Computational protocol: Heavy-Metal Concentrations in Small Mammals from a Diffusely Polluted Floodplain: Importance of Species- and Location-Specific Characteristics

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Protocol publication

[…] All data were collected at the Afferdensche en Deestsche Waarden (ADW), a moderately to heavily polluted embanked floodplain area along the river Waal, the main distributary of the Rhine in The Netherlands (Fig. ). The research area consists of lands inside and outside the summer dikes. The summer dikes are the lower inner embankments protecting agricultural areas in the floodplain against summer floods. Large parts of the floodplain are periodically flooded, on average once a year, predominantly in winter. The floodplain includes areas with and without agricultural activities. Those without agriculture, feature naturally developed vegetation and offer a wide range of habitats. Detailed descriptions of the research area are given in Wijnhoven et al. (, ). Fig. 1.Small mammals from the ADW floodplain were collected at 58 sites (Fig. ) between 2001 and 2003 using Longworth live traps in lines of 5 to 10 traps at each site. The traps were baited with apple, carrot, and rinsed meat and were stuffed with hay and tissue. Specifically for this study, there were three sessions of two 3-day trapping trials, all traps were checked twice a day in August 2002 and again in June and October 2003. Furthermore, all trapping casualties from monitoring studies were included, especially specimens trapped in winter and spring because mortality in that time of the year is higher (Wijnhoven et al. , ). Trapping locations were originally selected to monitor recolonisation of the floodplain after flood events (Wijnhoven et al. ), so they were chosen based on habitat characteristics (vegetation structure, soil type, and management type) without previous information on the levels of contamination. Therefore, trapping sites covered the whole range from nonflooded parts to flooded locations situated far from the nonflooded areas, and were expected to show a representative variation in contaminant levels for the study area. At each of the sites, three soil cores, prepared from three or five soil samples from a 1 m2 plot, were taken with line intervals of at least 10 m. A 5-g portion of soil from each sample was oven dried for 24 hours at 105°C. The total metal content of 0.2 mg dry weight (DW) substrate in a mixture of 3.0 ml 65% HNO3 and 1.5 ml 37% HCl was measured after microwave destruction using a MLS-1200 MEGA microwave oven (Milestone, Sorisole, Italy). The samples were topped up to 50 ml, after which the metal content was measured using inductively coupled plasma–atomic emission spectrometry (ICP-AES; Spectro Analytical Instruments, Kleve, Germany). The 0.01 M CaCl2-exchangeable fraction was determined as a measure of the potential metal solubility. A 6-g fresh-weight portion of substrate, to which 0.01 M CaCl2 had been added in a 1:10 (m[DW]/v) ratio, was mixed for 2 hours, after which the suspension was centrifuged at 12,000 rpm (5,000 x g) for 15 minutes. After the pHCaCl2 had been measured in the substrate suspension in 0.01 M CaCl2, the supernatant was filtered over a 0.45-μm pore filter. A pH of 2 was obtained by adding a few droplets of 65% HNO3, and the metal content of the sample was subsequently measured on the spectrometer.Fresh weights (FWs) of the mammals were determined, and the liver and kidneys of each specimen were weighed (FW). Parts of these organs and flank muscles were oven dried for 24 hours at 105°C, after which DWs were measured. The metal contents of the animal tissues were measured after microwave extraction of approximately 0.01 to 0.25 mg DW with HNO3 and HCl and analysed using ICP-AES as described previously. The metal concentrations in whole animals were calculated from the concentrations in the liver, kidneys, and muscle tissue. Concentrations in muscle tissue were assumed to reflect the concentration in the animal’s remaining tissues, i.e., the entire animal minus liver and kidneys. Calculations were based on the species-specific weighed average distributions in percentage DW for liver, kidneys, and other tissues (tissue-to-total body ratios) and the tissue-specific DW-to-FW ratios derived from our own data. […]

Pipeline specifications

Software tools MEGA, MUSCLE
Application Nucleotide sequence alignment
Organisms Mus musculus, Homo sapiens, Sorex araneus, Crocidura russula