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Aquatic Ecology 37: 159–167, 2003. © 2003 Kluwer Academic Publishers. Printed in the Netherlands.
A comparison of the fatty acid composition of Gammarus lacustris and its food sources from a freshwater reservoir, Bugach, and the saline Lake Shira in Siberia, Russia Olesia N. Makhutova 1,2, Galina S. Kalachova 1 and Michail I. Gladyshev 1,* 1
Institute of Biophysics of Siberian Branch of Russian Academy of Sciences, Akademgorodok, 660036, Krasnoyarsk, Russia; 2Krasnoyarsk State University, Svobodny av., 79, Krasnoyarsk, 660042, Russia; *Author for correspondence (e-mail:
[email protected]) Received 28 August 2002; accepted in revised form 10 February 2003
Key words: Fatty acid composition, Freshwater reservoir, Gammarus lacustris, gut content, Saline lake, Sediments, Seston Abstract We studied fatty acid (FA) composition in samples from bodies and intestinal contents of the littoral amphipod Gammarus lacustris Sars, from the Bugach freshwater reservoir. Simultaneously, samples of seston and bottom sediments were also collected from the reservoir during early August. There were no differences in FA composition of gut contents, seston and sediments of pebbly bottom. Seston was the main food source of Gammarus but some FAs Gammarus got from sediments. The FA composition of G. lacustris and seston from the Bugach freshwater reservoir were compared with those of the animals from the saltwater Lake Shira (Siberia). While FA composition of the two Gammarus populations differed significantly, those of seston were practically similar: the composition of long-chain unsaturated fatty acids, 20:53, 22:63 and 20:46, were significantly higher in animals from saline Shira Lake, whereas 16:1 and 16:0 were higher in the freshwater populations of amphipods from the Bugach freshwater reservoir. Taking into account the relevant literature data, we hypothesise that this difference in C16 acid might be a distinguishing characteristic of FA composition of freshwater and saltwater crustaceans. Introduction Lipid content and composition of aquatic organisms are known to be of great interest for pure and applied science, including aquaculture (e.g., Coutteau and Mourente (1997) and Goedkoop et al. (1998, 2000), Arts et al. (2001)). Lipid composition of food to a high degree determines food web interactions, individual and population growth (Brett and Muller-Navarra 1997). Müller-Navarra, Wacker, and Von Elert’s data (Müller-Navarra 1995; Müller-Navarra et al. 2000; Wacker and Von Elert 2001) suggest that the essential polyunsaturated fatty acids (PUFA), such as eicosapentaenoic (EPA, 20:53) and docosahexaenoic (DHA, 22:63), may be important for consumers of a higher trophic level, and this suggestion has been proven with Von Elert’s experiments (Von
Elert 2002). Nevertheless, field data on the FA composition are comparatively sparse, especially for freshwater ecosystems, thereby a few general regularities have been found. One of the most pronounced findings is that marine animals have significantly higher levels of EPA and DHA than freshwater species (e.g., Steffens (1997), but see Ahlgren et al. (1994)). For example, marine and salt lake crustacea have EPA levels of 10–30% of total fatty acids (Kattner et al. 1989; Napolitano and Ackman 1989; Nanton and Castell 1998; Gladyshev et al. 2000), while this value for freshwater crustacea is lower than 5% (Desvilettes et al. 1997; Gladyshev et al. 1999). These differences in lipid composition may be the result of several causes. Firstly, they may be attributed to lipid composition of food (Steffens 1997). Secondly, these differences may also represent the adaptation of or-
160 ganisms to salt water conditions, which is possible within one species. Thirdly, profound genetic differences between marine (salt water) and freshwater species may take place. To support some of the above suggestions, it should be useful to study salt and fresh water populations of the same species and their food sources. The aim of the present work was to study the FA composition of Gammarus lacustris Sars from a freshwater Siberian reservoir and compare it with that of another population of this species inhabiting a salt Siberian lake (Gladyshev et al. 2000), as well as seston and bottom sediments of these water bodies.
