢cation activityand ambient temperature [202^204]. However, inother shallow inshore environments nitrate inputsare more episodic events such as increased agricul-tural run-o¡ during winter and spring. In NorsmindeFjord, Jorgensen and Sorensen [162] showed thatthere were two seasonal maxima of denitrifying ac-tivity, one in May attributed to the deposition anddecomposition of a microalgal bloom and presenceof high nitrate concentrations in the water columnand one in the late autumn associated with increasedinputs of nitrate into the water column. In this sys-tem, nitrate concentrations in the sediment were con-trolled by the nitrate load in the overlying watercolumn. Where external nitrogen inputs are small

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nitri¢cation is the principal source of nitrate for de-nitri¢cation. Jenkins and Kemp [130] showed that inthe Patuxent Estuary there was a strong couplingbetween nitri¢cation and denitri¢cation. In this estu-ary s 99% of the nitrate produced by nitri¢cationwas reduced to dinitrogen during the spring. How-ever, during the summer this coupling decreased bytwo orders of magnitude, even though the denitri¢-cation potential was similar to that recorded in thespring, due to reduced nitri¢cation rates resultingfrom low O2 concentrations in the surface sediments.This seasonal pattern of high denitri¢cation rates inspring followed by lower rates in summer has alsobeen reported for Ochlockonee Bay in Florida [92].

In addition to seasonal changes in denitri¢cationrate, rates in intertidal and sub-tidal environmentsare also subject to signi¢cant change on a diel basisdue to the growth of benthic microalgae at the sedi-ment surface. During daylight hours O2 producedduring photosynthesis can di¡use into the surfacesediments and inhibit dissimilatory nitrate reduction.Jorgensen and Sorensen [162] showed that inhibitionof denitri¢cation was most pronounced in earlyspring when rates were reduced by as much as 60%in the light compared to dark controls. However,when calculated on an annual basis denitri¢cationrates were reduced by only 13% due to light inhib-ition. Similar ¢ndings have been reported for Lend-rup Strand sediment by Andersen et al. [161].

As described earlier in this review (Section 4) thepresence of macrophytes and infauna can exert aprofound in£uence on processes such as nitri¢cationand denitri¢cation. For example, Kemp and Murray[205] have calculated that oxygen released from theroots of P. perfoliatus was su¤cient to support V5times the rate of nitri¢cation in unvegetated sedi-ments. Equally macrophytes may stimulate denitri¢-cation by trapping readily degradeable organic detri-tus in the water column or releasing labile organiccarbon from the roots [200]. The exchange of nitratebetween the overlying water and sediment is alsosigni¢cantly in£uenced by the burrow-dwelling in-fauna. In the presence of di¡erent infaunal speciesthe nitrate £ux either increases or decreases. This hasbeen attributed to the substantial variation in thedegree of coupling and rates of nitri¢cation and de-nitri¢cation occurring in the burrows. For example,Henriksen et al. [154] showed that shallow burrowerssuch as Corophium volutator which have a higherventilation activity and large burrow volume com-pared to size stimulate nitri¢cation because oxygenis able to penetrate deeper into the burrow walls. Incontrast, deep burrowers such as N. virens usuallyshow a nitrate £ux into the burrow inferring a highrate of denitri¢cation. The infauna not only in£uencenitrogen cycling in marine sediments by producingburrows they also selectively concentrate organicmaterial as faecal pellets. Faecal pellets have longbeen recognised as sites of intense microbial activitywhich result in a high oxygen demand with the for-mation of reduced microniches [206,207]. These an-oxic microsites enable anaerobic processes such as

Table 7Denitri¢cation rates reported for coastal marine environments

System Denitri¢cation rate(mg N m32 day31)

