The importance of space and time when interpreting trophic structure from stable isotopes

Transkript

1 The importance of space and time when interpreting trophic structure from stable isotopes Master thesis in biology Joan Holst Hansen Marine Ecology, Department of Bioscience, Aarhus University August 2011 The Greenland Institute of Natural Resources Greenland Climate Research Center

2 Forord Denne specialerapport indeholder en generel indledning, som beskriver specialeprojektet og teorierne bag dette, samt et engelsk artikeludkast omhandlende fødenetstrukturen i det marine miljø ved Grønland. Artiklen er formateret til indsendelse til tidsskriftet Marine Ecology Progress Series. Specialet er lavet i samarbejde mellem Afdeling for Marin Økologi, Biologisk Institut, Aarhus Universitet, Grønlands Naturinstitut og Grønlands Klimaforskningscenter. Specialeprojektet har modtaget finansiel støtte af Grønlands Klimaforskningscenter og Forskningsrådet. Peter Grønkjær og Jens Tang Christensen fra Aarhus Universitet var interne vejledere, og Rasmus Berg Hedeholm og Kaj Sünksen fra Grønlands Naturinstitut var eksterne vejledere. Jeg vil hermed gerne rette en stor tak til alle fire for gode råd og grundig vejledning gennem hele specialeforløbet. En stor tak skal også lyde til Grønlands Naturinstitut samt hele besætningen på RV Pâmiut, som stillede laboratorieudstyr til rådighed og hjalp med indsamlingen af data. Endvidere vil jeg gerne benytte lejligheden til at udvise min store taknemmelighed til Rasmus Berg Hedeholm og Kaj Sünksen for at bidrage til et fantastisk ophold i Nuuk. Joan Holst Hansen Aarhus Universitet, august

3 Resumé Specialet omhandler fødenetstrukturen i de grønlandske farvande belyst ved hjælp af stabile kulstofog kvælstofisotoper (δ 13 C og δ 15 N). δ 15 N og δ 13 C er et valideret og effektivt værktøj til økologiske studier. δ 15 N værdien bruges som et integreret mål for en organismes trofiske niveau, da δ 15 N signalet i prædatorer typisk beriges med 3-4 i forhold til deres bytte. Samme berigelse ses ikke for δ 13 C signalet. δ 13 C bruges i stedet som et estimat for oprindelsen af den energi, der ernærer fødenettet (bentisk vs. pelagisk) og kan endvidere relateres til vandmassernes oprindelse. Specialeprojektet omhandler både en rumlig og en tidslig undersøgelse af det marine økosystem. Det rumlige forsøg blev udført langs den grønlandske vest- og østkyst (59 N 72 N). Tolv arter fra alle trofiske niveauer (grønlandshaj, havkat, hellefisk, torsk, rødfisk, håising, lodde, polartorsk, rejer, krill, zooplankton og POM) blev udvalgt til beskrivelse af fødenettet. Det tidslige forsøg blev udført i Nuuk Fjorden over en periode på otte måneder fra april til november. Her blev isotopsignaturen undersøgt for ni arter (hellefisk, helleflynder, torsk, rødfisk, håising, lodde, krill, copepoder og POM) fanget inden for et lille geografisk område. Undersøgelserne af isotopsignaturen viste, at der både er variationer på det rumlige og tidslige plan. Der var en signifikant effekt i δ 15 N på stor geografisk skala og studiet viste en parallelforskydningen af hele fødenettet med breddegraden. δ 15 N værdierne langs den grønlandske vestkyst ændres omkring 3 svarende til et helt trofisk niveau mellem det sydligste og nordligste undersøgte område. Det tidslige forsøg viste en signifikant forskel i δ 13 C med stigende værdier i sommerperioden. Ingen signifikant forskel i δ 15 N blev påvist i løbet af de otte måneder studiet varede. Isotopsignaturen for de undersøgte arter varierede altså på både et stort og lille geografisk område. Disse forskelle er af en størrelse som ikke alene kan skyldes ændret fødeadfærd, men sandsynligvis skyldes variationer i de abiotiske faktorer, såsom forskelle i vandets fysiske og biologiske egenskaber samt terrestrisk udledning til havet. Dette resultat har stor betydning for andre studier, som benytter stabile isotoper (δ 15 N og δ 13 C) til belysning af økologiske spørgsmål, idet manglende hensyntagen til rumlige og tidslige effekter kan have afgørende indflydelse på konklusionen. 2

4 Summary The thesis is concerned with the structure of the marine food web in Greenland using stable carbon and nitrogen isotopes (δ 13 C and δ 15 N). δ 13 C and δ 15 N is a validated and efficient tool when studying food webs. The δ 15 N value is used as an integrated measure of an organisms trophic level, because the signal of δ 15 N in predators are enriched by 3-4 compared to its prey. The same enrichment is not true for δ 13 C. The δ 13 C value is instead used as a proxy of the origin of the energy source of the food web (benthic vs. pelagic) and can also be related to the origin of the water masses. The thesis consists of both a spatial and a temporal study of the marine ecosystem. The spatial study was conducted along the Greenlandic West and East coast (59 N - 72 N). Twelve species (Greenlandic shark, wolffish, Greenland halibut, Atlantic cod, redfish, American plaice, capelin, polar cod, shrimp, krill, copepods and POM) were collected representing all trophic levels to describe the food web. The temporal study was executed in the Nuuk Fjord during an eight month sampling period running from April through November. In this study the isotopic signature was examined for nine species (Greenland halibut, Atlantic halibut, Atlantic cod, redfish, American plaice, capelin, krill, copepod and POM) sampled within a small geographic area. Comparing stable isotope signatures in the arctic marine food web demonstrated both spatially and temporally variations. On a large geographic scale the spatial study showed a significantly latitudinal effect in δ 15 N, revealing a parallel shift of the entire food web. δ 15 N values along the West coast of Greenland changed around 3 from south to north, corresponding to a full trophic level. In the temporal study a significant difference in δ 13 C values was demonstrated with increasing values during the summer. Regarding δ 15 N values no significant difference was shown during the sampling period. Accordingly the isotopic signature of the examined species varied both on a small and large geographic scale. These differences are of a size that not alone can be caused by chancing food preferences, but most likely are due to abiotic factors such as differences in physical and biological properties of the water masses as well as terrestrial inputs. This result will have a large influence on other ecological studies using stable isotopes (δ 13 C and δ 15 N) since such spatial and temporal effects can have a crucial influence on the conclusion if not included. 3

5 Indholdsfortegnelse Dansk introduktion FORORD... 1 RESUMÉ... 2 SUMMARY... 3 INDHOLDSFORTEGNELSE... 4 INTRODUKTION... 6 Arktiske fødekæder... 6 Stabile isotoper... 7 Trofisk fraktionering... 9 Regenereret produktion Begrænsninger ved brugen af stabile isotoper Det grønlandske system Det grønlandske fødenet Oceanografiske forhold Formål med specialet REFERENCER Article: Stable isotope variability in an arctic marine food web on a spatial and temporal scale ABSTRACT... 1 INTRODUCTION... 1 MATERIAL AND METHODS... 3 Spatial Study... 3 Field Sampling... 3 Temporal Study... 4 Field sampling... 4 Stable isotope preparation and analysis for both spatial and temporal studies... 5 Estimation of trophic level... 6 Estimation of relative stable carbon and nitrogen values... 7 Statistical analyses... 7 RESULTS... 7 Spatial Study... 8 Trophic level... 9 Temporal study Inshore-offshore comparison DISCUSSION

6 Spatial Study Temporal Study Inshore-offshore comparison CONCLUSION ACKNOWLEDGEMENTS REFERENCE TABLES AND FIGURES

7 Introduktion Til belysning af økologiske spørgsmål er det helt centralt, at skabe en forståelse af det givne systems struktur og interaktioner mellem de involverede arter. For at kunne gøre dette er det vigtigt at have en fundamental viden om de implicerede organismers fødeadfærd og indbyrdes afhængighedsforhold. Arktiske fødekæder Kort beskrevet er en fødekæde betegnelsen for hvem der spiser hvem. En fødekæde beskriver hvordan energien fra føden videregives fra primærproducenter gennem herbivorer og videre til carnivorer. Økologen Charles Elton (1927) var en af de første, som definerede en simpel fødekæde og opdagede at længden af en fødekæde ofte er begrænset til fire eller fem led (trofiske niveauer). Et eksempel på en fødekæde er den arktiske pelagiske fødekæde: Fytoplankton bliver spist af zooplankton såsom euphausider og copepoder. Disse zooplanktonarter er føde for en række carnivorer, heriblandt fisk, blæksprutter og bardehvaler som til sidst ender som føde for topprædatorer, såsom spækhuggeren (Fig. 1). Figur 1: Skematiseret eksempel af et arktisk fødenet. Efter 6

