Short-term post-mortality predation and scavenging and longer-term recovery after anoxia in the Northern Adriatic Sea

Bereits im Jahr 1990 lebte fast ein viertel (23 %) der Weltbevolkerung (ca. 1,2 Milliarden Menschen) in Kustengebieten (Nicholls & Small, 2002) und die Prognose ist, dass bis 2015 weltweit etwa 33 sogenannte „Megastadte“ mit jeweils mehr als 8 Millionen Einwohnern/Stadt in Kustennahe entstehen werden (UN/DESA, 2001). Die zunehmende Besiedlung dieser Gebiete durch den Menschen hat grosen Einfluss auf die Verschmutzung der Kustengewasser und die Zerstorung wichtiger Seichtwassserlebensraume. Besonders das Mittelmeer mit seiner hohen Biodiversitat gilt durch menschliche Aktivitaten als stark gefahrdet (Danovaro & Pusceddu, 2007). 
Die Nordadria weist mehrere Eigenschaften auf, die ein sensibles Okosystem ausmachen (Stachowitsch, 1986, 1991; Ott, 1992): geringe Wassertiefe (im Durchschnitt 30 m), Feinsediment-Boden, hoher Suswassereintrag (v.a. Po-Fluss, Italien), lange Residenzzeit des Wassers, hohe Primarproduktion sowie eine stabile Pyknokline wahrend der Sommermonate. 
Ein Grosteil des Meeresbodens der Nordadria ist von epibenthischen Lebensgemeinschaften besiedelt, deren Organismen in Form von sogenannten „multi-species clumps“ aggregiert sind. Fedra et al. (1976) hat diese Lebensgemeinschaft die Ophiothrix-Reniera-Microcosmus (O-R-M) community genannt, da die Gesamtbiomasse vom Schlangenstern Ophiothrix quinquemaculata, dem Schwamm Reniera spp. und der Seescheide Microcosmus spp. dominiert wird. Durch ihre enorme Filterleistung konnen diese Gemeinschaften grose Mengen an suspendiertem organischem Material in der Wassersaule abbauen (Ott and Fedra, 1977) und dienen daher als „naturliche Eutrophierungskontrolle“ (Officer et al., 1982). Eine Kombination von ungunstigen hydrologischen und meteorologischen Gegebenheiten (z.B. langanhaltende, stabile Stratifikation der Wassersaule durch ruhige Wetterbedingungen) kann im Spatsommer/Fruhherbst am Meeresboden, Hypoxie (Sauerstoffarmut) oder Anoxie (kein Sauerstoff), hervorrufen (Malej & Malacic, 1995). Solche Storungen konnen die Gemeinschaftsstruktur massiv verandern (z.B. reduzierte Artendiversitat bis hin zu Massensterben, (Stachowitsch, 1984, 1991) und die ursprungliche okosystemare Funktion schwachen (Wu, 2002; Helly & Levin, 2004; Diaz and Rosenberg, 2008; Riedel et al., 2008; Breitenburg et al., 2009; Levin et al., 2009; Middelburg & Levin, 2009; Zhang et al., 2010). 
Durch das Zusammenspiel von Wasserstratifizierung und dem einbringen enormer Mengen an Nahrstoffen (Eutrophierung) durch grose Flusse in Kustenregionen, werden die Auswirkungen von Phytoplanktonbluten verstarkt, und konnen in weiterer Folge zu Hypoxien/Anoxien am Meeresboden fuhren (Nixon, 1995; Rabalais et al. 2010; Zhang et al. 2010). Diaz and Rosenberg (2008) beschrieben mehr als 400 durch Uberdungung entstandene „Todeszonen“, die eine Flache von insgesamt mehr als 245.000 km² umfassen. Zudem fuhrt kommerzieller Fischfang mit Bodenschleppnetzen zu negativen Veranderung seichter Meeresokosysteme (Jennings & Kaiser, 1998; Thrush & Dayton 2002; Thrush & Dayton 2010). Auch in der Nordadria sind negative Auswirkungen von geringen Sauerstoff-konzentrationen im Meer auf die O-R-M-Gemeinschaft dokumentiert (Kollmann & Stachowitsch, 2001, Fuchs and Stachowitsch, 1995). Oft wird die erste Phase der Wiederbesiedelung nach einer Storung von juvenilen Stadien, mobilen Arten und schnellen Kolonisten dominiert (Pearson & Rosenberg 1978; Dayton et al., 1995). Zwei Hauptmoglichkeiten der Wiederbesiedlung konnen unterschieden werden: durch Ansiedlung freischwimmender Larven (grosflachig betroffenen Gebiete), sowie Migration mobiler Organismen (kleinflachige Gebiete) (Gunther, 1992; Pearson & Rosenberg, 1978; Whitlatch et al., 1998). Damit auf Weichboden, wie sie in der Nordadria typisch sind, jedoch neue „multi-species clumps“ entstehen konnen, werden Hartstrukturen (z.B. Muschel- oder Seeigelschalen) als Aufwuchssubstrat benotigt. Durch Sedimentation und Bodenschleppnetze konnen Schalenstucke jedoch mit Sediment uberlagert oder gewendet werden, und damit neu entstehende „multi-species clumps“ wieder zerstort (Stachowitsch and Fuchs, 1995). 
 