Reference

Randers Fjord 20^141 [159]a

Kysing Fjord 3^1109 [159]a

Delaware Inlet 20^40 [160]a

Lendrup Vig 40^715 [161]a

Norsminde Fjord 278^1401 [162]a

Norsminde Fjord 14^224 [163]d

Newport River Estuary 14^42 [164]a

Tomales Bay sub-tidal 14^98 [165]a

Tomales Bay mud £at 10^100 [166]a

Torridge River mud £at 2^19 [167]a

Torridge River marsh 8^198 [167]a

Chesapeake Bay, Z. marina 225^702 [168]a

Chesapeake Bay, non-vegetated 20^739 [168]a

Great Ouse Estuary 7^32 [169]a

Texel, Wadden Sea 3^185 [170]a

Southern North Sea 2^3 [91]a

Narragansett Bay 50^655 [171]b

Guadalupe Estuary 15^116 [172]b

Boston Harbour 6 10^412 [173]b

Massachusetts Bay 6 10^128 [173]b

Patuxent River Estuary 259^299 [130]c

Tokyo Bay 54^111 [174]c

Colne Point salt marsh 13^44 [175]c

Aarhus Bay 40^71 [176]d

Southern North Sea 3^4 [91]d

Arcachon Bay 0^59 [177]d

Etang de Prevost 1^153 [177]d

Colne Estuary 1^154 [178]d

Gulf of Finland 1^9 [179]d

Northern Baltic Proper 0^4.2 [179]d

aAcetylene block method.bN2 £ux method.c15N2 method.d15N isotope pairing method.

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denitri¢cation to take place in what are ostensiblyoxic surface sediments and may explain how thesetwo processes can occur in close proximity. Thesedata show that the infauna play an important rolein stimulating nitrogen cycling in coastal marinesediments.

Denitri¢cation is widely accepted as being thedominant process of nitrate reduction in most shal-low marine sediments [129,135,144,160,167]. How-ever, the alternative pathway of nitrate reduction,nitrate ammoni¢cation may also be important undercertain conditions. Few systematic studies of nitrateammoni¢cation have been undertaken and hence it isnot possible to evaluate how signi¢cant this processis in coastal marine systems. A number of studieshave demonstrated that heterotrophic bacteria withthe capacity to respire nitrate to ammonium arewidely distributed in marine sediments. They are pre-dominantly fermentative bacteria and include mem-bers of the genera Aeromonas, Vibrio [129,185], Clos-tridium [187,189] and Desulfovibrio [188]. Macfarlaneand Herbert [129] showed that in the Tay estuary,Scotland, populations of nitrate ammonifying bacte-ria in the surface sediments were in the order of 106^107 g dry wt. sediment31. Similar population den-sities of nitrate ammoni¢ers have been reported forother shallow marine ecosystems [208]. In organicallyrich sediments, such as those found in Mangoku-UraBay, Japan, nitrate ammoni¢cation accounted forV50% of the total nitrate reduced whereas in sedi-ments with a low C:N ratio the rates were lower (4^35%) and denitri¢cation was the dominant process[129,135,177,209,210]. Jorgensen [209] showed that inNorsminde Fjord, Denmark nitrate ammoni¢cationwas only a signi¢cant process during the late summerwhen the sediment was reduced all the way to thesurface. Under these conditions, nitrate ammoni¢ca-tion was maximal in the surface layer whereas at allother times activity was restricted to the deep sedi-ment layers and denitri¢cation was the predominantprocess of nitrate reduction. Laboratory studies byHerbert and Nedwell [3] demonstrated that nitrateconcentration plays a key role in determiningwhether nitrate is reduced to gaseous products orconserved as ammonium. These workers showedthat at low nitrate concentrations denitri¢ers wereoutcompeted by nitrate ammonifying bacteria where-as at high nitrate concentrations the converse was

true. These observations are consistent with ¢elddata obtained from the Colne estuary, England,which show that as the nitrate concentration in-creased denitri¢cation became the dominant process[211,212]. Similarly, Smith and co-workers [204]demonstrated that nitrate ammoni¢cation in a saltmarsh sediment decreased from s 50% to V4% ofthe total nitrate reduced when nitrate concentrationsincreased. These observations re£ect the di¡erentcompetitive abilities of denitri¢ers and nitrate ammo-nifying bacteria and hence selection of di¡erent ni-trate reducing communities under changing C:Nconditions in the ¢eld [213].

