Intra- and interspecific toxicity data for nematodes exposed to metals
Twenty-four hour median lethal concentration (24hr-LC50) toxicity tests were performed with five species of nematodes (Caenorhabditis elegans, Caenorhabditis briggsae, Pristionchus pacificus, Oscheius tipulae, and Oscheius myriophila) in response to copper chloride and zinc chloride. In addition, lethality tests were also performed with seven strains of C. elegans (N2 >1 year in culture, N2 newly acquired, N2 ancestral, ED3053, JU258, JU1171, and MY1) exposed to copper chloride. Nominal chemical concentrations were validated and analyzed according to U.S. Environmental Protection Agency method 6010 using inductively coupled plasma–atomic emission spectroscopy (ICP-AES). Files with raw toxicity data, LC50 calculations, and chemical concentration validation are all reported here. The results indicate that P. pacificus is the most sensitive species to both metals. Conversely, C. elegans is the least sensitive species to copper, but the second most sensitive to zinc, indicating that species relationships do not necessarily predict species sensitivity. We also found that the four wild strains of C. elegans are more sensitive than standard laboratory N2 strains to copper, especially the N2 strain that was cultured in the lab for >1 year, suggesting that long-term lab strains may be less sensitive to copper than their wild counterparts.
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Five species (Caenorhabditis elegans, Caenorhabditis briggsae, Pristionchus pacificus, Oscheius tipulae, and Oscheius myriophila) and six strains of C. elegans (N2, N2 ancestral, ED3053, JU258, JU1171, and MY1) were obtained from the Caenorhabditis Genetics Center (CGC). An additional N2 strain in culture for >1 year was obtained from the laboratory of Mary B. Kroetz at the University of South Alabama. All species and strains, except O. tipulae, were cultured in petri dishes with a 60 mm growth area at 15°C. These strains were fed once per week by transferring a subset of L4/J4 individuals to fresh plates with OP50 bacteria. O. tipulae was kept on plates with a 90 mm growth area at 15°C and was transferred to new plates weekly using the chunking method to accommodate the larger number of worms needed for transferring. All strains were age-synchronized for testing through bleaching. Range-finding LC50 tests were first conducted with four concentrations, five nematodes per well, and four replicates per concentration. Definitive LC50 tests were then conducted with six concentrations, ten nematodes per well, and four replicates per concentration. Four replicates of a control (containing K-medium only) were included in each test. Dilution series for toxicity tests with CuCl2 and ZnCl2 were made with complete K medium (1 L K medium, 1 mL cholesterol [5 mg/mL], 1 mL 1 M CaCl2, and 1 mL 1 M MgSO4). All species and strains were exposed for 24 hours and counted as alive if moving or dead if they did not move in response to probing with a wire pick. Each strain received one range-finding test and at least two definitive LC50 tests. Concurrent with experiments, separate solutions that bounded low, medium, and high nominal copper and zinc concentrations used in toxicity tests were analytically verified using a commercial laboratory testing service (Pace Analytical Services, St. Rose, Louisiana). Samples were prepared, stabilized through acidification within several hours of preparation, and analyzed according to U.S. Environmental Protection Agency method 6010 using inductively coupled plasma–atomic emission spectroscopy. The reporting limit ranged from 0.05 to 1 mg/L across samples depending on the concentrations tested (i.e., lower reporting limits were used for lower nominal concentrations) and was always at least 1 order of magnitude below the target nominal concentration. All dose-response models and statistical analyses were performed using RStudio. We used a log-logit model to estimate LC50 using the “glm” function of the “MASS” package in R. We used the 95% confidence interval of the combined LC50 for each strain to statistically compare LC50s across strains for both metals.