Mit einem an der Universitat Wien entwickelten Unterwassergerat (EAGU, Experimental Anoxia Generating Unit) welches mit einer Zeitrafferkamera sowie Messsensoren fur Sauerstoff, Schwefelwasserstoff, Temperatur und pH ausgestattet ist, wurden die Reaktionen (z.B. Verhalten, Interaktion, Mortalitat) benthischer Organismen hinsichtlich Hypoxie und Anoxie dokumentiert (Stachowitsch et al., 2007, Riedel et al., 2008a, Riedel et al., 2008b, Haselmair et al. 2010, Pretterebner et al. 2011). In dieser Studie wurde EAGU eingesetzt um die unmittelbaren Entwicklungen nach einer kunstlich induzierten Sauerstoffarmut zu untersuchen. Auf zwei jeweils 0.25 m² grosen Flachen wurde die Reihenfolge und die Arten der eintreffenden Rauber und Aasfresser beobachtet. Weiters wurde die langfristigere Wiederbesiedlung (bis zu 2 Jahren) benthischer Organismen auf denselben Flachen mittels einer Fotoserie dokumentiert. 
Die Auswertung der Bilder zeigt eine bestimmte Reihenfolge der eintreffenden Arten. Die Fische (Diplodus vulgaris, Gobius niger, Serranus hepatus, Pagellus erythrinus) kamen als erstes an, gefolgt von den ersten Einsiedlerkrebsen (Paguristes eremita) und den Schnecken (Hexaplex trunculus). Diese Reihenfolge wird mit den Fortbewegungsgeschwindigkeiten der jeweiligen Arten sowie deren Dichte erklart. Die Flachendeckung innerhalb der Untersuchungsquadrate durch sessile Organismen wurde vor dem Experiment berechnet (Experiment August: 1939 cm² und September: 631 cm²). Die toten Organismen wurden innerhalb von 7 Tagen (August-Experiment) und 13 Tagen (September-Experiment) nach der kunstlichen Anoxie grosteils konsumiert. Solange die toten Organismen vorhanden waren, war die Dichte der Aasfresser hoch. Bei Paguristes eremita konnte eine eindeutige Tag/Nacht-Rhythmus festgestellt werden. Dabei wurde beobachtet, dass wahrend der Nachtstunden die Individuenzahl stark abnahm. 
Auch zwei Jahre nach der kunstlich hervorgerufenen Anoxie konnte keine makroskopisch sichtbare Wiederbesiedlung der Areale festgestellt werden. 
Damit hebt diese Studie die hohe Empfindlichkeit von Lebensgemeinschaften auf Weichboden hervor und unterstreicht die dringliche Notwendigkeit einer Reduzierung anthropogener Storungen durch Eutrophierung und kommerziellen Fischfang mit Bodenschleppnetzen.

on biodiversity (Danovaro and Pusceddu, 2007;Coll et al., 2010).One major disturbance, coastal hypoxia and anoxia, has been exacerbated by eutrophication (Nixon, 1995;Zhang et al., 2010).No other environmental variable than dissolved oxygen has changed in shallow coastal seas and estuaries so drastically in such a short time (Diaz and Rosenberg, 2008;Diaz, 2001).Hypoxia adversely affects the community structure and trophodynamics of marine ecosystems (Gray et al., 2002), for example by eliminating sensitive species and promoting more tolerant species (Dauer, 1993;Wu, 2002).
The northern Adriatic is one of currently more than 400 eutrophication-associated dead zones worldwide, with a global area of more than 245 000 km 2 (Diaz and Rosenberg, 2008).As a shallow, semi-enclosed waterbody (mean depth 30 m) with water column stratification (Justić, 1991;Malej et al., 1995), soft bottoms, high riverine input mainly from the Po River, high productivity and long water residence time, it combines many of the features known to promote late summer hypoxia and anoxia (Stachowitsch and Avcin, 1988;Stachowitsch, 1991;Ott, 1992).These features, combined with anthropogenic eutrophication and massive marine snow events, qualify the northern Adriatic Sea as a sensitive marine ecosystem (Stachowitsch, 1986).It is at the same time one of the most heavily exploited and degraded seas worldwide (Lotze et al., 2010) and one of the most productive areas at several levels, from phytoplankton to fish, of the Mediterranean (Fonda Umani et al., 2005).Here, long-term decreases in the bottom-water oxygen content have been observed from the middle of the 20th century (Justić, 1991) until the early 1990s, which were connected with changes in and extensive mortalities of the epibenthic communities that characterize the soft bottoms (Stachowitsch, 1984;Justić et al., 1987;Ott, 1992).Fedra et al. (1976) named one such wide-ranging community, the community studied here, the Ophiothrix-Reniera-Microcosmus (ORM) community based on the dominant brittle stars Ophiothrix quinquemaculata (Delle Chiaje, 1828), sponges Reniera spp., and ascidians Microcosmus spp.This suspension-feeding community is aggregated into so-called multi-species clumps or bioherms, which show a wide and patchy distribution in the northern Adriatic Sea (Zuschin and Stachowitsch, 2009;McKinney, 2007).Introduction

Conclusions References
Tables Figures

Back Close
Full They consist of a shelly base (bivalve or gastropod shell) overgrown by sessile organisms (sponges, ascidians) which, in turn, serve as elevated substrates for vagile and hemi-sessile species such as brittle stars, holothurians, hermit crabs and gastropods (Fedra et al., 1976;Zuschin and Pervesler, 1996).Such aggregations are also a food source for epibenthic crustaceans and fishes in the northern Adriatic and North Sea (Hampel et al., 2004).Filter-and suspension-feeder communities are an important stabilizing compartment in the overall marine ecosystem by removing enormous amounts of suspended material from the waterbody (Ott and Fedra, 1977).Such benthic communities have been termed a natural eutrophication control (Officer et al., 1982) and thus provide crucial ecological goods and services.
The damage to or loss of such communities therefore has ecosystem-wide implications.During hypoxia and anoxia, the benthic organisms in the ORM community show a distinct sequence of behaviours and mortalities.This has been documented in situ during larger-scale anoxias (Stachowitsch, 1984) and has been confirmed and analysed in great detail experimentally using an underwater chamber on a small scale (0.25 m 2 ) on the seafloor in 24 m depth.These behaviours include emergence of infauna species (Riedel et al., 2008b), unusual interactions including predation (Riedel et al., 2008a;Haselmair et al., 2010), altered locomotion and activity patterns (Pretterebner et al., 2012), and a clear sequence of mortalities (Riedel et al., 2012).For an overview of these responses see Riedel et al. (2013).The immediate behavioural responses during the course of hypoxia and anoxia until mortality of the macrobenthos are therefore now well documented (sample 4 d film available at: http://phaidra.univie.ac.at/o:87923).The longer-term implications are less well known.The collapse of benthic communities and recurring disturbances -in this case mortalities and repeated hypoxia/anoxia along with marine snow events -raise fundamental issues of ecosystem stability and resilience.One approach to addressing these issues is to examine post-disturbance events, typically successions.Various stages of succession (opportunists, an ecotone point, a transition zone) have been described early on (Pearson and Rosenberg, 1978).Overall, events proceed in the direction of more Introduction

