MARINE ECOLOGY PROGRESS SERIESMar Ecol Prog Ser
Vol. 510: 275288, 2014doi: 10.3354/meps10959
Published September 9
INTRODUCTION
Jellyfish (here considered as Phylum Cnidaria,Class Scyphozoa) are ubiquitous components ofpelagic marine ecosystems, and marked changes intheir population size can be seen as a natural featureof healthy pelagic ecosystems (Graham et al. 2001).The combined effects of multiple anthropogenicstressors and/or climate change (Molinero et al. 2008,Richardson et al. 2009, Purcell 2012) may disturb thenatural oscillations of jellyfish blooming (Condon etal. 2013), leading to a significant increase in jellyfishpopulations in some coastal areas and large marineecosystems (Kogovek et al. 2010, Brotz et al. 2012).
In such pelagic ecosystems the role of jellyfishbecomes more important, shifting energy from fish tojellyfish (Uye 2011). As voracious predators theycompete with fish for food (Purcell & Arai 2001), andthrough predation on fish eggs (Gordoa et al. 2013)and fish larvae (Purcell & Arai 2001, Sabats et al.2010) jellyfish could even impede the recovery of fishstocks in heavily exploited marine ecosystems(Lynam et al. 2006). Furthermore, jellyfish popula-tions can bloom in a relatively short period of timeand can attain enormous biomasses. After the declineof such populations, organic material released intothe water column from the decomposing jellyfish tis-sue promote a microbially dominated food web (Con-
Inter-Research 2014 www.int-res.com*Corresponding author: [email protected]
Jellyfish biochemical composition: importance ofstandardised sample processing
T. Kogovek1,2,*, T. Tinta1, K. Klun1, A. Malej1
1National Institute of Biology, Marine Biology Station, Forna
Mar Ecol Prog Ser 510: 275288, 2014
don et al. 2011, Tinta et al. 2012), thus altering eco-system diversity and function. Large accumulationsof jellyfish carcasses at the sea bed are an importantsource of labile organic material for a wide spectrumof benthic macroorganisms (for a review see Lebratoet al. 2012). Nevertheless, the unconsumed jellyfishmaterial is decomposed by microbes, and oxygenconsumption together with remineralization productsthat accompany the decomposition may furtherimpact the benthic biota (Billett et al. 2006, West etal. 2009, Lebrato et al. 2012).
In order to understand the role of jellyfish in marinefood webs and in biogeochemical cycling, especiallyin regions with increasing jellyfish bloom events, theenergy stored and the trophic level occupied by jelly-fish need to be quantified. The pattern of energyflows and the efficiency of mass transfer through thefood web are reflected in the ratios of biomass be -tween different trophic levels, usually expressed asdry mass or carbon (C) and nitrogen (N) content. Dueto the patchy distribution of jellyfish and a lack ofsuitable sampling techniques (Purcell 2009), it is dif-ficult to estimate their biomass, therefore jellyfishwere often neglected in previous studies of trophicinteractions and were rarely included in ecosystemmodels (Pauly et al. 2009). The recent developmentof new techniques to assess jellyfish abundance/bio-mass such as hydroacoustics (Alvarez Colombo et al.2009), aerial observations (Houghton et al. 2006) androutine fishery surveys (assessing jellyfish abundanceas by-catch) (Brodeur et al. 2008) have enabled de -velopment of time series of the seasonal and spatialdistribution of jellyfish biomass and abundance. Nev-ertheless, very often in ecological models the role ofgelatinous organisms in the food web is simplified,considered as a single functional jellyfish group,assumed to have the same diet and to occupy thesame trophic level (Pauly et al. 2009). In the nearfuture, this problem might be overcome by an in -creasing number of studies that apply stable isotopesfor analysing the role of gelatinous plankton in plank -tonic food chains (see Pitt et al. 2009a for a review,Cardona et al. 2012, Syvranta et al. 2012, DAmbraet al. 2013a).
