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The relationship between benthic macroinvertebrates and environmental variables from 42 Italian lakes were analysed with the aim of developing a biotic index. In the investigated lakes, 570 species were found of which 373 belonged to the family of Chironomidae and 85 to the class of oligochaeta. Rare species, those present in less than 10 samples, were excluded from the analysis, leaving 57 species for data analysis. Multivariate analysis: canonical correlation analysis (CANON) and multivariate analysis of variance and covariance (MANCOVA) were carried out on a large database of 1060 sampling points, for which both environmental data (16 chemical and morphometric variables) and 57 species counts from soft bottom samples were available. In addition, a second dataset of 94 sites from small lakes sampled in 2005 in Northern Italy (Lombardy), was analysed for comparison. The data analysis (CANON) from the large database gave the following results: (1) the first canonical variate was related to conductivity, pH and alkalinity and accounted for 17% of the total variation; (2) the second canonical variate was related to total phosphorus, N-NH4 and dissolved oxygen, and accounted for 15% of the total variation; (3) lake maximum depth, volume and water temperature were related to the third canonical variate, which accounted for 14% of the source of variation. The analysis of the database from small lakes revealed that in the plane of the first two canonical variates dissolved oxygen was inversely correlated with the depth of sampling and that the second canonical variate showed an inverse relationship between transparency and nutrients. MANCOVA found evidence for differences between lake types and sampling years, confirmed the CANON results and allowed the quantification of the contribution of different target variables on species composition. A trophic status index was calculated ranking percentage oxygen saturation (O2 %sat), transparency (transp) and total phosphorus (TP). The means of O2 %sat, transp and TP weighted by the species abundances were calculated to have optimum values for each species; a benthic quality index was then obtained by multiplying the weighted means of O2 %sat, transp and TP by species abundances and dividing the product by the total number of specimens present at each station. A significant correlation was observed between trophic status and the benthic quality index. A good agreement was also observed between the indexes calculated using the large database and the second dataset on small lakes.

Changes in enzyme activities, metabolite concentrations, and membrane transport activity underlying the Chironomus riparius larvae adaptive response to anoxia were investigated. Trehalose, malate, and aspartate degradation and alanine accumulation were recorded. During anoxia exposure, there was a boost of antioxidant defenses as shown by an increase of the specific activity of the enzymes catalase, glutathione-S-transferase, glutathione peroxidase, glutathione-synthase, malic enzyme, and NADP-dependent isocitrate dehydrogenase. The ratio, glutathione reduced over glutathione oxidized, decreased. Except for alanine and catalase, the parameters return to their basal value when larvae are transferred to normoxic conditions. To test whether antioxidant defenses had protective effects on membrane functionality, L-leucine uptake into brush border membrane vesicles and membrane lipid peroxidation was measured. No difference between membranes prepared from larvae exposed to anoxia and control larvae was found. The amino acid alanine, when present inside the vesicles, trans-stimulated leucine uptake. This effect could represent a mechanism to stimulate amino acid uptake and catabolism in vivo when free alanine concentration increases during hypoxic periods.

Benthic macroinvertebrates are an essential component of freshwater environments. The ecotoxicological risk of benthic communities, estimated through the assay of biochemical markers, can be used as an early warning signal for environment alterations. In this work the activities of a number of enzymes, regarded as potential biomarker of exposure to pollutants (catalase, acetylcholinesterase, glutathione-S-transferase) was determined in homogenates of whole organisms. Specimens were collected in the rivers Taro and Ticino, northern Italy, in stony bottom reaches in five and three stations respectively. The orders of aquatic Insects Diptera, Plecoptera, Odonata, Ephemeroptera and Trichoptera, the Crustacea Amphipoda, and Annelida Oligochaeta were examined. Additional enzymes, such as NADP-dependent malate dehydrogenase, isocitric dehydrogenase, glucose-6-phosphate dehydrogenase, alcohol dehydrogenase, and lactate dehydrogenase were considered in the study. Results emphasize significant differences among taxa concerning the specific activity of most enzymes.

Leucine uptake into membrane vesicles from larvae of the midge Chironomus riparius was studied. The membrane preparation was highly enriched in typical brush border membrane enzymes and depleted of other membrane contaminants. In the absence of cations, there was a stereospecific uptake of l-leucine, which exhibited saturation kinetics. Parameters were determined both at neutral (Km 33 +/- 5 microM and Vmax 22.6 +/- 6.8 pmol/7s/mg protein) and alkaline (Km 46 +/- 5 microM and Vmax 15.5 +/- 2.5 pmol/7s/mg protein) pH values. At alkaline pH, external sodium increased the affinity for leucine (Km 17 +/- 1 microM) and the maximal uptake rate (Vmax 74.0 +/- 12.5 pmol/7s/mg protein). Stimulation of leucine uptake by external alkaline pH agreed with lumen pH measurements in vivo. Competition experiments indicated that at alkaline pH, the transport system readily accepts most L-amino acids, including branched, unbranched, and alpha-methylated amino acids, histidine and lysine, but has a low affinity for phenylalanine, beta-amino acids, and N-methylated amino acids. At neutral pH, the transport has a decreased affinity for lysine, glycine, and alpha-methylleucine. Taken together, these data are consistent with the presence in midges of two distinct leucine transport systems, which combine characters of the lepidopteran amino acid transport system and of the sodium-dependent system from lower neopterans.