Conclusions References
Tables Figures

Back Close
Full stable "normal" communities with an increase in the number of species, abundance and biomass (Rosenberg et al., 2002).Two basic recolonization strategies have been described: motile species can immigrate into denuded areas, in particular if the damage is rather small-scale or, when the affected areas are large, colonization depends mostly on post-larval settlement by pelagic recruits (Pearson and Rosenberg, 1978;Whitlatch et al., 1998;Norkko et al., 2010).The reestablishment of community structure on the soft bottoms of the northern Adriatic requires biogenic structures such as bivalve and gastropod shells or sea urchin tests as a substrate on which larvae of sessile and motile epifauna species can settle.Although such structures are abundant in the immediate post-mortality situation, sedimentation can cover them quickly and hamper new epigrowth.Compared to the mortality events, recovery is a much longer-term process (Stachowitsch, 1991;Stachowitsch and Fuchs, 1995).After bottom enrichment from a sulphite pulp mill in the Saltkallefjord, Sweden, the fauna in the central part of that fjord was examined in relation to oxygen deficiency and the presence of hydrogen sulphide.It took about 3 years for the top sediment to recover to a state suitable for macrofauna recolonization (Rosenberg, 1971).Intertidal communities from muddy sand habitats, for example, also have very slow biological and physical recovery rates (Dernie et al., 2003).Recovery of benthic communities after experimental trawling disturbances took more than 18 months (Tuck et al., 1998).
In the Gulf of Trieste northern Adriatic, Stachowitsch and Fuchs (1995) were unable to detect full recovery even after more than ten years, because recolonization was repeatedly interrupted by another source of disturbance, namely commercial fishing: the use of bottom trawls overturned or broke apart newly established multi-species clumps.Kollmann and Stachowitsch (2001) reported a negative influence in that elevated biogenic structures were either sheared off and uprooted (e.g.Pinna sp.shells) or overturned and crushed.Such effects are echoed elsewhere, with Rapido trawling significantly changing community structure and mean abundance of common taxa (Pranovi et al., 2000), or beam trawling altering the physical characteristics of the sea Introduction

Conclusions References
Tables Figures

Back Close
Full bottom (Kaiser and Spencer, 1996).The use of bottom gears reportedly increases oxygen consumption and nutrient concentrations, promotes phytoplankton primary production affects sessile epibenthic species and changes overall community structures (Riemann and Hoffmann, 1991).This includes potential long-term effects of fishing on bottom fauna (Jennings and Kaiser, 1998;Thrush and Dayton, 2002).
The reduction of long-lived suspension-feeding organisms is typically followed by a shift to a habitat dominated by detritus feeders, which can hinder the recovery of the suspension feeders by consuming them (Dayton et al., 1995).Such altered communities are often dominated by juvenile stages, mobile species and rapid colonizers (Pearson and Rosenberg, 1978;Dayton et al., 1995).In the Northern Adriatic Sea, for example, hermit crabs increased, while the designating community organisms such as brittle stars, sponges and ascidians decreased (Kollmann and Stachowitsch, 2001).
Such anthropogenic disturbance can lead to the loss of functional groups (top-down effect), compromising resilience and making ecosystems more vulnerable to additional threats such as pollution, and the capacity for self-repair is reduced (Folke et al., 2004).
All these circumstances make benthic communities a long-term "memory" of disturbances (Stachowitsch, 1992).The present work was designed to assess both the immediate, short-term predation and scavenging processes after disturbance (artificially induced anoxia) and the longer-term recovery processes in the northern Adriatic Sea.We evaluated the experimentally affected areas by examining time-lapse films and by analysing photographic series.In the former, predators and scavengers were counted image by image, in the latter, the area occupied by recolonizers was calculated as a measure of recovery.

Material and methods
The activities of seven mobile species were examined.These encompass all the predators and scavengers that entered the experimental quadrats after the induced anoxia.They included two invertebrates and 4 vertebrates: one gastropod, the Banded dye Introduction

Conclusions References
Tables Figures

Study site
The study site is located in the Gulf of Trieste, in the northern Adriatic Sea (45 • 32 55.68 N 13 • 33 1.89 E) off Cape Madona, Slovenia (Fig. 1).The experimental site is about 2 km offshore at a depth of 24 m, near the oceanographic buoy of the Marine Biology Station in Piran (Slovenia).This position was chosen to minimize disturbance and damage by commercial fisheries.The soft bottom here is dominated by the ORM-community described above.