The study site The first object of our investigation was a small reservoir situated 56°03⬘ N and 92°43⬘ E in the Bugach river (secondary tributary of the Yenisei river) called the Bugach reservoir. Its surface area is about 0.32 km 2, and its maximum depth is 7 m. During summer blue-greens dominate in the reservoir and high pH values of 7.7–9.5 are characteristic of the water. During the sampling period water temperature varied from 20.3 to 22.5 °C, chlorophyll concentrations were 69.9–107.7 µg l −1, Aphanizomenon flos-aquae (L) Ralfs (up to 67% of biomass) and Phormidium unicinatum (Ag) Gom (up to 28%) were dominants of phytoplankton. The second object of our investigation was a steppic salt lake, Shira, situated 54°30⬘ N and 90°14⬘ E. The lake has no outlet and water area is c. 35 km 2. In 1998 maximum depth was 24 m. Lake water is of a sulphate-chloride-sodium-magnesium type. In summer, salinity of the epilimnion was about 16 g l −1, and water temperature was up to 22 °C. The main physical, chemical and biological parameters are given in Table 1 (Gladyshev et al. 2001; Kalacheva et al. 2002). Absence of fish and a low plankton diversity are the characteristic features of the lake. Blue-green species Lyngbya contorta dominated in the pelagial and occurred in the littoral. In the littoral moderate summer blooms of Botryococcus occurred. The animals were collected in the littoral, at a depth of about 0.5 m. Water temperature during the sampling period in both lakes were similar and no conspicuous climatic differences occurred.
Table 1. Physical, chemical and biological parameters in the Bugach reservoir and Shira Lake (mg l −1). Parameter
Bugach M ± SE
Shira M ± SE
pH Chl t°C h max (m) NH 4+-N NO 3−-N PO 43−-P Cl − SO 42− Na K Ca Mg Cu Fe Mn Al Zn Pb Tot. miner.
9.0 ± 0.02 69.9–107.7** 21.4 7 0.1 ± 0.03 0.040 ± 0.018 0.02 ± 0.001 – – 39 ± 2.2 6.3 ± 0.70 39 ± 2.4 16 ± 0.4 2.29 ± 0.43** 0.29 ± 0.06 0.06 ± 0.010 0.3 ± 0.04 0.02 ± 0.002 2.26 ± 0.15** 325 ± 9.9
8.7 ± 0.04 1.1** 22 24 0.2 ± 0.10 0.004 ± 0.002 0.01 ± 0.004 2048 ± 91.8 8010 ± 270.0 2880 ± 74.5 37.4 ± 1.51 57 ± 1.3 969 ± 38.8 0.026 ± 0.001 0.12 ± 0.006 0.03 ± 0.001 0.2 ± 0.08 0.02 ± 0.002 n.d. 18623 ± 187.8
n.d. – not detected, ** µg l −1.
Methods Sampling in the salt Shira lake was carried out in August 1998. The samples from the Bugach reservoir, were collected on 1, 3 and 8 August, 2000. Briefly, Gammarus bodies were dissected, gut contents were extracted with dissecting needles under the stereomicroscope and collected on glass plates. The dissected bodies were separated from the guts. Seston was collected from the littoral water samples by vacuum filtration through membrane filters. Bottom sediments from the littoral were collected with a handle vessel under visual control to be sure that pebbles were collected with periphyton, although periphyton was only slightly developed. The pebbles were scraped and their coverage was added to the sandy part of the sediments for following lipid extraction. All the samples were pre-treated as described above within one–two hours after sampling. Just after the pre-treatments, lipids were extracted by homogenising the samples of bodies, gut contents, seston and sediments in cold methanol/chloroform (2:1, v/v); after addition of quantitative volumes of chloroform and water, two phases were allowed to separate over-
161 night (Bligh and Dyer 1959). The lower chloroform layer was then removed, dried by passing through anhydrous Na 2SO 4 and roto-evaporated at 35 °C. Then fatty acids were methylated and analyzed as described below. To convert fatty acids to methyl esters, 0.1 ml of benzene and 0.5 ml of methanol containing 2% H 2SO 4 was added to the dried extracts. The methylation was carried out at 75 °C for 2 hours. When the methylation had finished, 2 ml of distilled water were added to the mixtures and fatty acid methyl esters (FAMEs) were extracted three times with hexane. The hexane extracts were rinsed twice with distilled water and dried by passing through anhydrous Na 2SO 4 and roto-evaporated at 35 °C. FAMEs were analyzed and identified using a gas chromatograph–mass spectrometer (GC/MS, model GCD Plus, Hewlett Packard), equipped with 30 m × 0.25 mm HP-5 (5% diphenyl and 95% dimethylpolysiloxane) fused silica capillary column. Operating conditions were: helium as carrier gas, flow rate – 1.0 ml/min, on-column split or splitless injection, solvent delay – 3 min, column temperature programmed 100– 160 °C at 8 °C/min, then 160–230 °C at 8 °C/min, and 20 min isothermal, transfer line temperature 250 °C, ion source temperature 170 °C, electron impact mode 70 eV, scanning from 50 to 450 amu at 0.5 s/scan. Monounsaturation position are chemically determined using a dimethyl disulfide derivatization and GC/MS. Mass spectra of FAMEs and their dimethyldisulphide adducts were compared with those of authentic standards (SIGMA, SERVA) and with data reported in the literature. Methods of sampling and analyses are described in details elsewhere (Gladyshev et al. 2000). To determine double pond positions in polyenoic acids, GC-MS of dimethyloxazoline derivatives of FA was used. The dimethyloxazoline derivatives of FA were prepared as follows (Spitzer 1997): 0.2 ml of 2-amino-2-methyl-1propanol (Sigma, USA) was added to the saponified lipids, the flask was filled with helium, tightly closed and heated at 170–190 °C for 1.5 hour. Then the reaction mixture was diluted with distilled water, acidified and the dimethyloxazoline derivatives of FA were extracted by hexane:acetone (96:4, v/v). Standard errors (SE) and Student’s t-test were calculated (Campbell 1967). One-linkage cluster analysis was carried out according to Jeffers (1981), using Euclidean distances. Levels (% of total) of each FA were used as axes of multidimensional hyper-space.