In addition to dinitrogen and ammonium, nitrousoxide can also be produced as an end-product ofdissimilatory nitrate reduction. In denitri¢cation, ni-trous oxide is a true intermediate whereas in nitrateammoni¢cation it is a side reaction [180]. As dis-cussed previously (Section 4) nitrous oxide is alsoproduced as a side reaction of nitri¢cation. The rec-ognition that this gas plays a key role in both thestratospheric ozone and tropospheric heat budgethas stimulated research to identify the sources ofnitrous oxide production [214,215]. Estuarine andcoastal marine environments have been identi¢ed assources of atmospheric nitrous oxide on the basisthat in situ concentrations in the water are super-saturated relative to atmospheric levels [216,217].The production of nitrous oxide is controlled by sev-eral factors of which oxygen is considered the mostimportant. Jorgensen et al. [147] demonstrated thatat low O2 partial pressures (0^0.2 kPa) nitrous oxideproduction increased rapidly and was maximumunder anoxic conditions. These workers concludedthat denitri¢cation was the principal source of ni-trous oxide production. In a follow-up study, Jensenet al. [218] showed that nitrous oxide productionfollowed a diel cycle in a manner analogous to thatobserved for denitri¢cation [162]. They recordedmaximum emissions of nitrous oxide at night (0.4to 4 Wmol N2O-N m32 h31) whereas during theday rates decreased to between 30.4 to 0.4 WmolN2O-N m32 h31. Nitrous oxide production, resultingfrom denitri¢cation, was maximum in the surface to1 cm depth horizon and thus the emission patternwas inversely related to the presence of oxygen at thesediment surface. In the dark lack of oxygen produc-tion by the benthic microalgae enabled denitri¢ca-

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tion to occur close to the sediment-water interfacethereby facilitating the release of nitrous oxide tothe water. Daily average nitrous oxide emissionsfrom this sediment were V40 Wmol N2O-N m32

day31 during the winter months and early springwhereas no signi¢cant production was recorded insummer when the sediments were nitrate depleted.Robinson et al. [217] in a comprehensive study ofatmospheric N2O emissions from the hypernutri¢edColne estuary similarly concluded that benthic deni-tri¢cation and not nitri¢cation was the principalsource of N2O in this high nitrate estuary. Althoughthe estuarine water was always super-saturated withN2O no production could be demonstrated in thewater column. They elegantly demonstrated thatN2O emission £uxes from the surface of the tidallyexposed sediments decreased with time. This was aresult of the rapid turnover (6 40 min) of the sedi-mentary nitrate pool and its depletion when it wasno longer being recharged by nitrate from the over-lying water column. These workers estimated thatwhilst benthic N2O production was 6 2% of the ni-trate denitri¢ed in the Colne estuary it neverthelessrepresented a signi¢cant export of N2O to the atmos-phere. A further factor in£uencing nitrous oxideemissions from coastal marine sediments is nutrientloading. Seitzinger and Nixon [219] showed that in-

creased dissolved inorganic nitrogen loading to ma-rine mesocosms signi¢cantly increased nitrous oxideemissions. Similar results were obtained by Middel-burg et al. [220] for the Scheldt estuary. These work-ers showed that in the Scheldt estuary sediments ni-trous oxide £uxes appeared to respond linearly to anincreasing nitrogen load. Highest nitrous oxide £uxeswere recorded at the tidal freshwater member siteswhere total dissolved nitrogen may reach a concen-tration of 600 WM, whereas emissions were almostzero at the high salinity stations where nitrogen lev-els are low. At the most saline sites in£uxes of ni-trous oxide into the tidal £ats were recorded. Similarobservations have been recorded for other coastalsediments and have been attributed to the utilisationof nitrous oxide as a terminal e3 acceptor duringorganic matter degradation in the absence of nitrate[170,178,221,222]. This consumption of nitrous oxidein anoxic marine sediments is consistent with theiroverall high denitri¢cation activity [180].

In summary the preceding sections of this reviewhave amply demonstrated the complexity of nitrogencycling in shallow marine sediments and how indi-vidual microbiologically mediated processes aremodulated by physico-chemical and biotic factors.The complex nature of benthic nitrogen cycling isshown schematically in Fig. 2. The driving force

Fig. 2. Schematic representation of nitrogen cycling in coastal marine sediments.