8 Flere faktorer er dog medvirkende til, at fødekæder aldrig er så simple som dette eksempel. En fødekæde er ikke en isoleret enhed, men derimod sammenkædet med andre fødekæder i et større og mere kompliceret fødenet. Endvidere kan en art prædere på flere trofiske niveauer, og aldersbaserede fødeskift gør også at en fisk kan bevæge sig op gennem fødekæden efterhånden som de bliver større. Stabile isotoper Stabile isotoper er ikke-radioaktive atomer af samme grundstof. Stabile isotoper har samme antal protoner i kernen og derfor samme atomnummer, men har forskellige antal neutroner hvilket giver et forskelligt massetal. Mange grundstoffer har mere end én stabil isotop. Dette gælder for eksempel for grundstofferne kulstof (C), kvælstof (N), ilt (O), svovl (S) og hydrogen (H), som forekommer i atmosfæren, jorden, planter og dyr. Eksempelvis akkumuleres kulstof og kvælstof i væv og knogler hos dyr. Stabile isotoper har store anvendelsesmuligheder og kan blandt andet bruges til bestemmelse af en organismes forskellige fødekilder. Ikke kun i nulevende organismer, men også eksempelvis i fortidige mennesker hvor man ved hjælp af kulstof- og kvælstofisotoper i knogler kan se om føden stammer fra landjorden, havet eller søer (Szepanski et al. 1999; Arneborg et al. 1999). Andre anvendelsesmuligheder er iskernedatering, hvor iltisotoper bruges til at estimere tidligere tiders temperaturvariationer (Johnsen et al. 2001), estimering af trofisk niveau (Peterson & Fry 1987) eller identificering af forskellige biologiske processer (Lee et al. 2010). Marine økosystemer rummer ofte store og komplekse fødenet (Fig. 1), hvilket komplicerer evalueringen af disse. Ved hjælp af stabile isotoper, særdeles kulstof og kvælstof, er det dog muligt at undersøge strukturen og dynamikken af marine fødenet (Peterson & Fry 1987; France 1995a; Vander Zanden et al. 1999). Brugen af stabile kulstof- og kvælstofisotoper til undersøgelse af marine økosystemer startede da DeNiro og Epstein (1978 og 1981) fandt en sammenhæng mellem ratioen af den tunge og den lettere isotop af kulstof ( 13 C/ 12 C) og kvælstof ( 15 N/ 14 N) i en organisme og dens føde. Der er i løbet af de seneste årtier bygget på denne viden, og brugen af stabile isotoper til evaluering af fødenet er tiltaget kraftigt (Hobson & Wasenaar 1999). Forholdet mellem 13 C/ 12 C og 15 N/ 14 N betegnes δ 13 C og δ 15 N, og udtrykkes i promille ved følgende notation: δ ø hvor X er 13 C eller 15 N og R er den tilhørende 13 C/ 12 C eller 15 N/ 14 N ratio. 7

9 Grundet diskrimination mod de tunge isotoper ( 13 C og 15 N) i organismers biokemiske reaktioner akkumuleres de tunge isotoper op gennem fødekæden fra bytte til prædator i vævet. Berigelsen i isotopsammensætning mellem bytte og prædator, kaldet trofisk fraktionering, er estimeret i flere studier til værende 0-1 for δ 13 C og 3-4 for δ 15 N (DeNiro & Epstein 1978 og 1981; Peterson & Fry 1987; Post 2002; McCutchan et al. 2003; Sweeting et al. 2007a og 2007b). Forskellen i fraktioneringsraten mellem kulstof og kvælstof skyldes forskelle i de stofskiftereaktioner i organismen, som forårsager fraktioneringerne. Fraktioneringen af kulstof skyldes hovedsageligt afgivelse af isotopisk set lettere CO 2 i forhold til føden. Katabolisme af proteiner, lipider, og kulhydrater bevirker en diskriminering mod den tungere 13 C isotop når acetyl-grupper oxideres og isotopisk let CO 2 frigives fra organismen (Galimov 1985). Fraktioneringen i δ 13 C er som tidligere nævnt lille (0-1, Peterson & Fry 1987) men også variabel, og faktisk fandt DeNiro & Epstein (1978) lignende δ 13 C værdier mellem primærproducenter og primærkonsumenter, mens Sweeting et al. (2007b) fandt en fraktionering for fisk på 1-2. På grund af den lille fraktionering i δ 13 C mellem bytte og prædator er stabile kulstofisotoper ikke ideelle til bestemmelse af trofisk niveau som kvælstof er, men bruges i stedet til bestemmelse af organismers primære kulstofkilde (Vander Zanden & Rasmussen 1999, Post 2002). Dette skyldes en rumlig variation i sammensætningen af kulstof isotoper, hvorved det er muligt at anvende δ 13 C til bestemmelse af organismers levested samt eventuelle migrationsmønstre. Det er således muligt at undersøge om en organisme lever på land, i ferskvand nær kysten eller i vandsøjlen (France 1995b) eller i stille eller turbulent vand (Osmond et al. 1981). Eksempelvis undersøgte Vander Zanden & Rasmussen (1999) isotopsignaturen hos primærkonsumenter i søer og fandt en forskel i δ 13 C på cirka 4 mellem den littorale zone og den pelagiske zone, hvor primærkonsumenterne i den littorale zone havde mest berigede δ 13 C værdier. Desuden er det også muligt, at undersøge om en marin art er tilhørende en pelagisk vs. indenskærs eller bentisk vs. udenskærs fødekæde (France 1995b; Lawson & Hobson 2000). Dette skyldes, at δ 13 C værdien i bentiske organismers fødekilde har en anden isotopisk sammensætning end de pelagiske organismers, hvor pelagisk fytoplankton overvejende er kulstofkilden (Hobson 1999). Dette betyder at bentiske organismer har mere berigede δ 13 C værdier (mindre negative) end pelagiske organismer, da bentiske primærproducenter lever i mindre turbulent vand og dermed bliver diffusionslaget større, hvilket bevirker at mere 13 C assimileres af planten. Dette isotopsignal kan følges i andre organismer på højere trofiske niveauer i både den bentiske og pelagiske fødekæde. Modsat kulstofisotoper er fraktioneringen i kvælstof stor og foregår under kvælstofudskillelse (Peterson & Fry 1987), hvor den lettere 14 N isotop reagerer hurtigere end den tungere 15 N isotop. Derved vil en prædator have en højere δ 15 N værdi sammenlignet med dens føde. Fraktioneringen for akvatiske organismer i δ 15 N er 3-4 og en gennemsnitlig værdi på 3,4 er generelt accepteret (Post 2002). Der er dog ikke entydig konsensus omkring denne fraktioneringsgrad og flere studier har 8

10 foreslået en lavere kvælstoffraktionering. Sweeting et al. (2007a) fandt en fraktionering på 2,9 ved analyse af hele fisk og McCutchan et al. (2003) fandt ved undersøgelse af tidligere studier en lavere gennemsnitlig fraktioneringsværdi på 1,4 for organismer, som har levet af invertebrater, mens konsumenter der fik en proteinholdig føde havde en fraktionering på 3,3. Trofisk niveau På grund af denne store fraktionering i δ 15 N mellem prædatorer og deres bytte, er δ 15 N ideel til estimering af trofisk niveau. Dette kan gøres ved hjælp af følgende formel (Peterson & Fry 1987): δ æ δ δ hvor δ 15 N prædator er δ 15 N værdien for den pågældende art, Δδ 15 N er fraktioneringen i δ 15 N per trofisk niveau, δ 15 N basis er den gennemsnitlige δ 15 N værdi af den valgte art til basisniveau og TL basis er det trofiske niveau for den art (eks. 1 for fytoplankton, 2 for zooplankton under antagelse af, at det er herbivor zooplankton). Eksempelvis estimerede Nilsen et al. (2008) trofisk niveau for en række invertebrater og fisk i en fjord i det nordlige Norge. Dette blev gjort både ved hjælp af δ 15 N værdier og ovenstående formel samt til sammenligning ved brug af en økologisk model (ecopath). En ecopathmodel kan ved hjælp af relativ få og grundlæggende input om økosystemets komponenter (biomasse, produktion, konsumption) estimere trofisk niveau for hver komponent. Nilsen et al. (2008) fandt en god korrelation (r 2 = 0,72) mellem trofisk niveau beregnet ud fra δ 15 N og beregnet ved brug af ecopathmodellen. Dette verificerer brugen af stabile kvælstofisotoper til estimering af trofisk niveau, uden man har en detaljeret viden om den faktiske fødeadfærd af en organisme, hvilket kræver en relativ stor indsats og kendskab til det pågældende system og dets arter. Trofisk fraktionering Fraktioneringen i både kulstof og kvælstof er som nævnt ovenfor ikke konstant og kan variere afhængig af art, individets størrelse og kondition (Bode et al. 2007), temperaturen i det omgivende miljø (Barnes et al. 2007), væksthastighed (Trueman et al. 2005), vævstype og vævets omsætningsrate (Tieszen et al. 1983, Lorrain et al. 2002). Dette betyder blandt andet, at væv med en høj omsætningsrate (måneder til år), såsom knogler og muskelvæv, integrerer isotopsignaturen fra føden over en længere periode end lever- og gonadevæv, der har en høj omsætningsrate (dage til uger). Det er derfor vigtigt, alt efter hvor lang en tidsperiode man er interesseret i at undersøge, at udvælge den korrekte vævstype til undersøgelse af isotopsignaturen (O Reilly et al. 2002, Post 2002). Udover den 9