Results Forty FA species were identified in all freshwater samples from the Bugach reservoir (Table 2). In the bodies of G. lacustris there were 38 FAs. Saturated 16:0 acid was the dominant species. The other saturated acid, 18:0, monoenoic acids 16:1 (three isomers), 18:1 (two isomers), dienoic 18:26 and PUFA 20:53 had comparatively high levels. The markers of bacterial food, odd and branched acids, originated from the epiphytes, had remarkable levels: their sum was 4.38–5.31%, and among them 15:0 acid dominated. The body samples of 1 August and 8 August joined in one cluster (Figure 1), while the sample of 3 August unexpectedly separated from them due to a surprising detection of 18:4 acid, which was absent in any other sample (Table 2). FA compositions of intestinal tract contents, seston and bottom sediments from the Bugach reservoir, in general, differed significantly from that of Gammarus bodies and were comparatively close to each other and variable (Figure 1, Table 2). Saturated acids 16:0 and 18:0 were the dominant species, 14:0 and 20:0 also had high levels. Monoenoic 16:1 and 18:1 formed a considerable percentage of total FAs. Sum levels of bacterial acids were comparatively high: 6.5–12%, among them 17:0 dominated. 38 FA species were identified in all the samples from the salt Shira lake (Table 3). In the bodies of G. lacustris there were 23 FAs. Monounsaturated fatty acids (MUFA) were from 33.8% to 39.2%, among which 18:19 dominated. PUFAs were from 32.1% to 41.3%. EPA and DHA were the dominant species among PUFAs. The markers of bacterial food were absent. In the gut contents 23 FAs were identified. A high level (> 50% of total) of saturated acids 16:0 and 18:0 was characteristic of the FAs composition. The level of the other saturated even acids, C12–C22, was comparatively low. Monoenoic acids were primarily represented by oleic acid 18:19 with a level of 20%, by palmitoleic acid 16:1 (two isomers) and occasionally by 20:1. Dienoic acids were primarily represented by linoleic acid 18:26 and occasionally by 16:2. Among polyunsaturated fatty acids arachidonic acid 20:46 and 20:53 dominated (Table 3). Uneven and branched fatty acids, designated as bacterial, composed about 6% of total FAs of the gut contents. They were represented by 13:0, 15:0, 17:0, iso-15:0, anteiso-15:0 and iso-17:0 (Gladyshev et al. 2000). The Bugach data were compared with those for the salt Shira lake (Gladyshev et al. 2000). The Shira data
162 Table 2. Levels of fatty acids (% of total) in bodies of Gammarus lacustris, intestinal tract levels, seston and bottom sediments from the littoral of the Bugach pond, sampled in August 2000. Fatty acid
Gammarus body
Intestinal tract
Seston
Date August
1
3
8
1
3
8
1
3
8
1
3
8
12:0 i13:0 13:0 i14:0 14:0 ai15:0 i15:0 15:0 i16:0 16:0 ai17:0 i17:0 17:0 18:0 20:0 22:0 23:0 24:0 SAFA Sum FA* 14:1 15:1 16:19 16:17 16:16 17:1 18:19 18:17 18:16 20:19 22:1 MUFA 16:2 16:3 18:26 18:36 18:33 18:4 20:26 20:37 20:46 20:53 22:63 PUFA 3 6 3/6