Experimental set-up
Two different chamber configurations are available.One is the closed configuration, which creates anoxia by sealing a 50 × 50 × 50 cm volume off from the surrounding water with plexiglass walls.Another is the open configuration, in which the plexiglass chamber is replaced by an open frame of the same size to observe behaviour under normoxic conditions.Four removable tapered metal tips stabilize both configurations in the sediment.Centrally, on a lid on top of the chamber or frame, a digital time-lapse camera (Canon EOS 30D) with a zoom lens (Canon EFS 10-22 mm) is positioned, flanked by two exchangeable battery packs (9Ah Panasonic, Werner light power, Unterwassertechnik, Germany) and two flashes ("midi analog" series 11897; Subtronic, Germany).The battery packs, combined with a specially designed electronic control circuit, enable the equipment to be operated for about 72 h.Finally, a datalogger unit (PA3000UD, Unisense, Denmark) and four sensors for measuring oxygen, temperature Introduction

Conclusions References
Tables Figures

Back Close
Full and hydrogen sulphide are placed on the lid (for a full description see Stachowitsch et al., 2007).The present recovery experiments involved the following deployment protocol.The underwater device was initially positioned in its closed configuration over a representative multi-species clump (i.e. with both sponges and ascidians) on the sediment.After 72 h the plexiglass chamber was replaced by the open frame, i.e. the lid was briefly lifted and the chamber replaced by the open frame and the post-anoxia recovery experiment began.Within the next three days, post-mortality predation and scavenging were documented; the camera took images in 3-min intervals.

Short-term predation and scavenging
Two experiments were performed.In the "August" experiment, post-disturbance images (open configuration) were evaluated beginning on 15 August 2009 (10:35 a.m.) until 18 August 2009 (10:05 a.m.).The "September" experiment extended from 14 September 2009 (03:25 p.m.) until 17 September 2009 (10:49 a.m.).The August experiment yielded a total of 1430 images, the September experiment 1384 images.This corresponds to an overall documentation time of 71.5 h in August and 67.5 h in September.The images were also processed into time-lapse movies using the Adobe Premiere 6.5 program (August recovery time-lapse film available at: https://phaidra.univie.ac.at/detail object/0:262380).The fishes were analysed image by image: the data are summarized in 6-h steps.Because of the relatively slow movements of the hermit crabs and gastropods and the more gradual changes in their numbers, only every tenth image was evaluated for these groups.For every analysed image the present predator and scavenger species were recorded.For hermit crabs and gastropods, the different substrates (sponges, ascidians, sediment) they were on were recorded.Normally, the gastropods and hermit crabs are found only on the sediment surface.We therefore equate the dead and/or moribund organisms (sponges and ascidians) chosen by the organisms with their preferred prey item.The time and sequence of arrival/departure and the maximum number of individuals were recorded.Day/night Introduction

Conclusions References
Tables Figures

Back Close
Full activity (based on the time of sunrise and sunset, www.sun.exnatura.org) of the hermit crabs and the gastropods was analysed.Individual specimens of the same species could rarely be differentiated because colouration and sizes were typically very similar.Accordingly, the number of fish, gastropod and hermit crab individuals refers to the number of individuals visible over the respective time period.This number clearly overestimates the actual number of different individuals present because many individuals remained in the plot for extended periods.It does, however, capture the role that an individual or individuals played because they were present and exerted a scavenging or predatory influence.

Longer-term recovery
These experiments were conducted on the same two 0.25 m 2 plots used for the shortterm time-lapse camera study.Here, the pre-disturbance surface area occupied by living sponges and ascidians, as well as by other hard structures and lebensspuren, was calculated using CorelDraw9 and Excel 2010 based on the photos taken from the closed configuration.The corresponding surface areas were then measured again days to years after anoxia.For this purpose, photos were taken with a hand-held camera.
To better compare the two experiments, the times when the images were taken were categorized, e.g., 7-13 days is termed "10-days" in the following text.The data in the Introduction

Conclusions References
Tables Figures

Back Close
Full two sets of photographs taken after 1 yr and after 2 yr at each plot are averaged, and referred to as "1 year" and "2 years", respectively, unless stated otherwise.Time series analyses and cross-correlations were calculated to define the residence time of gastropods and hermit crabs in the plots.Graphs of time series analysis can be found in Blasnig (2012).A Chi 2 test was performed to determine the difference in the substrates chosen by gastropods and hermit crabs and between the experiments.For statistical analyses the program Past was used (Hammer et al., 2001).

Short-term scavenging after anoxia
Moribund and dead sponges and ascidians attracted fishes, hermit crabs and gastropods in a rapid and distinct sequence.Numerous fishes arrived in the first hours.They were followed by hermit crabs (Paguristes eremita), which showed a rapid initial increase in the number of individuals.Finally, the gastropods (only Hexaplex trunculus) appeared (Fig. 2), with a slow increase of individuals.number visible over this time period, not necessarily different individuals, see Sect. 2).

The fishes
In the September experiment, all four species were initially present and showed the highest abundance (individual per species) in the first 6-h time step: G. niger peaked with 80, D. vulgaris 18 and S. hepatus 13 individuals.Until the end of the experiment, the number of individuals decreased markedly.

The first 12 h: hermit crabs and gastropods
The first 12 h were evaluated separately to determine in more detail the order in which the individuals arrived and which organisms arrived first.In August, the first three Paguristes eremita were observed after 30 min and the number rapidly increased up to 19 individuals after 9 h.After 3 h the first Hexaplex trunculus arrived and the number of individuals slowly increased to 3 individuals after 8 h (Fig. 2a).In September, the first P. eremita appeared after 1 h, and numbers then increased up to 12 individuals after 6 h, before decreasing again to 9. Seven H. trunculus individuals survived the pre-anoxia deployment (Fig. 2b) and were present and visible from the start.After 30 min the first new gastropod entered the frame.Two hours later, 3 individuals again left the area (5 present), but thereafter the number increased steadily (maximum 17).