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for nitrogen mineralisation is the quality (C:N ratio),quantity, spatial distribution of the degradeable or-ganic matter in the sediments and the di¡usability ofthe decomposition products. These factors are notonly the major determinants of the amount of organ-ic nitrogen mineralised but also the respective quan-tities of ammonium, nitrate, dissolved organic N andnitrous oxide/dinitrogen released from the sedimentinto the overlying water column. The presence/ab-sence of rooted macrophytes and infauna togetherwith concentrations of oxygen, ammonium and ni-trate in the overlying water also play a role in deter-mining the ratios of these products by stimulating orinhibiting individual mineralisation processes. Theseoften opposing stimulations and inhibitions of theindividual processes which make up the nitrogencycle result in complex patterns of nitrogen mineral-isation that are di¤cult to interpret. As a conse-quence a number of investigators have employedmathematical models to examine the interrelation-ships between these di¡erent processes in order toidentify the parameter(s) which exert the greatestcontrol on the nitrogen cycle.

6. Modelling of nitrogen cycling in coastal marinesystems

A number of models have been developed to sim-ulate benthic nitrogen mineralisation [223^227].Many of the early models were constrained by thenumber of components that could be used and thislimited the spatial resolution of these systems. Inrecent years more complex models have been devel-oped. Blackburn [225] developed a systems dynamicmodel which linked reactants and products by di¡u-sion equations. However, this model also su¡eredfrom limited spatial resolution. More sophisticatedversions, based on the Cellmatics system, have nowbeen developed which yield a high spatial and tem-poral resolution [226,227]. Such models have beenused to simulate the e¡ects of increased rates of or-ganic matter loading to the surface of a marine sedi-ment on the e¥ux of nitrate, ammonium, DON anddinitrogen gas as well as nitri¢cation and denitri¢ca-tion rates [226]. The data obtained show that modestincreases in organic matter loading (21.6 mmol Cm32 day31) stimulated nitri¢cation by increasing am-

monium availability whereas at higher loading(s 64.6 mmol C m32 day31) nitri¢cation rates de-creased as a consequence of reduced oxygen penetra-tion into the surface sediment layer resulting fromstimulated respiratory demand. Since nitri¢cationwas reduced under these conditions denitri¢cationrates also declined due to nitrate limitation eventhough the prevailing anoxic conditions were opti-mum for nitrate respiration. A further consequenceof the decrease in ammonium oxidation rate at highorganic matter loadings (107 mmol m32 day31) wasthat ammonium e¥ux from the sediment increasedby V50%. In contrast at low organic matter loadingrates (7.2 mmol m32 day31) denitri¢cation rates werelimited by the availability of organic matter.

Simulations such as these are a valuable tool fortesting hypotheses and predicting the outcome ofmineralisation processes in sediments. A further ex-ample of the power of this experimental approachhas been the use of mathematical models to simulatethe e¡ects of the distribution of organic matter anddi¡erent organic matter loadings on nitri¢cation anddenitri¢cation rates [227^229]. Three distributionpatterns were simulated via surface deposition, lineardepth distribution and completely mixed to mimicactive bioturbation. Additional factors examined inthe model included di¡erent concentrations of O2,NO3

3 and NH�4 in the overlying water column andthe presence and absence of sul¢de di¡usion. It waspredicted that the mixed distribution of organic mat-ter would result in high rates of sulfate reduction,nitri¢cation and denitri¢cation in the absence of sul-¢de di¡usion (iron excess conditions) and this wascon¢rmed by the model. The explanation for this isthat organic matter oxidation in the deep sedimentwas coupled to sulfate reduction. Ammonium re-leased during this process was able to di¡use up-wards into the oxic surface layer where it was oxi-dised to nitrate and then denitri¢ed by downwarddi¡usion into the underlying anoxic zone. Underthese conditions carbon and nitrogen mineralisationprocesses are decoupled and since there was rela-tively little carbon in the oxic zone and sul¢de wassequestrated as iron sul¢de and pyrite more oxygenwas available for ammonium oxidation: hence thehigh nitri¢cation rates. When sul¢de was allowedto di¡use freely, simulating iron limiting conditions,it di¡used into the oxic zone and competed with the

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nitri¢ers for oxygen. As a consequence the depth ofoxygen penetration into the surface sediment waslimited and nitri¢cation and denitri¢cation rateswere correspondingly lower. These data have subse-quently been con¢rmed experimentally using sedi-ment cores amended with di¡erent organic matterloadings [4].