n.d. n.d. n.d. n.d. 1.93 0.73 0.21 1.48 0.38 39.16 0.44 0.29 1.08 8.77 0.81 0.45 n.d. 0.29 56.02 4.61 0.05 0.05 1.81 11.24 3.11 0.25 10.52 5.33 n.d. 0.12 0.04 32.52 0.77 0.73 4.23 0.55 1.33 n.d. 0.12 0.20 0.66 2.71 0.14 11.44 4.18 8.67 0.48
0.09 n.d. 0.05 0.09 3.44 0.99 0.40 1.84 n.d. 38.31 0.42 0.21 0.90 6.60 0.52 0.25 0.04 0.15 54.30 4.99 0.13 0.10 0.93 12.87 1.12 0.31 11.13 4.28 n.d. 0.09 0.08 31.04 0.83 0.48 5.65 0.46 1.60 1.05 0.10 0.11 0.66 3.54 0.17 14.65 5.31 7.99 0.66
n.d. n.d. n.d. n.d. 0.90 0.42 0.19 1.00 n.d. 28.22 0.45 0.37 1.46 10.95 1.58 1.18 0.25 0.82 47.79 3.89 0.05 0.07 0.67 9.04 0.95 0.42 16.93 5.92 n.d. 0.14 n.d. 34.19 0.65 0.51 10.30 0.50 0.71 n.d. 0.16 0.26 0.71 3.91 0.32 18.03 4.94 12.62 0.39
2.37 n.d. 0.49 0.64 9.14 1.96 1.10 3.78 0.55 41.12 1.05 0.36 2.01 17.69 5.49 4.23 n.d. n.d. 91.98 14.31 n.d. n.d. 0.17 2.64 1.10 n.d. 4.12 n.d. n.d. n.d. n.d. 8.03 n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 1.10 n.c.
0.34 n.d. n.d. 0.21 3.92 1.11 0.36 1.58 0.40 35.15 1.36 0.32 2.65 14.20 2.77 1.23 n.d. 0.37 65.97 12.25 n.d. n.d. 0.96 8.02 2.03 n.d. 13.31 6.18 n.d. n.d. n.d. 30.5 n.d. n.d. 3.54 n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 3.54 n.d. 5.57 n.c.
n.d. n.d. 0.02 0.05 1.24 0.47 0.25 1.13 0.41 27.64 0.93 0.44 2.59 15.00 1.47 0.74 n.d. 0.49 52.87 6.29 0.06 0.07 1.25 5.87 0.75 0.18 20.59 6.53 0.63 0.40 n.d. 36.33 n.d. n.d. 5.74 0.17 n.d. n.d. 0.61 n.d. 0.93 3.01 0.34 10.8 3.35 8.83 0.38
0.10 n.d. 0.06 0.14 3.57 1.42 0.54 2.26 0.41 38.55 0.48 0.35 1.29 9.56 0.81 0.33 n.d. 0.19 60.06 7.05 0.15 0.12 3.43 2.11 7.88 0.34 9.76 5.20 n.d. n.d. n.d. 28.99 0.70 0.42 5.95 0.43 1.65 n.d. n.d. n.d. 0.36 1.43 n.d. 10.94 3.08 14.62 0.21
0.33 n.d. 0.25 0.26 7.22 2.00 0.94 4.25 0.73 44.20 0.92 0.89 1.68 19.65 4.45 0.99 n.d. n.d. 88.76 12.25 n.d. n.d. n.d. 1.81 3.15 n.d. 0.80 5.06 n.d. n.d. n.d. 10.82 n.d. n.d. 0.42 n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 0.42 n.d. 3.57 n.c.
0.34 n.d. 0.30 0.26 8.51 1.68 0.74 3.18 0.69 49.32 0.90 0.43 1.50 9.05 0.84 0.14 n.d. n.d. 77.88 10.02 n.d. n.d. 1.35 2.96 4.24 n.d. 3.29 4.57 n.d. n.d. n.d. 16.41 0.42 1.45 1.49 n.d. 2.28 n.d. n.d. n.d. n.d. n.d. n.d. 5.64 2.28 5.73 0.40
0.56 0.07 0.11 0.28 6.85 1.99 0.47 1.90 0.53 45.42 1.28 0.45 3.12 8.79 1.17 0.29 n.d. n.d. 73.28 10.76 0.18 n.d. 1.71 6.74 3.51 0.15 7.70 7.07 n.d. 0.16 n.d. 27.22 n.d. n.d. 2.04 n.d. n.d. n.d. 0.07 n.d. 0.22 0.20 n.d. 2.53 0.20 5.84 0.03
0.65 0.10 0.20 0.36 7.25 2.44 0.60 2.32 0.62 42.93 1.42 0.44 2.81 6.50 0.85 0.21 n.d. 0.06 69.76 11.96 0.16 0.19 1.48 5.26 3.49 0.22 5.66 9.97 n.d. 0.19 n.d. 26.62 n.d. n.d. 1.64 n.d. n.d. n.d. 0.10 n.d. 0.58 1.19 0.10 3.52 1.29 5.35 0.24
0.09 n.d. n.d. 0.19 2.89 1.38 0.47 1.85 0.45 40.97 1.56 0.66 3.44 13.18 1.21 0.66 n.d. 0.20 69.20 10.09 0.05 n.d. 1.72 3.77 4.18 0.18 7.11 10.42 n.d. 0.09 n.d. 27.52 n.d. n.d. 1.49 n.d. n.d. n.d. n.d. n.d. 0.52 1.28 n.d. 3.29 1.28 6.19 0.19
n.d. – not detected, n.c. – not calculated, * sum of odd and branched FAs.