Day/night activity
In both experiments the number of Paguristes eremita varied considerably over time.
The values decreased in all three nights examined and increased conspicuously during daylight hours (Fig. 3).During the day the number of individuals in August increased to 24 (45.5 h after anoxia) and 33 (after 71 h), while at night, values fell to 6 and to 3 (after 38 h and 58 h, respectively).In September, the number of individuals during daylight hours peaked at 28 (17.5 h after the anoxia) and dropped to only 2 (after 37 h) and 3 individuals (after 62.5 h).The time series analysis for P. eremita showed highly significant peaks at 21.2 h (August), 28.4 h and 54 h (September), a significant peak at 12.4 h (August), and distinct but not significant peaks at 15.5 h and 57.2 h (August) and Introduction

Conclusions References
Tables Figures

Back Close
Full at 10.8 h (September) (data not shown).These peaks demonstrate a semidiurnal and diurnal periodicity.
Hexaplex trunculus showed a relatively slow but constant increase, levelling off somewhat over the last day.The peak number was 8 individuals at three periods between 46.5 and 65.5 h in August, and 17 individuals after 38.5 h in September.At the end of both experiments, 4 (August: 71.5 h) and 13 (September: 67.5 h) H. trunculus were still present in the plots.The time series analysis showed highly significant peaks at 18.5 and 38.1 (August) as well as 13.5 h, 27.0 h and 60.0 h (September).Visible but not significant peaks are present at 8.2 h, 9.7 h and 11.2 h (August) as well as at 18 h (September).These peaks point to a periodicity of 9 h and 12 h as well as multiples thereof.
The fishes show highly significant peaks at 9.9 h (August) as well as 12.3 h and 21.6 h (September) and visible but not significant peaks at 30.1 h (August) 6.4 h and 7.8 h (September).This points to a semidiurnal and a roughly diurnal periodicity.

Preferred substrates/prey items
The substrates that Paguristes eremita and Hexaplex trunculus chose were different and consistent in both experiments.Hermit crabs were observed on the sediment, on sponges and on ascidians, whereas H. trunculus occurred mainly on ascidians and to a lesser extent on the sediment (Fig. 4).All categories (i.e. compared species-experimental month pairs) are significantly different from each other.Importantly, there are bigger interspecific than intraspecific differences, i.e. the differences between the species (P.eremita in August and September versus H. trunculus in August and September) are bigger than between the months (P.eremita in August versus September and H. trunculus in August versus September) (Table 1).
The duration of the stay of 9 Paguristes eremita and 9 Hexaplex trunculus individuals was calculated.The individuals were chosen based on their recognizability (e.g.epigrowth on shell).This duration averaged 5 h and 19 min for P. eremita and 12 h and 9 min for H. trunculus.Thus, on average, the gastropods stayed nearly 7 h longer than Introduction

Conclusions References
Tables Figures

Back Close
Full the hermit crabs.In the August experiment, several P. eremita dragged off the ascidian Phallusia mammillata.Within 53 h, they dragged it (along with an attached anemone Cereus pedunculatus and the ascidian Microcosmus sp.) 8 cm in one direction, then 21 cm in the opposite direction, just outside the frame.The ascidian, which had become discoloured, was then partially consumed by the end of the film, with pieces being visibly torn off.

Sea anemones
The August experiment contained three sea anemones Cereus pedunculatus: two attached to ascidians (a Microcosmus sp. and P. mammillata), one next to a large sponge.All three survived the anoxia but showed extreme elongation and rotations.One was carried outside the frame by hermit crabs (attached to P. mammillata, see above), the second was also flipped out of the frame (attached to Microcosmus), probably by hermit crabs.
The third individual, immediately adjoining the sponge, fully emerged from the substrate and began to crawl away 66 h and 15 min after the end of the artificial anoxia (opening of plexiglass chamber).At the end of the short-term evaluation (film: 71.5 h, 18 August) the specimen was still positioned inside the frame.During the subsequent longer-term evaluation the same anemone was still alive and visible at that position on the image of 22 August.Two days later the anemone disappeared from the images.

Longer-term recovery
The surface area of each experimental plot (inside the chamber) was 2500 cm markedly by day 6 of the long-term experiment.The value dropped from 540 cm 2 to 155 cm 2 coverage (6 %) (Fig. 5a).The sessile fauna dropped from originally 1939 cm 2 to 12 cm 2 (0.5 %).In September, the initial coverage of sessile fauna accounted for 631 cm 2 (25 %) and of vagile fauna 663 cm 2 (26.5 %).As in August, both fauna groups decreased drastically and the trends in coverage by vagile fauna paralleled those of the sessile fauna.After 6 days, the sessile fauna fell to 286 cm 2 (11.5 %), the vagile fauna to 154 cm 2 (6 %) (Fig. 5b).In the "10-day" category (see Sect. 2), nearly the whole fauna was consumed (sessile) or had crawled away (vagile).Both 1 and 2 years after the start of the experiment, no macroscopic sessile epifauna was observed on either plot.The exception was one sea anemone, Cereus pedunculatus, in the September experiment; it survived anoxia and was still present in both following years.Shells (> 1 cm) and smaller shelly material ("coquina") were visible in both plots (Fig. 6a, b).Overall, in both experiments shell coverage (Fig. 6a) fluctuated but remained relatively stable with similar shell values in the beginning and the end of the two experiments ("2 years").More precisely, in August, shell cover increased from an initial 126 cm 2 to 185 cm 2 after 1 yr, with a drop after 2 yr to 62 cm 2 .In September, coverage decreased 1 and 2 yr after the anoxia, with a minimum of 46 cm 2 after "2 years".
In between, there was an increase to 139 cm 2 (at the 1-year-plus-10-day sampling).
Coquina coverage (Fig. 6b), in contrast, fluctuated more widely and clearly increased over the course of the experiments.Here, the two experiments were less similar, with values of the September experiment always exceeding those of August.
In the August experiment, no coquina was initially observed, but after 3-4 days the coverage increased.After "10-days" it fell again to 57 cm 2 , followed by a peak of 842 cm 2 after 1-yr-plus-10 days.Note, however, that 10 days earlier, the value was zero, showing the potential for major fluctuations even over short time periods.In the September experiment, values initially decreased, but then steadily increased, dropping after 1 year but peaking again at 1499 cm 2 after an additional 10 days (i.e.1year-plus-10-day sampling).Thus, values changed considerably within even a 10-day Introduction