Further runs of the model investigated the e¡ectsof di¡erent concentrations of oxygen, ammoniumand nitrate in the overlying water column in orderto quantify the proportion of nitri¢cation and deni-tri¢cation arising from the e¥ux of ammonium andnitrate from the sediment (Ns and Ds) and thesenitrogen species in the water column (Nw andDw). The simulations demonstrated that increasedwater column nitrate concentrations did not in£u-ence coupled nitri¢cation-denitri¢cation reactions inthe sediment. However, increased ammonium levelsin the overlying water stimulated coupled nitri¢ca-tion-denitri¢cation in the sediment as did elevatedoxygen concentrations. The converse was true whensul¢de was allowed to di¡use freely in the sediment.This is to be expected since increased ammoniumand oxygen availability stimulates nitrifying activityin the surface sediment and this in turn leads toincreased sediment denitri¢cation activity. Di¡usionof sul¢de into the oxic zone reduces oxygen avail-ability and inhibits nitri¢cation hence decreasing theamount of ammonium oxidised, and concomitantlythe rate of sediment denitri¢cation (Ds). However,when nitrate concentrations in the water columnare high it di¡uses into the sediment stimulating de-nitri¢cation (Dw). Since denitri¢cation is an anaero-bic process it is not surprising that rates should bemaximal under such conditions. These data highlightboth the complexity and apparent contradictions ofthe di¡erent processes involved in benthic nitrogencycling.

Thus, total rates of denitri¢cation are dependentupon diametrically opposing environmental condi-tions, viz sediment denitri¢cation (Ds) is dependentupon ammonium oxidation in the surface oxic zonewhilst denitri¢cation of water column nitrate (Dw) isdependent upon sediment anoxia and high nitrateconcentrations. The value of computer simulationssuch as those described above is that they may en-able environmental conditions that are conducive tohigh denitri¢cation rates to be identi¢ed thereby re-

ducing the amount of recycled nitrogen available tothe primary producers.

7. E¡ects of anthropogenic inputs on nitrogen cycling

Littoral ecosystems such as salt marshes, estuariesand inshore coastal waters are natural highly pro-ductive environments which in recent years havebeen subject to increased anthropogenic inputs ofnitrogen arising from such diverse sources as fertil-iser run-o¡, sewage discharges and aquaculture [230^233]. The net e¡ect of these elevated nitrogen inputsis to stimulate primary production [234,235]. Thesee¡ects have been most severe in areas such as shal-low embayments or where tidal £ushing is limited[236^238]. In general, increased nitrogen loadings(hypernutri¢cation) accompanied by excessive phyto-plankton growth (eutrophication) result in a sharpdecline in the rooted phanerogam communities dueto decreased water transparency and build up of epi-phytes [239^243]. Aquatic systems with extensivephytoplankton blooms frequently progress to sys-tems dominated by opportunistic, free £oating mac-roalgae belonging to the orders Ulvales or Clado-phorales (chlorophyta) and cyanobacterial blooms[236,243]. This excessive primary production in turnleads to high rates of production in the rest of thebiological food web in these ecosystems [235,244].The increased pool of autochthonous particulatematter produced results in intense microbial activitywhen it is deposited at the sediment surface[235,245]. As a result of these high decompositionrates oxygen demand in the sediment is high andmay lead to a temporary disappearance of dissolvedoxygen in the overlying water column with the con-comitant release of toxic sul¢de [236^238]. This phe-nomenon known as dystrophic crisis causes massmortality of the benthic macrofauna and ¢sh stocksin enclosed lagoons such as those found in the south-ern Mediterranean [235,237,238,242].