Sediments
163
Figure 1. Dendrogram of the cluster analysis of fatty acid composition (% of total FA) of fatty acids in Gammarus body (g1–g3), intestinal tract (i1–i3), seston (s1–s3) and bottom sediments (b1–b3, all numbers in chronological order, as in Table 1), from littoral of the Bugach reservoir, 2000 (B) and the Shira Lake, 1998 (S). 47-dimensional hyper-space; the axe represents Euclidean distances.
were used in the joint cluster analysis (Figure 1). Gammarus bodies and bottom sediments from the salt lake formed separate clusters, while intestinal tract contents and seston were in joint cluster with the freshwater samples. Hence, total FA compositions of the crustaceans from the freshwater reservoir and the salt lake differed significantly, while compositions of their main food (seston) were comparatively similar. In the Bugach reservoir sediments at the pebbly bottom were likely represented by freshly settled seston. To detect particular fatty acid species, whose levels differed significantly in Gammarus bodies from the Bugach and the Shira, fatty acid species were
compared using Student’s t-test. These acids are given in Table 4. For comparison, levels of the same FA species in seston, bottom sediments and intestinal tract are given, but all the differences were non-significant.
Discussion Our study of FA composition was carried out during a short period and evidently did not consider seasonal variations, which are known to be significant. Thereby, their comparison with data from other water
164 Table 3. Levels of fatty acids (% of total) in bodies of Gammarus lacustris, intestinal tract levels, seston and bottom sediments from the littoral of the Shira Lake, sampled in August 1998. Fatty acid
Gammarus body
Intestinal tract
Seston
Sediments
Date August
4
5
8
5
7
8
5
7
8
2
5
8
12:0 13:0 i14:0 14:0 ai15:0 i15:0 15:0 i16:0 16:0 ai17:0 i17:0 br17:0 17:0 18:0 20:0 21:0 22:0 23:0 24:0 SAFA Sum FA* 16:19 16:17 16:16 18:19** 18:17 18:16 20:19 20:17 MUFA 16:2 16:3 18:26 18:36 20:26 20:37 20:33 20:46 20:53 22:63 PUFA 3 6 3/6
n.d. n.d. n.d. 2.02 n.d. n.d. 0.78 n.d. 19.89 n.d. n.d. n.d. n.d. 5.26 0.32 n.d. 0.44 n.d. n.d. 28.71 0.78 0.31 6.41 n.d. 25.29 4.09 n.d. 1.33 0.88 38.31 n.d. 0.44 2.33 0.65 1.01 0.16 0.41 5.53 14.03 7.53 32.09 21.97 9.52 2.31
n.d. n.d. n.d. 1.07 n.d. n.d. n.d. n.d. 15.60 n.d. n.d. n.d. n.d. 5.12 0.41 n.d. 0.66 n.d. n.d. 22.86 n.d 0.19 3.90 n.d. 27.26 4.28 n.d. 0.95 2.56 39.14 n.d. 0.22 6.55 0.59 0.36 0.36 n.d. 5.30 14.90 8.96 37.24 23.86 12.80 1.86
n.d. n.d. n.d. 0.89 n.d. n.d. 0.66 n.d. 17.20 n.d. n.d. n.d. n.d. 4.51 0.38 n.d. 0.46 n.d. n.d. 24.10 0.66 0.34 3.23 n.d. 23.90 4.18 n.d. 1.95 0.19 33.79 0.44 0.73 7.07 0.56 1.06 0.29 0.24 5.92 15.74 9.23 41.28 25.21 14.61 1.73
0.31 0.34 n.d. 5.43 0.49 0.67 3.28 n.d. 38.54 n.d. 0.51 n.d. 2.53 16.51 1.62 0.34 1.38 n.d. 0.72 72.67 8.13 2.40 4.36 n.d. 14.27 1.83 n.d. n.d. n.d. 23.29 n.d. n.d. 2.00 n.d. n.d. n.d. n.d. 0.30 2.18 n.d. 4.48 2.18 2.30 0.95
0.09 0.23 n.d. 3.60 0.46 0.40 2.67 n.d. 34.51 n.d. 0.67 n.d. 1.67 16.63 1.68 0.44 0.93 n.d. 1.30 65.28 6.19 2.00 4.86 n.d. 17.52 1.97 n.d. n.d. n.d. 26.35 n.d. n.d. 2.59 n.d. n.d. n.d. n.d. 0.90 4.20 n.d. 7.69 4.20 3.49 1.20
0.13 0.10 n.d. 3.55 n.d. n.d. 1.80 n.d. 37.51 n.d. 0.68 n.d. 1.89 19.33 2.41 0.33 0.95 0.51 0.71 69.90 4.6 1.35 0.55 n.d. 18.99 4.79 n.d. n.d. n.d. 25.68 n.d. n.d. 3.57 n.d. n.d. n.d. n.d. 0.85 n.d. n.d. 4.42 n.d. 4.42 n.c.