11 høje omsætningsrate er væv som lever og gonader også ofte lipidholdige organer. Sammenlignet med kulhydrater og proteiner indeholder lipider mindre 13 C (Griffiths 1991; Sotiropoulos et al. 2004), hvilket betyder at en varierende mængde lipid i de forskellige væv påvirker δ 13 C værdierne (Lorrain et al. 2002). Derfor anbefaler flere studier at lipidekstrahere, hvis man laver δ 13 C analyser på lipidholdigt væv (Post 2002; Sweeting et al. 2007b). Med hensyn til δ 15 N er effekten af lipidekstrahering mere uklar, da tidligere studier har vist et lidt tvetydigt resultat, med ingen eller lille effekt af lipidekstrahering (Sweeting et al. 2006; Sweeting et al. 2007a; Søreide et al. 2006). Regenereret produktion I den eufotiske zone er de vigtigste former for uorganisk kvælstof ammonium (NH 4), nitrat (NO 3) og N 2 gas (Michener & Kaufman 2008). δ 15 N i primærproducenterne påvirkes af disse forskellige former af uorganisk kvælstof, som findes i vandsøjlen, samt af forholdet mellem mængderne af ny og regenereret kvælstof. Kvælstof som tilføres de frie vandmasser fra dybere vande, kvælstoffiksering og atmosfæren betegnes som nyt kvælstof. Da kvælstofkredsløbet er et delvist lukket kredsløb vil dette kvælstof atter indgå efter nedbrydning, nu som regenereret kvælstof. Regenereret kvælstof er således den plantetilgængelige kvælstof (ammonium og urinstof) i den eufotiske zone, der bliver gendannet i havet. Dette kan påvirke δ 15 N værdierne, da regenereret produktion oftest har højere δ 15 N værdier end ny produktion. Eksempelvis fandt Ostrom et al. (1997) i det kolde marine miljø ud for Conception Bay, Newfoundland, højere δ 15 N i partikulært organisk materiale (POM), grundet en drastisk berigelse i 15 N angiveligt på grund af højere δ 15 N i den regenererede produktion i området. Således var δ 15 N værdierne for ammonium i vandet højere end δ 15 N for nitrat. Det er dog ikke altid regenereret primærproduktionen giver berigede δ 15 N værdier, som eksempelvis Needoba et al. (2006) der fandt lavere δ 15 N værdier. Det er derfor altid vigtigt, at tage forbehold for disse eventuelle forskelle ved sammenligninger på tværs af studier. Begrænsninger ved brugen af stabile isotoper Brugen af stabile isotoper som et økologisk redskab indebærer en mængde fordele og muligheder. Disse er dog baseret på flere antagelser (eksempelvis fraktionering) og følgeligt flere begrænsninger. Når man bruger stabile kulstof og kvælstof isotoper til belysning af fødenetstruktur og energiflow, bliver resultatet som tidligere nævnt et integreret estimat af det samlede fødeindtag. Dette leder til en af de helt store begrænsninger, da det derved ikke er muligt direkte at udlede hvilke specifikke arter den undersøgte organisme har konsumeret. Til dette skal bruges en analyse af maveindholdet, hvilket dog kun giver et øjebliksbillede af den indtagne føde. En kombination af disse to metoder er derfor ofte den optimale tilgang til en detaljeret fødeanalyse, ligesom fedtsyreanalyse kan supplere billedet af fødeadfærden (Petursdottir et al. 2008) 10

12 En af de store udfordringer ved brugen af stabile isotoper er endvidere, at kunne sammenligne resultaterne på tværs af forskellige økosystemer. Ved sammenligninger på tværs af økosystemer giver δ 13 C og δ 15 N for én organisme alene ganske lidt information angående dens absolutte trofiske niveau eller den ultimative kulstofkilde (Kling et al. 1992, Post et al. 2000). Dette skyldes, at der er betydelig variation mellem økosystemer i δ 13 C og δ 15 N ved fødekædens basis (δ 13 C basis og δ 15 N basis). Uden et korrekt estimat af δ 13 C basis og δ 15 N basis i hvert økosystem er det ikke muligt, at bestemme om en given variation i δ 13 C og δ 15 N hos en organisme skyldes ændringer i fødenetstruktur og kulstofkilde eller ændringer i δ 13 C og δ 15 N ved basis af fødekæden. Variation i δ 13 C basis og δ 15 N basis skyldes forskelle i isotoprationen i det biologisk tilgængelige kulstof og kvælstof ved basis af fødekæden og variation i fraktioneringsraten. De fleste primærproducenter i marine økosystemer har en høj variation i δ 13 C og δ 15 N over tid, hvilket komplicerer deres direkte anvendelighed som indikatorer af δ 13 C basis og δ 15 N basis for organismer højere oppe i fødekæden (Cabana & Rasmussen 1996). På grund af sådanne forskelle i oprindelsen af kvælstof kan der forekomme både rummelige og tidslige variationer i basisniveauet hos primærproducenterne og følgelig også på højere trofiske niveauer. For at mindske variationen i basisniveau, vil det ofte være mere ideelt at bruge længerelevende organismer med en længere omsætningsrate i vævet. Dette kunne eksempelvis være muslinger, da disse som regel lever i flere år og er forholdsvis stationære, hvilket gør dem mindre sensitive overfor tidslige variationer. Alternativt kan herbivore zooplanktonarter bruges, da de ernærer sig direkte af det tilgængelige fytoplankton. Det vil især være nødvendigt hvis man arbejder i ikke-kystnære systemer hvor muslinger ikke er tilgængelige. Det grønlandske system Grønland har en mere end 2300 kilometer lang uafbrudt nord-sydgående kyststrækning og en række faktorer er varierende grundet denne store breddegradsgradient. Eksempelvis er mængden af lysindstrålingen i vandet meget varierende fra syd til nord, hvilket skyldes et faldende antal soltimer og en stigende mængde is med breddegraden. Også temperaturen varierer langs denne breddegradsgradient. Generelt er temperaturen faldende med stigende breddegrad, hvilket også er tilfældet med lufttemperaturen i Grønland, men ikke nødvendigvis for temperaturen i havet. Vandtemperaturen langs den Vestgrønlandske kyst er mere kompliceret og styres overordnet af to forskelligt tempererede dominerende vandmasser (se afsnittet om oceanografiske forhold) (Ribergaard 2011). Disse faktorer er medvirkende til, at også forårsopblomstringen af primærproduktionen i de grønlandske farvande er varierende langs denne breddegradsgradient. 11

13 Det grønlandske fødenet Det grønlandske fødenet er som andre arktiske fødenet domineret af relativt få arter (Roy et al. 1998, Allen et al. 2002). Eftersom marine organismer direkte eller indirekte er afhængige af det første led i fødekæden, primærproduktionen, har den rumlige og tidslige variation af primærproduktionen stor betydning for hele fødekæden. Primærproduktionen i de grønlandske farvande er afhængig af mængden af sollys, men selvfølgelig også af mængden af tilgængelige næringsstoffer. Dette betyder, at der er store sæsonmæssige variationer i primærproduktionen (Fig. 2). Denne store tidslige variation i primærproduktionen har som tidlige nævnt ikke kun betydning for organismer, som lever direkte af primærproduktionen. En stor del af primærproduktionen ender som føde for eksempelvis copepoder, som udgør grundlaget for fødenettene i de frie vandmasser, mens en anden del af primærproduktionen synker til havbunden og derved bidrager til at ernærer den bentiske fødekæde. Sæsonvariationen i primærproduktionen har altså stor betydning for hele fødenettet også for organismer på højere trofiske niveauer. Figur 2: Årlig variation i primærproduktionen ved Nuuk (64 N) i 2007 (Jensen & Rasch 2008). Oceanografiske forhold De hydrografiske forhold omkring den sydlige del af Grønland er karakteriseret af den Øst- og Vestgrønlandske Strøm (Fig. 3). Den kolde og lavsaline Østgrønlandske Strøm fra Polarhavet mødes med den tempererede Irmingerstrøm i Danmarksstrædet mellem Grønland og Island. Irmingerstrømmen er en gren af den Nordatlantiske Strøm, der udgøres af varmt og højsalint vand. Disse to havstrømme samles under vedvarende opblanding langs den grønlandske vestkyst i den Vestgrønlandske Strøm (Ribergaard 2010, Fig. 3), hvor den tungere og varmere Irmingerstrøm placeres under den Østgrønlandske Strøm. Den gradvise opblanding af de to vandmasser langs den grønlandske vestkyst resulterer i en opvarmning overfladevandet og vice versa for bundvandet. 12

14 Omkring Diskoøen (69 N) har den Vestgrønlandske Strøm aftaget i styrke og forgrener sig mod David Strædet, hvilket dermed resulterer i at vandets overfladetemperatur falder fra Diskoøen og længere nordpå. Dog varierer sammensætningen af komponenterne i den Vestgrønlandske Strøm fra år til år, hvilket er en medvirkende faktor til temperaturvariationer i vandmasserne langs den grønlandske kyst (Ribergaard 2010). Sådanne udsving i temperatur mellem år kan have stor betydning for organismer på alle trofiske niveauer i det marine økosystem. Det er blandt andet vist, at væksten hos flere arter i grønlandsk farvand påvirkes af disse rumlige forskelle i vandmassernes abiotiske faktorer. Hos hellefisk (Reinhardtius hippoglossoides) og lodde (Mallotus villosus) ses ændringer i væksten grundet temperaturvariationerne i vandmasserne. Således har hellefisk en faldende vækstrate med stigende breddegrad (Sünksen 2009), mens det modsatte vækstmønster ses for lodden (Hedeholm 2010) konsistent med det inverse temperaturmønster som følge af opblandingen. Udover disse overordnede mønstre giver den lokale bundtypografi og meget dybe fjorde en yderligere variation i vandmassernes input til forskellige områder (Mortensen et al. 2011). Figur 3: Havstrømme ved Grønlands Vest- og Østkyst (Pedersen & Smidt 2000). 13

15 Formål med specialet Kun ganske få studier har tidligere undersøgt den marine fødekæde langs Grønland. Størstedelen af disse har brugt mere traditionelle metoder, såsom maveanalyser. Enkelte analyser med stabile isotoper er blevet udført til belysning af den marine fødekæde langs vestkysten, men i disse studier har eventuelle ændringer i basisniveau langs breddegradsgradienten ikke været medtaget. Der er altså så vidt vides ingen tidligere studier, som har undersøgt eventuelle tidslige eller rumlige forskelle i isotopsignalet i den marine fødekæde ved Grønland. Det er netop disse problemstillinger jeg vil belyse i specialeprojektet. Resultaterne fra det rumlige forsøg langs den grønlandske vest- og østkyst vil give et detaljeret billede af det marine fødenet på et stort geografisk område, samt klarlægge om en ændring af basisniveau langs kysten er forårsaget af opblanding af vandmasser. Resultaterne fra det tidslige forsøg udført i Nuuk fjorden fra april til november, vil kunne belyse organismers eventuelle fødeskift samt ændringer i basisniveau i løbet af perioden. Resultaterne fra dette projekt har også stor betydning for andre studier, som benytter stabile isotoper (δ 13 C og δ 15 N) til belysning af økologiske spørgsmål. Dette gør sig ikke kun gældende for studier udført i det arktiske miljø, men for alle studier som undersøger fødenetstrukturer, da manglende hensyntagen til rumlige og tidslige effekter kan have afgørende indflydelse på konklusionen. 14