Conclusions References
Tables Figures

Back Close
Full period.With few exceptions (early period in August), coquina always covered a larger surface than larger shells.The coverage by lebensspuren (vagile fauna, endofauna burrow openings etc.) in the September experiment was high and varied considerably, ranging between 418 and 1360 cm 2 (data not shown).Insufficient visibility prevented evaluating lebensspuren on one date (6 days after removing the chamber) in September (in August, poor visibility prevented evaluation during the first 7 days after chamber removal).

Discussion
In the northern Adriatic, decreases in dissolved oxygen cause rapid mortalities of macroepibenthic communities (Stachowitsch, 1984), which are an important stabilizing compartment in this and other shallow marine ecosystems (Ott and Fedra, 1977).
Here, the recolonization of the benthic compartment is very slow (Stachowitsch, 1991), additionally hampered by harmful fishing activities.The recovery of marine ecosystems with slow successions could take 40 years or even longer (Jones and Schmitz, 2009).
The present study extends prior investigations on the behavioural responses (Stachowitsch et al., 2007;Haselmair et al., 2010;Pretterebner et al., 2012;Riedel et al., 2008aRiedel et al., ,b, 2013) ) and mortality sequences (Riedel et al., 2012) to artificial anoxia using a specially designed underwater device.It helps fill in the gap between ecosystem collapse and community recovery by examining short-term scavenging and longer-term recovery processes.

Short-term scavenging after anoxia
After the artificial anoxia, the moribund and dead organisms attracted predators and scavengers.This process resembles that described after damage done by benthic fisheries in the north Irish Sea (Jenkins et al., 2004), where most of the dead material was removed in the first days.The images revealed a clear sequence of Introduction

Conclusions References
Tables Figures

Back Close
Full predators/scavengers.The quick arrival of fishes is attributed to their swimming speed (Planes et al., 1997, e.g. Diplodus vulgaris: 12.3 cm s −1 ), which is clearly several hundred times faster than the next arriver, Paguristes eremita.These hermit crabs travel up to 21.6 m d −1 , with an average speed of 2.1 m h −1 (Stachowitsch, 1979); this is equivalent to 3.5 cm min −1 .Based on arrival time, Hexaplex trunculus may be the slowest of these three species The speeds of H. trunculus were not measured, but considering the speeds of other, smaller gastropods (e.g.Littorina littorea, 2.88-4.47cm min −1 Erlandsson and Kostylev, 1995) and the similar size of large P. eremita and adult H. trunculus, the speeds of the hermit crabs and gastropods may be similar.In such cases, arrival times would also depend on a species' density, with more abundant species having a greater probability of having individuals closer to the experiment plots.In the ORM-community in the northern Adriatic Sea, P. eremita has a density of 2.4 individuals m −2 (transect method: Pretterebner et al., 2012) , which is slightly higher than in an earlier quadrat sampling (1.9 individuals m −2 Stachowitsch, 1977).H. trunculus, in turn, has a density of 0.2 individuals m −2 in the northern Adriatic Sea (Wurzian, 1982).
Note, however, that our site is close to an oceanographic buoy, whose anchor chains are heavily overgrown with mussels, and detached mussels are found directly under the buoy.Accordingly, the density of H. trunculus, which feeds on such mussels, is very high in this area.Without this bias, we would expect the gastropods to arrive in greater numbers at a much later date.
Fishes were conspicuously abundant in the first six hours after opening the chamber.This is the time when the most food items were available, including smaller softbodied organisms freshly emerged from the sediment and cryptic fauna from sponges and other multi-species clumps.The fishes were dominated by Gobius niger, which feeds on polychaetes, amphipods, mysids and decapods (Richards and Lindeman, 1987).Pagellus erythrinus mainly preys on benthic organisms such as polychaetes, brachyuran crabs and benthic crustaceans (Fanelli et al., 2011), Serranus hepatus on invertebrates, mainly decapods (Labropoulou and Eleftheriou, 1997), and Diplodus vulgaris on benthic echinoids such as Echinocyamus pusillus (M öller, 1776) and