0.51 0.32 0.40 5.76 0.92 0.96 3.28 n.d. 42.92 n.d. n.d. n.d. 2.01 19.19 2.18 n.d. 1.46 n.d. n.d. 79.91 8.4 n.d. 3.46 n.d. 10.43 4.54 n.d. n.d. n.d. 18.30 n.d. n.d. 0.98 n.d. n.d. n.d. n.d. n.d. n.d. n.d. 0.98 n.d. 0.98 n.c.
0.63 0.55 0.38 3.43 0.86 1.02 4.25 n.d. 40.13 n.d. n.d. n.d. 3.12 16.22 2.18 0.89 1.23 n.d. 1.08 75.97 10.81 n.d. 7.75 n.d. 11.93 1.25 n.d. n.d. n.d. 20.93 n.d. n.d. 1.42 n.d. n.d. n.d. n.d. n.d. n.d. n.d. 1.42 n.d. 1.42 n.c.
0.86 0.52 0.49 1.09 0.91 1.18 4.50 0.49 39.59 n.d. 0.40 n.d. 5.21 18.10 1.84 n.d. 1.21 n.d. 1.08 77.47 14.56 n.d. 7.68 n.d. 12.19 1.30 n.d. n.d. n.d. 21.17 n.d. n.d. 1.77 n.d. n.d. n.d. n.d. n.d. n.d. n.d. 1.77 n.d. 1.77 n.c.
0.69 n.d. 0.64 4.66 1.80 2.37 2.64 0.45 20.60 0.52 0.55 0.67 0.80 4.36 2.42 0.30 4.22 1.04 6.65 55.38 11.13 2.07 23.09 n.d. 6.51 6.95 0.21 n.d. n.d. 38.83 n.d. 0.45 1.89 n.d. n.d. n.d. n.d. 1.42 3.12 n.d. 6.88 3.12 3.61 0.86
0.92 n.d. 0.34 1.41 1.68 2.13 0.85 0.87 13.78 1.24 0.75 0.81 0.60 4.98 2.45 0.34 4.59 1.68 2.55 41.97 11.43 6.38 9.35 n.d. 14.18 8.67 n.d. 1.07 n.d. 39.65 n.d. n.d. 6.96 0.38 n.d. 0.33 n.d. 1.09 2.55 n.d. 11.31 2.55 8.43 0.30
1.96 n.d. 0.64 4.00 2.65 3.83 1.20 1.27 17.87 1.14 0.88 0.75 0.75 4.15 1.60 0.30 2.47 0.84 4.73 51.03 15.07 8.01 11.65 n.d. 11.38 8.52 1.20 0.74 n.d. 41.50 n.d. n.d. 5.18 0.19 n.d. n.d. n.d. 0.41 0.96 n.d. 6.74 0.96 6.98 0.14
n.d. – not detected, n.c. – not calculated, * sum of odd and branched FAs, ** Sum of 18:19 and 18:33, 18:33 < 1%.
165 Table 4. Average levels (M,%) of dominant and essential fatty acids, their standard errors (SE) and values of Student’s t-test for bodies of Gammarus lacustris intestinal tract levels, seston and bottom sediments from the Bugach reservoir, 2000 and Shira Lake, 1998. Each mean is based on 3 samples.