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21 Stable isotope variability in an arctic marine food web on a spatial and temporal scale Joan Holst Hansen 1, Rasmus Berg Hedeholm 2,3, Kaj Sünksen 2, Jens Tang Christensen 1, Peter Grønkjær 1 1Marine Ecology, Department of Bioscience, Aarhus University, 8000 Aarhus C, Denmark 2Greenland Institute of Natural Resources, PO box 570, 3900 Nuuk, Greenland 3Greenland Climate Research Centre, PO box 570, 3900 Nuuk, Greenland Abstract Stable isotopes of carbon (δ 13 C) and nitrogen (δ 15 N) were used to examine trophic structures in an arctic marine food web on both a spatial and temporal scale. 12 species in total were examined around the Greenlandic West and East coast, from primary producers through Greenland shark. There was a significant latitudinal effect on δ 15 N values, with a difference of 2 to 4 between the most southern and northern areas. The temporal study was conducted on nine species in the Nuuk Fjord during an eight month long sampling period. A significant temporal effect was seen in δ 13 C values, with enriched values during the summer period. δ 15 N values did not change significantly with time, but an increasing tendency occurred at the end of the period (November). These significant differences in isotopic signatures on both a large and small geographic scale are most likely unrelated to feeding, but rather due to different physical and biological properties of the water masses, which would be reflected in the trophic signatures at the base of the food web and thereby also in higher trophic levels. Generally the results illustrate the importance of space and time when interpreting trophic structure from stable isotopes. Key words: δ 15 N, δ 13 C, Greenland, latitudinal effects, marine food web Introduction Analyses of nitrogen and carbon stable isotopes are a validated and commonly applied method to describe food web structure in marine ecosystems (e.g. Peterson & Fry 1987; Post 2002). The stable isotope ratios of carbon (δ 13 C) and nitrogen (δ 15 N) provide a time-integrated measure of an organisms trophic position and feeding ecology covering periods ranging from weeks to months, compared to the 1

22 traditional snap-shot picture provided by stomach content analyses, and have the potential to track energy flow through food webs (Hobson & Welch 1992; Hobson et al. 1995; Post 2002). The method is based on the principle that heavier isotopes ( 13 C and 15 N) accumulates from prey to predator (i.e. fractionation). The fractionation of nitrogen in consumers is on average between 3-4 relative to their diet (Post 2002; Søreide 2006). Thus stable nitrogen isotopes provide a good estimate of a consumers trophic position given a known baseline δ 15 N. Stable carbon isotopes are a less useful index of trophic position because the trophic fractionation of 13 C typically is less than 1 (DeNiro & Epstein 1978 and 1981; Vander Zanden & Rasmussen 2001; Post 2002). The stable carbon isotope ratio (δ 13 C) can however be useful when evaluating sources of primary production in marine systems as well as general patterns of inshore or benthic versus offshore or pelagic feeding preferences (France 1995a; Lawson & Hobson 2000). Benthic and inshore organisms in marine ecosystems in general have more enriched δ 13 C values as opposed to the more depleted δ 13 C values of pelagic and offshore organisms (France 1995a). Thus, the integrated use of δ 13 C and δ 15 N can provide valuable information on both food web structure and carbon source. Trophic level is estimated relative to a chosen food web base (δ 15 N base). This allows for comparison of segregated samples (spatially and/or temporally) in spite of differences in δ 15 N base. Therefore, if a proper δ 15 N base estimate is not available it is not possible to determine if variation in δ 15 N values between systems reflects differences in food web structure or baseline variation. δ 15 N base values are predominantly influenced by nitrogen limitation (Pennock et al. 1996), inter-specific differences in isotope fractionation (Needoba et al. 2003) and form of nitrogen assimilated (new primary production (nitrate) vs. regenerated production (ammonium), Ostrom et al. 1997). Because of these influencing factors both the baseline and the absolute nitrogen values will vary in time and space due to changing chemical and biological conditions (Cabana & Rasmussen 1996; Post 2002) and changes can be seen on a small geographic scale (Pentoja et al. 2002). The water masses in the coastal areas around Greenland are complex and consist of two main currents. These are the cold low saline East Greenland Current from the Polar Sea and the temperate saline Irminger Current which originates from the Atlantic Ocean. The two currents meet at the southern area of the Greenland East coast, with the heavier Irminger Current subducting the relatively low saline East Greenland current. The water masses round Cape Farewell, forming the West Greenland Current were the two water masses gradually mix flowing north (Ribergaard 2011). Petursdottir et al. (2008) showed that the δ 15 N value of Calanus finmarchicus caught in June in the Irminger Current of the Reykjanes Ridge south of Iceland was (mean ± SE) 3.5 ± 0.1, while Søreide et al. (2006) found a δ 15 N value of 6.4 ± 0.2 in September for C. finmarchicus in the Fram Strait west 2

23 of Svalbard. This clearly demonstrates that water masses around Greenland have different physical and/or biological properties, which is reflected in shifts in the isotopic baseline dependent on the degree of mixture between water masses. This in turn can be affected by large scale circulation patterns or local area bathymetry such as sill fjords. Hence differences in the isotopic baselines would be expected to vary on both large (Tamelander et al. 2009) and small geographical scale (Tamelander et al. 2006). Therefore, it is hypothesized that a significant latitudinal gradient in δ 15 N values would be found as illustrated in capelin by Hedeholm (2010) in Greenland. As Greenland has an extensive northsouth directed coast line with a number of relative large fjord systems with glacier run off that differ from offshore regions (Mortensen et al. 2011) it is an ideal place to study effects of geographical separation on food web structure and shifts in δ 15 N and δ 13 C values irrespective of changes in trophic position. The main objectives of this study were threefold. 1) Provide a novel detailed description of the marine food web on a large geographic scale along the Greenland coast with a quantification of the effect differences in sampling site may have on isotopic signatures. 2) Temporal variation in stable isotope structure and finally 3) importance of local area variation was described by comparing samples from the Nuuk fjord and adjacent offshore waters. Material and Methods The study has both a spatial and temporal component. The species and size categories selected for sampling in both aspects of the study were based on their ecological relevance in the Greenlandic ecosystem while also attempting to span as much as possible of the ecosystems trophic range. Spatial Study Field Sampling All samples were collected from the trawler RV Pâmiut (722 GRT) from June to August 2010 during the annual stratified-random bottom trawl surveys carried out by the Greenland Institute of Natural Resources in Greenlandic waters. Samples included seven teleost fish species: wolffish (Anarhichas lupus and Anarhichas minor), Atlantic cod (Gadus morhua (small (25-35cm) and large (45-55cm))), polar cod (Boreogadus saida) American plaice (Hippoglossoides platessoides), capelin (Mallotus villosus), Greenland halibut (Reinhardtius hippoglossoides), redfish (Sebastes mentella (small (15-25cm) and large (35-45cm))) and Greenland shark (Somniosus microcephalus), shrimps (Pandalus 3

24 borealis), krill (Thysanoessa raschii), copepods (Calanus finmarchicus and C. glacialis) and filtered POM giving a total of 14 sample groups (Table I and II). Fish were sampled in six predetermined areas at depths between 88 and meter along the Greenlandic coast from Upernavik (72 N) in west to Tasiilaq (66 N) in east (Fig. 1). For each of the predetermined areas it was attempted to sample five individuals of each sample group (Table I). To obtain the best representation of the within area variation, individuals were sampled from as many catches within each area as possible. Whenever, regardless of the six predetermined areas, a Greenland shark was caught the total length, weight and sex was determined and a muscle sample was taken near the dorsal fin before it was released. Shrimps, krill, copepods and filtered POM were collected at eleven different sites separated by approximately two latitudinal degrees (Fig. 1 and Table II). Shrimps and krill were taken from trawl hauls whereas copepods and filtered POM were sampled using a 500 µm plankton net and Niskin water bottle (5 L), respectively. The plankton net was lowered to 150 m and then slowly retrieved (20 m min -1 ) while the vessel was moving at 1 knot. This was done in the evening to minimize copepod gut content (Lampert & Taylor 1985; Head & Harris 1987). The content from the cod end was first filtered though a 2 cm sieve and afterwards a 200 µm sieve to remove jellyfish, chaetopods etc. and retain the larger copepods, respectively. The copepods were kept frozen on a filter until further analysis. The Niskin bottle was lowered to ten and twenty meters, respectively. Two liters from each depth was filtered though a 200 µm filter to remove large organisms (i.e. jellyfish, amphipods etc). Three liters of this water was then filtered onto a pre-combusted (450 C for 24 hours) GF/F filter (47 mm in diameter). All samples from all groups were immediately frozen at -20 C. A CTD profile was obtained at approximately every latitudinal degree along the Greenlandic west coast (60-71 N). Unfortunately the CTD malfunctioned on all East coast stations and at 62 N, 65 N and 66 N on the West coast. Temporal Study Field sampling Samples were collected in the Nuuk Fjord (64 N, Fig. 1) four times during an eight-month period from April to November 2010 and included six fish species (Atlantic cod (small and large), redfish, American plaice, Atlantic halibut, Greenland halibut and capelin), krill, copepods (C. finmarchicus) and filtered POM (Table III). Most fish species (except Greenland halibut) and phytoplankton were sampled at the same location, but logistic circumstances entailed that copepods and to some extent krill had to be sampled at a different site within the fjord (Fig. 1). Most fish species were caught using fishing rods, whereas undigested capelin were taken from cod stomachs immediately upon capture. Greenland 4