Increased nutrient loadings to shallow marine en-vironments and the accompanying stimulation ofprimary production have as might be expected aprofound e¡ect on benthic metabolism and nitrogenmineralisation processes. Since the progressive de-cline in seagrass meadows and their replacement byphytoplankton and opportunistic macroalgae such as

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Ulva not only increases the total quantity of organicmatter produced in these ecosystems but also itscomposition [96,236,246]. Fast growing macroalgaesuch as Ulva assimilate and store nitrogen in excessof their growth requirements. This `luxury' uptakecoupled with a rapid growth rate enables this macro-alga to outcompete other primary producers [247^249]. A consequence of the elevated nutrient contentof macroalgae such as Ulva is that their decomposi-tion is more rapid due to their high N:C ratios[96,97]. As a result, the replacement of seagrassmeadows by macroalgae changes not only the timingand rate of detritus production but also the decom-position rate. A less obvious e¡ect of the change inthe primary producer community is the oxygenationof the sur¢cial sediments. Rooted phanerogams pro-vide gas transport throughout the sur¢cial sedimentand hence contribute to its oxygenation. In contrast,£oating macroalgae do not and because of their mor-phology can promote physical strati¢cation of theoverlying water column [250]. The large thalli ofUlva act as a physical barrier separating the watercolumn into two layers, the upper one well oxygen-ated or even super-saturated and the lower one an-oxic and highly reduced [242]. Pelagic macroalgaesuch as Ulva can achieve biomass values s 1 kgdry wt m33 during the active growth phase in springand early summer [251]. In shallow coastal lagoonsdecomposition commences in mid-summer (July^Au-gust) when the growth of Ulva has ceased [242,251].This results in a rapid release of organic matter tothe sediment and overlying water and concomitantonset of anoxia. In the absence of oxygen, organiccarbon oxidation is coupled to sulfate reduction. Theremoval of oxygen and as a result of the abioticoxidation of sul¢de in the sediment whilst optimumfor denitri¢cation is inhibitory to nitri¢cation andhence nitrogen removal via coupled nitri¢cation de-nitri¢cation is also inhibited. Under these conditionsnitrogen can only be lost to the atmosphere throughdenitri¢cation using water column nitrate. In theshallow Sacca di Goro Lagoon, northern Italy,high rates of denitri¢cation (V1 g N m32 day31)have been reported to occur during the annual dys-trophic period and represent a signi¢cant loss of ni-trogen from this ecosystem [252]. Using benthicchambers incubated in the dark Viaroli and co-work-ers [238,252] showed that V14% of the original Ulva

nitrogen was released as total dissolved nitrogen(DON+DIN) over a 3 day incubation period butonly 6% of this was ammonium whilst in the lightthe values were 8% and 1% respectively. These datanot only indicate that mineralisation of Ulva nitro-gen is relatively slow but that the major decomposi-tion products were DON rather than DIN. Subse-quent experiments have con¢rmed that nitrogenrelease from decomposing Ulva was slow and after60 days did not exceed 50% of the initial N-content[253]. These data indicate that there is temporal un-coupling between the growth and decompositionphases in macroalgae such as Ulva.

In hyper-eutrophic coastal lagoons such as Saccadi Goro, northern Italy, and the Eè tang du Prevost,France, Ulva plays a central role in controlling thenitrogen cycle: in the active growth phase Ulva out-competes the phytoplankton for available nitrogenby virtue of its high a¤nity uptake systems for ni-trate and ammonium whilst during the decomposi-tion phase nitrogen regeneration is slow resulting ina low e¥ux of dissolved inorganic nitrogen to theoverlying water column [247,248]. The degree of ni-trogen limitation is further increased by the stimula-tion of denitri¢cation of water column nitrate duringthe dystrophic crisis period. Whilst it appears para-doxical that nitrogen could be the growth limitingnutrient in these systems which annually receive sub-stantial nitrogen inputs from anthropogenic sourcesthe bulk of it is sequestered in Ulva biomass andtherefore unavailable. The slow release of dissolvedinorganic nitrogen to the overlying water suggests aprolonged retention of nitrogen in the sediment overthe winter months which is then released during thefollowing spring when macroalgal growth is againinitiated. In these systems reduction of macroalgalgrowth can only be achieved by improving waterexchange or by reducing the nitrogen input in thein£ow streams and rivers.