Fatty acid
14:0 15:0 16:2 16:17 16:0 17:0 18:26 18:19 18:17 18:0 20:46 20:53 20:26 20:19 22:63
Gammarus body
Seston
Bugach
Shira
Bugach
Shira
M ± SE
M ± SE
M ± SE
M ± SE
2.1 ± 0.74 1.4 ± 0.24 0.8 ± 0.05 11.1 ± 1.11 35.2 ± 3.51 1.2 ± 0.16 6.7 ± 1.83 12.9 ± 2.04 5.2 ± 0.48 8.8 ± 1.25 0.7 ± 0.02 3.4 ± 0.36 0.1 ± 0.02 0.1 ± 0.01 0.2 ± 0.05
1.3 ± 0.35 0.5 ± 0.24 0.2 ± 0.15 4.5 ± 0.97 17.6 ± 1.25 0.8 ± 0.04 5.3 ± 1.50 25.5 ± 0.97 4.2 ± 0.06 5.0 ± 0.23 5.6 ± 0.18 14.9 ± 0.50 0.8 ± 0.22 1.4 ± 0.29 8.6 ± 0.53
3.4 ± 1.35 4.0 ± 0.37 n.d.
3.0*
6.4 ± 1.48 3.2 ± 0.57 0.4 ± 0.20 2.3 ± 0.34 44.0 ± 3.11 1.5 ± 0.11 2.6 ± 1.69 4.6 ± 2.67 4.9 ± 0.19 12.8 ± 3.45 0.1 ± 0.12 0.5 ± 0.48 n.d.
4.5* 15.8*
t 0.9 2.8 3.9* 4.4* 4.7* 2.0 0.6 5.6* 2.1 3.0* 27.1* 18.9*
Sediments
6.3 ± 1.42 40.9 ± 1.03 3.5 ± 0.94 1.4 ± 0.23 11.5 ± 0.55 2.4 ± 1.09 17.8 ± 0.87 n.d.
Intestinal tract
Bugach
Shira
t
M ± SE
M ± SE
1.5
5.7 ± 1.39 2.0 ± 0.15 n.d.
3.4 ± 0.99 1.6 ± 0.55 n.d.
5.3 ± 0.86 43.1 ± 1.29 3.1 ± 0.17 1.7 ± 0.16 6.8 ± 0.61 9.2 ± 1.05 9.5 ± 1.96 0.4 ± 0.11 0.9 ± 0.35 0.1 ± 0.03 0.2 ± 0.03 0.03 ± 0.03
14.7 ± 4.25 17.4 ± 1.98 0.7 ± 0.06 4.7 ± 1.49 10.7 ± 2.24 8.1 ± 0.55 4.5 ± 0.25 1.0 ± 0.3 2.2 ± 0.65 n.d.
1.1 – 2.7 1.0 2.1 0.7 2.5 2.3 1.4 –
n.d.
–
n.d.
–
n.d.
n.d.
–
n.d.
n.d.
–
0.6 ± 0.32 n.d.
t 1.4 0.8 – 2.2 10.9* 13.2* 2.0 1.7 0.9 2.5 1.7 1.8 – 1.4 –
Bugach
Shira
M ± SE
M ± SE
t
4.8 ± 2.32 2.2 ± 0.82 n.d.
4.2 ± 0.62 2.6 ± 0.43 n.d.
0.2
5.5 ± 1.56 34.6 ± 3.91 2.42 ± 0.20 3.1 ± 1.67 12.6 ± 4.74 4.2 ± 2.12 15.6 ± 1.06 0.31 ± 0.31 1.0 ± 1.00 0.2 ± 0.20 0.13 ± 0.13 0.11 ± 0.11
3.3 ± 1.36 36.9 ± 1.20 2.0 ± 0.26 2.7 ± 0.46 16.9 ± 1.39 2.9 ± 0.96 17.5 ± 0.92 0.7 ± 0.19 2.1 ± 1.21 n.d.
0.5 – 1.1 0.6 1.2 0.2 0.9 0.6 1.3 1.0 0.7 –
n.d.
–
n.d.