25 halibut were bought from the local fish market and were caught in the fjord using a long-line set at meters. Krill and copepods were collected at the inner part of the fjord with a 2 m in diameter and 600 µm mesh size MIK net and a MultiNet (Hydrobios) equipped with five 300 µm net. Samples of filtered POM were collected similarly to the spatial study. Stable isotope preparation and analysis for both spatial and temporal studies Total length, weight and sex of the fish were determined in the laboratory. Length of shrimps and krill were measured from the post-orbital notch to the posterior margin of the carapace and from the postorbital notch to the posterior end of the uropods, respectively. The weight of shrimp, krill and copepods were noted. Fish samples were prepared by removing white muscle tissue (10.31 ± 4.26 g wet weight, mean ± SD) dorsally from both sides of the fish, posterior to the dorsal fin which ensured that no bones were present in the sample. All skin was subsequently removed. White muscle tissue was used for stable isotope analysis because it best represents the isotopic signature of fish (Rounick and Hicks 1985; Hesslein et al. 1993) and does not require the removal of inorganic carbonates (Pinnegar & Polunin 1999). Muscle tissue was also dissected from shrimp and krill after the chitinous exoskeleton and the gut were removed. All samples were freeze-dried (Freeze dryer ALPHA 1-2/LD plus) to constant mass at -60 C for 24 hours. The dry samples were kept in an exsiccator containing silica gel until further analysis. In order to make stable isotope results from the lipid-rich copepods (Lee et al. 2006) comparable to other groups, lipids were removed using a chloroform-methanol solution (2:1) (Søreide et al. 2007). In order to confirm that this was necessary an initial experiment was performed at the Greenland Institute of Natural Resources. It was found that removing lipids had a significant effect on δ 13 C values and these significant values were then comparable to other groups. Whereas the removal of exoskeleton (using 10% HCL) had no significant effect compared to the non-treated samples on neither δ 13 C nor δ 15 N values. The dried muscle tissue from shark, fish, shrimps and krill was homogenized using a mortar and pestle. Approximately 1 mg (dry weight, dw) of sample (1.14 ± 0.10 dw, mean ± SD) was weighed into pre-weighted tin capsules (5x9 mm). Copepods with no visible stomach content were selected, and packed into the tin capsules as whole individuals. At least three copepods collected at the same time and place were pooled in one sample to obtain sufficient material for isotopic analysis. Filtered POM (3.09 ± 0.05 mg dw, mean ± SD) was carefully removed from the filter with needles to minimize the amount of GF/F filter mixed into the sample, and weighed into tin cups. 5

26 A total of 397 samples were analyzed in the spatial study (Table I and II) and 118 samples in the temporal study (Table III). Stable carbon and nitrogen isotope analyses were performed at the UC Davis Stable Isotope Facility in California, USA. Samples were combusted in a PDZ Europa ANCA-GSL elemental analyzer interfaced to a PDZ Europa isotope ratio mass spectrometer (Sercon Ltd., Cheshire, UK). The crimped tin capsules were introduced via a solid autosampler and combusted at 1000 C in a reactor packed with chromium oxide and silvered cobaltous/cobaltic oxide. Following combustion, oxides are removed in a reduction reactor (reduced copper at 650 C). A post-reactor gas chromatography (GC) column was kept at 65 C for separation of evolved N 2 and CO 2 before entering the IRMS. Stable isotopes are expressed in a δ notation as the deviation from international standards in parts per thousand ( ) according to the formula: δ where X is 13 C or 15 N and R is the corresponding ratio 13 C/ 12 C or 15 N/ 14 N. Standards for δ 13 C and δ 15 N were calibrated against Vienna PeeDee Belemnite and atmospheric air, respectively. Estimation of trophic level Trophic level (TL N) was estimated for all groups based on the following formula: δ δ δ where δ 15 N consumer is the δ 15 N of the species in question, Δδ 15 N is the enrichment in δ 15 N per trophic level, δ 15 N base is the average δ 15 N of the group chosen as the base of the food web and TL base is the trophic level of that group. Copepods (C. finmarchicus and C. glacialis) were chosen as the base assuming a strictly herbivorous diet (e.i. TL base = 2, Hobson & Welch 1992; Søreide et al. 2006; Nilsen et al. 2008). Filtered POM was not used as the values showed some irregularities (see results and discussion). A Δδ 15 N of 3.4 was applied for invertebrates which were based on analyzes of samples from the Barents Sea (Søreide et al. 2006) and a separate Δδ 15 N value of 3.2 for fish (Sweeting et al. 2007). The two different enrichment factors indirectly indicate variations in enrichment correlated to body size, diet (Fry et al. 1999), growth rate (Trueman et al. 2005) and tissue catabolism (Kurle & Worthy 2001; Olive et al. 2003). 6

27 Estimation of relative stable carbon and nitrogen values In order to detect changes in the isotopic signal across latitudes for the entire food web, comparable relative values of δ 13 C and δ 15 N were calculated. A single relative value was first estimated for each species in each area according to the formula: A single relative value was then calculated for each predetermined area as the mean of all species within that area: where is the relative value of δ 13 C or δ 15 N in any of the predetermined areas referred to as i. is the mean of either δ 13 C or δ 15 N for a given species in one of the areas. N is the number of areas. In this study N always equals six because there are six predetermined study areas. n is the number of species in area i, which in this study equals ten because if a species is not caught in every area it is not included in this calculation since this would bias the result. Statistical analyses All analyses were carried out using the statistical computing program R (R Development Core Team 2011). Standard parametric test were preceded by test for assumptions. When these were violated the data were either transformed or non-parametric statistics were applied. Filtered POM was excluded from all statistical analyzes as the measured δ 15 N values were unusually high in some of the sampling stations (see discussion). Results A total of 515 samples were taken along the Greenland coast from 72 N on the west coast to 66 N on the east coast (N = 397) and in the Nuuk Fjord (N = 118) during Stable carbon and nitrogen isotope ratios were analyzed for all samples representing 13 species and 15 sampling groups (Tables I-III). 7

28 Spatial Study Twelve species and 14 sampling groups were analyzed for δ 13 C and δ 15 N (Table I and II). Average stable isotope composition of the analyzed species ranged from to in δ 13 C and from 3.3 to 17.2 in δ 15 N. The most depleted δ 15 N values (mean ± SE) were found in filtered POM (6.43 ± 1.40 ) and copepods (7.76 ± 0.57 ) while Greenland shark (16.68 ± 0.25 ) followed by wolffish (13.35 ± 0.79 ) and Greenland halibut (12.87 ± 0.39 ) were the most enriched in δ 15 N. The strictly benthic organisms (i.e. Greenland shark, wolffish, American plaice and shrimp) were enriched in δ 13 C ( to ) compared with strictly pelagic organisms (copepod, krill, polar cod, capelin and Greenland halibut) which ranged from to (Fig. 2). There were large variations in δ 15 N values for filtered POM among the six different sampling areas. δ 15 N values ranged from (mean ± SE) ± 1.57 in area 1 to 3.26 ± 0.19 in area 6. It is also remarkable that the δ 15 N values of wolffish in all cases were higher on the West coast compared to δ 15 N values of Greenland halibut, but on the East coast the opposite pattern was found (Fig. 2). Furthermore, the mean δ 13 C values of polar cod on the East coast were depleted 2.1 compared to the mean values on the West coast (Fig. 2). There were significant regional differences in δ 15 N values among the six study areas along the Greenlandic coast (Two-way ANOVA, F 5,351 = 99.58, P < ) irrespective of the also significant species effect (Two-way ANOVA, F 13,351 = 91.41, P < ). Species specific tests showed a significant effect of area (i.e. latitude) on all species (P 0.026) except for Greenland shark (P = 0.35), which could be due to its limited number of areas included (N = 3, Table I). For most species the δ 15 N values decreased from north (area 1) to south (area 4) on the West coast and increased slightly on the East coast (area 5 and 6). A clear example of this pattern was seen in shrimps (Fig. 3). However, a few species did diverge (small cod, capelin, polar cod and wolffish) from this general pattern. For instance, the δ 15 N values of capelin did not decline gradually from north to south. Rather, the δ 15 N value (mean ± SD) of capelin declined significantly (Kruskal-Wallis, χ 2 = 19.01, P < ) from a high value in area 1 and 2 (12.64 ± 0.12 ) to a lower mean value in area 3, 4, 5 and 6 (9.58 ± 0.77 ). Also, wolffish showed a different pattern in δ 15 N. The values of δ 15 N did not increase on the East coast, but rather continued to decline from high values (mean ± SD) in the most northern area on the West coast (16.11 ± 0.44 ) to the lowest value in the northern area on the East coast (11.25 ± 0.51 ). Comparing δ 15 N values on the West coast separately for each species, it was found that the δ 15 N values for most species did not differ significantly among two adjacent areas, but for most species there was a significant difference between areas which were not adjacent (Table IV). Similar to δ 15 N values there was a significant effect of area on δ 13 C values in most species (ANOVA, F 4-5, , P 0.04), but not in capelin (ANOVA, F 5,24 = 0.53, P = 0.75) and small redfish (Kruskal- 8