In more hydrodynamic marine environments suchas estuaries which receive considerable nitrogen in-puts from fertiliser run-o¡ and sewage dischargesfrom the adjacent land areas denitri¢cation may ac-count for between 20 and 50% of the annual nitro-gen input into these systems. Denitri¢cation has beenproposed as a `natural control' of coastal eutrophi-cation [170,216,211]. However, nutrient mass balancedata are only available for a limited number of es-

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tuaries [173,254]. As a consequence it is di¤cult toassess the role of denitri¢cation in mitigating thee¡ects of increased anthropogenic nitrogen loadsinto coastal marine environments. Nowicki and co-workers [173] carried out an extensive 3 year study ofdenitri¢cation in Boston Harbour and MassachusettsBay using the N2 £ux method. They recorded highestdenitri¢cation rates in summer in Boston Harbour(6 10^412 mg N m32 day31) and in spring and au-tumn in Massachusetts Bay (6 10^128 mg N m32

day31) coincident with phytoplankton maxima. De-nitri¢cation rates correlated with temperature, or-ganic carbon content and benthic macrofaunal activ-ity. Whilst recorded sediment denitri¢cation rateswere high relative to other US east coast estuaries,the annual integrated denitri¢cation loss for BostonHarbour was only 8% of the annual total nitrogenload. One explanation for the low removal of nitro-gen by denitri¢cation is the relatively short waterresidence time, 2^10 days in the harbour. Recentstudies have postulated that the magnitude of nitro-gen loss through denitri¢cation in estuaries is a func-tion of the water residence time [163,181]. Data pre-sented in Fig. 3 show that in systems with shortresidence time viz Norsminde Fjord, Boston Har-bour and Ochlockonee Bay little denitri¢cation oc-curs and the bulk of the nitrogen load is exportedo¡shore whereas in estuaries, lagoons and bays withlow £ushing rates signi¢cant removal of nitrogen oc-curs. Currently little is known about the couplingbetween estuaries and the adjoining continental shelf

areas and the relative role of denitri¢cation in theeconomy of these systems. Few direct measurementsof denitri¢cation for shelf sediments are availableand more comprehensive studies are required to as-sess the magnitude of denitri¢cation as a permanentsink for nitrogen inputs in these systems before thesigni¢cance of this process can be fully appreciated[255].

8. Concluding remarks

The past decade has seen very major advances inour understanding of the physico-chemical and bi-otic factors regulating nitrogen cycling in shallowcoastal marine sediments. Much of this is due tosigni¢cant improvements in experimental methodol-ogies resulting in greatly improved spatial and tem-poral resolution of the individual processes thatmake up the benthic nitrogen cycle. In particular,the development of microelectrodes for measuringO2, nitrate, nitrous oxide, sul¢de and light irradiancein sediments with a spatial resolution of 6 0.1 mmrepresents an important technological advance. Theapplication of microsensors has led to new insightsinto the spatial distribution of the di¡erent processesinvolved in nitrogen mineralisation and how they areregulated by oxygen, light, sul¢de and carbon avail-ability. A further signi¢cant advance has been thedevelopment of sophisticated computer models tosimulate the complex interactions which occur dur-ing nitrogen cycling in marine sediments. Such anexperimental approach enables the analysis of inter-actions at a resolution that would be unobtainableby other methods. Mathematical models can alsoplay an invaluable role in designing ¢eld experimentsby identifying the key parameters and processes wor-thy of more detailed investigation. The recent trendto integrated research programmes studying nitrogencycling in whole ecosystems involving multi-discipli-nary research teams has been scienti¢cally extremelyproductive and has identi¢ed the important roleplayed by the infauna and macrophyte roots inbenthic nitrogen cycling. These new insights intohow organic matter is mineralised in shallow coastalsediments provide a ¢rm foundation on which todevelop e¡ective management programmes for theseenvironmentally sensitive ecosystems. They also pro-

Fig. 3. Percentage of annual nitrogen load denitri¢ed in variousestuarine and coastal marine systems expressed as a function ofmean water residence time. From Nixon et al. [181].

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vide the basis for remediation programmes to reha-bilitate seagrass communities in lagoons currentlydominated by pelagic macroalgae and/or cyanobac-terial blooms.

Acknowledgements

The author is indebted to Dr. B. Lomstein, Dr.D.T. Welsh and Professor J.A. Raven and the anon-ymous referees for their critical reading of the manu-script and their constructive comments.

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Nitrogen cycling in coastal marine ecosystems

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