–
* significant difference, n.d. – not detected.
bodies and populations is limited. Nevertheless, we can compare our data with those collected in the same period, the beginning of August 1998, at the same water temperature of 20–22 °C, in saline Siberian lake Shira (Gladyshev et al. 2000). Hence, our attempt to compare the short-period data on the two populations seems to be valid. FA composition of the freshwater and saltwater G. lacustris differed significantly (Table 4). Levels of PUFA, namely EPA and DHA, in bodies of the freshwater Gammarus were several times lower than those in the saltwater population. These differences may be
the result of several causes. Firstly, our finding is in good agreement with the point of view that salt water animals have significantly higher levels of EPA and DHA than freshwater species (e.g., Steffens (1997)). For example, marine and salt lake crustacea have EPA level about 10–30% of total fatty acids (Kattner et al. 1989; Napolitano and Ackman 1989; Nanton and Castell 1998), while this value for freshwater crustacea is lower than 5% (Desvilettes et al. 1997; Gladyshev et al. 1999). Secondly, perhaps Gammarus from Bugach reservoir are fattier than Gammarus from Shira lake and contains a lot of TAG, which is char-
166 acterized by a high level of SUFA. However, our results are not conclusive. It is necessary to continue our investigation. It should be emphasised that we consider only crustacea. Other arthropods could have a different FA composition. For instance, freshwater aquatic larvae of insects have comparatively high levels of EPA, up to 18%, probably because they overwinter as larval stages (Goedkoop et al. 2000). According to our data (Gladyshev et al. 1999), larvae of Chironomus plumosus had an EPA level four times higher than that of Gammarus. Nevertheless, it must be mentioned, that in contrast to literature data, cited above, and our findings, Arts et al. (2001) reported very high contents of EPA and DHA in bodies of freshwater G. lacustris. The percentage of some acids, listed in Table 4, such as 16:0, 16:1, in bodies of the freshwater Gammarus were on average significantly higher than those of the saltwater population. They were also higher than levels of those acids reported for other marine and saltwater crustaceans, including amphipods (Clarke et al. 1985; Kattner et al. 1989; Napolitano and Ackman 1989; Coutteau and Mourente 1997; Nanton and Castell 1998). Thereby, a comparatively high level of 16:1 and 16:0 of freshwater crustacea might be regarded as a general tendency, together with the well known differences in 20:53, 22:63 and 20:46 level, confirmed in our study. In contrast to the finding for C16 FAs, there was no such general tendency for C18 acids. The comparison of C18 levels in marine crustaceans reported by authors referred to above, gave no explicit pattern: the levels were both higher and lower compared to those of the freshwater species. Meanwhile, levels of oleic acid in the Artemia nauplii (Coutteau and Mourente 1997) was equal to that of the saltwater Gammarus and thereby higher than its level in the freshwater Gammarus. As mentioned in the Introduction, these differences in lipid composition, reported in the literature, cited above and revealed in our study, may be the result of several causes. According to our data, in contrast to the bodies compositions, FA composition of seston and bottom sediments in the freshwater reservoir and the salt lake were comparatively similar: there were no significant differences in FA levels (Table 4). Thus, the differences in Gammarus FA composition may not be attributed to seston and sediments food composition. It is interesting to remark that together with the differences in lipid composition of marine and fresh
water multicellular animals, there are some data concerning similarity of fatty acid spectra of some marine and freshwater phytoplankton species, e.g., diatoms (Erwin 1973). These data support our finding on seston. On the basis of the present study the question arises as to how did the two populations of Gammarus maintain the levels of essential PUFAs, namely 20:53? According to our data (Gladyshev et al. 2000), G. lacustris from the salt lake Shira selectively consumed some particles, contained EPA, from bottom sediments. The selective consumption of PUFArich bottom sediment particles by zoobenthos was also reported by Goedkoop et al. (2000). The populations from the freshwater Bugach may support the necessary level of EPA, which was comparatively low, by consuming 18:33 from seston and converting it in 20:53 by relevant desaturases and elongases. In any case, these two population of the same biological species evidently had different levels of the essential PUFAs in August. Causes of these differences between salt- and freshwater species and their stability during all the growing season are unclear at present, but levels of PUFAs in food sources of these two populations were found to be similar.
Conclusions The freshwater G. lacustris had high levels of 16:1 and 16:0 acids compared to literature data on saltwater crustaceans. Taking into account the relevant literature data, we hypothesise that this difference might be a general tendency of FA levels of freshwater and saltwater crustaceans, together with the well known differences in 20:53, 22:63 and 20:46 levels, confirmed in our study. On the basis of comparison of FA levels in intestinal tracts, seston and bottom sediments, it could be concluded that seston was the main food source for the Gammarus but some FAs were obtained from sediments.
Acknowledgements We used the GS-MS of Joint Equipment Unit of Krasnoyarsk Scientific Centre of Siberian Branch of Russian Academy of Sciences. We are grateful to Dr Vadim E. Panov who confirmed the Gammarus lacus-
167 tris identification and to Dr G. Ahlgren for helpful comments. The work was supported by grant No. REC-002 of the US Civilian Research & Development Foundation for the Independent States of the Former Soviet Union (CRDF) and Ministry of Education of Russian Federation.
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