29 Wallis, χ 2 = 9.28, P = 0.10). For most species δ 13 C values were declining from area 3 to 6 whereas no clear trend was found for the northern area on the West coast. An example of this pattern of δ 13 C along the coast was seen in shrimps (Fig. 3). In order to reveal the difference between areas across all species, relative values of δ 15 N and δ 13 C were calculated. The regional differences were also reflected in the relative δ 15 N values calculated for each area which differed significantly (ANOVA, F 5,54 = 28.86, P < , Fig. 4). On the West coast the highest relative δ 15 N value were in the most northern study area (area 1) being 29% higher than the most southern study area (area 4). Hence, along the investigated 1300 km gradient on the Greenlandic West coast there was a positive relationship between latitude and relative δ 15 N values (linear regression, F 1,38 = 141.5, r 2 = 0.79, P < , Fig. 4). On the East coast the two relative δ 15 N values did not differ significantly from each other (Tukey s post hoc test, P = 0.55), neither did the relative values from the East coast (areas combined) differ from the most southern area on the West coast (Tukey s post hoc test, P > 0.97) (Fig. 4 and Table VI). The relative δ 13 C values also differed significantly between areas (ANOVA, F 5,54 = 5.92, P = ), but this was only caused by area 3 having a significant lower relative δ 13 C value than areas 1, 5 and 6 (Tukey s post hoc test, P < 0.02, Fig.4). The relative δ 13 C values of area 1, 2 and 3 increased with latitude, like the relative values of δ 15 N. However, the relative δ 13 C value of area 4 is approximately the same as the relative δ 13 C value of area 5 on the East coast. There is an almost linear relationship among area 1, 2 and 3 (linear regression, F 1,28 = 10.37, r 2 = 0.27, P = 0.003) and between area 4, 5 and 6 (linear regression, F 1,28 = 5.80, r 2 = 0.17, P = 0.02, Fig.4). Trophic level Greenland shark was the apex predator with a mean trophic level (mean ± SE) of 4.77 ± 0.22 followed by wolffish (3.74 ± 0.16) and Greenland halibut (3.60 ± 0.18, Table V). There was a significant difference in trophic level between species (ANOVA, F 12,341 = 84.75, P < ) and there was also an area effect (ANOVA, F 5,341 = 31.91, P < ). However, this effect was primarily caused by West and East coast differences, with 77% (37 of 48) of the significant species comparisons between areas being West and East coast related. The only species not being significantly different between the coasts were krill (ANOVA, F 5,34 = 1.46, P = 0.23). On the west coast only small cod (ANOVA, F 2,12 = 13.17, P > ), polar cod (ANOVA, F 3,16 = 15.67, P < ) and shrimp (ANOVA, F 3,16 = 13.09, P > ) showed a significant difference between areas. The estimated trophic level of polar cod and shrimp were both significantly higher in area 3 and 4 compared to area 1 and 2 (Tukey s post hoc test, P 0.03). The same pattern was not found for small cod. Here only area 3 had a higher value of trophic level, which was significantly different from both area 2 and 4 (Tukey s post hoc test, P 0.01). 9

30 Temporal study 118 muscle samples divided between nine species and ten sampling groups were analyzed for δ 13 C and δ 15 N during April, June, August and November 2010 in the Nuuk Fjord (Table III). Average isotopic values ranged from to in δ 13 C and from 8.40 to in δ 15 N (Fig. 5). Mean δ 15 N values differed significantly between species (ANCOVA, F 8,96 = 15.76, P < ) but there was no effect of time-of-catch (ANCOVA, F 1,96 = 0.16, P = 0.69). However, a significant interaction term (ANCOVA, F 8,96 = 2.97, P = 0.005), indicates that some species do vary in δ 15 N during the eight month long sampling period. Thus, δ 15 N values for small cod increased from in April to in November (ANOVA, F 1,16 = 13.23, P = 0.002). Similarly, large redfish (ANOVA, F 1,18 = 9.74, P = 0.006) and krill (ANOVA, F1,13 = 6.66, P > 0.03) increased from to and 9.53 to 10.16, respectively during the sampling period, but this trend was not true for all species. Atlantic halibut and large cod showed almost no variation while American plaice showed a decrease in the middle of the sampling period (Fig 6). δ 13 C values varied both between species (ANCOVA, F 8,104 = 46.30, P > 0.001) and as a result of time-ofcatch (ANCOVA, F 1,104 = 11.12, P = 0.001) with a tendency of increasing values. There was no significant interaction (ANOVA, F 8,96 = 0.44, P = 0.90) (Fig. 6). This general tendency is for example shown by small cod and krill where δ 13 C values (mean ± SD) of small cod increasing from the lowest value in June ( ± 0.32 ) to the highest value in November ( ± 0.34 ). Likewise, krill showed a slight increase from ± 0.54 to ± 0.17 (mean ± SD) during the sampling period. Inshore-offshore comparison Comparing species caught in- (Nuuk Fjord) and offshore (area 3) at approximately the same time (June and July 2010) there was a significant area effect (i.e. in- vs. offshore) with regard to both δ 13 C (ANOVA, F 1,51 = 9.36, P = 0.004) and δ 15 N (ANOVA, F 1,51 = 13.03, P = ) values. Also, there was a significant species effect in both δ 13 C (ANOVA, F 7,51 = 31.93, P < ) and δ 15 N (ANOVA, F 7,51 = 22.51, P < ). In general values were highest inshore, however significant interactions terms (δ 13 C: ANOVA, F 7,51 = 7.81, P < and δ 15 N: ANOVA, F 7,51 = 3.27, P = 0.006) show that the pattern is not similar for all species. In the case of δ 13 C values, this was caused by copepods being the only value higher offshore compared to inshore (Fig.7). Regarding δ 15 N values, they were for all species higher inshore compared to offshore, but not significantly in all species (Fig. 7). It is noteworthy that δ 15 N values (mean ± SE) for copepod (9.98 ± 0.18 ) and krill (9.96 ± 0.19 ) inshore were approximately the same, while there was a difference of 2.3 in δ 15 N values between copepod (7.26 ± 10

31 0.28 ) and krill (8.92 ± 0.46 ) offshore. Values of δ 13 C were likewise higher (i.e. less negative) for all species, except copepods, inshore compared to offshore (Fig. 7). Especially δ 13 C values (mean ± SE) of American plaice caught inshore ( ± 0.48 ) were higher than δ 13 C values offshore ( ± 0.13 ). Comparing trophic level for each species caught in- and offshore there was an area effect (Kruskal- Wallis, χ 2 = 11.08, P = ) as well as a species effect (Kruskal-Wallis, χ 2 = 43.90, P < ). The mean trophic level were for all species higher offshore compared to inshore, which properly is a result of higher δ 15 N baseline values (i.e. copepods) offshore. Hence, the compared food webs is fairly similarly in composition (Fig. 2 and 5). Even though the estimated mean trophic level were higher offshore compared to inshore only four out of the eight species sampled differed significantly (krill, ANOVA, F 1,8 = 8.00, P = 0.022; American plaice, ANOVA, F 1,6 = 34.05, P = 0.001; small cod, ANOVA, F 1,8 = 38.51, P = and large cod, ANOVA, F 1,8 = 6.59, P = 0.033, Fig 8). Discussion Spatial Study δ 15 N values differed significantly between six predetermined areas along the Greenland West and East coast being positively correlated with latitude. For all species (except Greenland shark) δ 15 N values increased between 2 and 4 (Fig. 2) from the most southern area to the northern study area on the West coast, which is equivalent to a full trophic level (Post 2002). On the East coast the overall pattern was the same (Fig. 3), but a few species have a reversed pattern of decreasing δ 15 N values with latitude (i.e. wolffish, polar cod, small cod and Greenland halibut, Fig. 2). These represent very different ecosystem components, and the reason for the discrepancy is unknown but could be related to migration patterns or actual feeding differences between areas. The same latitudinal gradient was not evident in δ 13 C values. The general pattern of δ 13 C values were not gradually increasing but instead divided in two with increasing values from area 1 to 3 on the West coast and decreasing values from area 4 to 6. A rather large difference of up to 1.5 appears between area 3 and 4 (Fig. 3). The latitudinal gradients can be explained by a multitude of factors. This includes a change in food consumption with changing latitude (Sweeting et al. 2005), but given the consistent shift across the entire food web in especially δ 15 N values the explanation must be of a more fundamental nature. Supporting this is the fact that the estimated trophic level for each species does not change consistently with latitude, but stays fairly constant across study areas (Table V). In addition to some 11

32 base line induced coastal differences in trophic level, only three species (shrimp, polar cod and small cod) differed significantly in trophic level between some of the West coast areas (Table V). So although it is not possible to totally disregard an effect of small feeding differences across latitudes, we do not believe they contribute significantly to the large latitudinal shifts in δ 15 N values. More likely, changes are a result of changing isotope values at the base of the food web with changing degree of latitude which then cascades through the food web. The baseline isotope values are potentially influenced by physical, chemical and biological properties of the currents (Pantoja 2002), terrestrial input (Sherwood & Rose 2005) and primary producer species composition and bloom progression (Tamelander et al. 2009). The combined effect of these direct and indirect effects on the primary produces will change the values on all higher trophic levels. The currents along the Greenlandic west coast are a mixture of the cold low saline East Greenlandic Current coming from the north and the temperate saline Irminger Current branching of from the Gulf Stream (Ribergaard 2011). The two currents meet at the southern Greenland East coast and then merge gradually, homogenizing the water masses moving north along the West coast. If the water masses differ in isotope signal in the biological available nutrients this will be reflected in POM and consequently in higher trophic levels. Hence, the parallel shift of the food web may well be related to shifts in the relative contributions of the two dominant water masses around Greenland. Two previous studies conducted in the two main currents dominating the waters around Greenland (East Greenland Current and the Irminger Current) supports the notion of differing water masses influencing δ 15 N and δ 13 C (Table VII). Sarà et al. (2009) measured stable isotopes in the Irminger Current on the Reykjanes Ridge south of Iceland, whereas Hobson et al. (1995) performed stable isotope analyses in the East Greenland Current (Table VII). Comparing species, or ecologically similar species, between areas demonstrates relatively large differences in δ 15 N values, with species caught in the Northeast Water Polynya (East Greenland Current) having similar or higher values than those caught in the Irminger Current. However, all mean δ 15 N values in the most northern area on the West coast (area 1) were all higher compared to mean δ 15 N values in the northern area on the East coast (area 6) opposite of what intuitively to expect given the simplified current pattern described here, suggesting a complex mixing pattern or some other contributing factor. However, given the differences in water masses, it surely in some way contributes to the pattern described here. This gradual mixing of water masses could also possibly be related to the migrational pattern of the sampled species. Wolffish is considered a stationary species (Riget & Messtorff 1988) and displays a very large latitudinal gradient (3.2 ) whereas the more migratory behavior seen in for instance Greenland shark (Skomal & Benz 2004) could serve to dilute the effect of latitudinal related 12