Despite the fact that numerous data on jellyfishbody elemental composition exist (compiled in Lucaset al. 2011), reliable and uniform protocols for jelly-fish sample processing are not established. Until veryrecently the methodology for sample processing andbiomass estimation for jellyfish was the same as thatfor crustacean zooplankton, despite the difference inbiochemical composition between the 2 groups. Thejellyfish body contains high water and low organic
content compared to crustacean plankton. Moreover,dried jellyfish samples contain a large amount of saltsin relation to their organic content. Dry mass (DM)and ash free dry mass (AFDM) of jellyfish vary inrelation to ambient salinity (Hirst & Lucas 1998).Additionally, the residual water bound in dry jellyfishmaterial gives an overestimation of organic materialand ash content (Larson 1986). Thus, DM and AFDMoffer a poor representation of jellyfish biomass. Mostcommonly, dry mass is determined by drying thematerial at 60C until constant mass (Harris et al.2000). The effect of the prolonged exposure of pro-tein-rich jellyfish tissue (Quensen et al. 1981, Malejet al. 1993, Lucas 1994, Pitt et al. 2009b) to elevatedtemperatures is unknown. Omori (1978) showed aslight loss of organic matter in 4 zooplankton species(a chaetognath Sagitta nagae and 3 copepods speciesCalanus sinicus, Pleuromamma xiphias and Acartiatonsa) during oven-drying at 60C until constantmass, interpreting it as volatilization of some lipidsand amines, as well as the denaturing of proteins.Furthermore, preservation methods and processingmight modify the stable isotope composition of thesamples. Several studies demonstrated that freezing,the most commonly used method for sample preser-vation, does not affect stable nitrogen and carbonisotopes values in fish, octopus, anemone, bivalves,kelp and algae (Kaehler & Pakhomov 2001, Carabelet al. 2009). However, the opposite was suggested byFleming et al. (2011) in the case of gelatinous plank-ton: while freezing did not affect 13C, the prolongedstorage of frozen Aurelia aurita medusae resulted inthe enrichment in 15N. Preservation in ethanol affectedthe 15N in some species of invertebrates (Carabel etal. 2009, Fleming et al. 2011), while there was no sig-nificant change in nitrogen isotope ratio in fish, octo-pus and kelp (Kaehler & Pakhomov 2001). In additionto preservation techniques, sample processing mightalso alter sample stable isotope ratios. For example,dehydration of samples leads to enrichment in 15N inoven-dried compared to freeze-dried samples (Cara-bel et al. 2006). Furthermore, lipids are known to bedepleted in 13C (DeNiro & Epstein 1978) and chemi-cal extraction of lipids prior to analysis might berequired to account for variable lipid content in tissues (e.g. Syvranta & Rautio 2010, DAmbra etal. 2013b). Together, these observations indicatethat the effect of sample processing on biochemicalcomposition is taxon specific.
The main objective of our study was to establish areliable and uniform protocol for the determinationof jellyfish biomass (dry mass, ash free dry mass) andbiochemical composition. In order to achieve this,
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several methods commonly used in zooplankton eco -logy as well as to determine jellyfish biomass, ele-mental, stable isotope and amino acid pool composi-tion were compared and assessed. We introduce anew desalination method into jellyfish sample pro-cessing. Finally, we propose a pre-analytical process-ing protocol for jellyfish according to selected analysis.
MATERIALS AND METHODS
In order to establish a suitable protocol for the bio-chemical analysis of gelatinous organisms, severalindividual sample processing steps were examined.Apart from conventional methods used for zooplank-ton analysis, some alternatives were proposed. Fig. 1shows a schematic representation of each individualstep.
Jellyfish homogenate (Step 1). Medusae were col-lected seasonally in coastal waters from a boat or apier. The organisms were deposited in a plastic con-tainer filled with ambient water and stored at ashaded place. The jellyfish were transported to thelaboratory within 1 h after collection. Bell diameter(to 1 mm accuracy) and wet mass (to 1 g accuracy)were determined before storing specimens at 30C(each medusa separately in a plastic zip-lock bag).Chrysaora hysoscella, Pelagia noctiluca and Rhizo -stoma pulmo medusae (3 to 5 medusae per species)with an average wet mass ( SD) of 464 g (487), 534 g(186) and 2881 g (1407), respectively, were homo -genized using an Ultra-Turrax TP 18/10 mechanical
homogenizer (Janke & Kunkel). Each homogenatewas divided into 2 sub-samples of the same wet massand dried (see Step 2). The organic content in Aureliaaurita was relatively low compared to other speciesinvestigated in this study; therefore, the homogenatewas not divided into sub-samples. Instead, 10 me -dusae of similar wet mass (43 21 g) were unfrozenbefore they were homogenized individually and thewhole volume of each sample was processed and usedfor further analysis. In order to compare medusaebiochemical composition on a higher taxonomic level,individuals of Hydromedusae Aequorea forskaleawere collected (41 23 g, n = 34) in addition to 4scyphozoan jellyfish.
Drying (Step 2). One set of homogenate sub-sam-ples (see Step 1) was poured into pre-weighed alu-minium containers and oven dried (OD) at 60C untilconstant mass (Step 2a). The time required to reachthe constant weight was 4 to 7 d. The OD sampleswere cooled to room temperature in a desiccator andweighed twice per day. The other set of sub-sampleswas stored at 30C until freeze-drying (FD samples,Step 2b). The samples were dry after 7 d. Dry masswas determined (to 0.01 g accuracy) for each set ofsub-samples (from Steps 2a and 2b), cal