Conclusions References
Tables Figures

Back Close
Full Psammechinus microtuberculatus (Blainville, 1825), but also decapods and bivalves (Pallaoro et al., 2006).P. eremita and H. trunculus apparently mainly fed on material that remained after the fishes had already been present for several hours.Based on their positions, this was mainly ascidians (Phallusia mammilata, Microcosmus sp.) and sponges.The time-lapse camera approach was unable to provide direct evidence that the fishes also fed on sponges or ascidians, i.e. the act of feeding was never captured in an image, although the fish were often positioned in an oblique angle with the mouth facing down.Ascidians rely on chemical (Lindquist et al., 1992) and physical defences (e.g.tunic toughness: Koplovitz and McClintock, 2011) against predation, although the efficiency and strategies vary greatly between species (Tarjuelo et al., 2002).These mechanisms may still be partly effective in freshly dead individuals, making them unattractive/unsuitable for fishes as the quickest post-anoxia arrivals.(Young, 1989), for example, observed ascidians being eaten by gastropods, which insert their proboscis into a siphon of the ascidians and consume them.Our evaluation showed some H. trunculus on the siphon of Microcosmus sp., but other individuals were also positioned on other parts of the ascidians.
To better determine which food sources the crabs and the gastropods prefer, we evaluated the substrates on which they positioned themselves, equating the chosen dead or moribund species with the preferred prey.In some cases, these substrates were clearly eaten and reduced in size, although we cannot exclude that smaller species associated with the sponges or ascidians may have been preyed upon.While both species feed on sponges and ascidians, H. trunculus was much more frequently positioned on Microcosmus.Thus, the interspecific differences were greater than intraspecific differences.The August images also showed P. eremita dragging off and consuming an ascidian (Phallusia mammillata).H. trunculus, in contrast, crawled up the ascidians and typically remained there for several hours: on average, the gastropods stayed within the frame more than twice as long as the crabs.This is in line with Sawyer et al. (2009), who documented 10.5 h, 5.2 h and 2.9 h for feeding and manipulating the mussel prey in three selected H. trunculus individuals in this community.Stachowitsch

Conclusions References
Tables Figures

Back Close
Full (1979) observed the activity behaviour of P. eremita in the Gulf of Trieste and recognized two types of movement interruptions: (1) short stops related to feeding and investigating structures on the sediment and (2) longer pauses during the night hours reflecting a resting period.This was evident in our study by a semidiurnal and diurnal periodicity of presences.Moreover, an explanation for why only few individuals remained in the plot near their prey items at night might be that, in their dormant night phase, crabs do not aggregate or remain near larger prey items because it would increase the risk that they themselves would be consumed, along with the prey, by other, larger (fish) predators/scavengers.The same may hold true for H. trunculus based on the 9-and 12-h periodicity indicated by the time series analysis.
Tolerance to hypoxia not only improves survival but may -depending on the behaviour and tolerance of both predator and prey -enable also more successful predation during and after low-oxygen events (Pihl et al., 1992).Molluscs, for example, are generally considered to be more tolerant to hypoxia than many other invertebrate groups (Vaquer-Sunyer and Duarte, 2008).H. trunculus is among the more tolerant species in this soft-bottom community (Riedel et al., 2012) and also in the present study, 7 individuals survived the oxygen depletion during the September experiment.Although none of the P. eremita survived the experimental anoxia, hermit crabs were identified as more tolerant crustaceans (Riedel et al., 2012), improving their opportunities for predation during and after hypoxia.Depending on the severity of hypoxia, both species may be able to take advantage of more vulnerable prey.Such altered predator-prey relationships can affect community structure.
The sea anemone Cereus pedunculatus (August plot) crept away from its original position 66 h and 15 min after the chamber was opened.Anemones, although normally firmly attached to a substrate, are known to be able to detach and move, providing a mechanism to leave unfavourable conditions (Rittschof et al., 1999).The hermit crab symbiont Calliactis parasitica (Couch, 1842) allows Dardanus arrosor (Herbst, 1796) to detach it mechanically by using specific behaviours, whereby the anemone then reattaches itself to the new shell (Ross, 1979).In the present experiment, the movement

Conclusions References
Tables Figures

Back Close
Full of C. pedunculatus is intriguing because it occurred in the re-oxygenated environment rather than during anoxia.One explanation is that the high density of feeding hermit crabs may have disturbed the anemone and caused it to release the attachment and move.An alternate explanation would be that the adjoining dead sponge caused unfavourable conditions.Accordingly, the disturbance experienced after the return of normoxic conditions was apparently more severe than the effects of anoxia: in none of the many other experiments that evaluated behaviour did anemones ever move away during severe hypoxia or anoxia (with or without H 2 S conditions),, although they showed a series of severe stress reactions including body contractions, rotation and extension (Riedel et al., 2008b(Riedel et al., , 2013)).