33 differences in δ 15 N values give little north-south difference (0.8 ). However, too little is known about the migration of the sampled species to make any conclusions. The parallel shift in δ 15 N values with latitude across groups could also be related to the length of the productive open-water period (Blicher et al. 2007). In high-arctic areas this period is identical with the ice-free period, while diminishing day lengths have a pronounced effect further south were ice-free periods are longer. The productive open-water period is here defined according to Blicher et al. (2007) as the annual number of days with open water and a minimum day length of six hours. There is a significant relationship between the productive open-water period and the relative δ 15 N values in the four West coast areas and a similar tendency in the two East coast areas (Fig. 9). Hence, the shorter the productive open-water period is this higher is the relative δ 15 N values. A possible explanation could be that a longer period with ice cover results in an increasing primary production intensity, whereby nutrients are exhausted at a faster rate which ultimately could lead to more regenerated production and subsequently increasing δ 15 N values (Wainright & Fry 1994). Another explanation could be that a longer ice cover period stabilizes the water column, either due to the ice cover or the outlet of freshwater which would cause a stronger stratification thereby decreasing possible upwelling also leading to more regenerated primary production (Wu et al. 1997). Exploring the results from the POM analyses should help in interpreting the results. We have nevertheless chosen to exclude them from most analyses as the variation in mean POM δ 15 N values between areas was large, ranging from 3.3 to 10.8, and often exceeded other groups being biologically impossible (Fig. 2). However, given the low intra-area variation in isotope values and general high analyses precision, measurement error does not appear to be a factor. Rather, some POM samples were taken close to or over the shelf edge (Fig. 1) and δ 15 N values in these samples were generally higher (mean ) compared to the values of POM sampled on the shelf ( ). CTD profiles taken simultaneously with POM samples clearly demonstrate that different water masses are present (Fig. 10), but whether these cause the differences in δ 15 N values is unknown, as other factors such as changes in POM composition or seasonal progression could give similar differences (Tamelander et al. 2009). Thus, as there clearly is some spatial effect on the results on both a large and small spatial scale (see also the inshore-offshore section) so whatever the ultimate cause of this latitudinal related pattern in isotopic values (irrespective of feeding behavior) is, it must be taken into account when addressing hypotheses by means of stable isotope analysis covering a noticeable geographical range (Møller, 2006, Riget et al. 2007), especially when considering a few number of species that does not reveal any geographical effect. Lastly, the vast latitudinal gradient in the present study is naturally associated with a similar temperature gradient. Barnes et al. (2007) showed in a laboratory study that rising temperature can have a negative effect on δ 15 N values. In Greenland benthic fish will generally experience colder 13

34 temperatures moving north (Sünksen et al. 2009) while the reverse may be the case for the strictly pelagic species such as capelin, at least on the West coast (Hedeholm 2010) but as the potential effect of temperature is small (i.e per 1 C for δ 15 N assuming linearity, Barnes et al. 2007) compared to the differences in isotope values and temperature observed here (Fig. 2 and 10) it offers little explanation in the present study. Temporal Study During the eight month sampling period from April to November there was an overall effect of Day-ofyear on δ 13 C. The most enriched and depleted values of δ 13 C were obtained in June and August and April and November, respectively (Fig. 5 and 6) with only small cod showing a different pattern. Greenland halibut and large redfish experienced the largest peak in δ 13 C with a difference of about 1 in the summer period. The increasing δ 13 C values in the summer period are consistent with other studies investigating primary production, invertebrates and fish (Sarà et al. 2002; Vizzini & Mazzola 2003). This increase in δ 13 C during the summer period could indicate a shift in feeding preference towards a more benthic diet. However, generally evidence of isotopic temporal variability is confined to relatively short-lived primary producers and consumers (e.g. Nordström et al. 2009; Vizzina & Mazzola 2003) and the finding of temporal changes of this magnitude are unexpected, as muscle turnover is relatively slow and feeding induced changes exceeds that normally associated with small feeding shifts (Bootsma et al. 1996). Hence, a substantial influence from differences in the isotopic baseline also appears to be of importance in the inshore area which could be due to differences in water masses, currents and terrestrial input example due to increasing glacial ice melt. Although there was no significant time effect on δ 15 N irrespective of species, the general pattern of mean δ 15 N showed an increasing trend at the end of the sampling period (Fig. 5 and 6) perhaps indicating a beginning of a winter peak similar to δ 13 C. Given the time laps in the response of muscle tissue (weeks to months, Hesslein et al. 1993) this could be caused by an intensified summer feeding period following a winter period with lower feeding intensity. In order to demonstrate this potentially annual isotopic pattern a longer and maybe more intense sampling period must be conducted. Like in the spatial study a large variation occurred in δ 15 N values of POM. The largest difference occurred between the first two sampling periods with a variation of 4. Again this large different in isotopic signatures of primary production is most likely caused by changes in physical and chemical properties of the water masses during the sampling period, hence the water masses and conditions of the currents in the Nuuk Fjord change between summer and winter and increasing freshwater runoff during summer (Mortensen et al. 2011). It is however also possible that the large difference is due to variation in species composition of the primary production in the different sampling periods. 14

35 Inshore-offshore comparison When comparing stable isotope data measured offshore (area 3) and inshore (Nuuk Fjord) almost all comparable species (except copepods) had higher δ 13 C and δ 15 N values inshore than offshore. This is in agreement with the observation that inshore species have a more enriched δ 13 C values compared to the more depleted offshore species, as benthic primary production generally exhibit less δ 13 C fractionation during carbon fixation than do pelagic phytoplankton (Vander Zanden & Rasmussen 1999, Fig. 7). The pattern in δ 15 N values agrees with those of Sherwood & Rose (2005) which on a similar geographic scale also found lower δ 15 N values offshore, but is in contrast with result represented by France 1995b. One explanation of the higher δ 15 N values inshore could be a high degree of vertical mixing caused by physical processes involving ocean currents, wind and tides leading to high levels of nutrient availability and productivity in the inshore waters (Sherwood & Rose 2005). Because of the relatively large difference in the base of the food web (i.e. copepods) between inshore and offshore the estimated trophic level offshore is shifted upwards for all species, but maintains the same relative pattern (Fig. 8). Conclusion Comparing stable isotope signatures in the arctic marine food web demonstrated large differences on both a large and small geographic scale. The spatial study showed a significantly latitudinal effect, revealing increasing δ 15 N values with latitude. Also a temporal effect was demonstrated, even though only significant in δ 13 C. In order to accurately demonstrate the importance of a temporal change in δ 15 N values a more balanced and intensive sampling may be required. These differences in isotopic signature both spatial and temporal are probably caused by abiotic factors, differences in physical and biological properties of the water masses as well as terrestrial inputs. These results will have a large influence on comparative studies if such differences are not considered when studying isotopic signatures. Acknowledgements The authors acknowledge the staff at the Greenland Institute of Natural Resources for permission to use laboratory facilities and sampling assistance. Also, we are thankful to the staff at RV Pâmiut and Sanne Kjellerup and Rasmus Swalethorp (DTU Aqua, DK) for sampling assistance. The study received 15

36 financial support from the Greenland Institute of Natural Resources and the Danish Agency for Science, Technology and Innovation and is a part of the Greenland Climate Research Centre. 16

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43 Table I. Sampling from six areas along the Greenland coast (Fig. 1). Mean total length, standard deviation (SD) and number (N) sampled are illustrated for each species in each area. Species Size interval (cm) Area 1 Area 2 Area 3 Area 4 Area 5 Area N N N N West coast West coast West coast East coast 16 June 11 July 11 June-21 July July 5-8 August N West coast 1-5 July Total length (mean cm ± SD (N)) N East coast August Anarhichas sp. (Wolffish) ± 9 (4) 42 ± 4 (5) 33 ± 1 (5) 52 ± 5 (5) 50 ± 0 (1) 48 ± 8 (4) Boreogadus saida (Polar cod) ± 3 (5) 11 ± 1 (5) 12 ± 0 (5) 15 ± 2 (5) 11 ± 0 (1) 12 ± 0 (5) Gadus morhua (Atlantic cod), small ± 0 (5) 31 ± 1 (5) 28 ± 0 (5) 28 ± 3 (5) 29 ± 2 (5) Gadus morhua (Atlantic cod), large ± 1 (5) 48 ± 3 (6) 46 ± 0 (5) 50 ± 2 (5) 49 ± 0 (5) Hippoglossoides platessoides (American plaice) ± 1 (5) 31 ± 2 (5) 25 ± 1 (5) 23 ± 1 (5) 25 ± 2 (5) 30 ± 0 (5) Mallotus villosus (Capelin) ± 0 (5) 13 ± 0 (5) 13 ± 1 (5) 12 ± 1 (5) 14 ± 1 (5) 12 ± 0 (5) Reinhardtius hippoglossoides (Greenland halibut) ± 2 (5) 40 ± 0 (5) 45 ± 3 (5) 45 ± 7 (4) 43 ± 3 (2) 41 ± 1 (5) Sebastes sp. (Redfish), small ± 0 (5) 17 ± 8 (5) 16 ± 8 (5) 16 ± 0 (5) 23 ± 2 (4) 22 ± 1 (5) Sebastes sp. (Redfish), large ± 1 (3) 38 ± 2 (5) 38 ± 2 (5) 38 ± 3 (5) 39 ± 2 (5) 37 ± 1 (5) Somniosus microcephalus (Greenland shark) Any 270 ± 0 (1) 445 ± 0 (1) 449 ± 17 (6) 23

