DOI: 10.5061/dryad.4f4qrfjcm
title: Longitudinal variation in the nutritional quality of basal food sources and its effect on invertebrates and fish in subalpine rivers

description: 1. There is growing recognition of the importance of food quality over
 quantity for aquatic consumers. In streams and rivers, most previous
 studies considered this primarily in terms of the quality of terrestrial
 leaf litter and importance of microbial conditioning. However, many recent
 studies suggest that algae are a more nutritional food source for riverine
 consumers than leaf litter. To date, few studies have quantified
 longitudinal shifts in the nutritional quality of basal food resources in
 river ecosystems and how these may affect consumers. 2. We conducted a
 field investigation in a subalpine river ecosystem in Austria to
 investigate longitudinal variations in diet quality of basal food sources
 (submerged leaves and periphyton) and diet source dependence of stream
 consumers (invertebrate grazers, shredders, filterers and predators, and
 fish). Fatty acid (FA) profiles of basal food sources and their consumers
 were measured. 3. Our results indicate systematic differences between the
 FA profiles of terrestrial leaves and aquatic biota, i.e., periphyton,
 invertebrates and fish. Submerged leaves contained very low proportions of
 long-chain polyunsaturated fatty acids (LC-PUFA), which were conversely
 rich in aquatic biota. While the FA composition of submerged leaves
 remained similar among sites, the LC-PUFA of periphyton increased
 longitudinally, which was associated with increasing nutrients from
 upstream to downstream. 4. Longitudinal variations in periphyton LC-PUFA
 were reflected in the LC-PUFA of invertebrate grazers and shredders, and
 further tracked by invertebrate predators and fish. However, brown trout
 (Salmo trutta) contained a large proportion of docosahexaenoic acid (DHA,
 22:6ω3), a LC-PUFA almost entirely missing in basal sources and
 invertebrates. The fish accumulated eicosapentaenoic acid (EPA, 20:5ω3)
 from invertebrate prey and may use this FA to synthesize DHA. 5. Our
 results provide a nutritional perspective for river food web studies,
 emphasizing the importance of algal resources to consumer somatic growth
 and the need to account for the longitudinal shifts in the quality of
 these basal resources.
description: This study was carried out in the
 subalpine River Ybbs catchment, Austria (47°45'N, 15°12'E). The
 area has a temperate climate with evenly distributed precipitation over
 the year. The primary land use in the catchment is forestry, with alpine
 meadows and agriculture constituting only a small area in the lower
 catchment. All study streams were selected in the upper catchment,
 <i>Weiße Ois</i>, which had little or no human disturbances in
 the study reaches or their upstream catchments. The substrata of study
 streams were cobbles, and the underlying geology was primarily limestone.
 Nine study streams were chosen, with consideration of different levels of
 riparian canopy cover (shade <i>vs </i>open), altitude
 (associated with temperature) and nutrient concentrations (dissolved
 inorganic nitrogen (DIN) and soluble reactive phosphorus (SRP)), including
 2 headwater streams with shaded canopy, 2 headwater streams with open
 canopy, 2 midstream sites with open canopy and 3 downstream sites with
 open canopy.
 Basal food sources, i.e., submerged leaves and
 periphyton, and their consumers, i.e., macroinvertebrates and fish were
 collected from 9 streams in October 2016. All samples were collected along
 a 20-m reach from each stream. Submerged leaves were picked from water by
 hand. Periphyton and macroinvertebrates were sampled using a 1.5 m x 1.5 m
 quadrat. Three replicated samples of periphyton were collected from three
 quadrats, respectively, and each periphyton sample was collected from five
 different cobbles by scraping them with brushes. Macroinvertebrates
 clinging to those cobbles within each quadrat were washed into a white
 tray, identified to Genus and assigned to FFG (Cummins & Klug,
 1979). Macroinvertebrate grazers in the study streams included
 <i>Ecdyonurus </i>sp., and <i>Baetis</i> sp.;
 shredders were<i> Nemoura</i> sp., <i>Leuctra</i>
 sp., <i>Allogamus</i> sp<i>., </i>and
 <i>Potamophylax</i> sp.; The only filterer collected was
 <i>Hydropsyche</i> sp.; predators included
 <i>Rhyacophila </i>sp., <i>Plectrocnemia </i>sp.,
 <i>Perla</i> sp., <i>Perlodes</i> sp., and
 <i>Isoperla</i> sp.. A separate invertebrate sample was
 preserved for further taxonomic identification. Fish samples were
 collected by electrofishing, anesthetized and killed <i>in
 situ</i> according to the Federal Act on the Protection of Animals,
 Austria (http://www.ris.bka.gv.at). Total body length (mm) of each
 collected fish was measured.  Three
 fish species were collected in the study catchment, including Brown trout
 (<i>Salmo trutta</i>), Rainbow trout (<i>Oncorhynchus
 mykiss</i>) and European Bullhead (<i>Cottus gobio</i>).
 However, only juvenile brown trout were abundant and occurred in most
 sampling sites, and therefore this species was used for FA analyses. Their
 body lengths ranged from 55 to 218 mm.
 All samples were placed in zip-lock
 plastic bags, stored on ice and kept in the dark in a portable freezer in
 the field. The dorsal muscle tissue of fish samples was extracted
 immediately for FA analyses when back in lab. All FA samples were placed
 in a -80 °C freezer until further processing, and the separate
 invertebrate sample was preserved in 75% ethanol for invertebrate
 identification.
 All FA samples of fish, macroinvertebrates,
 periphyton and leaves were freeze-dried (Virtis Genesis Freeze Dryer).
 After freeze-drying, each sample was homogenized with a glass rod and/ or
 a food processor. A dry mass of each invertebrate sample (5-7 mg), each
 fish sample (5-7 mg), each periphyton sample (10 mg) and each leaf sample
 (50 mg) was used for lipid extraction and FA methylation according to the
 methods described in Guo et al. (2016). Fatty acid methyl esters (FAME)
 were analysed using a gas chromatograph (Thermo Trace) equipped with a
 temperature-programmable injector and an autosampler. FAME were separated
 by a Supelco<sup>TM</sup> SP-2560 column (100 m, 25 mm i.d.,
 0.2 um film thickness), identified by comparison of their retention times
 with known standards (37-component FAME mix, Supelco 47885-U; Bacterial
 Acid Methyl Ester Mix, Supelco 47080-U) and quantified with reference to
 seven-point calibration curves based on known standard concentrations. FA
 compositions were expressed as percentage values relative to total FA
 (FA%).