Longer-term recovery
Even 2 years after anoxia, no macroepibenthic recovery was observed in either of our experimental plots.Benthic recolonization is scale-dependent and can involve larval settlement (extensively damaged areas) and immigration by vagile organisms, which may be more important for small-scale recovery (Pearson and Rosenberg, 1978;G ünther, 1992;Whitlatch et al., 1998).Past mortalities in the northern Adriatic Sea ranged from restricted sites (several km 2 ; Stachowitsch, 1992) to thousands of square kilometres (Ott, 1992).The worldwide second largest anthropogenic dead zone is in the Gulf of Mexico, covering a mean area of 17 000 km 2 (Turner et al., 2008).Accordingly, larval settlement would be expected to define recolonization -a long-term process.
However, the extensive loss of the characteristic aggregations of sessile sponges and ascidians (multi-species clumps as larval producers) during mortalities in the northern Adriatic points to the difficulty in restoring community structure and function.Moreover, recovery from human disturbances can take longer than from natural disturbances (Jones and Schmitz, 2009).Additionally, also the high sedimentation rate and fishing activities may hinder successful epigrowth.
In our small-scale experiment, both recolonization strategies are conceivable.Although the experimental area was tiny compared with past hypoxia events, no recovery Figures was recorded despite immediately adjoining multi-species clumps and vagile organisms.Importantly, settlement and growth of sessile organisms depend on the presence of adequate substrates.In the northern Adriatic, these are typically gastropod and bivalve shells or echinoderm tests (Zuschin and Stachowitsch, 2009, and references herein).Although we recorded shells in our plots, they were apparently not large enough, sufficiently exposed, or available at larval settlement times.This is supported in the percent coverage by shells: it fluctuated and ultimately declined after two years.Small-scale factors that affect availability include manipulation by vagile species or use as camouflage by the sea urchins (Riedel et al., 2008b); large-scale factors include sedimentation and resuspension, which bury small or flat shells.Fishery gear such as dredges and bottom trawls resuspend enormous amounts of sediment, creating major turbidity and sedimentation events (M.Stachowitsch, personal observation).Such gear also overturned shells, crushed multi-species clumps and sheared off structures projecting from the bottom (e.g.pen shells), impacting newly settled invertebrates growing on them (Stachowitsch and Fuchs, 1995).Finally, the passage of large ships near the study site can also resuspend sediment.All these factors help explain the great variation in the visible coverage by shells, impacting larval-based recolonization.Such unstable conditions in the top sediment layer are also indicated by the increasing amount of coquina.Finally, one short-cut to the establishment of multi-species clumps in this community -the deposition of heavily encrusted gastropod shells by hermit crabs (Stachowitsch, 1980) -did not take place within our two plots in the two years examined.Anoxia-related disturbance and recovery in the northern Adriatic has been described as "rapid death -slow recovery" (Stachowitsch, 1991): most organisms die shortly after anoxia is attained, but recovery takes years.The ORM-community showed little recovery 10 years after the collapse in 1983; although certain vagile fauna such as hermit crabs increased, larger multi-species clumps did not develop.This was attributed to repeated anoxia and other disturbances such as dredging and trawling damage (Stachowitsch and Fuchs, 1995;Kollmann and Stachowitsch, 2001).Recovery is therefore Figures also determined by the frequency of disturbance.After benthic trawling the recovery time for slowly growing sponges could take 8 yr (Kaiser et al., 2006); sponge recolonization in Alaska, without further disturbances, could take decades (Rooper et al., 2011).It is therefore unsurprising that we did not observe any recovery of sessile organisms during our experiment.
Vagile fauna is an important part of this macrobenthic community.The rapid decrease of sessile organisms (moribund or dead sponges and ascidians) within the first 6 days (August) and 13 days (September) after anoxia is attributed to their removal or consumption by the mobile animals that entered the experimental plots (or that survived inside the plots (H.trunculus in September experiment)).The subsequent decline in food items was correlated with a corresponding drop in these organisms.The coverage of the plots by vagile fauna after 1 and 2 years consisted mainly of P. eremita, H. trunculus, Psammechinus microtuberculatus and Ophiothrix quinquemaculata.Kollmann and Stachowitsch (2001) consider that lebensspuren are helpful parameters to quantify community status and activity of mobile forms.Our lebensspuren observations in the September plot revealed a coverage of between 418 and 1360 cm 2 .This is equivalent to 17 and 54 % of the whole plot and points to abundant vagile fauna in our study area.Such high densities may also keep larval recruits low: grazing on and manipulation of the shells that the multi-species clumps initially require to grow on (Zuschin et al., 1999) could also help explain the lack of recovery.
As opposed to recolonization and succession, where using small-scale experiments to predict larger-scale responses may not be possible (Zajac et al., 1998), we consider our results on scavenging and predation in the small plots to be valid for larger spatial scales.The distinct sequence of arrivals (fish, followed by hermit crabs and finally gastropods) -related to the relative speeds of the organisms -could be expected on the large scale as well.In wide-ranging anoxia in the northern Adriatic Sea and elsewhere, the process would be delayed: immediate immigration would be slower due to the greater distances involved (Stachowitsch et al., 2012)  Full et al. (1995) established that coastal marine ecosystems are the world's most endangered habitats, and the Mediterranean is no exception, with documented effects Figures Back Close Full Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | In the August 2009 experiment (71.5 h) we observed three different species, Diplodus vulgaris, Serranus hepatus and Gobius niger, and in the September 2009 experiment (67.5 h) additionally Pagellus erythrinus.In both experiments, the maximum number of individuals per species was present in the first six hours.G. niger was most abundant and showed a slowly decreasing trend, while the other species decreased rapidly to very low numbers.A maximum of three species were visible at the same time in a single image.In the first 6 h of the August experiment, all three species were present: G. niger with 83, D. vulgaris 35 and S. hepatus 26 individuals.The two latter species then decreased to very low numbers or zero in the successive 6-h periods.G. niger also decreased with time, but never fell below 22 observed individuals per 6 h (i.e. total Introduction Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | the August experiment, 1939 cm 2 (77.5 %) were covered by sessile organisms and 540 cm 2 (21.5 %) by vagile fauna (most of the latter represented by hermit crabs and gastropods; total values can exceed 100 % because animals on living substrates were counted separately).After the increase in vagile forms during the shortterm observations (gastropods and hermit crabs, see above), this fauna decreased Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | , but the sequence would presumably be the same as observed in the present experiments.Scaling up Figures Back Close Full Screen / Esc Printer-friendly Version Interactive Discussion Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | our small-scale experiments therefore points to alarming long-term effects and calls for intense management measures to reduce further pollution and physical destruction of shallow marine environments.Discussion Paper | Discussion Paper | Discussion Paper | Norkko, J., Norkko, A., Thrush, S. F., Valanko, S., and Suurkuukka, H.: Conditional responses to increasing scales of disturbance, and potential implications for threshold dynamics in softsediment communities, Mar.Ecol.Prog.Ser., 413, 253-266, 2010.4371 Officer, C. B., Smayda, T. J., and Mann, R.: Benthic filter feeding: a natural eutrophication control, Mar.Ecol.Prog.Ser., 9, 203-210, 1982.Discussion Paper | Discussion Paper | Discussion Paper |

Fig. 5 .
Fig. 5. Coverage of sessile (dead and living) and vagile fauna after anoxia in the August 2009 (above) and September 2009 (below) plot.Note different scales on y-axis.Start of experiment defined as 0.1; values measured inside closed chamber (original coverage).

Table 1 .
Chi 2 test for differences between categories of chosen substrates/prey by Paguristes eremita and Hexaplex trunculus in the two experiments (2009).