44 Species Table II: Sampling from eleven stations along the Greenland coast. Mean length, standard deviation (SD) and number (N) sampled are illustrated for each species at each location. Only N is illustrated for Calanus sp. because the individuals were not measured. At all stations the krill species was Thysanoessa raschii except at station 10 were it was Meganyctiphanes norvegica. Station 1 71 N West coast 4 July Station 2 69 N West coast 15 June-11 July Station 3 66 N West coast Station 4 64 N West coast July Station 5 62 N West coast July Station 6 61 N West coast July Length (mean cm ± SD (N)) Station 7 60 N West coast 28 July Station 8 60 N East coast 5-6 August Station 9 62 N East coast 7 August Station N East coast 11 August Station N East coast August Calanus sp. N = 3 N =3 N =3 N =3 N =3 N =3 N =3 N =3 N =3 N =3 N =3 Krill 2.0 ± 0.2 (5) 1.5 ± 0.1 (5) 1.8 ± 0.1 (5) 1.9 ± 0.1 (5) 1.9 ± 0.3 (5) 1.8 ± 0.1 (5) 1.9 ± 0.1 (5) 1.9 ± 0.1 (5) 1.8 ± 0.1 (5) 1.8 ± 0.1 (5) Filtered POM N = 2 N = 2 N = 2 N = 2 N = 2 N = 2 N = 2 N = 2 N = 2 N = 2 N = 2 Pandalus borealis 2.6 ± 0.1 (5) 2.3 ± 0.1 (5) 2.6 ± 0.1 (5) 2.5 ± 0.1 (5) 2.1 ± 0.2 (5) 2.2 ± 0.2 (5) 2.4 ± 0.4 (4) 2.7 ± 0.2 (5) 24

45 Table III. Samples caught in the Nuuk Fjord. All fish species were sampled at the same location except R. hippoglossoides, while krill and copepods were sampled in a nearby bay (see text and Fig. 1). Species Position ( N; W) Size interval (cm) Sampling depth (m) April June August November Length (mean cm ± SD (N)) Gadus morhua (Atlantic cod), small 64.25; ± 1 (3) 35 ± 3 (5) 34 ± 4 (5) 42 ± 2 (5) Gadus morhua (Atlantic cod), large 64.25; ± 2 (5) 55 ± 3 (4) 55 ± 3 (5) 51 ± 2 (5) Hippoglossus hippoglossus (Atlantic halibut) 64.25; ± 0 (1) 48 ± 10 (2) 49 ± 0 (2) 45 ± 0 (1) Hippoglossoides platessoides (American plaice) 64.25; ± 1 (5) 36 ± 2 (2) 34 ± 5 (5) 37 ± 2 (5) Mallotus villosus (Capelin) 64.25; ± 0 (4) 12 ± 1 (2) Reinhardtius hippoglossoides (Greenland halibut) 64.34; Unknown (2) Unknown (2) Unknown (2) Sebastes sp. (Redfish), large 64.25; ± 1 (5) 41 ± 3 (5) 39 ± 4 (5) 42 ± 2 (5) Calanus finmarchicus 64.45;50.25 N = 3 N = 3 Thysanoessa raschii 64.45; ± 0.1 (5) 1.4 ± 0.1 (5) 1.2 ± 0.1 (5) Filtered POM 64.21; N = 1 N = 1 N = 1 N =1 25

46 Table IV. Species specific comparisons of δ 15 N values in the different areas (see Fig 1). indicate a significant difference (P < 0.05), ns indicates no significant difference between the species in the compared study areas and indicate species absence in one/both areas. Shaded section is comparisons between west and east coast. Species Comparison of different areas 1 vs. 2 1 vs. 3 1 vs. 4 2 vs. 3 2 vs. 4 3 vs. 4 1 vs. 5 1 vs. 6 2 vs. 5 2 vs. 6 3 vs. 5 3 vs. 6 4 vs. 5 4 vs. 6 5 vs. 6 Anarhichas sp. (Wolffish) ns ns ns ns ns ns Boreogadus saida (Polar cod) ns ns ns ns ns ns ns ns ns ns Gadus morhua (Atlantic cod), small ns - - ns Gadus morhua (Atlantic cod), large ns - - ns ns ns Hippoglossoides platessoides (American plaice) ns * ns ns ns ns ns Mallotus villosus (Capelin) ns ns ns ns ns ns ns Reinhardtius hippoglossoides (Greenland halibut) ns ns ns ns ns ns ns ns ns ns ns ns ns ns Sebastes sp. (Redfish), small ns ns ns ns ns Sebastes sp. (Redfish), large ns ns ns ns ns ns ns ns Calanus sp. (Copepod) ns ns ns ns ns ns ns ns ns ns ns ns ns ns Thysanoessa raschii (Krill) ns ns ns ns ns ns Pandalus borealis (Shrimp) ns ns ns ns ns ns ns ns 26

47 Table V. Trophic level estimated according to Nilsen et al. (2008) and Søreide et al. (2006). Calanus sp. were used as baseline in trophic level estimation. The mean trophic level is an unweighted mean across areas. Species Trophic Level Area 1 Area 2 Area 3 Area 4 Area 5 Area 6 Mean Calanus sp. (Copepods) Thysanoessa raschii (Krill) Pandalus borealis (Shrimp) Mallotus villosus (Capelin) Sebastes sp. (Redfish), small Boreogadus saida (Polar cod) Gadus morhua (Atlantic cod), small Sebastes sp. (Redfish), large Hippoglossoides platessoides (American plaice) Gadus morhua (Atlantic cod), large Reinhardtius hippoglossoides (Greenland halibut) Anarhichas sp. (Wolf fish) Somniosus microcephalus (Greenland shark)

48 Table VI. Comparison of the relative δ 13 C and δ 15 N values in study areas along the Greenlandic West and East coast. indicate a significant difference (P < 0.05) while ns indicate no difference between the two areas in question (P > 0.05). Area 2 Area 3 Area 4 Area 5 Area 6 δ 13 C δ 15 N δ 13 C δ 15 N δ 13 C δ 15 N δ 13 C δ 15 N δ 13 C δ 15 N Area 1 ns ns Area 2 ns ns ns ns Area 3 ns ns Area 4 ns ns ns ns Area 5 ns ns 28

49 Table VII. A comparison of different ecosystems in different water masses, based on the current study and literature review. Data from Iceland is by Sarà et al. (2009), Northeast Water Polynya is by Hobson et al. (1995) and Greenland Area 1 and 6 is from the current study (Fig. 1). a indicate standard error instead of standard deviation, - indicate data not available. All values from Hobson et al. (1995) are lipid extracted. Species Iceland, Irminger Current Northeast Water Polynya, East Greenland Current Greenland, Area 1, West Greenland Current Stable isotope values ( ± SD) Greenland, Area 6, East Greenland Current δ 15 N δ 13 C δ 15 N δ 13 C δ 15 N δ 13 C δ 15 N δ 13 C Anarhichas sp./icelus bicornis 13.1 ± ± ± ± ± ± ± ± 0.5 Boreogadus saida 13.7 ± ± ± ± ± ± 0.2 Gadus morhua (Atlantic cod), small 12.8 ± ± ± ± 0.1 Hippoglossus platessoides/lycodes rossi 12.0 ± ± ± ± ± ± ± ± 0.2 Mallotus villosus/micromesistus poutassou 10.8 ± ± ± ± ± ± ± ± 0.5 Sebastes sp. (redfish), small 11.6 ± ± ± ± ± ± ± ± 0.3 Calanus sp. 3.5 ± 0.1 a ± - a 8.2 ± ± ± ± ± ±0.2 Krill/Themisto sp ± ± ± ± ± ± ± ± 0.3 POM 5.2 ± ± ± ± ± ± ± ±

50 Figure 1. The southern part of Greenland. The offshore squares represent the study areas in which all fish species were sampled. The gray circles mark the stations where shrimps, krill, copepod, POM and CTD-measurements (CTD-measurements were only sampled on the West coast) were sampled. The line indicates the 500 meter depth contour and thereby the shelf edge. The small map shows the Nuuk fjord system with Nuuk city, fish (except Greenland halibut) and POM station (open square), sampling station of copepod and krill (black circle) and the sampling site of Greenland halibut (black triangle) marked. 30

51 Figure 2. δ 13 C and δ 15 N ( ) values (mean ± SE) along the Greenlandic coast for the analyzed species: Filtered POM (POM), copepod (COP), krill (KRI), shrimp (SHR), capelin (CAP), polar cod (POL), small redfish (RES), large redfish (REL), American plaice (APL), small cod (ATS), large cod (ATL), Greenland halibut (GHL), wolffish (WOF) and Greenland shark (GSK). 31

52 Figure 3. The general pattern of δ 15 N ( ) and δ 13 C values (mean ± SE) along the Greenlandic west (area no. 1, 2, 3 and 4) and east coast (area no. 5 and 6), represented by shrimp. Note the difference in scale on the y-axis. 32

53 Figure 4. The calculated relative values (± SE) of δ 13 C and δ 15 N against latitude ( N). Note the difference in scale on the y-axis. 33

54 Figure 5. δ 13 C and δ 15 N ( ) values (mean ± SE) in the Nuuk Fjord for the analyzed species: Filtered POM (POM), copepod (COP), krill (KRI), capelin (CAP), small cod (ATS), large cod (ATL), redfish (large) (REL), American plaice (APL), Greenland halibut (GHL), Atlantic halibut (AHL)). 34

55 Figure 6. Mean index values of δ 13 C and δ 15 N for each sampling date in the Nuuk Fjord during the sampling period in Mean index values are calculated based on the mean values of each species in each sampling period divided by the mean of the given species at all sampling dates, and then a mean value of all species at each sampling dates were calculated. The mean index values of δ 13 C and δ 15 N is in this way centered around one. All δ 15 N data points are displaced two days from the original sampling day. Error bars represent standard error. 35

56 Figure 7. Inshore and offshore comparison of δ 15 N and δ 13 C ( ) values (mean ± SE). indicate significant difference (P 0.05). 36

57 Figure 8. Mean trophic level (± SE) comparison among species inshore and offshore. indicate a significant difference (P < 0.05). 37